12132019

1

12132019

2

We are going 2024Artemis ndash Greek Goddess of the Hunt the Moon and VirginityTwin of Apollo

Sources of Radiation Exposure

Man-made vs natural radiation

Man-made

radiation

52

Natural

background

radiation

48

Nuclear medicinex-rays

91

Consumer

products

65

Occupational

1

Fallout

1

Nuclear fuel cycle 05

Mettler 2007

Brooks 2010

12132019

3

Ionizing radiation that

will affect biology

Galactic Cosmic

Radiation (GCR)

Solar Particle Events

(SPEs)

Interplanetary space radiationWhat are we going to encounter beyond LEO

Limits of life in space as

studied to date

125 days on a lunar round trip

15 years in low Earth orbit on ISS

Galactic Cosmic Radiation - GCR

12132019

4

Deep Space Radiation

bull Galactic cosmic rays (GCR)mdashhighly-ionizing radiation (Linear Energy Transfer - LET)

bull Comprised of high-energy protons and heavy ions

bull Solar particle events (SPEs) primarily energetic protons (85) helium (14) and high-energy (E) and high-charge (Z) ions called HZE particles (1)

8

12132019

5

DNA Damage

Credit NASA

12132019

6

water food oxygenfuel avoid solar particle events (SPEs)

11

Deep space missions will require significant countermeasures likely both technological and biomedical to protect the crew from the effects of chronic radiation exposure

Human missions will need

12

Autonomous Biosensors for Deep Space

Organisms that can survive for extended periods with minimal life support technology and must function reliably with intermittent communication with Earth

12132019

7

13

12132019

8

Saccharomyces cerevisiae

Saccharo ndash Sugar

Myces ndash Fungus

Cerevisiae ndash Beer(Sugar fungus that makes beer)

15

bull Winemaking Baking and Brewingbull Eukaryoticbull Highly studied and understoodbull Genome has been sequencedbull Genetics are easily manipulatedbull Very easy to maintain in the lab and in space

BioSentinel Mission

NASA Ames Research Center

Northern Ontario School of Medicine

SNOLAB Sudbury

NASA Ames Research Center

Cubesatb

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

2

We are going 2024Artemis ndash Greek Goddess of the Hunt the Moon and VirginityTwin of Apollo

Sources of Radiation Exposure

Man-made vs natural radiation

Man-made

radiation

52

Natural

background

radiation

48

Nuclear medicinex-rays

91

Consumer

products

65

Occupational

1

Fallout

1

Nuclear fuel cycle 05

Mettler 2007

Brooks 2010

12132019

3

Ionizing radiation that

will affect biology

Galactic Cosmic

Radiation (GCR)

Solar Particle Events

(SPEs)

Interplanetary space radiationWhat are we going to encounter beyond LEO

Limits of life in space as

studied to date

125 days on a lunar round trip

15 years in low Earth orbit on ISS

Galactic Cosmic Radiation - GCR

12132019

4

Deep Space Radiation

bull Galactic cosmic rays (GCR)mdashhighly-ionizing radiation (Linear Energy Transfer - LET)

bull Comprised of high-energy protons and heavy ions

bull Solar particle events (SPEs) primarily energetic protons (85) helium (14) and high-energy (E) and high-charge (Z) ions called HZE particles (1)

8

12132019

5

DNA Damage

Credit NASA

12132019

6

water food oxygenfuel avoid solar particle events (SPEs)

11

Deep space missions will require significant countermeasures likely both technological and biomedical to protect the crew from the effects of chronic radiation exposure

Human missions will need

12

Autonomous Biosensors for Deep Space

Organisms that can survive for extended periods with minimal life support technology and must function reliably with intermittent communication with Earth

12132019

7

13

12132019

8

Saccharomyces cerevisiae

Saccharo ndash Sugar

Myces ndash Fungus

Cerevisiae ndash Beer(Sugar fungus that makes beer)

15

bull Winemaking Baking and Brewingbull Eukaryoticbull Highly studied and understoodbull Genome has been sequencedbull Genetics are easily manipulatedbull Very easy to maintain in the lab and in space

BioSentinel Mission

NASA Ames Research Center

Northern Ontario School of Medicine

SNOLAB Sudbury

NASA Ames Research Center

Cubesatb

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

3

Ionizing radiation that

will affect biology

Galactic Cosmic

Radiation (GCR)

Solar Particle Events

(SPEs)

Interplanetary space radiationWhat are we going to encounter beyond LEO

Limits of life in space as

studied to date

125 days on a lunar round trip

15 years in low Earth orbit on ISS

Galactic Cosmic Radiation - GCR

12132019

4

Deep Space Radiation

bull Galactic cosmic rays (GCR)mdashhighly-ionizing radiation (Linear Energy Transfer - LET)

bull Comprised of high-energy protons and heavy ions

bull Solar particle events (SPEs) primarily energetic protons (85) helium (14) and high-energy (E) and high-charge (Z) ions called HZE particles (1)

8

12132019

5

DNA Damage

Credit NASA

12132019

6

water food oxygenfuel avoid solar particle events (SPEs)

11

Deep space missions will require significant countermeasures likely both technological and biomedical to protect the crew from the effects of chronic radiation exposure

Human missions will need

12

Autonomous Biosensors for Deep Space

Organisms that can survive for extended periods with minimal life support technology and must function reliably with intermittent communication with Earth

12132019

7

13

12132019

8

Saccharomyces cerevisiae

Saccharo ndash Sugar

Myces ndash Fungus

Cerevisiae ndash Beer(Sugar fungus that makes beer)

15

bull Winemaking Baking and Brewingbull Eukaryoticbull Highly studied and understoodbull Genome has been sequencedbull Genetics are easily manipulatedbull Very easy to maintain in the lab and in space

BioSentinel Mission

NASA Ames Research Center

Northern Ontario School of Medicine

SNOLAB Sudbury

NASA Ames Research Center

Cubesatb

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

4

Deep Space Radiation

bull Galactic cosmic rays (GCR)mdashhighly-ionizing radiation (Linear Energy Transfer - LET)

bull Comprised of high-energy protons and heavy ions

bull Solar particle events (SPEs) primarily energetic protons (85) helium (14) and high-energy (E) and high-charge (Z) ions called HZE particles (1)

8

12132019

5

DNA Damage

Credit NASA

12132019

6

water food oxygenfuel avoid solar particle events (SPEs)

11

Deep space missions will require significant countermeasures likely both technological and biomedical to protect the crew from the effects of chronic radiation exposure

Human missions will need

12

Autonomous Biosensors for Deep Space

Organisms that can survive for extended periods with minimal life support technology and must function reliably with intermittent communication with Earth

12132019

7

13

12132019

8

Saccharomyces cerevisiae

Saccharo ndash Sugar

Myces ndash Fungus

Cerevisiae ndash Beer(Sugar fungus that makes beer)

15

bull Winemaking Baking and Brewingbull Eukaryoticbull Highly studied and understoodbull Genome has been sequencedbull Genetics are easily manipulatedbull Very easy to maintain in the lab and in space

BioSentinel Mission

NASA Ames Research Center

Northern Ontario School of Medicine

SNOLAB Sudbury

NASA Ames Research Center

Cubesatb

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

5

DNA Damage

Credit NASA

12132019

6

water food oxygenfuel avoid solar particle events (SPEs)

11

Deep space missions will require significant countermeasures likely both technological and biomedical to protect the crew from the effects of chronic radiation exposure

Human missions will need

12

Autonomous Biosensors for Deep Space

Organisms that can survive for extended periods with minimal life support technology and must function reliably with intermittent communication with Earth

12132019

7

13

12132019

8

Saccharomyces cerevisiae

Saccharo ndash Sugar

Myces ndash Fungus

Cerevisiae ndash Beer(Sugar fungus that makes beer)

15

bull Winemaking Baking and Brewingbull Eukaryoticbull Highly studied and understoodbull Genome has been sequencedbull Genetics are easily manipulatedbull Very easy to maintain in the lab and in space

BioSentinel Mission

NASA Ames Research Center

Northern Ontario School of Medicine

SNOLAB Sudbury

NASA Ames Research Center

Cubesatb

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

6

water food oxygenfuel avoid solar particle events (SPEs)

11

Deep space missions will require significant countermeasures likely both technological and biomedical to protect the crew from the effects of chronic radiation exposure

Human missions will need

12

Autonomous Biosensors for Deep Space

Organisms that can survive for extended periods with minimal life support technology and must function reliably with intermittent communication with Earth

12132019

7

13

12132019

8

Saccharomyces cerevisiae

Saccharo ndash Sugar

Myces ndash Fungus

Cerevisiae ndash Beer(Sugar fungus that makes beer)

15

bull Winemaking Baking and Brewingbull Eukaryoticbull Highly studied and understoodbull Genome has been sequencedbull Genetics are easily manipulatedbull Very easy to maintain in the lab and in space

BioSentinel Mission

NASA Ames Research Center

Northern Ontario School of Medicine

SNOLAB Sudbury

NASA Ames Research Center

Cubesatb

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

7

13

12132019

8

Saccharomyces cerevisiae

Saccharo ndash Sugar

Myces ndash Fungus

Cerevisiae ndash Beer(Sugar fungus that makes beer)

15

bull Winemaking Baking and Brewingbull Eukaryoticbull Highly studied and understoodbull Genome has been sequencedbull Genetics are easily manipulatedbull Very easy to maintain in the lab and in space

BioSentinel Mission

NASA Ames Research Center

Northern Ontario School of Medicine

SNOLAB Sudbury

NASA Ames Research Center

Cubesatb

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

8

Saccharomyces cerevisiae

Saccharo ndash Sugar

Myces ndash Fungus

Cerevisiae ndash Beer(Sugar fungus that makes beer)

15

bull Winemaking Baking and Brewingbull Eukaryoticbull Highly studied and understoodbull Genome has been sequencedbull Genetics are easily manipulatedbull Very easy to maintain in the lab and in space

BioSentinel Mission

NASA Ames Research Center

Northern Ontario School of Medicine

SNOLAB Sudbury

NASA Ames Research Center

Cubesatb

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

9

NASA Ames Collaboration

bull Collaboration with NASArsquos BioSentinelresearch group

bull Understand effects of ionizing radiation in deep space (high dose) as well as deep underground in the absence of background radiation (low dose)

17

S cerevisiae (budding

yeast)

What is BioSentinel

Ionizing radiation

DNA double strand breaks

bull Yeast cells are similar to human cells including in terms of DNA damage and repair

Yeast cells will act as biosensors to measure

the effects of deep space radiation

bull Deep space radiation environment cannot be precisely mimicked on Earth

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

10

Launch

Main objective develop a tool with autonomous life support technologies to

study the biological effects of the space radiation environment at different orbits

bull First biological study beyond low Earth orbit (LEO) in almost 50 years

bull First biological 6U CubeSat to fly beyond LEO

bull Far beyond the protection of Earthrsquos magnetosphere

bull Will compare the biological effects of different radiation environments

(EM-1 ISShellip)

BioSentinel mission

Launch

Secondary

payload

deployment (L+4-

5 hrs)

Lunar

transit

(3-7 days)Lunar Transfer

amp Fly-by

BioSentinel

escapes into a

heliocentric orbit

Distance to ISS ~ 350 km

Distance to the Moon ~385000 km

Distance at 6 months ~40000000 km

Launch of Exploration Mission - 1

Space Launch System (SLS) rocket

Methodology

Yeast Strainsbull Saccharomyces cerevisiae Wild Type (YBS71-1)

W303 (NASA Yeast)

bull Saccharomyces cerevisiae RAD51 Δ (YBS72-1)

W303 (NASA Yeast)

ndash RADiation sensitive

ndash RAD51 assists in repair of double stranded DNA breaks

20

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

11

NASA BioSentinel

bull Genetically engineered Saccharomyces cerevisiae

bull Goal is to study DNA damage and repair mechanisms in the deep space radiation environment beyond LEO

bull Simplistic colorimetric endpoint quantifies changes in metabolism due to accumulated DNA damage

22

Long-term survival ndash 32 weeks RT then 64 weeks 4oC at 0 RH

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

12

23

Models of risk

LNT

Epidemiology

Hormesis

Threshold

Natural

Background

RadiationSub-background

Bio

log

ical R

isk

Radiation Dose

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

13

Chemico-Biological InteractionsVolume 301 March 2019

Tharmalingam S Sreetharan S Brooks

AL Boreham DR Re-evaluation of the

Linear No-threshold (LNT) Model Using

New Paradigms and Modern Molecular

Studies 54-67

Zarnke AM Tharmalingam S Boreham

DR Brooks AL BEIR VI Radon The Rest

of the Story 81-87

Ricci PF Tharmalingam S Ionizing

Radiations Epidemiology Does Not

Support the LNT Model128-140

Scott BR Tharmalingam S The LNT

Model for Cancer Induction is Not

Supported by Radiobiological Data 34-53

Hypothesis

Natural background radiation is essential

for life and helps to maintain the stability of

our genome

Prolonged exposure to a sub-background

environment will be detrimental to living

systems

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

14

SNOLAB

2 km

6800 level

NoriteRock

Creighton Mine

9 Shaft

SNOLAB

2 km underground (6 km

water equivalent)

Class 2000 clean room

50 million times less cosmic

radiation

Air filtration of 50 m3 s-1 for

radon reduction

Radiation ResearchSNOLAB Focus Issue

Volume 188 October 2017

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

15

REPAIR Project

REPAIR Project

Researching the Effects of the Presence

and Absence of Ionizing Radiation

Cell culture

Life Sciences Laboratory2019

Biological Safety Cabinet

Microscope

Cell counter

Water bath

Centrifuge

Tissue culture incubator

UPS

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

16

Advanced Tissue Culture Incubator

31

Background Radiation Characterization

Radon

0 2 4 6 8 10 12 14 16 18 200

5

10

15

75150

Time (days)

Rad

on

co

ncen

trati

on

(B

q m

-3)

Underground (12345 plusmn 1353 Bq m-3)

Glovebox (079 plusmn 093 Bq m-3)

Surface (367 plusmn 214 Bq m-3)

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

17

Methodologybull Determining survival with Biosentinal assay

and colony-forming formation

bull Transcriptomic analysis will be performed at the optimal time point using RNA sequencing (RNA-Seq) with next-generation sequencing (NGS) technology

34

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

18

35

Deep Space Travel

bull Complex environment intrinsic to space travel will likely exacerbate radiation-induced tissue injury

bull Microgravity

bull High CO2

bull 23 - 53 mm Hg vs 03 mm Hg on Earth

bull Confined living space

bull Circadian disruption bull 16 sunrises and sunsetsday

bull Altered physiological processesbull Poorly understood impact on nutritional requirement

36

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

19

Deep Space Travel - Countermeasures

bullBone demineralization

bullCataracts

bullCancer

bullCardiovascular disease

bullCognitive decline

bullEpigenetic Modification

37

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

20

Vineland Estate Riesling

STS 118 Endeavor

Doses to Astronauts2 mSv ndash Apollo 11

2000 mSv - Mars

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

21

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

22

NASA Johnson Complex Dietary Countermeasure

The Dietary Countermeasure

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

23

NASA Johnson Complex Dietary

Countermeasure Collaboration

bull Designed to offset key

processes associated

with elevated ROS

ndash Oxidative stress

ndash Inflammation

ndash Impaired glucose

metabolism

ndash Membrane deterioration

ndash Mitochondrial

dysfunction

Mouse Model

bull Transgenic Growth Hormone Mice (TGM)

bull Model of elevated free radical processesndash Elevated superoxide radical production

ndash Increased lipid peroxidation

bull Model of accelerated agingndash Average lifespan 12 months

ndash Premature onset of senescence

Model for Astronauts

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

24

The Results

bull Increased lifespan by 28 in TGMbull 13 in normal siblings

bull Almost eliminated sarcopenia cataracts arthritis alopeciabull All markers of oxidative damage andor inflammation

bull Increased mobility and activity levels

bull Protected sensory and cognitive functionsbull Normalized ROS and inflammatory processesbull Enhanced mitochondrial function

bull ETC complex activity increased 200 ndash 400

bull Eliminated metastases in P53 cancer-prone micebull Eliminated paralysis in EAE model of MSbull Protected cognition in 3xTg AD model

47

B

Chromosome Aberrationsbull Spectral Karyotyping (SKY) used to examine all

chromosomes for structural aberrations

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

25

Genomic Instability

bull Used Spectral Karyotyping (SKY) to determine type and amount of

chromosome aberrations in TGM compared to age-matched normals

Chromosome Aberrations

Percent Structural Chromosome Aberrations

0

10

20

30

40

50

60

70

Untreated Norm Untreated TGM Suppl Norm Suppl TGM

Experimental Groups

Pe

rce

nt

Str

uc

tura

l A

be

rra

tio

ns

0Gy

2Gy

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

26

Cognition

0

5

10

15

20

25

30

35

40

45

50

UT Normal UT TGM Suppl Normal Suppl TGM

Experimental Groups

o

f T

ria

ls t

o L

earn

Brain Cell Study

bull Old TGM have ~44 of brain cells

remaining

ndash No increase in apoptosis

ndash Most acellular region corresponds with the

nucleus septi medialis

bull one of the main origins of cholinergic fibres

innervating the hippocampus

bull Countermeasure completely abolishes

brain cell loss

bull Responsible for retention of learning

ability

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

27

Brain Cell Density

bull 12 mo Old normal mice do not have significant brain cell loss compared to young mice

bull Untreated old TGM have less than 50 of young number of brain cells

bull Supplemented 12 mo old TGM have brain cell density equivalent to young mice

Brain Cell Number in Old Female Mice

0

20000

40000

60000

80000

100000

120000

140000

UT Norm Suppl Norm UT TGM Suppl TGM

Experimental Groups

Cell

Nu

mb

er

Glucose

Metabolism

in Brain

TGH untreated

TGH diet

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

28

FDG Activity in Brain

0

005

01

015

02

025

03

035

04

Normal TGM

Experimental Group

Ac

tiv

ity

Unsupp Supp

Cranial Irradiation Project

bull 10Gy gamma dose to head only

bull Countermeasure treated micebull Pre-IR fed 30d prior to WBI to end of studybull Post-IR fed 24h post-IR to end of study

bull Animals assessed for cognitive function and olfactory acuity

bull 30d and 120d post-irradiation

bull 10Gy mice took significantly longer to find scent lure

bull Countermeasure (MDS) completely protected olfaction

bull Protection effective whether fed before or after irradiation

56

Time to discover peanut butter smell

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

29

Next Steps

bull Efficacy of Countermeasure in high LET irradiation is being examined

bull Manufacturing of human formulation complete

bull NASA Johnson human clinical trials for efficacy and tolerability starting in 2020

bull Gamma knife surgery on benign brain tumours

bull Human efficacy to see if supplement can offset shortlong-term effects

bull Head-to-head formulation comparison

bull NASA Ames collaborationbull Microgravity study at Ames Research Center

bull ISS studies

57

Acknowledgements

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

30

Thank You

59

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew

12132019

31

Depart Arrive of Flights Avg dose (uSv) Avg Time (hr) Avg Dose Rate (uSvhr)

1) Epidemiologic studies of pilots and aircrew Boice JD Jr Blettner M Auvinen A Health Phys 2000 Nov79(5)576-84

ldquoEpidemiologic studies conducted over the last 20 y provide little consistent evidence linking cancer with radiation exposures from air travelrdquo

2) Cosmic radiation in commercial aviation Bagshaw M Travel Med Infect Dis 2008 May6(3)125-7 Epub 2007 Nov 26

ldquoEpidemiological studies of flight crew have not shown conclusive evidence for any increase in cancer mortality or cancer incidence directly attributable to ionising radiation exposurerdquo

3) Epidemiological studies of cancer in aircrew Hammer GP Blettner M Zeeb H Radiat Prot Dosimetry 2009 Oct136(4)232-9 Epub 2009 Jul 16

ldquoIonising radiation is considered to contribute little if at all to the elevated risks for cancers among aircrew whereas excess ultraviolet radiation is a probable cause of the increased melanoma riskrdquo

Epidemiology of Aircrew