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5/22/2013

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Mechanisms of cardiotoxicity of targeted therapeutics:Focus on kinase Inhibitors

Thomas ForceCenter for Translational Medicine

Cardiology DivisionTemple University School of Medicine

Philadelphia

“STATE-OF-THE-ART PAPER”

Cancer Genetics and the

Cardiotoxicity of the Therapeutics

JACC 2013

Hind Lal

Kyle Kolaja

Thomas Force

Outline

1. Brief intro on kinase inhibitors (KIs) and cancer.

2. Why can they cause cardiotoxicity:On- and off-target toxicity

3. How should it be addressed?

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Dysregulation of tyrosine kinases in cancer: Bcr-Abl, the Philadelphia chromosome, CML and ALL

Bcr

Abl

Bcr

AblPP

BcrAbl

Kinase domain

ATP SubstratesP

Oligomerizationdomain

Kinase domain

Transformation (cancer)

Signaling pathways (Ras/ERK; PI3-K)

Bcr-Abl and imatinib (Gleevec)

Bcr

Abl

Bcr

AblPP

BcrAbl

Kinase domain

ATP SubstratesP

Oligomerizationdomain

Kinase domain

Signaling pathways (Ras/ERK; PI3-K)

imatinib

Transformation (cancer)

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Name Target Company Class FDA Approval

Name Target Company Class

The TKI market: Kinase inhibitor patents: 1988-2005

~ 10,000 compounds currently in development

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Table 1. TKIs and mAbs in Cancer

Agent Class TK target(s) Malignancies Cardiotoxicity/ (Rate)/Other

imatinib (Gleevec)

TKI ABL1/2, PDGFR/, KIT CML, Ph+ B-ALL, CMML, HES, GIST

Y / (low)*

dasatinib (Sprycel)

TKI ABL1/2, PDGFR/, KIT, SRC family CML Y / (low to mod)* / QT prolongation

Nilotinib

(Tasigna)

TKI ABL1/2, PDGFR/, KIT CML Unknown

sunitinib (Sutent)

TKI VEGFR1/2/3, KIT, PDGFR/, RET, CSF-1R, FLT3

RCC, GIST Y / (mod) / hypertension, hypothyroidism

Lapatinib

(Tykerb)

TKI EGFR (ErbB1), HER2 (ErbB2) HER2+ breast cancer N

sorafenib (Nexavar)

TKI c-/B-Raf, VEGFR2/3, PDGFR/, KIT, FLT3

RCC, melanoma Y / (low?)* / ACS / hypertension

gefitinib (Iressa)

TKI EGFR (ErbB1) NSCLC N *

erlotinib (Tarceva)

TKI EGFR (ErbB1) NSCLC, pancreatic cancer, N *

Temsirolimus (Torisel)

novel mTOR (indirect- binds to FKBP12 and complex inhibits mTOR)

RCC N *

trastuzumab (Herceptin)

mAb HER2 (ErbB2) HER2+ breast cancer Y / (

bevacizumab (Avastin)

mAb VEGF-A Colorectal cancer, NSCLC Y / (low to mod)* / arterial thrombosis

cetuximab (Erbitux)

mAb EGFR (ErbB1) Colorectal cancer, squamous cell carcinoma of head/neck

N *

Panitumumab

(Vectibix)

mAb EGFR (ErbB1) Colorectal N *

Rituximab (Rituxan)

mAb CD20 B cell lymphoma Unknown

alemtuzumab (Campath)

mAb CD52 B-cell CLL; Y (in patients with mycosis fungoides/Sezary syndrome)

lestaurtinib TKI JAK2/FLT3 Unknown

pazopanib TKI Multi-targeted RCC Unknown

vandetanib TKI VEGFR/EGFR NSCLC Unknown

cediranib TKI VEGFR NSCLC Unknown

alvocidib TKI CDK CLL Unknown

enzastaurin KI PKC B-cell lymphoma Unknown

mAb, humanized monoclonal antibody; TKI, tyrosine kinase inhibitor; * effect on LV function has not been determined and therefore these represent best.

Adapted from Lal et al. JACC 2012

Tyrosine kinase targets in malignant hematologic disorders

Tyrosine kinase Cancer

ABL CML/ALL/AMLARG AMLALK ALCLFGFR1 aCMLFGFR3 MMFLT3 AML

c-FMS MDS/AMLNTRK3 AMLPDGFR HES/SMPDGFR CMML; AMLJAK2 PCV/ET/IMFc-KIT AML/SMSYK MDS

# Prediction of therapeutic efficacy in some cases is based on pre-clinical studies or invitro data showing inhibition of the tyrosine kinase.

Tyrosine kinase targets in solid tumors

ALK IMTEGFR NSCLC; ovarian;

SCCHN; RCC; C-RHER2 Breast; lung

EGFR3 Clear cell sarcomac-KIT GIST;SCLC;sarcomac-MET SCLC;gastric;

melanoma; renalNTRK1 PTCPDGFR Glioblastoma; GIST

osteosarcoma;RET MEN-2A/BVEGFR-1/2/3 NSCLC; breast;GIST

Renal;C-R; prostate


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Tyrosine kinase targets in solid tumors

ALK IMTEGFR NSCLC; ovarian;

SCCHN; RCC; C-RHER2 Breast; lung

EGFR3 Clear cell sarcomac-KIT GIST;SCLC;sarcomac-MET SCLC;gastric;

melanoma; renalNTRK1 PTCPDGFR Glioblastoma; GIST

osteosarcoma;RET MEN-2A/BVEGFR-1/2/3 NSCLC; breast;GIST

Renal;C-R; prostate


Last count: ~150 kinases proposed as targets in cancer

Is cardiotoxicity inevitable?

Yes

On-target toxicity

Targeting the PI3-Kinase pathway in cancer

RTKs (EGFR, HER2, c-Kit, PDGFRs, Met, etc)

PI3K(P110α)

WortmanninLY294002 PTEN

AKTPDK1 mTORC2

mTORC1

Rapamycin

S6K 4E-BP

Translation &Cell growth

HIF1α

TSC1/2

AMPKLKB

Energy stress

GSK3

Ras-Raf-ERK-RSK

Growth Factors

?

AMP

Cheng and Force, Circ Res: 2010

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The intersection of genetic and chemical genomic screens identifies GSK-3 as a

target in human AML

Banerji et al. JCI 2012

GSK-3 KO and MI: Deletion leads to increased mortality post-MI due to increased rupture

Post MI Survival:GSK3 alpha KO

0 20 40 600

20

40

60

80

100

MI-WT (n=33)

MI-KO (n=33)

Sham-WT (n=8)Sham-KO (n=7)

p=0.0418

Days Post MI

Pe

rce

nt

su

rviv

al

(WT-MI VS KO-MI)

Lal et al. Circulation 2011

Deletion of GSK-3 exaggerates cardiac fibrosis and extracellular matrix remodeling post-MI in the

remote myocardium

P<0.01

P<0.01

N=5 N=5

N=7 N=6

Col

-1/r

RN

A

WT KO

Sha

mM

IF

ibro

sis

(%)

P<0.001

N=3 N=3N=4 N=4

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Off-target toxicity: Selectivity as the key issue

Kothe et al. Biochemistry 2007

ATP

Thus it is relatively easy to make an ATP competitive inhibitor…

ATP

Type I inhibitors

But the high conservation creates a key problem with these agents: lack of selectivity

and “off-target” effects

The kinome:

500+ protein kinases in the kinome

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Imatinib

Dasatinib

Bosutinib

Bantscheff et al. Nat. Biotech. 2007

DDR1

Non-kinase targets also identified including NQO2

Inherent non-selectivity of TKIs- Drugs targeting Abl

Double-Edged Sword of the New Cancer Therapeutics

Comment on Montani et al.

“Pulmonary arterial hypertension in patients treated by dasatinib”

Circulation 2012

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Nilotinib and arterial occlusive disease

Types of TKIs-designing more selective agents

Type I

DFG out (Type II)

Type III

Type I target active conformation, DFG-in: erlotinib, dasatinib, sunitinibType II target inactive conformation, DGF-out: imatinib, sorafenib, vatalinibType III employ binding sites and mechs of regulation that are unique to a specific kinase (PD and UO comounds targeting ERKs.

Is cardiotoxicity unavoidable?

Yes: on-target and off-target effects are inescapable…At least at this point in time

But it is not a class effect and the cardiotoxicity of individual agents can often be understood

at the molecular level

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1. Last count: ~150 kinases proposed as targets in cancerand our understanding of their role in the heart is limited

2. Poor selectivity of the current crop of agents leading to many targets being inhibited, any one of which could be the culprit.

Predicting problematic agents:Key roadblocks

Table 2. Evidence from experimental models suggesting cardiotoxicity of TKIs

by TK target.

Abbreviations: WT, wild type; KO, knockout- gene deleted; D/N, dasatinib, nilotinib; ER, endoplasmic reticulum; CMP, cardiomyopathy; HGF, hepatocyte growth factor (ligand for Met); FGFR fibroblast growth factor receptor; MI, myocardial infarction. See text for other abbreviations.

TK target(s)

TKIs Model Cardiac phenotype of model References

ERBB2 trastuzumab lapatinib

ERBB2 KO: + TAC

Spontaneous dilated CMP; worsened heart failure with pressure load; enhanced anthracycline sensitivity.

26, 27

VEGF VEGFRs

sunitinib sorafenib

bevacizumab

WT: VEGF Trap + TAC

Pathologic remodeling in response to pressure load.

46, 47, 69

KIT imatinib/D/N sunitinib sorafenib

1) W/WV mouse (KIT-deficient) + MI

2) WT: Arterial injury + imatinib

1) Adverse remodeling post MI due to reduced homing of mesenchymal stem cells to sites of injury; 2) Reduced stenosis post arterial injury.

53-55

Raf-1/B-Raf

sorafenib Raf-1 KO and dominant negative

+ TAC

LV dilatation and CHF with pressure load. 59, 60

PDGFRs imatinib/D/N sunitinib sorafenib

WT: MI + Administration

of PDGF

Reduced injury (ischemic protection). 48-50

JAK2 lestaurtinib STAT3 KO: MI; aging;

anthracycline administration;

pregnancy

Increased ischemic injury; reduced capillary density with aging; increased anthracylcine toxicity; peri-partum cardiomyopathy.

64, 65

Abl/Arg imatinib/D/N WT: imatinib Decline in LV function; induction of ER stress. 9, 38

Met (HGF

receptor)

N/A WT: MI or CMP models +

administration of HGF

Reduced fibrosis in MI and CMP models. Neoangiogenesis with HGF.

70 and refs. therein

FGFR1/3 N/A Cell culture models: Administration of

FGF

Enhanced proliferation of cardiomyocytes and cardiac-resident stem cells.


Chen et al. Circulation, 2008

Table 1 Kinase Role of kinase in heart/vasculature Models

used Other notes References

Raf-1/B-Raf Anti-apoptotic; preserves LV fxn in setting of stress. Pheno: 1) LV dysfxn and HF in the absence of additional stress (KO); 2) reduced hypertrophy but LV dysfxn due to cell death (DN-TG)

KO DN-TG

KO: effects mediated by ASK-1; Raf inhibits ASK-1 via a non-kinase-dependent mechanism. Raf mutations account for some cases of Noonan syndrome (HCM phenocopy).

[103]

PI3-K (p110 ) Physiologic heart growth; cardiomyocyte survival

TG (CA/DN);

KI

DN-TG: greater LV dysfunction with TAC. CA-TG improved LV function

[33] [104] [105]

PI3-K (p110 ) Regulates contractility and pathologic hypertrophy

KO KO: protected from isoproternol induced injury

[106, 107] [108]

PDK1 Cardiomyocyte survival/ -arenergic responsiveness

KO Cardiac-specific KO : heart failure and DCM

[109]

Akt Central regulator of cardiomyocyte survival, growth, and metabolism

CA/DN/KO

Akt1 promotes physiologic and suppresses pathologic hypertrophy; Akt2: Pro-survival; insulin sensitivity

[110] [111]

mTOR mTORC1:Central regulator of protein synthesis; inhibition key to energy preservation under stress; mTORC2 regulates Akt activation

KI Rapamycin Rx well-tolerated. Blocks cardiac hypertrophy. Will be used in combination regimens in cancer. Long-term Rx inhibits mTORC2 and Akt.

[112]

AMPK Sensor of energy stress. Inhibits mTORC1, preserving energy stores. KO of 2: increased hypertrophy/LV dysfunction w TAC

TG/KO Activated by tumor suppressor LKB1. Activated mutant leads to glycogen storage hypertrophic myopathy.

[113] [114]

Aurora kinases M-phase regulators KIs KI expected to disrupt 1) CPC proliferation, 2) karyokinesis and 3) any cytokinesis of cardiomyocytes, resulting in cell death. KIs have been assoc w cardiotoxicity.

[120] [121]

Force and Kolaja,Nat Rev Drug Disc:2011

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Inhibition definitely or likely badKinasesVEGFRsPDGFRsRaf-1 / B-RafPI3-K / PDK1 / Akt / Pim / SGK /GSK-3 ERKsLKB1/ CamKK / AMPKCDKsAurora kinasesPLKsHer2c-KitJak2FAKDMPKLTKPKG

Non-kinasesPTENHSPs

Inhibition may be goodROCK1/2PKGCaMKIIGRK2Ask1LTKCDK4/6DMPKPKCPKC

Adapted from: Force and Kolaja: Nat Rev Drug Disc 2011

Kinase targets in cancer

This is not a class effect:Kinase Targets: Unlikely primary role in heart

Target Drugs Evidence for role in heart

EGFR cetux/erlot/etc. Role in transducing Ang II signals

RET sunitinib Hypothyroidism (CHF)

FLT3 sunit/soraf/cep701 Not expressed in c-myo

FMS sorafenib Not expressed

ROS imat/sunit/soraf Not expressed

NTRK1/3 Not expressed

ALK Not expressed

SYK ? Anti-aggregatory (platelets)

Vignette: Sunitinib and the trend toward multi-targeted inhibitors: Folkman

Targets of sunitinib (known): VEGFRsPDGFR /c-KitFLT3 CSF-1RRET

Cancers: *RCC *GIST

BreastNSCLCHepatocellularNeuroendocrineColon

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Change in LVEF from baseline in patients treated with sunitinib

-55%

-50%

-45%

-40%

-35%

-30%

-25%

-20%

-15%

-10%

-5%

0%

5%

10%

15%

20%

Patients (n=36)

LV

EF

Ch

ang

e (

EF

%)

fro

m B

asel

ine

CHF requiring hospitalization, and/or EF drop > 15EF% in 19% of pts.

Chu et al. Lancet:2007

Cardiotoxicity at the ultrastructural level

Kerkela et al. Clin Trans Sci:2009

Control Sunitinib

Mitochondrial damage in sunitinib-treated mice


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Sunitinib-induced hypertension

6%

31%

42% 42%

47%

14%

31%

36%

48%50%

0%

8%11%

17% 17%

0%

10%

20%

30%

40%

50%

60%

0 1 2 3 4

Cycle

% pts with grade I HTN

% pts on antihypertensives

% pts with grade III HTN


Sunitinib-induces apoptosis in vivo, but only in the setting of hypertension


Are the effects on the vasculature critical to the anti-cancer effects of VEGFR/PDGFR-targeted therapies?

Also reported with axitinib

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VEGFR2VEGFR2

VEGF

PDGF

HIF Activation/Stabiliation

Cell SurvivalAngiogenesisProstaglandin ProductionNO production

TKI

Bevacizumab(Anti-VEGF)

Ramucirumab(Anti-VEGFR2)

TKI (FDA Approved)SunitinibSorafenib, PazopanibAxitinibVandetanibRegorafenib

Afliberecept(VEGF Trap)

FDA-Approved Inhibitors

In Clinical Trials

TKI (In clinical trials)CediranibSemaxanibTorceranib, BrivanibTivozanibCabozantinib

Recovery of LV function with w/d of sunitinib therapy and ACEIblocker

Sunitinib-induced LVEF Decline and Immediate Recovery

0%

10%

20%

30%

40%

50%

60%

70%

80%

Baseline EF Greatest ChangedLVEF

Immediate Recovery

Eje

cti

on

Fra

ctio

n

*

MH Chen, unpublished

Recovery of LV function with w/d of sunitinib therapy and ACEIblocker

Sunitinib-induced LVEF Decline and Immediate Recovery

0%

10%

20%

30%

40%

50%

60%

70%

80%

Baseline EF Greatest ChangedLVEF

Immediate Recovery

Eje

ctio

n F

ract

ion

*

LVEF is NOT an adequate marker for cardiotoxicity or for recovery from same

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Resolution of cardiotoxicity at the ultrastructural level

Kerkela et al. Clin Trans Sci:2009

What is/are the target(s), inhibition of which leads to toxicity: VEGFRs, PDGFRs, AMPK, other?

30

40

0ImatinibVehicle

20

10

% K

i67+

50

60

30

40

0ImatinibVehicle

20

10

%

-act

inin

+

50

60

KIs not only target cardiomyocytes:Imatinib blocks differentiation of c-Kit+ CRSCs in culture

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BrdU Analysis

1 week Post-MI (n=5)• 1 BrdU+/16 myocyte

nuclei1 week Post-

MI+Sorafenib (n=2)

0

1

2

3

4

5

6

7

8

9

MI MI+Sorafenib

Per

cent

Brd

U+

(%

)

%BrdU+ Myocytes

0

5

10

15

20

25

30

MI MI+Sorafenib

Per

cent

Brd

U+

(%

)

%BrdU+ Non-Myocytes20 um

20 um

BrdU

DAPI

20 um

Actin

20 um

0

100000

200000

300000

400000

500000

600000 Control 500 nM 2 uM 5 uM

Total cell count

Effects on stem cells (2): Imatinib blocks proliferation of c-Kit(-) cardiac SP cells in culture

0

100000

200000

300000

400000

500000


0

20

40

60

80

100

PI- ANNEX-

CONTROL 0.5 uM 2 uM 5 uM

0

5

10

15

20

PI- ANNEX+

CONTROL 0.5 uM 2 uM 5 uM

Total cell count

Imatinib blocks proliferation of c-Kit(-) cardiac SP cells in culture

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0

100000

200000

300000

400000

500000


Total cell count

Mechanisms: 1) BCRP inhibition and 2) an additional mechanism related to kinase inhibition

Proliferation assay

0

40

80

120

WT WT + 2uMGleevec

Bcrpko Bcrpko +2uM Gleevec

Fo

ld d

iffe

ren

ce

Started with 10K P5 FVB cells cultured for 6days

*

*

#†

*: p<0.05 vs WT control#: p<0.01 vs Bcrpko†: p<0.01 vs WT+2uM Gleevec

Apoptosis (Annexin-V only)

0

2

4

6

WT WT + 2uM Gleevec Bcrpko Bcrpko + 2uM Gleevec

*

*: p=0.05 vs WT control#: p<0.01 vs WT control†: p<0.05 vs Bcrpko@: p<0.01 vs WT+2uM Gleevec

#

†@

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Mechanisms: 1) off-target BCRP inhibition and 2) an additional mechanism related to kinase

inhibition (on-target)

What do we need (Part 1)?

Better pre-clinical models

Cell Growth and ProliferationMaintenance of Sarcomere StructureCell Survival and RepairAngiogenesis

TKI: lapatinibneratinib, afatinib

Trastuzumab

Trastuzumab-DM1

Pertuzumab

NRG-1, 2, 3, 4HB-EGFEpiregulinBetacellulinEGFTGF-AmphiregulinEpigen

HB-EGFEpiregulinBetacellulinEGFTGF-AmphiregulinEpigen

Cell Growth and ProliferationMaintenance of Sarcomere Structure

AMPK Activation

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Phenotype of erbB2 Conditional Knock-out Mouse:On-target toxicity

erbB2-floxed erbB2-CKO

Crone SA, et al., Nature Medicine 8: 459-465 (2002)

Abnormal cardiac development and LV dilation with erbB2 knock-out

Phenotype of erbB2 Conditional Knock-out Mouse:On-target toxicity

erbB2-floxed erbB2-CKO

But the pre-clinical models are not always (or even mostly) predictive

Zebrafish as a pre-clinical model?

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We need better pre-clinical models: Sorafenib induces cardiomycyte loss in zebrafish

TG: cmlc2::DsRed-nuc

4dpf fish hearts (20x)

N=16

Cheng et al. Circ Res, 2011

EM

3 m

ed

ium

DM

SO

0.5

uM

1.0

uM

2.0

uM

5.0

uM

0.5

uM

1.0

uM

2.0

uM

5.0

uM

0.5

uM

1.0

uM

2.0

uM

5.0

uM

0

20

40

60

80

100

[Sorafenib] [Sunitinib] [Gefitinib]

5uM Sorafenib

5uM Sunitinib

Medium/DMSO

5uM Sunitinib5uM Gefitinib

Su

rviv

al %

Fish survival rate at 5dpf, treated at 2dpf

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Tabel 1: Videomicroscopic measurements at 5dpf in embryos that were treated at 2dpf with vehicle or 0.5uM TKIs..

n Long-axis Short-axis

EDD ESD FS=(EDD-ESD)/EDD

Wall thickness


vehicle 6 98.7±1.8

40.7±2.9

0.59±0.03

40.4±1.0

62.4±1.9

24.0±1.9

0.61±0.04

gefitinib 6 98.0±5.0

39.2±1.3

0.56±0.03

39.9±3.5

55.5±2.5

22.4±1.6

0.59±0.03

sorafenib 12 102.9±5.4

69.7±6.4

*

0.33±0.03 *

27.1±2.2 *

61.3±3.7

37.7±2.8

*

0.39±0.02 *

sunitinib 12 117.1±4.1 * #

78.6±4.5

*

0.23±0.03 *

23.9±1.0 *

60.8±2.5

42.6±2.1

*

0.30±0.03 * #

* denotes sorafenib or sunitinib vs vehicle or gefitinib; # denotes sunitinib vs sorafenib

Cheng et al. Circ Res, 2011

Tabel 1: Videomicroscopic measurements at 5dpf in embryos that were treated at 2dpf with vehicle or 0.5uM TKIs.



Wall thickness


vehicle 6 98.7±1.8

40.7±2.9

0.59±0.03

40.4±1.0

62.4±1.9

24.0±1.9

0.61±0.04


39.2±1.3

0.56±0.03

39.9±3.5

55.5±2.5

22.4±1.6

0.59±0.03


69.7±6.4

*

0.33±0.03 *

27.1±2.2 *

61.3±3.7

37.7±2.8

*

0.39±0.02 *

sunitinib 12 117.1±4.1 * #

78.6±4.5

*

0.23±0.03 *

23.9±1.0 *

60.8±2.5

42.6±2.1

*

0.30±0.03 * #


Table 1: Videomicroscopic measurements at 5dpf in embryos that were treated at 2dpf with vehicle or 0.5uM TKIs.



Wall thickness


vehicle 6 98.7±1.8

40.7±2.9

0.59±0.03

40.4±1.0

62.4±1.9

24.0±1.9

0.61±0.04


39.2±1.3

0.59±0.03

39.9±3.5

55.5±2.5

22.4±1.6

0.59±0.03


69.7±6.4

*

0.33±0.03 *

27.1±2.2 *

61.3±3.7

37.7±2.8

*

0.39±0.02 *

sunitinib 12 117.1±4.1 * #

78.6±4.5

*

0.30±0.03 *

23.9±1.0 *

60.8±2.5

42.6±2.1

*

0.30±0.03 * #


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0 2 4 6 8 10 12 140

20

40

60

80

100

Day

Pe

rce

nt s

urv

iva

l

Sham

MI+30 mg/kg/d SorafenibMI

Sham+30 mg/kg/d Sorafenib

MI+40 mg/kg/d Sorafenib

Sorafenib Decreases Survival of Mice Over 2 Weeks After MI Injury

100%

62.7%

23.3%p = 0.004

7.5%p < 0.000

Biomarkers

• TnI /TnT: Validated in setting of anthracycline use and suggestive data for trastuzumab (Daniella Cardinale; Dan Lenihan)

• BNP

• PET imaging

• Metabolomics


Gerszten and co-workers

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What do we need (Part 3)?• A full selectivity profile of all new agents (currently ~ 250 kinases)

• Greater selectivity of agents

• A better understanding of function of kinases in the heart

• Increased use of KI re-design strategiesa. Avoid bystanders with no/little role in cancerb. Dial down inhibition of kinases mediating toxicity (if not central to tumor progression)

Re-design of imatinib: Comparable tumor efficacy vs GIST with no LV dysfunction

Fernandez et al. J Clin Invest : 2007

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Effects of Cdk4/6 inhibition on anthracycline-induced cardiotoxicity


A little luck

Dox

Dox

Dox+ Cdk4/6 inhib

Full and complete co-operation between cardiology, oncology, toxicology, funding agencies, industry,

and non-profits… ?


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Human diseases caused by mutations in protein kinases: the other side of the coin

Potential uses of KIs in CV disease

Sunitinib in HCM phenocopy- AMPK mutation

Sorafenib in Leopard/Noonan syndrome- Ras/Raf mutations

Imatinib in Pulmonary Hypertension- Phase II study

Sorafenib in Pulmonary Hypertension

AcknowledgmentsJefferson Medical College CHB; DFCI; BWHRisto Kerkela Ming Hui ChenAdam Dicker Tammy ChuGabor KariUlrich RodekTempleRon VagnozziHui ChengSteve Houser; Jason Duran; Cat Makarewich

Kyle KolajaMD AndersonJB DurandAarif KhakooDan LenihanEd Yeh

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Effects on stem/progenitor cells

30

40

0V

20

10

Eje

ctio

n F

ract

ion

(%

) 50

60

Effects of Cdk4/6 inhibition on anthracycline-induced cardiotoxicity

p < 0.001 V vs. D

KI D D/KI

p < 0.001 D vs. D/KI

What has happened in past three years?

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What has happened in the past three years?

• HFA of the ESC published a guidelines/white paper (Eur J HF)

Eschenhagen and Shah

• HFSA committee developing guidelines/white paper-

Lindenfeld, Lenihan, Chen

• NCI convened two panels- a) detection and mgmt of HTN and b) cardiotoxicity panel for multi-targeted (VEGFRs +)

Steingart and Maitland

• The concept of cardio-oncology has emerged.

Durand, Lenihan, Sawyer, Schocken, many others

Acridine Orange staining at 3dpf, treated at 2dpf

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28

20 uM

10 uM

5 uM

2 uM

1 uM

Control

Zfish treated with VEGFR 2 small molecule inhibitor

Quantification of ventricular wall thickness in zebrafish treated with TKIs

0.3

0sunitinibControl

0.2

0.1

Ven

tric

ula

r w

all t

hic

knes

s

0.4

sorafenib

0.5 M

**

* p<0.01 vs. Control

Control

EGFR MO

AG1478

Courtesy G. Kari, A. Dicker , U. Rodeck- KCC

5/22/2013

29

Control cSP verapamil

50nM 500nM 1μM 2μM

+ Gleevec+ Gleevec

Effect of Gleevec on CSP (N=2)

0

0.5

1

1.5

Control 50nM 500nM 1_M 2_M Verapamil

Sunitinib induces ATP depletion

t = 8h

AMPK activity in vivo- ACC

5/22/2013

30

Abl c-Kit


Re-engineering of imatinib

imatinib WBZ-4

Re-engineering of imatinib

GIST (c-Kit) CML (Abl)


Selectivity profiles

5/22/2013

31

Sunitinib induces cytochrome c release

Chu et al. Lancet: 2007

LV myocardial volume (mass) normalized to BW

Wolf et al. Leuk Res 2010

5/22/2013

32

LV myocardial volume (mass) normalized to BW

Loss of 19% of LV mass after correction for body weight

Effects on cardiac resident stem/progenitor cells?

Differentiation of c-Kit+ CRSCs in culture

Red: CRSCs

Green: -actinin

Blue: DAPI

30

40

0ImatinibVehicle

20

10

% K

i67+

50

60

30

40

0ImatinibVehicle

20

10

%

-act

inin

+

50

60

Imatinib blocks differentiation of c-Kit+ CRSCs in culture

5/22/2013

33

Off-target inhibition of AMPK by sunitinib (Invitrogen)

eEF2K mTORC1

AMPK

eEF2

P

ACC

Translationinitiation

Translationelongation

P

Fatty acidbiosynthesis

AMP/ATP

Fatty acidoxidation

Energy homeostasis

AMPK and mTORC1/eEF2

eEF2K mTORC1

AMPK

eEF2

P

ACC

Translationinitiation

Translationelongation

P

Fatty acidbiosynthesis

AMP/ATP

Fatty acidoxidation

Energy homeostasis

AMPK and mTORC1/eEF2

sunitinib

5/22/2013

34

Partial rescue of sunitinib cardiotoxicity with AMPK-CA

The TKI market: Kinase inhibitor patents: 1988-2005

On-target toxicity of multi-targeted inhibitors

5/22/2013

35

Impaired flow reserve, reduced vessel number, and hypoxia in the cKO

Anthracyclines

30

40

0

dnPI3KNTg

20

10

Fra

ctio

nal

sh

ort

enin

g (

%)

50

60

Effects of inhibiting PI3-K (p110) on the heart’s response to hemodynamic stress

Sham

TAC

*

* p < 0.05 vs. sham

McMullen and Jay; Cell Cycle 2007

5/22/2013

36

Are the effects on the vasculature critical to the anti-cancer effects of VEGFR/PDGFR-targeted therapies?

Sunitinib

Proliferation assay

0

40

80

120

WT WT + 2uMGleevec

Bcrpko Bcrpko +2uM Gleevec

Fo

ld d

iffe

ren

ce

Started with 10K P5 FVB cells cultured for 6days

*

*

#†

*: p<0.05 vs WT control#: p<0.01 vs Bcrpko†: p<0.01 vs WT+2uM Gleevec

5/22/2013

37

Table 2. Evidence from experimental models suggesting cardiotoxicity of TKIs

by TK target.

Abbreviations: WT, wild type; KO, knockout- gene deleted; D/N, dasatinib, nilotinib; ER, endoplasmic reticulum; CMP, cardiomyopathy; HGF, hepatocyte growth factor (ligand for Met); FGFR fibroblast growth factor receptor; MI, myocardial infarction. See text for other abbreviations.

TK target(s)

TKIs Model Cardiac phenotype of model References

ERBB2 trastuzumab lapatinib

ERBB2 KO: + TAC

Spontaneous dilated CMP; worsened heart failure with pressure load; enhanced anthracycline sensitivity.

26, 27

VEGF VEGFRs

sunitinib sorafenib

bevacizumab

WT: VEGF Trap + TAC

Pathologic remodeling in response to pressure load.

46, 47, 69

KIT imatinib/D/N sunitinib sorafenib

1) W/WV mouse (KIT-deficient) + MI

2) WT: Arterial injury + imatinib

1) Adverse remodeling post MI due to reduced homing of mesenchymal stem cells to sites of injury; 2) Reduced stenosis post arterial injury.

53-55

Raf-1/B-Raf

sorafenib Raf-1 KO and dominant negative

+ TAC

LV dilatation and CHF with pressure load. 59, 60

PDGFRs imatinib/D/N sunitinib sorafenib

WT: MI + Administration

of PDGF

Reduced injury (ischemic protection). 48-50

JAK2 lestaurtinib STAT3 KO: MI; aging;

anthracycline administration;

pregnancy

Increased ischemic injury; reduced capillary density with aging; increased anthracylcine toxicity; peri-partum cardiomyopathy.

64, 65

Abl/Arg imatinib/D/N WT: imatinib Decline in LV function; induction of ER stress. 9, 38

Met (HGF

receptor)

N/A WT: MI or CMP models +

administration of HGF

Reduced fibrosis in MI and CMP models. Neoangiogenesis with HGF.


FGFR1/3 N/A Cell culture models: Administration of

FGF

Enhanced proliferation of cardiomyocytes and cardiac-resident stem cells.


Chen et al. Circulation, 2008

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