Advanced Glycation End Products in Diabetic Kidney

Disease

Grishma Parikh, MDMount Sinai School of Medine

5th Annual Friedman Fellows Symposium11/17/2012

AGEs

• Advanced Glycation End products (AGEs)– Formed by non-enzymatic glycosylation of

proteins and lipids– Potent inducers of inflammation and ROS– Source: • Exogenous : Food and Cigarettes• Endogenous : Normal metabolism and Aging

– Excess: Linked to major chronic diseases

333H.Vlassara: Annals of Medicine, 2008

Glucose+Proteins

LipidsRNA/DNA

A

HO· ROS B

RO·

C

HbA1c

Formation of AGEs (Oxidants)

AGEs toxicity is due both to the release of ROS and alterations of proteins and lipids.

AGEs

MG CMLDry heat

Striker 3

AGEs Regulate Oxidant Stress

Diet is a major source of excess oxidants (AGEs).

Koschinsky, T., Vlassara et al. PNAS 1997

ROS, AGEs, NOS E.R. Stress

Inflammation, Proliferation, Apoptosis or Cell Death

NAPDH ox, PKC, Mitochondrial Oxidant

Stress etc.Glucose, Fatty

Acids

Exogenous Oxidants (AGEs)

Food Intake

ROS

What is the Source of AGEs?

Oxidant (AGEs) Content ofCommon Foods

This is not “starvation”, it is French cooking!!

17MASHED1522Potato: fried

502SUSHI1348Salmon: broiled

1011STEWED5245Chicken: broiled

2000STEWED5367Beef: broiled

Low AGE diet (U/mg)Regular diet (U/mg)

AGE Formation Depends on Temperature of Cooking

A 50% Reduction may be sufficient clinically.

What is the effect of restriction of AGEs in diet?

Striker et al.88

Diabetic Subjects: AGE-Restricted Diet (x 4-6 wks)Reduces Inflammatory Markers

-20

-10

0

10

**-50

-25

0

25

50

*

hsCRP TNF α VCAM -1

%

fro

m b

asel

ine

-50

0

50

100

**

H.Vlassara, Proc.Nat.Acad.Sc: ‘O2

Type 2 Diabetes Trial: AGE Restriction and Insulin Resistance (x4 mos)

AGE-Restriction Reduces Plasma Insulin and Leptin

Vlassara group: Db Care, 2011

Plasma Insulin (μU/ml)

0

5

10

15

20

25

Pre- Post-

*

Leptin (ng/ml)

0

10

20

30

40

*

Pre- Post-

Type 2 Diabetes Trial: AGE Restriction (4 mo)

Vlassara group: Db Care, 2011

AGER1

0

50

100

150

200

250

pre post

*

NL

ADIPONECTIN

0

2

4

6

8

10

12 *

pre post NL

SIRT-1

0

100

200

300

400

500 *

pre post NL

AGE-Restriction Normalizes anti-oxidant defenses in T2D: i.e. AGER1, SIRT-1 and Adiponectin

Renal Function and AGEs

• The kidneys are a primary site for metabolism and excretion of oxidants

• Decreased renal function: increased OS and inflammation (TNFα) due to reduced anti-oxidant defenses

Striker et al.

Uribarri, J, Vlassara , H, JAGS, 2009

• AGEs and Renal Function are Linked

• Oral AGEs Influence Systemic AGEs at All Levels of Renal Function

Renal Function and Serum AGEs

(years)

0

50

100

20 40 60 80 100

150

200r = - 0.447p = 0.0001

eGFRCr vs. Aging (years) eGFRCr vs. Serum CML

0

10

20

eGFRCr

0 50 100 150 200

r = - 0.647p = 0.0001

30

40

50

60

NORMAL :

sAGE

(u/m

l)

0

1015202530

hours0 20 40

5

High AGE Low AGE

Serum60

30

0 20 40hours

Urine

Serum/Urine AGEs in Diabetes after Meals

GFR= 84, proteinuria GFR= 100, μ/alb

hours

0

60

30

20 40

DIABETIC

0

202530

0 20 40

5

hours

10sAGE

(u/m

l)

15

AGE excretion can be reduced prior to clinically significant changes in GFRT. Koshinsky. H.Vlassara PNAS 1997

14

Urine CML (nmoles/24 hr)

0.0

4,000.0

8,000.0

12,000.0

16,000.0

20,000.0

1st 5th visit

*p<0.05

AGE Restriction Restores Urinary Excretion of AGEs (active and filtration)

Vlassara group, 2009

AGE-Restriction Effect on Oxidant Excretion (MG) and/or filtration (CML) in T2D

This also applies to Normal Adults!!

Urine MG (nmoles/24 hr)

0.00

100.00

200.00

300.00

400.00

1

* p<0.025

5th visit

T2 Diabetic Nephropathy and AGE restriction (4 mo)

Vlassara et al: PNAS 2002; JCEM, 2009; Diab Care 2-12

% Change-100 -50 0 50 100

RAGE

8-iso

sCML

TNFα

VCAM-1

sMG

AGER1

AGE-Restricted Diet Restores Anti-ox (AGER1) Levels, Promotes Renal Elimination of AGEs, and Reduces OS/Inflammation (i.e.TNFα)

Lowering oxidant intake allows kidneys to excrete AGEs, which reduces OS and Inflammation.

Standard Diet AGE-Restricted

AGE-Restricted Diet Prevents Diabetic Nephropathy in NOD Mice

Restriction of AGEs – not nutrients or blood glucose – can Prevent CKD in T1D-Susceptible Mice

Peppa, M.: Diabetes J., 2005

Ways to Decrease AGE Uptake

• Change AGEs in food– Western diet is rich in AGEs and NOT EVERYONE

CAN FOLLOW THE DIET• Sequester AGEs in the gut and eliminate them

in the stool– Sevelamer is a non-absorbable polymer that was

found to reduce HbA1c and lipids, similar to effects seen in patients treated with a low-oxidant diet. This suggested it might remove AGEs

20

40

60

80

100 *

AGE AGE +pH=1

AGE +pH=7.0

%

Bou

nd 1

25 I-

AGE-

BSA

0

Sevelamer

Binding of AGEs in vitro:Sevelamer Sequesters AGEs in a pH-dependent manner

1) Sevelamer carbonate treatment would result in a 20% decrease in serum AGEs within 8 weeks.

Therefore we proposed a Pilot Cross-over Study in 20 T2D patients with CKD Stages 2-4 with the following Hypotheses:

2) Renvela treatment would decrease serum inflam/ROS markers by 10% within 8 weeks.

No Medications were changed, all participants were on drugs to control BP, many were on statins, insulin, etc..

Vlassara group, CJASN, 2011

% Change

25

0

-50

TNFα

-25

8-iso

sCM

L

TNFα

sMG

*

8-iso

sCM

L

sMG

***

50

CaC03 (CONTROL)

-75

-100

75

100

AGER

1SI

RT1

*

*

FGF2

3

*

AGER

1SI

RT1

SEVELAMER

SEVELAMER Lowers OS and Inflammation and Restores AGER1, SIRT1 and FGF23 mRNA in T2D with CKD

METABOLIC CHANGES: Sevelamer Carbonate Lowers HbA1c and Plasma Lipids in T2D with CKD

Vlassara group, CJASN, 2011

LDL-

Chol

10

0

-20

Tota

l Cho

l

-10

HbA1

c

Trig

**

*

20

CaC03 SEVELAMER

Tota

l Cho

lLD

L-Ch

ol

HbA1

cTr

ig

*

% C

hang

e

Striker et al. 22

Conclusions Pilot Trial• Sevelamer lowered serum (and cellular) AGE

levels, by sequestering AGEs in the gut• Sevelamer lowered markers of Oxidative Stress

and inflammation and decreased FGF23• Sevelamer lowered HgbA1c, and lipids (t-chol,

triglycerides, even though most were on statins)

• These data suggest that Sevelamer reduces AGEs as one primary target

• A longer/larger study is now being conducted to validate these results

Striker et al.

Proposed New Study of Sevelamer: Examine the Effects of Sevelamer Carbonate on

Cardio-Metabolic Risk Factors and Selected Aspects of Kidney Function in Type 2 Diabetics

with Stage 2-4 CKD

Hypothesis: Sevelamer carbonate (Renvela) will lower inflammation and oxidative stress in the body by absorbing AGEs in the gut and removing them from the body.

Study Design• 6 month prospective, comparative study• 120 patients randomized to receive, in parallel, either:

1. Sevelamer carbonate (1600 mg tid) 2. Calcium carbonate (1200 mg tid)

• Endpoints• Primary: HbA1c and AGEs• Secondary: markers of glucose metabolism,

inflammation, oxidative stress

Dietary AGE content is assessed

Recruitment

• MSSM: • 89 randomized

• Beth Israel: • 49 randomized

• Sponsor: Genzyme-Sanofi• Industry sponsored (investigator-initiated)

Striker et al.

Summary• AGEs elevate levels of inflammatory markers and oxidative stress• The diet is a major source of oxidants (AGEs) in DM.• The kidneys are the major sites of disposal of oxidants, (and

AGEs), so treatment/prevention of CKD is critical• Major internal and external sources of ROS/Infl and AGEs can be

managed with current medications

• Sevelamer reduce oxidants (AGEs) and improves anti-oxidants, lipids, FGF23 and glucose control in T2D

Many Thanks to Team AGE

MSSM• Gary Striker• Helen Vlassara• Nephrology: Jaime Uribarri,

John He, Shirisha Guthikonda

• Endocrinology: Ronald Tamler

Beth Israel New York• Endocrinology: Leonid

Poretsky, Augustin Busta, Carla Romero, Yun Feng

• Nephrology: Nikolai Harbord

STUDY COORDINATORS AND MANY OTHER COLLABORATTORS…………