1,5 Anhydroglucitol and the Monitoring of Postprandial Glucose Control Steven D Wittlin M.D. U of Rochester School of Medicine and Dentistry

Importance of Post- Prandial Hyperglycemia

The Growing Epidemic of Diabetes

Diabetes is Not Well- Controlled Data from NHANES and DCCT

Duration of daily metabolic conditions Breakfast Lunch Dinner0:00 am4:00 amBreakfast Postprandial PostabsorptiveFasting Monnier L. Eur J Clin Invest 2000;30(Suppl. 2):3–11

Relationship between FPG and 2-hr post-challenge PG in type 2 diabetes patients: NHANES III Distribution (%) HbA 1c Fasting plasma glucose <7 mmol/l 7–7.8 mmol/l >7.8 mmol/l <7.8 mmol/l 7.8–11.1 mmol/l >11.1 mmol/l Erlinger TP, Brancati FL. Diabetes Care 2001;24:1734–8 Distribution (%) HbA 1c 2-hr post-load plasma glucose

Relationship between HbA1C, FPG and 2-hr post-challenge PG in type 2 diabetes patients: NHANES III Erlinger TP, Brancati FL. Diabetes Care 2001;24:1734–8

Correlation between plasma glucose levels after OGTT and standard mixed meal Wolever TMS et al. Diabetes Care 1998;21:336–40 r=0.97

Relationship between HbA1C, FPG and 2 h. PPG Van Haeften T et al Metabolism 2000

Woerle HJ et al Arch Intern Med. 2004;164: Relative Changes in FPG and 2-h PG as HbA 1c Increases Plasma Glucose (mg/dL) = HbA 1c versus 2hppg = HbA 1c versus FPG r = 0.55 y = 47.1 x -109 r = 0.48 y = 12.0 x +30 HbA 1c (%)

Relationship between FPG and 1 st -Phase Insulin Release ( van Haeften T et al Metabolism 2000 )

Relationship Between Diabetes Status and 1 st Phase Insulin Release ( van Haeften T et al.Metabolism 2000 )

As Patients Get Closer to A1C Goal, the Need to Successfully Manage PPG Significantly Increases Adapted from Monnier L, Lapinski H, Collette C. Contributions of fasting and postprandial plasnma glucose increments to the overall diurnal hyper glycemia of Type 2 diabetic patients: variations with increasing levels of HBA(1c). Diabetes Care. 2003;26:

Post-load Hyperglycemia and The Metabolic Syndrome –0.13 – * Fasting plasma glucose 0.20* * 0.21* 0.24* 0.25* 2-hr plasma glucose C-reactive protein Cellular fibronectin Tissue plasminogen activator (TPA) Free fatty acids Triglycerides Waist: hip ratio Body mass index (BMI) Age Yudkin JS. Lancet 2002;359:166–7 *p<0.05; p<0.01

Controlling Postprandial Glucose Prospective trial of fasting vs pc control in 164 pts w/ Type 2 DM Prospective trial of fasting vs pc control in 164 pts w/ Type 2 DM Forced titration to target first FBS < 100 and then, 90 min pc < 140 if not achieved previously Forced titration to target first FBS < 100 and then, 90 min pc < 140 if not achieved previously Results: Results: HbA1C fell from 8.7 % to 6.5% HbA1C fell from 8.7 % to 6.5% Only 64% of patients achieving FPG < 100 only reached HbA1C < 7% Only 64% of patients achieving FPG < 100 only reached HbA1C < 7% 94% of patients w/ pc < 140 reached HbA1C < 7% 94% of patients w/ pc < 140 reached HbA1C < 7% Decreased pc BG accounted nearly twice as much as FBS for fall in HbA1C Decreased pc BG accounted nearly twice as much as FBS for fall in HbA1C If HbA1C < 6.2%, pc accounted for ~ 90% If HbA1C < 6.2%, pc accounted for ~ 90% If HbA1C > 8.9%, pc accounted for ~ 40% If HbA1C > 8.9%, pc accounted for ~ 40% Woerle HJ et al Diabetes Research and Clinical Practice 2007

Contribution of Postprandial BG to HbA1C Woerle HJ et al Diabetes Res Clin Pract Jan 19

Glycemic Excursions Predict Oxidative Stress Monnier L et al JAMA. 2006;295:

Responses to a Mixed Meal in Diabetic and Non-Diabetic Individuals (Singhal P et al AJP 2002 )

Endogenous Glucose Production After a Mixed Meal in Diabetic and Non-Diabetic Individuals (Singhal P et al AJP 2002 )

Routes of Post-Prandial Glucose Disposal

Post-Prandial Glucose Metabolism : Mechanism Study of 11 normal volunteers after a standard test meal Study of 11 normal volunteers after a standard test meal Triple isotope technique and indirect calorimetry Triple isotope technique and indirect calorimetry Intravenous Intravenous Tritiated GlucoseTritiated Glucose C-14-labelled bicarbonateC-14-labelled bicarbonate Oral Oral Deuterated GlucoseDeuterated Glucose Woerle HJ et al.Am J Physiol Endocrinol Metab 284: E716-E725, 2003

Post-Prandial Glucose Disposal ( Woerle Hans J et al AJP Endo Metab 2003 )

Changes in Postprandial Glucose Metabolism in Type 2 DM Use triple isotope technique and indirect calorimetry Use triple isotope technique and indirect calorimetry DM pts had: DM pts had: increased overall glucose release increased overall glucose release Increased gluconeogenesis and glycogenolysis Increased gluconeogenesis and glycogenolysis ~90% of the increased glucose release occurred in the first 90 min post-prandial ~90% of the increased glucose release occurred in the first 90 min post-prandial In DM glucose clearance and oxidation were reduced In DM glucose clearance and oxidation were reduced Non-oxidative glycolysis was increased Non-oxidative glycolysis was increased Net splanchnic glucose storage was reduced ~ 45% d.t. increased glycogen cycling Net splanchnic glucose storage was reduced ~ 45% d.t. increased glycogen cycling Woerle HJ et al Am J Physiol Endocrinol Metab 2006

Effect of Pre- or Post-Meal Exercise on Glycemic Control (Yamanouchi K et al Diab Res & Clin Pract, Oct 2002 )

The Glycemic Index

Physiologial Consequences of a High Glycemic –Index Meal

Summary of Studies of Low vs High Glycemic Index Diets

Effects of 2h Post-Prandial Glucose +/- Vitamins C +E on Flow-Mediated Dilatation ( Title LM et al JACC Dec 2000 )

Relative risk for death increases with 2-hour blood glucose irrespective of the FPG level <6.16.1– –11.0 <7.8 Fasting plasma glucose (mmol/l) 2-hour plasma glucose (mmol/l) Hazard ratio Adjusted for age, center, sex DECODE Study Group. Lancet 1999;354:617–621

Effect of Acarbose on CVD in Patients with IGT ( STOP-NIDDM) ( Chiasson J-L et al JAMA July 2003 )

Effect of Acarbose on Development of HBP in Patients with IGT (STOP-NIDDM ) ( Chiasson J-L et al JAMA July 2003 )

Summary Postprandial glycemia plays a clinically important role in the complications of diabetes Postprandial glycemia plays a clinically important role in the complications of diabetes Postprandial glycemia is a major contributor to overall glycemic control ESPECIALLY in moderately-well to well controlled patients Postprandial glycemia is a major contributor to overall glycemic control ESPECIALLY in moderately-well to well controlled patients

So How Can We Assess Post-Prandial Glucose Control Clinically ?? Frequent fingersticks Frequent fingersticks HbA1C HbA1C Fructosamine Fructosamine Continuous Glucose Monitoring Systems Continuous Glucose Monitoring Systems Historical Historical Real-time Real-time 1,5 Anhydroglucitol 1,5 Anhydroglucitol

So How Can We Assess Post- Prandial Glucose Control Clinically ?? Frequent fingersticks Frequent fingersticks HbA1C HbA1C Fructosamine Fructosamine Continuous Glucose Monitoring Systems Continuous Glucose Monitoring Systems Sensor-Augmented Insulin Pumps Sensor-Augmented Insulin Pumps

A New Idea ! 1,5 Anhydroglucitol

History of 1,5AG ,5AG was discovered in plant of Polygala Senega. 1973Presence in human body was reported. 1977Decrease of plasma 1,5AG concentration with uremia and diabetes mellitus was reported Blood 1,5AG was determined in Japan. After this, research on relationship between diabetes mellitus and 1,5AG has become active.

Blood test measuring 1,5-anhydroglucitol (1,5-AG) 1,5-AG is a monosaccharide (similar to glucose structure) 1/40 of glucose concentration – healthy human blood Primary Source of 1,5-AG – Food Further distributed to skin, muscle, and other tissues/organs Reabsorbed very efficiently through kidney (urinary excretion is 1/20 of total amount in body) Large Body Pool of 1,5-AG (6-7 times > Blood) Not metabolized much in the body (metabolic turnaround rate at least 3 days) 1,5-AG urinary excretion remarkably increases with hyperglycemia!

The structure of 1,5-anhydroglucitol (1,5AG) O OH HO OH O HO D-glucose 1,5-anhydro-D-glucitol (1-deoxyglucose)

Fully Automated Enzymatic Method for 1,5 AG Assay ( Glycomark ) Fukumura Y et al Clin Chem 1994 HRP=Horseradish peroxidase ; PROD= pyranose oxidase; HTB=3 hydroxyriiodobenzoic acid ; 4AAP= 4 aminoantipyrine

Specificity of Assay The following dont interfere in concentrations up to 10 grams/L : The following dont interfere in concentrations up to 10 grams/L : Sorbitol Sorbitol Mannitol Mannitol Sucrose Sucrose Lactose Lactose Maltose Maltose Fructose Fructose

1,5 AG Content of Foodstuffs in Japanese Diet Yamanouchi T et al Am J Physiol 263: E268-E

1,5 Anhydroglucitol Specimen Requirements

Oral Supply 1,5AG (5-10mg/day) Blood stream Tissues Internal Organs ( mg) Kidney Urinary excretion (5-10mg/day) Oral Supply 1,5AG (5-10mg/day) Blood Stream (1,5-AG Level Lower) Tissues Internal Organs ( mg) Kidney Urinary excretion (INCREASED) A. Normoglycemia B. Hyperglycemia Glucose Blocks Reabsorption Physiology of 1,5-AG Buse JB et al Diab Tech & Ther (3) :

1,5 AG Kinetics in Humans Yamanouchi T et al Am J Physiol 263: E268-E

Urinary Excretion of Glucose and 1,5 AG Fluctuate in Parallel in Rats Yamanouchi T et al Am J Physio : E423-E427

Urinary Excretion of Glucose and 1,5 AG Fluctuate in Parallel in Rats Yamanouchi T et al Am J Physio : E423-E427

But…1,5 AG Does Not Fluctuate with Variations in Plasma BG in Nephrectomized Rats !! Yamanouchi T et al Am J Physio : E423-E427

Fructose, Mannose and 1,5 AG Share A Common Transport Mechanism Yamanouchi T et al Biochim et Bipophys Acta : 89-95

Fructose, Mannose and 1,5 AG Share A Common Transport Mechanism Yamanouchi T et al Biochim et Bipophys Acta : 89-95

Renal Tubular Absorption of Glucose and 1,5 AG Normal Glucose active transporter Fructose, mannose active transporter Glucose 1,5AG Filtration at glomerulus Reabsorption at renal tubule urine

Renal Tubular Absorption of Glucose and 1,5 AG Hyperglycemia Glucose active transporter Fructose, mannose active transporter Glucose 1,5AG Filtration at glomerulus Reabsorption at renal tubule urine

Renal Tubular Absorption of Glucose and 1,5 AG Stickle D and Turk J. Am J Physiol Endocrinol Metab 273: E821-E830, 1997

Linearity of 1,5 AG Assay Nowatzke W et al Clin Chim Acta 2004

Recovery Time of 1,5 AG in Treated Patients with Type 2 DM Yamanouchi T et al Jpn. J. Clin. Med. 47: , 1989

Histograms of serum 1,5AG concentrations in Japanese healthy subjects ,5AG (µg/mL) 26.6±7.2 Male (n=332) Frequency 21.5±6.0 Female (n=207) Frequency ,5AG (µg/mL) 24.6±7.2 Male and Female (n=539) Frequency ,5AG (µg/mL)

Distribution of 1,5 AG in a Healthy US Population Nowatzke W et al Clin Chim Acta 2004

The mean 1,5AG levels in healthy subjects during 2 years (n=245) mean±SD

Diurnal change of plasma glucose and 1,5AG : healthy :IGT Others: diabetes B: breakfast L: lunch D: dinner BLD

Histogram of serum 1,5AG concentrations Healthy (n=539) 24.6±7.2 µg/mL Diabetes (n=808) 7.3±7.1 µg/mL Serum 1,5AG (µg/mL)

Serum 1,5AG levels in healthy subjects, Impaired- glucose-tolerance, diabetes mellitus, and various other disorders ,5AG (µg/mL) Healthy subjects (n=539) IGT (n=451) DM (n=808) Non-DM (n=238) P<0.001 Yamanouchi T et al Diabetes 1991; 40: 52-57

1,5AG Index 1,5AG x UG = , 5AG x UG Plasma 1,5AG (µg/mL) N =47 Log y = log x R = (P < ) Plasma 1,5AG (µg/mL) Urinary glucoses (g/day)

1,5-AG Physiology Implication Because 1,5-AG levels fluctuate according to glucosuria, the response is much more rapid than glycemic markers based on the glycation process (A1C). Responds Rapidly and Sensitively to Glycemic Changes

Correlation between glycemic control markers and past fasting plasma glucose (FPG) Distribution of 1,5AG to FPG shows hyperbolic dispersion different from HbA 1C and fructosamine. Therefore, Spearman's rank correlation coefficient is used here to make comparison between groups. Measurement point of FPG 1, 5AGHbA 1C Fructosamine same time 1 week ago 2 weeks ago 3 weeks ago 4 weeks ago r = r = r = r = r = r = 0.27 r = 0.53 r = 0.81 r = 0.51 r = 0.72 r = 0.80 r = 0.73 r = months ago 3 months ago r = -0.39r = 0.73 r = 0.65

Glycemic control markers 1,5A G Fructosamine HbA 1C Blood glucose Weeks before measurement

0 Renal excretion of glucose (g/day) Plasma glucose (mg/dL) Weeks Changes in various glycemic control markers in NIDDM patient with poorly controlled glycemia after starting insulin treatment HbA 1C (%) Plasma 1,5AG (µg/mL) Lente 12u 8 6

Clinical Parameters for 1,5 AG 1,5-AG serum concentrations in normal humans vary widely (10-40 ug/ml) Little change day to day because of large body pool relative to daily intake and metabolic inertness Few normal subjects show an alteration in 1,5-AG level in the normal range during 2-3 years When hyperglycemia occurs (glucosuria), 1,5-AG serum levels fall rapidly (1-2 days) Individual variance in renal threshold does not appear to seriously influence clinical utility of 1,5-AG (glucose fluctuates more widely than individual renal threshold variance) Exception – Gestational Diabetes

Post-load glucose measurements in OGTTs correlate well with 1,5-AG in subjects with IGT R= R= ,5 Anhydroglucitol is a better indicator than A1C of postprandial blood glucose levels in IGT subjects N = 211 Yamanouchi T et al., Clinical Science 2001

Correlation between 1,5AG and HbA 1c Cut-off value HbA 1c (Normal range %) 1,5AG (µg/mL) (%)

Weeks Mean plasma glucose (mg/dL) ,5AG (µg/mL) HbA 1C (%) 7 1,5AG, HbA1C, and mean plasma glucose values during a 92-weeks period in a patient with Type 1 DM

Changes of 1,5AG and HbA1C values during a 13- months period in a patient with Type 2 DM

Glycemic control in Type 2 DM patients before and after the study treatment Before after

Measurement of 1,5AG and HbA1c between October and April, around the new year, in 17 patients with Type 2 DM

1,5AG, HbA 1c, and FPG levels in NIDDM patients, well controlled with diet, OHA, CIT or MIT Diet OHA CIT MIT Mean±SE Fasting Plasma Glucose (mg/dL) HbA 1c (%) 1,5AG (µg/mL) * * *p<0.05:Diet vs. OHA and MIT **p<0.05:CIT vs. OHA and MIT OHA: oral hypoglycemic agents therapy CIT: conventional insulin therapy MIT: multiple insulin therapy

GlycoMark Evaluates Daily Glycemic Excursions in Moderately-Well Controlled Patients HbA1c showed no significant differences among all groups *Plasma 1,5-AG in diet group significantly higher than OHA and MIT groups (P<0.05) **Plasma 1,5-AG in CIT significantly lower than diet, OHA, and MIT groups (P<0.05) N = 76 well-controlled type 2 diabetes patients OHA=oral hypoglycemic agents, CIT=Conventional Insulin Therapy, MIT=Multiple Insulin Injection Therapy Kishimoto et al.Diabetes Care 1995)

Objective: Monitor glycemic control following changes in antidiabetic medication Objective: Monitor glycemic control following changes in antidiabetic medication 56 type 2 diabetic patients treated with oral hypoglycemic agents for 4 weeks 56 type 2 diabetic patients treated with oral hypoglycemic agents for 4 weeks After 4 weeks, treatment discontinued in half of patients and monitored for 2 more weeks After 4 weeks, treatment discontinued in half of patients and monitored for 2 more weeks 1,5-AG, Glucose, A1C, and Fructosamine were measured 1,5-AG, Glucose, A1C, and Fructosamine were measured Clinical usefulness of serum 1,5-AG in monitoring glycemic control Yamanouchi T et al., The Lancet 1996

Serial Changes in A1C in Newly Diagnosed Type 2 Diabetes Patients Group A – 28 patients who continued treatment for 6 weeks Group B – 28 patients who discontinued treatment after 4 weeks No Significant Difference Yamanouchi T et al.Lancet 1996

Serial Changes in 1,5-AG in Newly Diagnosed Type 2 Diabetes Patients Group A – 28 patients who continued treatment for 6 weeks Group B – 28 patients who discontinued treatment after 4 weeks P< ,5 Anhydroglucitol detected slight change in glycemia Yamanouchi T et al.Lancet 1996

Time course of normalization of Plasma 1,5AG levels in NIDDM weeks 1,5AG (µg/mL) Yamanouchi T et al Diabetes :

McGill J et al Diabetes Care 2004

FDA Study – Longitudinal Changes Time point Statistic 1,5AGug/mlA1C%Fructosamineumol/LGlucosemg/dL Baseline Mean Mean Visit 2 (2 weeks) Mean Mean Mean% Change Mean% Change3.0*57.9% % * *-11.7%187.4*-16.7% Visit 3 (4 weeks) Mean Mean Mean% Change Mean% Change3.7*94.7%8.8*-7.4%340.0*-17.2%181.4*-19.4% Visit 4 (8 weeks) Mean Mean Mean% Change Mean% Change5.0*163.2%8.2*-13.7%317.5*-22.7%172.6*-23.3% *p<0.05 vs. baseline McGill J et al Diabetes Care 2004

FDA Study – Longitudinal Changes McGill J et al Diabetes Care 2004

Assessing the Role of 1,5 AG for Monitoring Post-Prandial Glycemic Excursions K Dungan, J. Buse, J Largay, M Kelly, E Button, S Kato, S. Wittlin University of North Carolina University of Rochester Dungan K et al Diabetes Care; June 2006

Study to Evaluate 1,5 AG/Glycemic Excursions as Determined by Continuous Glucose Measurements Moderately-Controlled Patients (n=34) with A1Cs between 6.5 and 8.0 monitored over 7 days Comparing 1,5 AG, Fructosamine,and Hemoglobin A1C levels to Glycemic excursions above the renal threshold (> 180 mg/dl) Glycemic excursions measured by CGMS Objective: To demonstrate the relationship between serum 1,5-AG (relative to A1C and fructosamine) and the occurrence of postprandial hyperglycemia as reflected by CGMS in suboptimally controlled patients with diabetes Dungan K et al Diabetes Care; June 2006

Study Methodology Dungan K et al Diabetes Care; June 2006

Postprandial Variables AUC-180 – measure of total area above 180 mg/dl for (mg/dl*Day) Average Postmeal (Maximum) Glucose (mg/dl) – maximum height of each postmeal glucose excursion for breakfast, lunch, and dinner Postprandial Index (PI) – 4 variable combination (max glucose levels for post-breakfast, lunch, dinner and AUC days) Dungan K et al Diabetes Care; June 2006

Correlation AUC-180 vs. Glycemic Assay AUC-180^ (mg/dl*Day) Avg. A1C A1C-End Interval* Avg. 1,5- AG 1,5-AG- End Interval Avg. FA FA- End Interval Interval 1 & 2 Interval 1 & 2N=34 R = 0.36 p =0.02 R = 0.35 p =0.02 R = p = R = p = R = 0.33 p =0.03 R = 0.38 p =0.01 Interval 1 N=34 R = 0.23 p =0.09 R = 0.22 p =0.11 R = p =0.02 R = p =0.02 R = 0.16 p =0.18 R = 0.12 p =0.25 Interval 2 N=33 R = 0.35 p = 0.02 R = 0.34 p =0.03 R = p =0.008 R = p =0.005 R = 0.37 p = 0.02 R = 0.39 p = 0.01 ^AUC-180=area under the curve for glucose greater than 180mg/dL as determined by CGMS software. *End-interval is visit 2 for interval 1, visit 3 for interval 2 and total (1 & 2). 1,5-AG Correlated Better than A1C or Fructosamine to AUC-180 Dungan K et al Diabetes Care; June 2006

Avg. Maximum Postmeal Glucose vs. Avg. Glycemic Assay 1,5-AG Correlated Better than A1C or Fructosamine to ALL Postmeal Max Values Avg. A1C Avg. 1,5-AG Avg. FA Avg. Postmeal Max (Breakfast) N=20 R = 0.12 p = 0.31 R = p =0.05 R = p =0.494 Avg. Postmeal Max (Lunch) N=23 R = 0.19 p = 0.19 R = p =0.15 R = 0.06 p = 0.39 Avg. Postmeal Max (Dinner) N=22 R = 0.25 p = 0.13 R = p = R = 0.35 p = 0.06 Combined Postmeal Max (Breakfast, Lunch Dinner)- Multiple Regression N=19 R = 0.25 R = R = 0.36 Dungan K et al Diabetes Care; June 2006

Postprandial Index vs. Average Glycemic Assay 1,5-AG Correlated Better than A1C or Fructosamine to the Postprandial Index Postprandial Index (Multi- variate-PI) N=19 Avg. A1C Avg. 1,5- AG Avg. FA R=0.36R=0.58R=0.36 *Postprandial Index is the conglomerate multivariable analysis using AUC-180 and post-meal maximum glucose values as the independent variables. Dungan K et al Diabetes Care; June 2006

Postprandial Index vs. A1C/1,5-AG Assay Ratio A1C/1,5-AG Ratio Correlated Better than A1C or 1,5-AG independently to the Postprandial Index Combination of 1,5-AG and A1C are more predictive of postprandial hyperglycemia Postprandial Index (Multi- variate-PI) N=19 Avg. A1C Avg. 1,5- AG Avg. A1C/Avg. 1,5-AG Ratio R=0.36R=0.58R=0.66 *Postprandial Index is the conglomerate multivariable analysis using AUC-180 and post-meal maximum glucose values as the independent variables. Dungan K et al Diabetes Care; June 2006

Average and Premeal Glucose vs. Glycemic Assay Fructosamine and A1C correlated better than 1,5-AG to both average glucose and premeal glucose variables Avg. A1C Avg. 1,5-AG Avg. FA Average Glucose – CGMS Sensor N = 34 R = 0.27 p = 0.26 R = p = 0.23 R = 0.40 P = 0.04 Combined Premeal (Breakfast, Lunch Dinner) - Multiple Regression N=19 R = 0.42 R = R = 0.45 Dungan K et al Diabetes Care; June 2006

1,5 AG as Adjunct to A1C to Reflect Postprandial Hyperglycemia 1,5 AG is indicative of differing postmeal glucose levels in moderately controlled patients – despite similar A1C levels! GlycoMark (1,5-AG) Range 0-6 N=17 A1C (%) Mean 1,5-AG (ug/ml) Mean Total AUC-180 Glucose 1 PostMeal Glucose-Max Mean (mg/dl) BreakfastN=9 LunchN=10 DinnerN=9 Higher Postprandial Variables GlycoMark (1,5-AG) Range 6-18 N=16 A1C (%) Mean 1,5-AG (ug/ml) Mean Total AUC-180 Glucose 1 PostMeal Glucose-Max Mean (mg/dl) Breakfast N=11 PostMeal Glucose-Max Mean (mg/dl) Lunch N=13 N=13 PostMeal Glucose-Max Mean (mg/dl) DinnerN=13 Lower Postprandial Variables Dungan K et al Diabetes Care; June 2006

GlycoMark Monitors Postprandial Hyperglycemia Dungan K et al. Diabetes Care (June 2006) Patients were sorted by glycemic excursions as measured by CGMS (AUC-180) and subdivided into two populations – bottom 50 th percentile (17 patients) and top 50 th percentile (17 patients). Authors Conclusions 1,5-AG (GlycoMark) assay reflects glycemic excursions, often in the postprandial state, more robustly than other established glycemic assays. 1,5-AG was reflective of varying postmeal glucose levels, despite similarities in A1Cs. In clinical practice, A1C and 1,5-AG may be used sequentially, first employing the A1C assay to identify patients who are moderately controlled and then using the 1,5-AG assay to determine the extent of postprandial glycemic excursions.

52 year old female with type 1 DM A1C 7.43% 1,5-AG 12.4mcg/dL PPG max 195 mg/dL 49 year old male with type 2 DM A1C 7.27% 1,5-AG 4.5mcg/dL PPG max 235 mg/dL Representative Patients

Even though A1C levels are similar, 1,5 Anhydroglucitol levels are indicative of differing postmeal glucose levels in moderately controlled patients (A1C 6.5 to 8.0%). Dungan K et al Diabetes Care; June 2006

UNC/Rochester Study Conclusions In a subset of moderately controlled patients (A1C 6.5 to 8.0), significant postprandial hyperglycemia was present 1,5-AG reflects postprandial hyperglycemia more robustly than established glycemic assays At similar A1C levels, there may be variability in postprandial hyperglycemia –which is reflected by 1,5- AG levels!!

UNC/Rochester Study Clinical Possibilities 1,5-AG may be used in combination with A1C for better predictability of postprandial hyperglycemia than either assay alone A Two-Step Sequential Process Might be Used: 1) Use A1C to identify patients who are moderately controlled (A1C 6.5 to 8.0) 2) Use1,5-AG to determine extent of postprandial hyperglycemia

1,5 Anhydroglucitol Ranges and Diabetes Assessment GlycoMark values < 10 ( in A1C range 6.5 – 8.0) indicate presence of postprandial hyperglycemia GlycoMark (ug/ml) Assessment of Diabetes 14 or higher Normal (healthy) 10.0 – 13.9 Well-Controlled 6.0 – 9.9 Moderately Uncontrolled Moderately Uncontrolled 2.0 – 5.9 Poorly Controlled 1.9 or lower Very Poorly Controlled

1,5 AG (ug/ml) Approximate Mean Postmeal Maximum Blood Glucose (mg/dl) > 12 < < 2 > 290 1,5-AG and Postmeal Glucose Levels 1,5-AG and Postmeal Glucose Levels

1,5 Anhydroglucitol as Comprehensive Adjunct to A1C Diagnostic Algorithm Managing Short-Term Glucose Control Managing Postprandial Glucose Control (PPG) 1,5 AG (ug/ml) DiabetesA1C > 10 Well- Controlled – 10* Moderately Controlled Poor Control < 2 Very Poor Control > 10 1,5 AG (ug/ml) Approximate Mean Postmeal Maximum Blood Glucose (mg/dl) > 12 < < 2 > 290

Performance of 1,5 Anhydroglucitol in Recent Drug Trials

Sitagliptin and 1,5 Anhydroglucitol Evaluated efficacy and tolerability of sitagliptin in Japanese patients with T2DM over 12 weeks Evaluated efficacy and tolerability of sitagliptin in Japanese patients with T2DM over 12 weeks Initial A1C levels to 10.0% Initial A1C levels to 10.0% Randomized to sitagliptin (n=75) or placebo (n=76) Randomized to sitagliptin (n=75) or placebo (n=76) Stein P et al ADA Poster 537-P

Sitagliptin and 1,5 Anhydroglucitol Change from Baseline to Study End – Comparison of Mean Values PlaceboSitagliptin 100mg 1,5-AG (μg/mL)Baseline Week A1C (%)Baseline Week Stein P et al ADA Poster 537-P

Sitagliptin and 1,5 Anhydroglucitol Sitagliptin and 1,5 Anhydroglucitol Change from Baseline to Study End PlaceboSitagliptin 100mg Between Group Comparison LS95% ClLS95% ClLS Difference 95% Cl A1C 0.41(0.26, 0.5)-0.65(-0.80, 0.50)-1.05*(-1.27, -0.84) 1,5- AG -0.33(-1.05, 0.38)4.45(3.73, 5,17)4.78*(3.76, 5.80) *P value <0.001 Change in Postmeal Glucose Compared to A1C and 1,5-AG % Changes (Baseline to Study End) Absolute Change in 2 hour postmeal glucose (mg/dL) A1C Absolute % change 1,5-AG Absolute % change Sitagliptin 100mg % 83% Placebo %-7.3% Stein P et al ADA Poster 537-P

Miglitol and 1,5 AG Clinical Drug Trial – T. Yamanouchi (University of Teikyo) Placebo N=84 Miglitol N=158 1,5-AG (μg/mL)Baseline4.5 Week * A1C (%)Baseline7.3 Week * Comparison of Mean Values After 4 weeks, mean 1,5-AG was 9.0 ug/ml (p<0.001) compared to baseline *p<0.001

PPG more than 180 Mean Maximum PPG PPG more than 180 Mean Maximum PPG Before Breakfast After Breakfast Before Lunch After Lunch Before Dinner After Dinner Before Breakfast After Breakfast Before Lunch After Lunch Before Dinner After Dinner BG Patient 1: Age 75, female, type2 DM Patient2:Age 73, female, type2 DM Data from Dr Mori in Japan

Pramlintide and 1,5 AG Objective: To assess 1,5-AG as a marker of PPG control in Pramlintide-treated patients with type 1 diabetes (T1DM) Objective: To assess 1,5-AG as a marker of PPG control in Pramlintide-treated patients with type 1 diabetes (T1DM) Initial A1C levels to 8.0% Initial A1C levels to 8.0% Randomized to Pramlintide (n=18) or placebo (n=19) Randomized to Pramlintide (n=18) or placebo (n=19) Twenty-nine week study Twenty-nine week study Lush C et al.AACE 2007 Meeting (Poster 296)

Pramlintide and 1,5 AG Pramlintide and 1,5 AG Comparison of Changes in Values from Baseline to Week 29 Placebo (n=19)Pramlintide (n=18)Pramlintide vs. Placebo 2-hr PPG excursions /- 7.6 mg/dL / mg/dL P < Body Weight /- 0.7 kg-2.0 +/- 1.2 kgP < 0.01 A1C / %0.18 +/ % NS 1,5-AG ug/ml Percent Change / /- 8 % / /- 16 % P<0.05 P<0.01 Lush C et al.AACE 2007 Meeting (Poster 296)

Pramlintide and 1,5 AG Conclusions Pramlintide, as an adjunct treatment for T1DM patients on intensive insulin therapy, led to improved PPG and significant reduction in body weight. Pramlintide, as an adjunct treatment for T1DM patients on intensive insulin therapy, led to improved PPG and significant reduction in body weight. Despite similar reductions in A1C, the change in 1,5 AG levels was consistent with improvement in PPG control in pramlintide-treated subjects, as measured by SMBG. Despite similar reductions in A1C, the change in 1,5 AG levels was consistent with improvement in PPG control in pramlintide-treated subjects, as measured by SMBG. 1,5-AG, as a complement to A1C, may be a useful marker of PPG control. 1,5-AG, as a complement to A1C, may be a useful marker of PPG control. Lush C et al.AACE 2007 Meeting (Poster 296)

Exenatide and 1,5 Anhydroglucitol Objective: To assess 1,5-AG as a marker of PPG control in Exenatide-treated patients with type 2 diabetes (T2DM) Objective: To assess 1,5-AG as a marker of PPG control in Exenatide-treated patients with type 2 diabetes (T2DM) 144 Patients 144 Patients Initial A1C levels – 8.2 +/-1% Initial A1C levels – 8.2 +/-1% Randomized to Exenatide (5 or 10 ug) or placebo Randomized to Exenatide (5 or 10 ug) or placebo Thirty week study Thirty week study Kendall D, Holcombe J et al ADA & EASD 2007 Annual Meetings

Exenatide and 1,5 Anhydroglucitol Exenatide (5 ug) Exenatide (10 ug) 1,5-AG ug/ml Percent Change /- 0.6* / /- 0.6 ** / A1C % / /- 0.1 ** Correlations: Changes from baseline 1,5-AG vs. HbA1C: r = ; P < ,5-AG vs. fasting plasma glucose (FPG): r= -0.54; P < When grouped as HbA1C change tertiles patients with larger HbA1C changes from baseline had larger 1,5-AG changes from baseline. 1,5 AG changes were more robust than HbA1C changes Comparison of Changes in Values from Baseline to Study End * P < 0.05; ** P < 0.01 Kendall D, Holcombe J et al ADA & EASD 2007 Annual Meetings

Summary 1,5 Anydroglucitol appears to be a more robust indicator of glycemic excursions than either HbA1C or Fructosamine 1,5 Anydroglucitol appears to be a more robust indicator of glycemic excursions than either HbA1C or Fructosamine It is currently FDA-approved and clinically available; might a home kit be of clinical utility !!?? It is currently FDA-approved and clinically available; might a home kit be of clinical utility !!?? 1,5 AG responds more rapidly and sensitively than either HbA1C or Fructosamine 1,5 AG responds more rapidly and sensitively than either HbA1C or Fructosamine 1,5 AG may be a useful clinical adjunct and indicator for monitoring moderately well-controlled patients with diabetes 1,5 AG may be a useful clinical adjunct and indicator for monitoring moderately well-controlled patients with diabetes More clinical trials are necessary and underway to explore how effective this tool can be and to define other areas in which it may be limited or most helpful More clinical trials are necessary and underway to explore how effective this tool can be and to define other areas in which it may be limited or most helpful

1,5 Anhydroglucitol – Key References Dungan K et al Diabetes Care : McGill, J. et al. Circulating 1,5 Anhydroglucitol Levels in Adult Patients With Diabetes Reflect Longitudinal Changes of Glycemia: A U.S. Trial of the GlycoMark assay Diabetes Care 2004 Buse, J. et al. Serum 1,5-Anhydroglucitol (GlycoMark): A Short-Term Glycemic Marker. Diabetes Technology and Therapeutics 2003; 5: Dworacka M. et al. 1.5-Anhdro-D-glucitol: A Novel Marker of Glucose Excursions. International J. of Clinical Practice 2002; Supplement 129:40-44 (Eli Lilly Symposium) Kishimoto M. et al. 1,5-Anhydroglucitol Evaluates Daily Glycemic Excursions in Well-Controlled NIDDM. Diabetes Care 1995; 18(8): Matsumoto, K. et al. Effects of Voglibose on Glycemic Excursions, Insulin Secretion, and Insulin Sensitivity in Non-Insulin-Treated NIDDM Patients. Diabetes Care 1998; 21(2): Yamanouchi T. et al. Estimation of Plasma Glucose Fluctuation With a Combination Test of HbA1c and 1,5-AG. Metabolism 1992; 8: Yamanouchi T. et al. Clinical usefulness of serum 1,5-anhydroglucitol in maintaining glycaemic control. Lancet 1996;347: Yamanouchi T. et al. Post-load glucose measurements in oral glucose tolerance tests correlate well with 1,5-AG in subjects with impaired glucose tolerance. Clinical Science 2001;101:

Many Thanks To: Mary Kelly RN…Who is asked to do it all..and succeeds !! Mary Kelly RN…Who is asked to do it all..and succeeds !! John Buse John Buse Kathleen Dungan Kathleen Dungan Eric Button Eric Button Shuhei Kato Shuhei Kato

Questions ??