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HbA1c : glycosylated hemoglobin

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A presentation to physicians on using Hba1c in clinical practice.

A presentation to physicians on using Hba1c in clinical practice.

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  • 1. Glycosylated hemoglobin HbA1c Mathew John MD, DM, DNB Endocrinologist Providence Endocrine & Diabetes Specialty Centre
  • 2. Some terms • A1c : Glycated hemoglobin = glycosylated hemoglobin= glycohemoglobin(US) • IFCC: International Federation of Clinical Chemistry • NGSP: National Glycohemoglobin Standardisation Programme • DCCT : Diabetes Control and Complications Trial • ADAG : A1c derived average glucose
  • 3. Hemoglobin HbA0(α2 ß2) HbA2(α2δ2) HbF(α2γ2) 90 % HbA1 HbA1c Non ezymatically glycosylated form of human hemoglobin, taking place under physiological conditions, at a specific site on the protein
  • 4. Terminology • Hb: hemoglobin • HbA1: is a series of glycated variants resulting from attachment of various carbohydrates to N terminal valine of Hb • Glycation results in increased negative charge and hence runs fast on electrophoresis systems Pickup & Williams , Textbook of Diabetes
  • 5. GHb: glycated hemoglobin 1. HbA1a1: fructose 1,6 diphosphate N terminal valine 2. HbA1a2: glucose 6 phosphate N terminal valine 3. HbA1b: unknown carbohydrate N terminal valine 4. HbA1c: (60-80%): attachment of glucose to N terminal amino acid valine of the beta chain of hemoglobin Total glycated Hb: HbA1c+ sugar Non N terminal sites
  • 6. Amadori rearrangement of glucose molecule.
  • 7. Relationship of HbA1C to Risk of Microvascular Complications Diabetes Control and Complications Trial (DCCT) 15 13 Retinopathy Relative Risk (%) Nephropathy 11 Neuropathy 9 Microalbuminuria 7 5 3 1 6 7 8 9 10 11 12 HbA1C (%) Skyler JS. Endocrinol Metab Clin North Am. 1996;25:243-254.
  • 8. Factors affecting HbA1c Falsely elevated values • HbF or HbG • Uremia ( BUN > 85 mg/dl) • Hypertriglyceridemia( cation exchange +, EP-) • Alcohol • High bilirubin( cation exchange+, HPLC+) • Aspirin • Splenectomy, Aplastic anemia
  • 9. Factors affecting HbA1c Falsely low HbA1c • HbC, HbS • Hemolytic anemias • Pregnancy • Acute/ Chronic blood loss • Vitamin E/C • Dapsone • Severe nephropathy ( shorten RBC survival) Glycated hemoglobin monitoring BMJ 2006 ; 333;586-8
  • 10. Age specific targets Age HbA1c target <6 years 7.5 –8.5 % 6-12 years < 8% 13-19 years <7.5 % Silverstein J, Diabetes Care; 28;2005
  • 11. Current HbA1c recommendations Normal IDF ADA AACE A1c* <6% <6.5% <7% <6.5% Preprandial <100 <110 90-130 <110 Postprandial <140 <155 <180 <140
  • 12. What does HbA1c represent ?
  • 13. Patients with variable diurnal profiles can have the same A1c Roger Mazze DIABETES TECHNOLOGY & THERAPEUTICS Volume 10, Supplement 1, 2008
  • 14. Relationship between FPG, PPG and HbA1c 80 Postprandial Fasting Hyperglycemia Contribution (%) 60 40 20 0 1 2 3 4 5 (<7.3) (7.3-8.4) (8.5-9.2) (9.3-10.2) (>10.2) Monnier L, Diabetes Care 2003;26 HbA1c quintiles
  • 15. ADAG study A1c Derived Average Glucose • Define the mathematical relationship between A1c and average glucose levels • 507 subjects : 268 with type 1 diabetes, 159 with type 2 diabetes and 80 non diabetic subjects • A1c at end of 3 months compared with average glucose during the previous 3 months • From 2 day CGMS 4 times+7 point SMBG 3 times/week Nathan D Diabetes Care 31:1-6, 2008
  • 16. ADAG study • Approx 2700 values/subject in 3 months • Linear regression analysis between A1c and AG values provided the tightest correlations AG (mg/dl) = 28.7X A1C-46.7 ( R2 0.84, P 0.0001)
  • 17. ADAG study Estimated average glucose ( e AG) mg/dl mmol/L DCCT 135 170 205 240 275 310 Nathan D Diabetes Care 31:1-6, 2008
  • 18. Hba1c represents more recent sugars
  • 19. Mean blood sugars vs. ADAG ADAG MBG ADAG : A1c Derived Average Glucose
  • 20. Methods of measuring HbA1c • Ion exchange chromatography : low pressure HPLC • Electrophoretic methods • Immunoturbimetric methods • Affinity methods • Chemical methods: e.g thiobarbituric method • Electrospray iontophoresis • Mass spectroscopy • Reversed phase HPLC
  • 21. Can we use HbA1c for diagnosis of diabetes ?
  • 22. Cut offs Fasting plasma glucose cut offs for definition of IGT and DM Normal IGT Type 2 diabetes 100 mg/dl 126 mg/dl
  • 23. Diagnosis of diabetes • Diagnosis of diabetes has always been glucose centric : based on FBS, 2 hr post glucose , RBS • National Diabetes Data Group (NDDG) 1979 : relied on distributions of glucose levels • Based on their association with decompensation to “overt” or symptomatic diabetes FPG > 140 mg/dl PPG > 200 mg/dl
  • 24. 1997 1997, the Expert Committee on the Diagnosis and Classification of Diabetes Mellitus refocused attention on the relationship between glucose levels and the presence of long-term complications as the basis for diagnosis of diabetes The Expert Committee on the Diagnosis and Classification of Diabetes Mellitus. Report of the Expert Committee on the Diagnosis and Classification of Diabetes Mellitus. Diabetes Care 1997;20:1183– 1197
  • 25. Expert Committee on the Diagnosis and Classification of Diabetes Mellitus Committee recommended that the FPG cut point be lowered to 126 mg/dl (7.0 mmol/l) so that this cut point would • Represent a degree of hyperglycemia that was “similar” to the 2HPG value and diagnosis with either measure would result in a similar prevalence of diabetes in the population • Introduced the concept of IFG and IGT The Expert Committee on the Diagnosis and Classification of Diabetes Mellitus. Report of the Expert Committee on the Diagnosis and Classification of Diabetes Mellitus. Diabetes Care 1997;20:1183– 1197
  • 26. Pathophysiologic cut offs Looked at 3 studies which compared glycemia to risk of retinopathy • Egyptian population (n 1,018) • Pima Indians (n 960), • U.S. National Health and Nutrition Examination Survey (NHANES) population (n 2,821)
  • 27. FPG/PPG /HbA1c vs. Retinopathy U.S. National Health and Nutrition Examination Survey (NHANES) population (n 2,821)
  • 28. Current use of HbA1c • Monitor long term glycemic control • Adjust therapy • Assess the quality of diabetes care • Predict the risk for the development of complications
  • 29. HbA1c for diagnosis of diabetes • HbA1c correlates with retinopathy • There was a stronger correlation between A1C and retinopathy than between fasting glucose levels and retinopathy • Similar correlation between A1c and Retinopathy has been seen in DCCT/ UKPDS trials • 1997 Expert Committee recommended against using A1C values for diagnosis in part because of the lack of assay standardization
  • 30. 2009 :International Expert Committee Report on the Role of the A1C Assay in the Diagnosis of Diabetes DIABETES CARE, VOLUME 32, NUMBER 7, JULY 2009 Recommends that HbA1c be adopted as one of the diagnostic criteria for diabetes
  • 31. What has happened between 2003 and 2009 ? Advances in instrumentation and standardization, the accuracy and precision of A1C assays at least match those of glucose assays International Expert Committee Report on the Role of the A1C Assay in the Diagnosis of Diabetes. DIABETES CARE, VOLUME 32, NUMBER 7, JULY 2009
  • 32. New reference standards • International Federation of Clinical Chemistry (IFCC) • Measures “ pure” A1c • Pure A1c: N-[1deoxylfructos-1-yl]) hemoglobin beta chain, abbreviated as DOF hemoglobin • Expressed as mmol/mol of Hb • HbA1c of 5% would now be about 33 mmol/mol, and an 8% A1C would be about 58 mmol/mol.
  • 33. Pitfalls with glucose measurement • The measurement of glucose itself is less accurate and precise than most clinicians realize • 41% of instruments have a significant bias from the reference method that would result in potential misclassification of 12% of patients • Potential preanalytic errors owing to sample handling • Lability of glucose in the collection tube at room temperature International Expert Committee Report on the Role of the A1C Assay in the Diagnosis of Diabetes. DIABETES CARE, VOLUME 32, NUMBER 7, JULY 2009
  • 34. Advantages of HbA1c • HbA1c is stable after collection • New reference method to calibrate all A1C assay instruments should further improve A1C assay standardization in most of the world between- and within-subject • Coefficients of variation have been shown to be substantially lower for A1C than for glucose measurements • The variability of A1C values is also considerably less than that of FPG levels, with day-to-day within-person variance of 2% for A1C but 12–15% for FPG International Expert Committee Report on the Role of the A1C Assay in the Diagnosis of Diabetes. DIABETES CARE, VOLUME 32, NUMBER 7, JULY 2009
  • 35. Advantages of HbA1c • Convenience for the patient and ease of sample collection for A1C testing • Relatively unaffected by acute (e.g., stress or illness related) perturbations in glucose levels International Expert Committee Report on the Role of the A1C Assay in the Diagnosis of Diabetes. DIABETES CARE, VOLUME 32, NUMBER 7, JULY 2009
  • 36. Cut off of HbA1c for diagnosis of diabetes • Cut offs at which the prevalence of retinopathy increases • NHANES data and DETECT 2 study
  • 37. DETECT 2 study Prevalence of retinopathy by 0.5% intervals and severity of retinopathy in participants aged 20–79 years. NPDR, nonproliferative diabetic retinopathy. Adapted with permission from (S.C., personal communication). 19,000 subjects from nine countries The glycemic level at which the prevalence of “any” retinopathy begins to rise above background levels and for the more diabetes-specific “moderate” retinopathy, was 6.5% when the data were examined in 0.5% increments
  • 38. Cut off of HbA1c • A1C level of 6.5% is sufficiently sensitive and specific to identify individuals who are at risk for developing retinopathy and who should be diagnosed as diabetic • A1C level is at least as predictive as the current FPG and 2HPG values.
  • 39. Should we use of HbA1c to diagnose diabetes in our set up ?
  • 40. Limitations • Cost may preclude routine use FBS + PPBS: Rs. 60/ - HbA1c : Rs. 275/ - • Standardized methods and instrumentation POC instruments • Hemoglobin variants • Any condition that changes red cell turnover, such as hemolytic anemia, chronic malaria, major blood loss, or blood transfusions • A1C levels appear to increase with age
  • 41. Limitations • Discordance with standard diagnostic criteria • The prevalence of diabetes in some populations may not be the same when diagnosis is based on A1C compared with diagnosis with glucose measurements, and one method may identify different individuals than the other. • Ethnic variations in HbA1c at same glucose levels exists
  • 42. “ Prediabetes” • Once A1c is used to diagnose diabetes, “ prediabetes” or IGT/ IFG may be obsolete • HbA1c between 6 and 6. 5 % : higher risk for developing diabetes more effective interventions
  • 43. Practical considerations • POC instruments are not to be used to make this diagnosis • Always confirm using the same tests • Intermethod variability is reported to still be a potential source of inaccuracy
  • 44. Point of care instruments • DCA Vantage • Nycocard • In2it (BioRad) • A1cNow( Bayer)
  • 45. Methods for HbA1c The better and best Electrospray iontophoresis Mass spectrometry HPLC CV : 2-3 % Immunoassay methods CV 5-6 % •Point of care ( POC) Instruments • Colorimetry
  • 46. BioRad D10 • A1C quantitation in the presence of HbS, HbC and HbF • Optimized to minimize interference from carbamylation, lipemia and labile A1C • Traceable to the IFCC reference method • NGSP Certified
  • 47. Words of wisdom • HbA1c and mean glucose corroborate abnormal glucose metabolism, but it requires self monitoring ( or CGMS) to detect the location and magnitude of the abnormalities • HbA1c and SMBG should be considered together, with each complementing the information provided by the other Peacock I J Clin Path 1984
  • 48. HbA1c for all patients ? Added Glimiperide 1 month 2 mg/day HbA1c Started On Metformin Glargine 14 1000 mg/day units/day 3 months
  • 49. HbA1c for all patients ? Glargine dose 18 units/day HbA1c Glargine 14 units/day Metformin 1000 mg/day Glimiperide 2 mg/day
  • 50. 2010 Consensus Statement on the Worldwide Standardization of the HbA1C Measurement • HbA1c test results should be standardized worldwide • The IFCC reference system for HbA1c represents the only valid anchor to implement standardization • HbA1c results are to be reported by clinical laboratories worldwide in SI units (mmol/mol, no decimals) and derived NGSP units (%, one decimal) DIABETES CARE, VOLUME 33, NUMBER 8, AUGUST 2010
  • 51. High Commission Labs Pvt Ltd, 69, Park Road, NY Name: Kuttapan J, 45 yrs Male HbA1c: 8.0% ( Biorad D10 variant 11) eAG2: 183 mg/dl IFCC HbA1c3 : 58mmol/mol 1.Biorad D10 is a DCCT aligned method 2.e AG are derived from ADAG study by Nathan et al. Nathan D Diabetes Care 31:1-6, 2008 3.IFCC A1c is estimated from a regression equation . From Jeppsson J-O, Clin Chem Lab Med 2002;40:78-8
  • 52. Thank you