Genetics of the Hemoglobinopathies & Newborn Screening for the Hemoglobinopathies

1. Genetics of the Hemoglobinopathies & Newborn Screening for the Hemoglobinopathies 张咸宁 zhangxianning@zju.edu.cn Tel：13105819271; 88208367 Office: C303, Teaching Building 2016/03

2. Required Reading Nussbaum RL, et al. Thompson & Thompson Genetics in Medicine, 8th ed. Elsevier，2016，pp195-214； ● Clinical Case Studies-42. Sickle Cell Disease，44. Thalassemia

3. Part I. Genetics of the Hemoglobinopathies血红蛋白病

4. Learning Objectives • To review the normal structure-function relationships of Hb and expression of globin genes • To examine the hemoglobinopathies as disorders of Hb structure, or α- or β-globin gene expression • To explore the influences of compound heterozygosity and modifier genes on hemoglobinopathy phenotypes

5. Compound heterozygote（复合杂合子）：An individual, or a genotype, with two different mutant alleles at the same locus • Compound heterozygote ≠homozygote≠ Double heterozygote

6. Molecular Disease（分子病） A disease in which there is an abnormality in or a deficiency of a particular molecule, such as hemoglobin in sickle cell anemia 酶蛋白病（enzymopathy）：Any of a group of congenital disorders caused by an inherited defect in a single specific enzymethat results in a disruption or abnormality in a specific metabolic pathway

7. Hemoglobinopathies血红蛋白病 • Disorders of the human hemoglobins • The most common single gene disorders in the world • WHO: 5% of the world’s population are carriers for clinically significant hemoglobinopatihies • Well understood at biochemical and molecular levels

8. HbA: α2β2 / HbF: α2γ2 • Globular tetramer • MW 64.5 kD • α-Chain • 16p13.3 • Polypeptide length of 141 amino acids • β-Chain • 11p15.4 • Polypeptide length of 146 amino acids

9. Globin Tertiary Structure

10. Gene cluster基因簇: A group of adjacent genes that are identical or relatedPseudogene假基因: DNA sequence homologous with a known gene but is non-functional

13. Developmental Expression of Globin Genes and Globin Switching（发育时珠蛋白基因的表达和珠蛋白的转换）

15. Globin Gene Developmental Expression and Globin Switching • Classic example of ordered regulation of developmental gene expression • Genes in each cluster arranged in • Same transcriptional orientation • Same sequential order as developmental expression • Equimolar production of α-like and β-like globin chains

16. The normal human Hbs at different stages of development

18. Clinic Disease: Influences of Gene Dosage and Developmental Expression • Dosage • 4 - vs. 2 -globin alleles per diploid genome • Therefore, mutations required in 4 -globin alleles compared with 2 -globin alleles for same 100% loss of function • Ontogeny（个体发育） •  expressed before vs.  expressed after birth • Therefore, -chain mutations have prenatal consequences, but -chain mutations are not evidenced even in the immediate postnatal period

19. Genetic disorders of Hb 1. Structural variant: alter the globin polypeptide without affecting its rate of synthesis 2. Thalassemia: reduced rate of production of one or more globin chains 3. Hereditary persistence of fetal hemoglobin (HPFH. 遗传性胎儿血红蛋白持续症) : a group of clinically benign conditions, impairing the perinatal switch from γ- to β-globin synthesis

20. There are >400 structural variants

23. HbS is the first variant to be discovered (1949) Its main reservoir is Central Africa where the carrier rate approximates 20%. (Heterozygous advantage) Approximately 8% of African-Americans will carry one sickle gene.

24. Heterozygote Advantage • Mutant allele has a high frequency despite reduced fitness in affected individuals OR: • Heterozygote has increased fitness over both homozygous genotypes e.g. Sickle cell anemia.

26. Thalassemia: An imbalance of globin-chain synthesis • Hb synthesis characterized by the absence or reduced amount of one or more of the globin chains of Hb • α-thalassemia • β-thalassemia

28. Varius forms of α-Thalassemia

29. Hb Bart’s (hydrops fetalis水肿胎儿)

32. Hb Bart’s胎儿水肿综合征最常见于中国和东南亚

33. β-thalassemia:underproduction of the β-chain. ●β-thal trait (β+/ β orβ0 /β) : .asymptomatic (β+:reduced;β0:absent) ●β-thal intermedia (β+/ β+ ): . moderate anemia ●β-thal major (β0 /β0 orβ+/β0 or β+/ β+ ) : . severe anemia during the first two years of life . hepatosplenomegaly . growth failure . jaundice . thalassemic facies

35. Thalassemias can arise in the following ways: • One or more of the genes coding for hemoglobin chains is deleted. • 2. A nonsense mutation that produces a shortened chain. • 3. A frameshift mutation that produces a nonfunctional chain. • 4. A mutation may have occurred outside the codingregions.

36. β-globin gene andβ-thalassemia

37. Part II. Newborn Screening for the Hemoglobinopathies

38. Learning Objectives • To review the evolving principles of newborn screening • To examine newborn screening (NBS) for the hemoglobinopathies • To understand the appropriate response to a positive hemoglobinopathy NBS • To appreciate the role of clinical follow-up for the hemoglobinopathies

39. Population-Based Screening

40. Genomic Medicine • Principles:“5P” Medicine • Change from current paradigm with emphasis on acute intervention • Will rely on strategies from preventive medicine and public health

41. Genetic Screening • Population-based approach to identify individuals with certain genotypes known to be • Associated with a genetic disease, or • Predisposition to a genetic disease • Disorder targeted may affect • Individuals being screened, or • Their descendents

42. Objective of Population Screening • To examine all members of the population designated for screening • Carried out without regard for family history • Should not be confused with testing for affected individuals or carriers within families ascertained because of a positive family history

43. Genetic Screening • Important public health activity • Will have increasingly significant role with availability of more and better screening tests for • Genetic diseases • Diseases with an identifiable genetic component • Critical strategic hurdle for implementation • Venue in which to capture 100% of target population

44. Principles of Newborn Screening (NBS)

45. NBS • Public health governmental programs • Population screening for all neonates • Intervention • Prevents or at least ameliorates consequences of targeted disease • Cost-effective • Controversial • Not simply a test, but a system

46. Criteria for Effective NBS Programs • Treatment is available. • Early institution of treatment before symptoms become manifest has been shown to reduce or eliminate the severity of the illness. • Routine observation and physical examination will not reveal the disorder in the newborn – a test is required.

47. Criteria for Effective NBS Programs • A rapid and economical lab test is available that is highly sensitive (no false- negatives) and reasonably specific (few false-positives). • The condition is frequent and serious enough to justify the expense of screening; that is, screening is cost-effective.

48. Criteria for Effective NBS Programs • The societal infrastructure is in place • To inform the newborn’s parents and physicians of the results of the screening test, • To confirm the test results, and • To institute appropriate treatment and counseling.

49. Evolving NBS Criteria • Treatment available – Not always • Example: Tandem Mass Spectrometry (MS/MS)串联质谱法 • Analogy: Childhood cancer (75% survival) and protocol-driven iterative improvements • Pre-symptomatic treatment effective – No • Example: For rarer hemoglobinopathies may not have accurate knowledge of natural hx

50. Evolving NBS Criteria • Clinical ascertainment not effective, so test required – Not always • Example: G6-PD deficiency and kernicterus（核黄疸） • Problem: Clinical ascertainment is never 100% • Rapid and effective lab test available – No • Example: Severe combined immunodeficiency (SCID) • Problems: Limited federal funding for test development until recently, and low cost and margin limit corporate interest