Hemoglobinopathies

Matthew M. Heeney, MD Associate Chief – Hematology Director – Sickle Cell Program Boston Children’s Hospital

THE FAIRMONT COPLEY PLAZA HOTEL BOSTON, MA. SEPTEMBER 23-28, 2018 Faculty Disclosure

Matthew M. Heeney, MD

Personal financial interests in commercial entities that are relevant to my presentation(s) or other faculty roles: • Astra Zeneca Consultant, Clinical Trial funding • Pfizer Clinical Trial funding • Micelle Biopharma Consultant, Clinical Trial funding • Novartis Consultant Objective

• Review the basic genetics of globins • Review – Qualitative disorders of • • Other Hemoglobinopathies – Quantitative disorders of hemoglobin • Thalassemias

3 Hemoglobin

• Four globular proteins (globins) – 2 α-like globins – 2 β-like globins • Four heme groups – One per globin chain

– Reversibly bind O2 (CO2, NO) • Hb synthesis must be balanced and coordinated • All components are labile and toxic – globins, heme, iron

4 Globin Genes

Nathan and Oski's Hematology of Infancy and Childhood, 7th Ed.

5 Globin Protein Synthesis

Nathan and Oski's Hematology of Infancy and Childhood, 7th Ed.

6 Hemoglobin tetramers

• are distinguished by globin composition: % nl adult

HbA α2β2 97-98%

HbA2 α2δ2 2-3%

HbF α2γ2 ≤2% S HbS: α2β 2 0% C HbC: α2β 2 0%

HbH: β4 0%

Hb Barts: γ4 0%

7 Global distribution of hemoglobin disorders • Distribution of thalassemia & sickle cell disease mirror worldwide distribution of malaria prior to 20th century. • Hypothesis (Haldane and others): heterozygous forms confer fitness - Thalassemia trait, sickle trait, G6PD etc.. protective against death from cerebral falciparum malaria. “Benefit” of trait

WHO - Control of Hereditary diseases. 1996 outweighs homozygous risk.

8 Disorders of Hemoglobin

Qualitative disorders – Structural variants or hemoglobinopathies – Production of abnormal globin chains

Quantitative disorders – Thalassemias – Decreased (imbalanced) production of normal globin chains

9 Qualitative Disorders of Hemoglobin SICKLE CELL DISEASE

10 1910

First description of sickle cell anemia in a West Indian dental student with “peculiar elongated and sickle-shaped” red blood cells.

James B. Herrick Presbyterian Hospital Chicago, Illinois

Herrick JB. Peculiar elongated and sickle-shaped red blood corpuscles in a case of severe anemia. Archives of Internal Medicine 1910. 6(5): 517-211

11 1949

Established sickle cell anemia as a genetic disease in which affected individuals have a different forms of Linus Pauling hemoglobin in their California Institute of Technology blood Pasadena, CA

Pauling L et al. Sickle Cell, A Molecular Disease. Science. 110 (2865): 543–548.

12 1956

Vernon Ingram and J.A. Hunt working at MIT, discovers the single change that causes hemoglobin to sickle

Sickle cell anemia became the 1st genetic Vernon Ingram disorder whose molecular basis was , known.

Ingram VM. A Specific Chemical Difference between Globins of Normal and Sickle-cell Anemia Hemoglobins" Nature. 178 (4537): 792–794

13 Epidemiology

Most common single gene disorder in African Americans 1/375 homozygous affected 1/12 are heterozygous carriers (~8%)

Also affects other ethnicities: India Middle East Hispanic

U.S. Prevalence: 80,000 – 120,000 U.S. Incidence: ~2000 live births annually

14 Sickle cell and malaria

Distribution of endemic malaria Distribution of sickle cell allele

Muntuwandi at en.wikipedia

15 Molecular Pathophysiology - Polymerization

The sickle mutation is a single amino acid substitution at position 6 of β globin.

Results in a hydrophobic region that is exposed in the deoxygenated state.

Adapted from Rotter MA et al. Biophys J. 2005 Oct;89(4):2677-84..

16 Bunn HF. NEJM. 1997 Sep 11;337(11):762-9

17 Delay time

• Delay time: period during which Hb is deoxygenated, but not yet polymerized • If passage through the capillaries exceeds the delay time, Hb will aggregate and initiate sickling.

18 Cellular Pathophysiology

Polymerization leads to: • Distortion of cell shape • Damage to RBC membrane • Abnormal permeability • Irreversible sickling Premature hemolysis = Anemia Impairment of RBC flow = Infarction

19 Hemolysis and Nitric Oxide depletion

• ↓ NO • Endothelial dysfunction

20 Dichotomization of Pathophysiology?

Adapted from Kato GJ et al. Blood Rev. 2007 Jan; 21(1): 37–47.

21 Manwani D and Frenette PS Blood. 2013:122(24):3892-3898

22 Vaso-occlusion Intra-cellular dehydration

Endothelial Activation Hemolysis Inflammation Reperfusion Injury

Nitric Oxide consumption

23 23 Sickle cell genotypes

Genotype Characteristics Hb SS Anemia : Hb 7 - 9 g/dL “Sickle cell anemia” Smear: Irreversibly sickled cells Electrophoresis: Hb S, Hb F

Hb SC Anemia: Hb 9 - 11 g/dL Smear: Sickle and target cells Electrophoresis: Hb S and Hb C

Hb S/β0 thalassemia Anemia: Hb 7 - 9 g/dL Smear: Microcytosis with sickle and target cells Electrophoresis: Hb S, Hb F

Hb S/β+ thalassemia Anemia: Hg > 10 g/dL Smear: Microcytosis with sickle and target cells Electrophoresis: Hb S, Hb F, and small %Hb A

24 Primary pathological processes in SCD

Vaso-occlusion Chronic Hemolysis • Pain episodes / “crises” • Cholelithiasis • Acute chest syndrome • Folate deficiency • Avascular necrosis • Cardiomegaly • Splenic sequestration • High-output heart failure (mostly in children) • Liver disease from iron overload (with repeated • Renal insufficiency transfusions) • Proliferative retinopathy • Non-hemorrhagic stroke (HbSC > HbSS) • Pulmonary hypertension • Excess fetal loss • Leg ulcers • Priapism

25 Infection in SCD

• Functional asplenia  Sepsis from encapsulated organisms

AA SS

• Osteomyelitis Salmonella and Staphylococcus aureus • Aplastic crisis (Parvovirus B19) • Hepatitis B and C (from transfusion)

26 Acute chest syndrome (ACS)

• Clinical syndrome of – Fever, – Respiratory symptoms (e.g. hypoxia, tachypnea) – New pulmonary infiltrate

27 Acute chest syndrome (ACS)

• 2nd most common cause of hospitalization. • Most common cause of death in sickle cell anemia. • Increased mortality with poorly controlled asthma • Prevent atelectasis – Control chest pain, incentive spirometry

• O2 and trial of bronchodilators • Often associated with atypical organisms (e.g. chlamydia, mycoplasma). – Empiric broad-spectrum antibiotics + macrolide • Simple or exchange transfusion may be life-saving

28 Stroke

Ohene-Frempong et al Blood. 1998; 91(1):288-94.

29 Stroke

Ohene-Frempong et al Blood. 1998; 91(1):288-94.

30 Stroke Risk Assessment Transcranial Doppler ultrasound (TCD) is a reproducible, non-invasive technique to predict 1° stroke risk in children < 170 cm/sec “Normal” 170 – 199 cm/sec “Conditional” > 200 cm/sec “Abnormal”

31 Stroke Risk Assessment

R.R. of stroke with Abnormal TCD = 44 (95% CI = 5.5 - 346)

Adams et al. Ann Neurol 1997; 42:699 -704

32 Transfusion therapy for stroke prevention

Effect of chronic Effect of chronic transfusion on 1° transfusion on 2° stroke rate in an stroke probability. asymptomatic patients with elevated TCD. (“STOP” Trial)

Effect on stroke rate Incident rate of stroke of discontinuing in SS children living in chronic transfusion in California (CA), asymptomatic before and after patients with publication of the elevated TCD from STOP Trial STOP Trial. (“STOP2” Trial)

Adapted from: Platt OS. Hematology 2006.. 2006:54-57 (ASH Education Book 2006)

33 Acute Transfusion therapy

• Goal – maximize O2-carrying capacity • Post –transfusion Hb should not exceed 10-11 g/dL

Platt OS. Hematology 2006.. 2006:54-57 • Transfuse phenotypically matched blood for minor antigens C, E and Kell • Allo-sensitization should be reassessed 1-3 months after episodic transfusions

34 Chronic transfusion therapy

Benefits: • Protection from CVA, ACS Risks • Iatrogenic Fe overload • Allo-sensitization • Unknown Infectious Risk

35 Perioperative management of SCD

• Preoperative period – Admit to hospital 12 to 24 hours before surgery for hydration. – Treat obstructive lung disease with bronchodilators. – Simple transfusion to a Hb 10-11g/dL except for minor operations. • Intraoperative period – Maintain oxygen therapy with pulse oximetry. – Maintain hydration. – Prevent hypothermia. – Monitor blood loss and replace blood when necessary. • Postoperative period – Pulse oximetry, IV fluids, incentive spirometry. – Monitor for development of acute chest syndrome.

36 Role of fetal hemoglobin in sickle cell disease

• Sickle cell phenotype only occurs after ~ 6months of age • Hemoglobin F levels are inversely correlated with disease severity / mortality • Biochemical evidence that gamma globin disrupts sickle globin polymerization

37 Hydroxyurea

• The only FDA approved agent for sickle cell disease. • Old drug – New indication. • first synthesized in 1869 • anti-neoplastic since the 1960’s • mechanism of action unclear. • ribonucleotide reductase inhibitor. • Mechanism • ↑ Hemoglobin F • ↓ HbS polymerization and hemolysis • ↑ Hemoglobin • ↓ White Blood Cells (“Side-Effect”?)

38 Hydroxyurea – Clinical Trials

MultiCenter Study of Hydroxyurea (MSH) (1992-1995) • Adults • ↑ Hemoglobin F; ↓ Hemolysis & Anemia; ↓ white blood cells • ↓ pain crises by 40%; ↓ acute chest syndrome by 50%; ↓ transfusions by 50%

Charache S et al. N Engl J Med 1995;332:1317-1322. MSH Follow-Up (1996-2001) • 40% reduction in mortality after 9 years of follow-up

Steinberg MH et al JAMA. 2003;289(13):1645-1651 MSH Follow-Up (1996-2010) • after 17.5 years of follow-up Steinberg MH et al. Am J Hematol 2010;85: 403–408.

39 MSH Follow-up – Cumulative Mortality

Steinberg MH et al. Am J Hematol 2010;85: 403–408.

40 Who should get Hydroxyurea? Indications: a) HbSS, HbSβ0 + ?? HbSC. b) Frequent pain crises? c) Acute chest syndrome?

https://www.nhlbi.nih.gov/sites/www.nhlbi.nih.gov/files/sickle-cell-disease-report.pdf

41 Chronic management of SCD

• Hydroxyurea • Folate • Consideration of chronic transfusion

• Renal and cardiac follow-up • Management of iron burden • Routine eye examination

42 Stem Cell Transplantation

- CVA risk - Tx related mortality - ACS risk - Infertility - inexorable accrual of - Secondary malignancy chronic end organ damage - cGVHD

Complications Complications of SCA of SCT

43 Investigative therapies - sickle cell disease • HbF induction with new agents • Shift oxygen dissociation curve – Small molecule allosteric modifier • Interference with endothelial/selectin adhesion – Anti-selectin • Reverse endothelial/vascular dysfunction – Omega fatty acids • Anti-platelet agents

– P2Y12 receptor antagonists • Gene therapy – Introduce normal or interfering globins

Zaidi AU, Heeney MM. Pediatr Clin North Am. 2018 Jun;65(3):445-464.

44 Sickle cell trait

• Present in about 8% of African Americans • Diagnosed by hemoglobin electrophoresis • Generally clinically silent – isosthenuria generally develops with age – occasional hematuria 2˚ to papillary necrosis – ?sudden death upon profound dehydration/exertion • No restrictions indicated for general activity • Screening requirement for NCAA athletes

45 Qualitative Disorders of Hemoglobin OTHER HEMOGLOBIN VARIANTS

46 Clinically significant hemoglobin variants

• Unstable: congenital Heinz body hemolytic anemia

• High O2 affinity: familial erythrocytosis

• Low O2 affinity: familial cyanosis

• M-hemoglobins: familial cyanosis

47 Unstable Hemoglobins

• Amino acid residues of globin oxidize and precipitate too readily • Form inclusions (Heinz bodies) that damage erythrocyte membrane. • May see abnormal “smeared” band on electrophoresis • Cause hemolytic anemia • Patients often benefit from splenectomy.

• e.g. Hb Koln; Hb Hasharon; Hb Zurich

48 Hemoglobins with altered oxygen affinity

Increased O2 affinity: • Hb Syracuse • HbF

Decreased O2 affinity: • Hb Kansas

Diagnosis: • May be ‘silent’ on electrophoresis.

• VBG with co-oximetry & P50.

49 Hemoglobins with altered oxygen affinity

P50 • P50 describes the affinity of a given Hb for oxygen.

• P50 is the PO2 at which the Hb is 50% saturated with oxygen. • As the P50 ↓, oxygen affinity ↑.

• Hb A 26.5 mmHg • Hb F 20 mmHg • Hb S 34 mmHg

50 Methemoglobinemia I • Methemoglobin: – Oxidation of hemoglobin iron from ferrous Fe2+ to ferric Fe3+ – Methemoglobin levels normally maintained at <3% by methemoglobin reductase (NADH-dehydratase)

• Methemoglobinemia results in increased O2 affinity, poor tissue oxygen delivery, and cyanosis. • Congenital methemoglobinemias – M-hemoglobins: globin mutations – Inherited defects in the methemoglobin reductase • Toxic methemoglobinemia – Nitrites, trinitrotoluene, aniline

51 Methemoglobinemia II

• Diagnosis of methemoglobinemia

– Unexplained cyanosis with normal PaO2 – Characteristic absorption peaks at 630 and 502 nm – Pulse oximetry gives inaccurate reading of 85% for blood with 100% methemoglobin – Blood is brownish color • Treatment – Methemoglobin levels >30%, patients start to have symptoms of oxygen deprivation – M hemoglobins and deficiencies of reductase usually do not need treatment. Can use oral methylene blue for cyanosis. – Emergency treatment: methylene blue 1-2 mg/kg IV infused rapidly, repeat at 1 mg/kg after 30 minutes if necessary

52 Quantitative Disorders of Hemoglobin THALASSEMIAS

53 Thalassemia: an international problem

Nathan and Oski's Hematology of Infancy and Childhood, 7th Ed.

54 Thalassemia: an international problem

• The most common global genetic disorder. • Thalassemia trait: >240 million people • Thalassemia intermedia syndromes: >2 million people • Hb E/thalassemia • 3-gene alpha thalassemia • Homozygous /compound heterozygous thalassemia syndromes: Hundreds of thousands

Nathan and Oski's Hematology of Infancy and Childhood, 7th Ed. • ~ 1,000 in N America

55 What are thalassemia syndromes?

• Hereditary anemias caused by mutation/deletion of one or more α or β globin genes. – α thalassemia caused by a defect in α globin gene(s) – β thalassemia caused by a defect in β globin gene(s) • Imbalanced expression of α and β globin leads to: – Decreased functional hemoglobin, resulting in anemia – Excess of α or β globins, cause membrane toxicity and intramedullary hemolysis of erythroid precursors • Anemia results in erythroid drive with “ineffective erythropoiesis” that stimulates increased iron absorption.

56 Thalassemia syndromes • Any imbalance of the “perfect” 4:2 ratio of α to β globin • β globin to α globin protein ratio determines severity

Nathan and Oski's Hematology of Infancy and Childhood, 7th Ed.

57 Quantitative Disorders of Hemoglobin β-Thalassemias

59 β-Thalassemia Mutations

IVS IVS β-globin gene

pre mRNA

mRNA

β+-thalassemia: Mutations cause defects in splicing or mRNA expression • Still allows some normal processing β0-thalassemia: Nonsense mutations in coding region, partial gene deletions • No normal processing of mRNA

60 β-Thalassemia Mutations

Nathan and Oski's Hematology of Infancy and Childhood, 7th Ed.

61 β-Thalassemia pathophysiology

Primary defect: Deficient synthesis of β-globin Consequence: Excess α-globin precipitates in erythroid cells

Bone marrow Blood Spleen

• Ineffective erythropoiesis • Anemia • Sequestration • Expansion of bone marrow spaces • Hypochromic red cells • Organ enlargement • Osteoporosis

62 β-Thalassemia trait

• Mild anemia and indirect hyperbilirubinemia • Microcytosis, hypochromia and poikilocytosis; RBC stippling, target cells.

• Increased HbA2 (>3.5%) or (rarely) HbF • Normal serum iron. • No clinical sequelae.

63 β-Thalassemia trait Hb electrophoresis

HbA (α2β2)

HbA2 (α2δ2)

β-thal trait Normal

64 Homozygous β-Thalassemia (“major”)

• Transfusion dependent anemia  allosensitization, blood borne pathogens. • Marked marrow expansion – Cortical thinning/Osteoporosis  fractures. • Iron overload (Transfusional/Absorption) – Hepatic  cirrhosis. – Cardiac  cardiac failure, arrhythmia. – Endocrine • Pancreas  diabetes. • Thyroid  hypothyroidism, growth failure. • Pituitary  delayed puberty, gonadal failure. • Extramedullary hematopoiesis/Hepatosplenomegaly • Hemolysis

65 β-Thalassemia Bone changes

Behrman: Nelson Textbook of Pediatrics, 16th ed

66 β-Thalassemia Bone changes

Kindness David Nathan

67 β-Thalassemia major Treatments

• Anemia / Ineffective erythropoiesis – Chronic monthly transfusion to keep Hb > 10g/dL – Novel HbF inducers (e.g. HDAC inhibitors – Differentiating agents (e.g. TGF-β ligand trap - Luspatercept) • Complications of transfusion – Endocrinopathies • Hormone replacement – Osteoporosis • Vitamin D • Osteoclast inhibitors (e.g. bisphosphonates) – Iron Overload • No physiologic way for the body to reduce iron. • Chelation therapy

68 How do we get the iron out?

Deferiprone (Ferriprox)

Deferoxamine (Desferal)

Deferasirox (Exjade/Jadenu)

Overnight SubQ infusion

69 Properties of Chelators

Property Deferoxamine Deferiprone Deferasirox Typical Dose 25 – 60 75 – 99 20 – 40 14-28 (mg/kg/day) Route SC or IV infusion Oral tablet Tablet for Film coated oral tablet, suspension sprinkles Dosing frequency Over 8 - 24 hours 3 times daily Once daily Adverse Effects Local reactions Gastrointestinal Gastrointestinal Audiologic Neutropenia/Agranul Rash Ophthalmologic ocytosis Rise in creatinine Bone abnormalities Arthralgia Elevated hepatic Elevated hepatic enzymes enzymes Liver Iron removal +++ ++ +++ Cardiac Iron Removal ++ +++ ++ (continuous infusion) Challenges Adherence with Weekly blood count GI side effects may parenteral route monitoring limit optimal dosing Adapted from Kwiatkowski JL, Ann N Y Acad Sci. 2016 Mar;1368(1):107-14

70 Goals of Chelation

• Maintain a “safe” level of iron – Prophylaxis: Prevent excess iron accumulation – Treatment • Remove excess stored iron • Reverse iron-related organ dysfunction • Detoxify iron through binding to non-transferrin bound, labile iron – Continuous chelator exposure is optimal

Adapted from: Kwiatkowski JL, Hematology Am Soc Hematol Educ Program. 2011;2011:451-8.

71 Chelation adjustments based on MRI results

UnacceptablyUnacceptably high high IntensifyIntensify chelation chelation ConsiderConsider combination combination chelation chelation

Moderately high Intensify chelation

Acceptable Continue current

Acceptable LowContinue –monitor current for Adverse effects

Low – monitor for adverse events

Ware, Kwiatkowski, Pediatric Clin N A, 2013 Survival by Availability of Chelation Therapy

Transfusion-Dependent Patients With Thalassemia Major

Year of Assessment 1985-97 1980-84 1.00 1975-79

1970-74

0.75

1965-69 0.50 1960-64

(N=1073) 0.25 Survival probability Survival P<0.00005

0 0 5 10 15 20 25 30

Age (y) Adapted from Borgna-Pignatti et al. Haematologica. 2004;89:1187. Survival Proportional to Chelation Adherence

100 80%-100% 60%-80%

75

40%-60% % Deferoxamine Compliance Thalassemia major 50 Torino 1965-1997

20%-40%

25 Cumulative survival Cumulativesurvival (%) 0%-20% 0 0 10 20 30 40 Years

Adapted from Gabutti and Piga. Acta Haematol. 1996;95:26. Monitoring Iron/Chelation Status • Serum ferritin – Advantages -Can be measured with every visit, widely available. – Disadvantages -Loose correlation with liver (body) iron. -Inaccurate with inflammation, abnormal liver fxn. • Liver biopsy – Disadvantages -Invasive. • MRI – Can interrogate many organs (liver, heart, pancreas, etc.)

– Hepatic R2 or R2* Obtain annually, reported as mg/g dry weight liver. • LIC > 15 mg/g dry weight predict poor prognosis

– Cardiac T2* -Obtain annually, reported in ms (lower is worse) • < 20 ms predicts increased risk of arrhythmias/cardiac failure • < 6 ms predicts high risk cardiac failure in next year

St. Pierre et al, Blood 2005;105:855; Wood, Blood 2005 106:1460; Saliba AN et al, J Blood Med. 2015; 6: 197–209.

75 Hepatic and Cardiac Iron by T2* MRI

Andy Powell, MD - Boston Children’s Hospital β-Thalassemia genotypes/phenotype

S. L. Thein, ASH Hematology 2005

77 β Thalassemia Intermedia Syndromes

• Wide phenotypic variability. • “Strong” β+ allele(s) may allow patient to be transfusion independent. • But brisk and ineffective erythropoiesis persists with all the end-organ damage similar to thalassemia major, but later than in life (e.g. growth failure, osteoporosis, exuberant iron absorption, etc.). • Some of the sickest North American thalassemia patients are adults with thalassemia intermedia. Hemoglobin E - A Special Case

• βE is a mutated β globin that is both mildly unstable and produced at decreased rate because of a splicing defect.

Normal β-globin IVS-1 IVS-2

βE IVS-1 IVS-2

79 Hemoglobin E - A Special Case • HbE trait (βNβE): mild microcytosis, no anemia • HbE disease (βE βE): + microcytosis, mild anemia • HbE/beta-thalassemia (β0 βE): thalassemic phenotype (intermedia or major)

• HbE trait is present in 15-30% of the population in regions of Laos, Cambodia, Vietnam, and southern China

80 Investigative therapies - β Thalassemia

• Reduction of ineffective erythropoiesis – JAK2 inhibitors and TGF-β ligand traps • Decrease iron overload – hepcidin agonists, erythroferrone inhibitors • Reduction of Oxidative stress – activators of Foxo3, inhibitors of HO-1 • Gene therapy – Introduce normal or interfering globins

Makis A et al. Am J Hematol. 91:1135–1145, 2016.

81 Quantitative Disorders of Hemoglobin α-Thalassemia

82 α-Thalassemia Pathophysiology

• Primary defect: Deficient synthesis of α-globin. • Consequence: Excess β-globin leads to formation of

β globin tetramers (β4 = HbH). • HbH is relatively unstable & soluble, but under certain conditions (e.g. oxidant stress, older RBCs) HbH precipitates leading primarily to extravascular hemolysis in the spleen. • Concentrated in Southeast Asia, Malaysia, & southern China.

83 α-Thalassemia Mutations

• α-thalassemia is most frequently a result of deletions involving one or both α globin genes. • Less commonly caused by non-deletional defects.

Nathan and Oski's Hematology of Infancy and Childhood, 7th Ed.

84 α-Thalassemia

• 4 primary clinical conditions of increasing severity • Genotype-phenotype correlation has been only partly clarified Genotype Lab feature Severity Silent carrier α-/αα Normal 0 α-thalassemia trait α-/α- or αα/-- ↓ MCV, ↓MCH, 1+ minimal anemia Hb H disease α-/-- ↓↓ - ↓↓↓ MCV and 2+ MCH, hemolytic anemia (7-10g/dL), mild jaundice, Hb Barts/HbH Hydrops fetalis --/-- Profound anemia, 4+ Hb Barts/HbH

85 α-Thalassemia Diagnosis

• Silent carrier state (- α / α α) can only easily be diagnosed in newborn period by presence (1–2%) of

HbBarts (γ4)on Newborn screen and RBC indices and Hb electrophoresis are normal in the adult. • α-thalassemia trait (- - /αα or - α / - α) show greater increase (5– 6%) of HbBarts on Newborn screen and microcytic hypochromic RBC indices, with normal

HbA2 and HbF in the adult. • HbH Disease (- - / - α) microcytic hypochromic

anemia Hb (7-10g/dL), reduced (<2%) HbA2 and variable amounts of HbH (up to 30%)

86 Thalassemia Peripheral Blood Smear

Hb H disease: α-/-- 87 α-Thalassemia Management

• Silent carriers and α-thalassemia trait generally do not need any treatment. • HbH disease management is influenced by its marked phenotypic variability. – Most individuals with HbH disease are clinically well and require no treatment. – Avoid oxidant drugs (same as G6PD deficiency). – Hemolytic or aplastic crises may require transfusion support.

89 Hemoglobinopathies overview

LCR ε Gγ Aγ δ β Chr 11

HS -40 ζ2 ζ1 α2 α1 θ Chr 16 β-thalassemia Genotype Lab feature Severity Hb A: α2β2 0 + Hb A2: α2δ2 β-thal trait β /β or β /β ↓MCV 1+ Hb F: α2γ2 Hb S: codon 6 β-globin mutation β-thal β+/β+ or β0/β+ Moderate anemia 2+ Hb C: different codon 6 β-globin mutation intermedia Hb E: β-globin splice variant - thal equivalent 0 0 Hb H: β4 β-thal major β /β Severe anemia 4+ Hb Barts: γ4

Unstable: Heinz body hemolytic anemia α-thalassemia High O affinity: familial erythrocytosis 2 Genotype Lab feature Severity Low O2 affinity: familial cyanosis M-hemoglobins: familial cyanosis Silent carrier α-/αα Normal 0

α-thal trait α-/α- or αα/-- ↓MCV 1+ Sickle cell genotypes: SS SC Hb H disease α-/-- Hemolysis, HbH 2+ SD S/β thal Hydrops fetalis --/-- Hb Barts 4+

90 Take Home Messages

• Quantitative disorders of globin synthesis are Thalassemias – Imbalanced production of globins result in ineffective erythropoiesis/hemolysis. – Wide phenotypic spectrum, with severe forms resulting severe anemia, bone changes and endocrinopathies. These complications can be limited by adequate transfusions and chelation therapy. • Qualitative disorders disorders of globin synthesis are Hemoglobinopathies – Production of abnormal globin proteins which can be

unstable, have altered O2 affinity or other presentations.

91 Acknowledgements

Ben Ebert, MD, PhD Ellis Neufeld, MD, PhD

92 Supplemental References

• Rund D, Rachmilewitz E. Beta-thalassemia. N Engl J Med. 2005; 353:1135-46. Galanello R, Cao A. Alpha Thalassemia. Genet Med. 2011; 13:83–88. • Neufeld, EJ. Update on Iron Chelators in Thalassemia. Hematology Am Soc Hematol Educ Program. 2010: 451-455. Brittenham, GM. Iron-Chelating Therapy for Transfusional Iron Overload. N Engl J Med 2011; 364:146-156. Marsella M et al. Transfusional iron overload and iron chelation therapy in thalassemia major and sickle cell disease. Hematol Oncol Clin North Am. 2014 Aug;28(4):703-27. • Steinberg MH, Nagel RL. Unstable Hemoglobins, Hemoglobins with Altered Oxygen Affinity, Hemoglobin M, and Other Variants of Clinical and Biological Interest. In: Steinberg MH, Forget BG, Higgs DR, Weatherall DJ, editors. Disorders of Hemoglobin - Genetics, Pathophysiology, and Clinical Management. Cambridge: Cambridge University Press; 2009. p. 589-606. Globin Gene Server - HbVar Database (http://globin.cse.psu.edu/) • Management of Sickle Cell Disease (http://www.nhlbi.nih.gov/health/prof/blood/sickle/sc_mngt.pdf).Yawn BP et al. Management of sickle cell disease: summary of the 2014 evidence-based report by expert panel members. JAMA. 2014 Sep 10;312(10):1033-48

93 Background

• Deoxygenation results in non-covalent polymerization of hemoglobin, distortion of erythrocyte shape, and unfavorable rheological properties leading to vaso-occlusion. • Leukocytes, platelets, coagulation cascade also contribute to VOC. • Hypoxia-Reperfusion leads to endothelial activation, inflammatory state and further VOC. Dover GJ, Heeney MM. Sickle Cell Disease. Nathan and Oski’s Hematology of Infancy and Childhood. 2008; 949-1014 Platt OS. Sickle cell anemia as an inflammatory disease. J Clin Invest. 2000 August 1; 106(3): 337–338.

• SCD patients have chronically elevated levels of multiple inflammatory mediators • This milieu is attractive for use of anti-inflammatories / corticosteroids Black LV, Smith WR. Are Systemic Corticosteroids an Effective Treatment for Acute Pain in Sickle Cell Disease? ASH Education Book 2010:416-41

94 Background

• Acute chest syndrome (ACS) – 1. New CXR infiltrate 2. Fever 3. Pulmonary symptom(s) – Significant cause of morbidity and a leading cause of mortality.

Johnson CS. The acute chest syndrome. Hematol Oncol Clin North Am. 2005 Oct;19(5):857-79 • Etiology is multifactorial – Infectious (bacterial, atypical bacteria, viral) – Infarctive (VOC, atelectasis) – Inflammatory (asthma, fat embolism) Vichinsky EP et al . Causes and outcomes of the acute chest syndrome in sickle cell disease. National Acute Chest Syndrome Study Group. N Engl J Med. 2000 Jun 22;342(25):1855-65 • Asthma and ACS – higher prevalence of airway hyper-responsiveness and obstructive lung disease than expected. Overlapping or co-morbid conditions? – Trial of bronchodilators often recommended in ACS. Nordness ME, et al. Asthma is a risk factor for ACS and cerebral vascular accidents in children with sickle cell disease. Clin Mol Allergy. 2005; 3: 2. Field JJ, DeBaun MR. Asthma and sickle cell disease: two distinct diseases or part of the same process? ASH Education Program. 2009:45-53.

95