Hemoglobinopathies & Inherited Disorders of Coagulation

red blood cells

(erythrocyte)

• carries oxygen (hemoglobin)

• biconcave discoid in shape

• no nucleus

• mitochondria

• lifespan: 120 days

anemia

• having low red blood cells or less than normal hemoglobin

• hemolytic anemia: when RBCs break down

hemoglobin

• 4 subunits: two alpha globin chains and two beta (or beta-like) globin chains

• a, β, γ, δ, ε and ζ

• carry and deliver oxygen from lungs to tissues

heme iron

• oxygenation and deoxygenation occurs at the heme iron

• heme iron is covalently linked to a histidine at specified residues on alpha and beta globin chains

  • mutations → result in unstable hemoglobin

qualitative disorders

• point mutations, deletions, fusion, or elongation of Hb chains

• >400

• ½ are clinically significant

• ex.) sickle cell, hemoglobin C, hemoglobin E

• make hemoglobin tetramer unstable (hemolytic anemia)

• altered oxygen transport

  • increased or decreased oxygen affinity

  • formation of methemoglobin (incapable of binding oxygen)

quantitative disorders

• imbalance in alpha-beta chain ratio

• ex.) thalassemia, hemoglobin Barts, hemoglobin H

sickle cell disease

• Gene(s): HBB

• Inheritance Pattern: autosomal recessive

• Features:

  • hemoglobin SS

  • carrier rate is 10% in African Americans

  • deoxygenation of RBCs → polymerization of hemoglobin → RBCs in a rigid, sickle shape

  • sickle cells are more likely to be damaged and to block small blood vessels

  • affected individuals have chronic pain, fatigue, weakness, heart and lung problems, complications during pregnancy

• Treatments:

  • hydration, aggressive fever control, pain management

  • pharmacologic: hydroxyurea, L-glutamine

  • chronic simple or exchange transfusions (risk for iron overload and need for chelation therapy)

  • bone marrow transplant (standard of care w/ matched sibling donor)

  • gene therapy: Lyfgenia (>12 y/o)

hemoglobin C disease

• Gene(s): HBB

• Inheritance Pattern: autosomal recessive

• Features:

  • beta-globin structural variant: decrease in solubility → crystals form → hemolytic anemia (no sickling)

  • HbS/HbC compound heterozygotes: mild sickle cell disease

• Electrophoresis Patterns:

  • disease: HbC 95%, HbF <7%

  • trait: HbA 60%, HbC 40%

hemoglobin E

• Gene(s): HBB

• Inheritance Pattern: autosomal recessive

• Features:

  • beta-globin structural variant: activates cryptic splice site → reduced synthesis of beta-globin

  • clinically asymptomatic/mild anemia

  • Hb E/beta-thal (most common B-thal in Asian populations)

• Electrophoresis Patterns:

  • disease: HbE 90%, HbF

  • trait: HbA, HbE 30%

thalassemia

• absent or decreased production (quantitative) of normal hemoglobin due to diminished synthesis or stability of one or more globin chains

• decreased synthesis of globin chain → unbalanced alpha:beta ratio → normally produced chain is in excess → can’t form a tetramer → precipitates → damages red cell membrane → hemolytic anemia

• 5% of the world’s population may have at least one thalassemia allele

alpha thalassemia

• Gene(s): HBA1, HBA2

• Inheritance Pattern: autosomal recessive

• Features:

  • affects both fetal and adult hemoglobin

    • deletions, point mutations, mutations in LCR

    • common (non-deletional) mutation: constant spring (associated with HbH disease); fetal onset of symptoms

  • deletion of alpha gene → decreased alpha globin → accumulation of unpaired beta-globin chains → unstable tetramers

• Molecular Diagnosis:

  • >250 mutations but deletions are most common

    • most commonly 3.7 (3.8) kb deletion

    • HBA2-HBA1 fusion gene

    • several different breakpoints

  • most common diagnostic approach is. . .

    • targeted common deletion analysis

    • sequencing

    • full gene MLPA deletion analysis

hemoglobin Bart’s

(- - / - -)

• severe microcytic anemia

• hydrops fetalis

• incompatible with life

hemoglobin H disease

(a-/ - -)

• moderate microcytic anemia

• hemolytic anemia

• splenomegaly

alpha thalassemia minor/trait

(aa/- -) or (a-/a-)

• mild microcytic anemia

• two cis carrier parents are at risk for having a fetus with hemoglobin Bart’s (25%)

silent carrier for alpha thalassemia

(aa/a-)

• normal or mildly decreased hemoglobin

• normal or mildly decreased MCV

• usually asymptomatic

beta thalassemia

• Gene(s): HBB

  • usually single nucleotide variant (>200 mutations)

• Inheritance Pattern: autosomal recessive

• Features:

  • decreased beta globin → accumulation of unpaired alpha-globin chains → alpha-globin precipitate → decreased red cell production and hemolysis

  • synthesis of HbA (a2b2) reduced, HbA2 (a2s2) increased; HbF increased

  • prevalent in Mediterranean countries, the Middle East, Central Asia, India, Southern China, Northern Africa

• Molecular Diagnosis:

  • >200 point mutations or small insertions or deletions

  • 95% caused by point mutations

  • abnormal RNA transcription, processing or stability → reduced expression (B+) or result in complete absence (B0) of beta-globin

  • one altered gene copy: carrier

  • both altered gene copies: affected

  • severity correlates w/ the number of functioning hemoglobin chains

• Treatments:

  • transfusion dependent individuals → regular transfusions

    • iron overload → cardiomyopathy, liver fibrosis, and endocrine dysfunction

    • iron chelation: subcutaneous or oral agents

    • Reblozyl (luspatercept) (>18 y/o): erythroid maturation agent

    • BMT: prior to the development of iron overload

  • non-transfusion dependent individuals → may never require transfusion or may require one periodically (during pregnancy or acute infections)

  • gene therapy: Zynteglo (lentiviral vector modified beta-globin gene)

beta thalassemia major

• severe variants

• fatigue, weight loss, severe anemia, iron overload, congestive heart failure

• jaundice

• bone deformities

• extramedullary hematopoiesis → frontal bossing, prominent maxilla, splenomegaly, growth restriction

• increase of HbF, HbA2

beta thalassemia intermedia

• mild variants

• moderate anemia

• non-transfusion dependent

• jaundice, splenomegaly, microcytosis

beta thalassemia minor

• may have mild anemia

• microcytosis

hereditary persistence of fetal hemoglobin

• Gene(s): mutations in the beta- or alpha-globin gene clusters or the gamma promoter gene region

  • regulatory mutation

• Features:

  • inherited disorder of increased HbF in adults

    • HbA typically replaces HbF by 6-12 months

    • deletions or point mutations lead to continued expression of gamma-globin (switch from gamma to beta and delta doesn’t occur → HbF synthesis continues) h

  • usually clinically asymptomatic (even in homozygotes)

  • patients who coinherit a hemoglobinopathy (HbSS or B-thal) can have milder phenotype due to increased HbF being naturally produced

    • a patient with HbSS disease who also has HPFH can have up to ~20-30% HbF production essentially converting them to a sickle cell trait phenotype

hemophilia A

• Gene(s): F8 on Xq28

• Inheritance Pattern: X-linked recessive

• Features:

  • deficiency of factor VIII → cofactor for factor IXa, which converts factor X to Xa

  • 1 in 5,000 males

  • 100% penetrant in males

  • 30% of female carriers have decreased clotting activity and are at risk for bleeding

  • clinically indistinguishable from hemophilia B

    • spontaneous hemarthroses: bleeding in the joints → chronic arthritis

    • muscle hematoma (from minor injury)

    • intracranial bleeding

    • post-operative bleeding

    • easy bruising

hemophilia B

• Gene(s): F9 on X26-27.3

• Inheritance Pattern: X-linked recessive

• Features:

  • deficiency of factor IX (Christmas factor) → activates factor X

  • 1 in 20,000

  • 100% penetrant in males

  • 30% of female carriers have decreased clotting activity and are at risk for bleeding

  • clinically indistinguishable from hemophilia A

    • spontaneous hemarthroses: bleeding in the joints → chronic arthritis

    • muscle hematoma (from minor injury)

    • intracranial bleeding

    • post-operative bleeding

    • easy bruising

  • Treatment:

    • gene therapy (Hemagenix) approved in 2022

Von Willebrand Disease

• Gene(s): VWF

  • quantitative or qualitative defect

  • common, typically mild

  • often undiagnosed

• Inheritance Pattern: autosomal dominant or recessive (depends on the type)

• Features:

  • type 1: partial quantitative deficiency of normal VWF

    • treatable w/ DDAVP (desmopressin)

  • type 2: qualitative deficiency of defective VWF

  • type 3: complete quantitative deficiency of VWF

  • VWF carries factor VIII

  • VWF activates the platelet plug in clotting

  • individuals may experience prolonged cutaneous or mucosal bleeding (nosebleeds, GI bleeding, gum bleeding); bruising without recognized trauma; menorrhagia; prolonged bleeding following surgery, trauma, or childbirth

• Diagnosis:

  • hemostasis factors + VWF antigen

  • molecular testing does not identify all cases such as large deletions

Factor V Leiden

• Gene(s): F5

  • eliminates the cleavage site in Factor Va for activated protein C → actor factor V → increased fibrin → more clotting

• Inheritance Pattern: autosomal dominant

• Features:

  • most common known hereditary predisposition to venous thrombosis

    • ~20% of patients w/ first episode

  • increased risk for venous thromboembolism (VTE)

    • 5-10x in heterozygotes → lifetime risk is 10%

    • 50-100x in homozygotes → lifetime risk is ~100%

    • modest increased risk for recurrent VTE

  • may increase risk for pregnancy loss by two-to-threefold

    • may also increase risk for pre-eclampsia, intrauterine growth restriction, and placental abruption → precise risk unknown

  • heart attack, arterial thrombosis, and ischemic stroke are not associated

prothrombin thrombophilia

• Gene(s): F2

  • gain of function → increased production factor II (prothrombin)

• Inheritance Pattern: autosomal dominant

• Features:

  • second most common venous thrombosis risk factor

  • increased risk for venous thromboembolism (VTE) and pulmonary embolism

    • 2-4x in heterozygotes

    • 80x in homozygotes → lifetime risk is ~100%

MTHFR

(methylene tetrahydrofolate reductase)

• Features:

  • decreased MTHFR activity → hyperhomocysteinemia

    • affected by serum folate levels

    • HCY: risk factor for venous thrombosis and associated with other cardiovascular diseases (coronary artery disease) but data is conflicting

    • common polymorphism exist

• ACMG Recommendations:

  • polymorphism genotyping should not be ordered as part of clinical evaluation for thrombophilia or recurrent pregnancy loss

  • MTHFR polymorphism genotyping should not be ordered for at-risk family members

• Severe MTHFR (autosomal recessive)

  • high HCY, low methionine

  • microcephaly, developmental delay, neonatal seizures, premature thrombotic events