Other rare red blood cells disorders Batrice GULBIS, M.D., PhD 1 - - PowerPoint PPT Presentation

Other rare red blood cells disorders

Béatrice GULBIS, M.D., PhD

BHS training course 2017

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Objectives of the training course

• Rare red blood cells disorders:

Inherited, haemolytic (anaemia)

– Inheritance and epidemiology – Clinical diagnostics aspects – Useful diagnostic tests, algorithms – Therapeutics

BHS training course 2017

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Rare or very rare anaemias

• About 90 rare types of

anaemia have been described

• Among them ± 80% are

inherited

• www.enerca.org
• www.orpha.net

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Rare or very rare anaemias

• Haemoglobinopathies (Unstable haemoglobin

variant)

• Red blood cells membrane defects
• Red blood cells enzymes defects
• Rare iron metabolism related anaemia
• Aplastic anaemia
• Acquired haemolytic anaemia and

erythropoietic defects

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Rare or very rare anaemias

• Prevalence

– How much of condition at a certain period of time

• Incidence

– Rate of occurrence of new cases in a specified period

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Rare haemolytic (anaemias) Epidemiology

• Few international (European) registries (CDA)
• Estimated incidence:

1 2 3 4 5 6 7 10 100 1000 /106 live births Congenital Dyserythropoietic Anaemia(CDA) Hereditary Spherocytosis (HS) Paroxysmal Nocturnal Haemoglobinuria (PNH) Rare RBC enzymopathies

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Rare haemolytic (anaemias) Epidemiology in your practice

• Age of onset ?

PNH Affects particularly young adults HS – CDA – RBC Enzyme disorders – Hbpathies From birth to adulthood

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Rare haemolytic (anaemias) Epidemiology in your practice

• Who is affected?
• Influence of ethnic origin

– No, but – effect of consanguinity (homogamy)

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Rare haemolytic (anaemias) Epidemiology in your practice

Relationship to each other Degree relatives Proportion genes in common

Brothers and sisters, non-identical twins, parents and children First 50% Uncles and aunts, nephews and nieces, grandparents and half brothers/sisters Second 25% First cousins, half-uncles/aunts half-nephews/nieces Third 12.5%

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Rare haemolytic (anaemias) Epidemiology in your practice

• Who is affected?
• Gender

– No, but – Some X-linked (G6PD)

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Very rare anaemia Inheritance and epidemiology

• Mode of inheritance?

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Very rare anaemias Inheritance and epidemiology

Autosomal dominant inheritance 50% Autosomal recessive inheritance 25%

http://www.genetics.edu.au/

Hereditary spherocytosis CDA III Hereditary spherocytosis And other membranopathies RBC enzymopathies (except G6PD) CDA

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Very rare anaemias Inheritance and epidemiology

X-linked inheritance

http://www.genetics.edu.au/

25% X-linked inheritance G6PD Rare cases of CDA

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Very rare anaemias Inheritance and epidemiology

• X-linked inheritance

– Random X chromosome inactivation:

• a woman carrier can …

Mutated

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Very rare anaemias Inheritance and epidemiology

X-linked inheritance

http://www.genetics.edu.au/

25% X-linked inheritance

XrXr

This is a girl…

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Very rare anaemias Inheritance and epidemiology

• Hardy-Weinberg law – two alleles

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Very rare anaemia Inheritance and epidemiology

• Examples

– Haemophilia A:

• Affected males: q= 1/10,000

>>> affected females q²= 1/100.106

– G6PD deficiency

• Affected males: q= 1/50

>>> affected females q²= 1/2500

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Rare haemolytic (anaemias) Inheritance

• Selection (advantage)
• Mutation rate

– G6PD deficiency as a life long haemolytic anaemia is rare

• Modulator genes

– Spectrin alpha Low Expression Prague

• …

Rare haemolytic (anaemias)

• Not only during childhood
• If X-linked inheritance, not only boys
• If autosomal recessive inheritance, importance
• f consanguinity (homogamy)

BHS training course 2017

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Objectives of the training course

• Rare red blood cells disorders:

Inherited, haemolytic (anaemia)

– Inheritance and epidemiology – Clinical diagnostics aspects – Useful diagnostic tests, algorithms – Therapeutics

BHS training course 2017

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Rare haemolytic (anaemias) In your practice

• Only signs and symptoms of anaemia?

Clinical diagnostics aspects Extravascular haemolysis +++

Chronic haemolytic (anaemia) of variable severity and/or Splenomegaly and/or Jaundice and/or Biliary lithiasis

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Clinical diagnostics aspects Extravascular haemolysis +++ Signs and symptoms depend on the degree of decompensation of anaemia

– Variable age at diagnosis, but

• HS ± 65% with neonatal icterus
• Each “stress” on the RBCs = decompensated anaemia

– Birth – Infection (i.e. Parvovirus B19) – Pregnancy

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Rare haemolytic (anaemias)

• Only signs and symptoms related to red

blood cell pathology?

– Congenital dyserythropoietic anaemias

Acetylcholinesterase Adenosine deaminase Adenylate kinase Aldolase (ALD) Bisphosphoglycerate mutase Catalase NADH-cytochrome b5 reductase Enolase Galactokinase Galactose-4-epimerase -Glutamylcysteine synthetase Glucose phosphate isomerase (GPI) Glucose-6-phosphate dehydrogenase Gluthathione peroxidase Gluthathione reductase Glutathione synthetase Glutathione-S-transferase Glyceraldehyde 3-phosphate dehydrogenase Hexokinase In bold= involved in haemolysis Lactate dehydrogenase Monophosphoglycerate mutase Multiple inositol polyphosphate phosphatase NADPH diaphorase Phosphofructokinase (PFK) Nucleoside phosphorylase Phosphoglucomutase 6-Phosphogluconate dehydrogenase 6-Phosphogluconolactonase Phosphoglycerate kinase (PGK) Phosphoglycolate phosphatase Phosphomannose isomerase Pyrimidine-5’-nucleotidase Pyruvate kinase Ribosephosphate isomerase Superoxide dismutase Transaldolase Transketolase Triosephosphate isomerase (TPI)

Rare chronic haemolytic (anaemias) RBC enzymopathies

Non-haematological manifestations

• Neuromuscular disease – TPI, GPI

deficiency

• Myopathy (myoglobinuria) – PFK, PGK,

ALD deficiency

• Impaired granulocyte function – TPI, GPI

deficiency

• Mental retardation – ALD, PGK deficiency
• Rhabdomyolysis – ALD deficiency
• Psychomotor retardation – ALD deficiency
• Cardiomyopathy – TPI deficiency
• Increased susceptibility to infection – TPI

deficiency

• ....

Van Wijk and van Solinge, Blood 2005

Rare haemolytic (anaemias)

• First signs, symptoms: splenomegaly, jaundice,

biliary lithiasis

• Episode of haemolytic « crisis » (i.e. infection)
• Not only haemolysis i.e. TPI deficiency, CDA I

BHS training course 2017

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Objectives of the training course

• Rare red blood cells disorders:

Inherited, haemolytic (anaemia)

– Inheritance and epidemiology – Clinical diagnostics aspects – Useful diagnostic tests, algorithms – Therapeutics

BHS training course 2017

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Erythropoiesis? Haemolysis? RBC indices Immunomediated? Inherited or Acquired?

Haemolytic (anaemia) Algorithm

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Family? Personal history? Coombs (DAT) test

• Unconj. Bili.

Haptoglobin LDH Reticulocytes (IRF) WBC and platelet counts? MCV MCH MCHC

First Level Laboratory Testing

• General markers of haemolysis
• ↑ Unconjugated bilirubin
• ↑ Reticulocytes (absolute values) or unrelated with  level of Hb
• ↓ Haptoglobin
• Exclusion of acquired anaemias
• Negative DAT Autoimmune haemolytic anaemia
• Negative CD55
• Negative CD59

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PNH

Red Blood Cell Morphology

BHS training course 2017

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Unremarkable /anisocytosis Distinct morphological abnormalities RBC enzyme defect CDA RBC membrane disorders

 Ask to the lab: RBC morpholgy

Unstable Hb

RBC enzymopathies

BHS training course 2017

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A red blood cell enzyme disorder should be assumed in cases of persistent normocytic haemolytic anaemia in which hemoglobin abnormalities and antiglobulin reactions have been excluded, spherocytes are absent, and

• smotic fragility is normal

RBC Enzyme activity DNA analysis

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CDAs

Ineffective erythropoiesis (reticulocytes < 150,000/mm³) DNA analysis Bone marrow Electrophoresis RBC membrane proteins

100 200 300 400 500 600 700 5 10 15

Retic. 10³

Hb g/dL

C D A II

Unstable haemoglobin variant

• Destabilization of Hb tetramers

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Heinz bodies Stability tests Isopropanol /heat Separation

• f Hb

fractions/ DNA

Red Blood Cell membrane disorders RBC Morphology

BHS training course 2017

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Distinct morphological abnormalities Spherocytes Elliptocytes Elliptocytes /Poikilocytes Stomatocytes

• Not always high % of abnormal cells
• Spherocytes: AHAI, G6PD deficiency haemolytic crisis, etc.

Hereditary spherocytosis (HS) and misdiagnosis of “Stomatocytosis”

MCV MCHC Osmotic fragility EMA binding test Ektacytometry Hereditary spherocytosis N or  N or   (+*) Altered osmotic gradient curve Dehydrated Stomatocytosis N or  ↑ / (-) Left-shifted osmotic gradient curve

Overhydrated Stomatocytosis ↑↑ ↓ / (-) Right-shifted osmotic gradient curve

BHS training course 2017

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DNA analysis * Might be negative

Diagnosis: the future

• Gene panel – « Mendeliome »

– Paper No: 2433 ASH meetinfg dec 2016 - « Using a next

generation sequencing panel to discover the obscure causes of hereditary hemolytic anemia » A. M. Agarwal

Br J Haematol. 2016 Sep;174(5):806-14.

– Detection of new pathogenic mutations in patients with congenital haemolytic anaemia using next-generation sequencing. Int J Lab Hematol. 2016 Dec;38(6):629-638.

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Diagnosis

• Unexplained or

doubtful diagnosis haemolytic anaemia?

• A panel
• f 28 genes
• r more

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Rare haemolytic (anaemias)

• Algorithm for diagnosis exist
• Final diagnosis: need collaboration with expert

centres

• New perspectives with NGS

BHS training course 2017

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Objectives of the training course

• Rare red blood cells disorders:

Inherited, haemolytic (anaemia)

– Inheritance and epidemiology – Clinical diagnostics aspects – Useful diagnostic tests, algorithms – Management - Therapeutics

BHS training course 2017

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Overall management

– Supportive measures:

• Folate therapy is recommended in severe and moderate

forms of haemolytic anaemia

• red cell transfusions may be required in severely

anaemic cases i.e. aplastic crisis, infections, and pregnancy.

– Iron ferritin levels should be monitored particularly in the presence of co-inherited HFE gene mutations associated to hereditary hemocromatosis and chronic RBC transfusions. – Gallstones formation occurs frequently in RBC membrane defects and requires periodical abdominal ultrasound monitoring.

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G6PD deficiency Treatment

• Prevention of crisis

– To avoid the trigger

• Transfusion if necessary

(+ Folic acid)

BHS training course 2015

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G6PD deficiency

Drugs that trigger haemolysis

Br J Haematol. 2014 Feb; 164(4): 469–480.

• No drug is tested , no test is available
• In many cases haemolysis was probably triggered by the infection and not the drug.
• Possible haemolysis
• Dose related,
• Combination of drugs
• Co-morbodity

G6PD deficiency

Factors that influence severity of haemolysis

• Drug dose

– The same dose of a drug gives the same level of haemolysis

• RBC ageing

– The oldest RBC have the least G6PD activity and are the first to haemolyse – If a drug is given at a high dose, almost all RBC will haemolyse (ageing as well as young). If a lower dose is given, only the ageing RBC will be concerned. For the next dose, RBC with high amount of G6PD will not be concerned.

BHS training course 2011

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New drug in PK deficiency

• AG-348

– Pharmacologic activator of red cell PK – Allosteric activator of PK-R that activates wild-type and a range of mutated PK-R enzymes in vitro, increases PK-R activity and restores ATP levels in RBC from patients with PK deficiency ex vivo – Oral administration

BHS training course 2017

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New drug in PK deficiency

• Phase 2 clinical trial

– Start date June 2015 – Expected end date May 2019 – Enrollment of patients > 18 y.o. – Multicentre, randomized

• Primary outcome

– Safety, tolerability, clinical efficacy

BHS training course 2017

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Pyruvate kinase deficiency

• Rare cases of PK deficiency are treated by stem

cells transplantantion

BHS training course 2017

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Extravascular haemolysis: Splenectomy

• Before any decision

– Diagnosis done – Balance between benefit and complications

• Infections
• Thromboembolic complications

– Acute splenic and vein thrombosis » Role of prophylactic antithrombotic therapy? » At least one Doppler ultrasound study on day 7 after splenectomy – Late cardiovascular events » Arterial and venous thromboembolism » Pulmonary arterial hypertension (see increase in

platalets number and non removal of senescent and damaged RBC)

BHS training course 2017

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Guidelines

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BHS training course 2017

Splenectomy RBC enzymopathies

• Heterogeneous aetiology : response difficult to

predict

• Beneficial in most patients who are transfusion-

dependent or do not tolerate anaemia (grade 2 recommendation, grade C evidence)

• In Pyrimidine -5’-nucleotidase deficiency:

insufficient data for giving recommendations

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Haematologica 2017;102: 1304-

Splenectomy CDAs

• CDA I:

– Insufficient data for giving recommendations

• CDA II:

– Considered if transfusion-dependent or do not tolerate anaemia

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Haematologica 2017;102: 1304-

Splenectomy Unstable Hb

• Severe anaemia and/or splenomegaly +++

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Haematologica 2017;102: 1304-

Splenectomy Disease specific recommendations

Hereditary spherocytosis: Related on severity of the disease

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Severity Hb g/dL Reticulocytes X 103/mm3 Bilirubin mg/dL Splenectomy Severe < 8 > 400 >3 Yes (delay > 6 y.o.) Moderate 8-12 > 240 2-3 Individually tailored Mild 11-15 120 - 240 1-2 No Haematologica 2017;102: 1304- (grade 2 recommendation, grade C evidence)

Splenectomy Hereditary spherocytosis

• Median increase in Hb > 3 g/dL
• Decrease bilirubin and reticulocytes within reference

range + cholecystectomy if symptomatic gallstones When not severe, « individually tailored » splenectomy is related to: – Decrease quality of life (jaundice+++) – Symptomatic and or painful splenomegaly

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Haematologica 2017;102: 1304-

Splenectomy Hereditary stomatocytosis (both dehydrated and overhydrated)

• Contraindicated (grade 2 recommendation, grade C

evidence)

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Haematologica 2017;102: 1304-

Splenectomy

RBC enz. CDA I/ CDA II Unstable Hb HS Stomatocytosis Severe cases PK:

• Insuf. Data

Severe cases Splenomegaly +++ Severe cases Quality of life Splenomegaly +++ Contraindicated

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Grade 2 recommendation, grade C evidence Haematologica 2017;102: 1304-

Rare haemolytic (anaemias)

• Overall management is common to all diseases:

extravascular haemolysis

• New drugs are in the pipeline
• Place of splenectomy has been defined but it is

based mostly on experience of experts

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• Red Blood Cell Disorders Committee

– www.bhs.be

• Clinical cases:
• www.enerca.org
• New European group:
• www.eurobloodnet.eu

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Iron overload

BHS training course 2015

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