in brief about evaluation of pancytopenia in children and aplastic anemia

1. APLASTIC ANEMIA
DR NAHAR KAMRUN
PEDIATRIC HEMATOLOGIST AND ONCOLOGIST
CMH, BD

2. PANCYTOPENIA

3. • Pancytopenia is a medical condition in which there is reduction in the number
of cell all lineage.
• Pancytopenia is a combination of anaemia, leucopenia and
thrombocytopenia in peripheral blood.
 TC of WBC <4X109/L
 Hb: < 13.5 g/dl (male) or < 12 mg/dl (female)
 Platelets: <150X109/L
• If only two cell lineage are lower than normal it termed bicytopenia

4. PANCYTOPENIA
Anemia --------------------- fatigue
Thrombocytopenia -------------------- bruising/bleeding
Neutropenia …………………….. infection
Presentation varies with degree of cytopenia
Red blood cells, which carry oxygen to the tissues from the lungs
White blood cells, which fight infection
Platelets, which seal damaged blood vessels to prevent bleeding

5. PANCYTOPENIA – AETIOLOGY
Bone marrow failure
 Stem cell defect.----- Aplastic anaemia
 Ineffective hemopoiesis
 Megaloblastic anaemia (B12, folate)
 AIDS
 Myelodysplastic syndromes
 Bone marrow suppression ---- Cytotoxic drugs
 Bone marrow infiltration
 Haematological malignancy (leukemia, lymphoma, myelofibrosis)
 Non-haematological malignancy
 Inherited metabolic diseases
 Myelofibrosis
 Primary - clonal haematologic abnormality
 Secondary to malignancy
 Haemophagocytic Syndromes
 Congenital: Fanconi’s, Schwasmann-Diamond, Folate uptake deficiency,
Marble bone disease
Peripheral consumption
• Hypersplenism
• Combined immune cytopenia

6. BM FAILURE
• Bone marrow failure may manifest as a single cytopenia (e.g., erythroid,
myeloid, or megakaryocytic), or as pancytopenia.
• It may present with a hypoplastic or aplastic marrow or result from
invasion of the bone marrow by non-neoplastic (e.g., storage cells) or
neoplastic cells.
• It may be either congenital or acquired.
• Mitochondrial disease also may present with BM failure eg Pearson syn,
Wolfram syn and various types of sideroblastic anemia.

7. BONE MARROW FAILURE – CLASSIFICATION
CONGENITAL
• FANCONI ANEMIA
• DYSKERATOSIS CONGENITA
• SHWACHMAN DIAMOND SYNDROME
• CONGENITAL AMEGAKARYOCYTIC
THROMBOCYTOPENIA
• DIAMOND BLACKFAN ANEMIA (DBA)
• APLASTIC ANEMIA WITH CONSTITUINAL
CHROMOSOMAL ANOMALIES
ACQUIRED
• IDIOPATHIC – 70% CASES
• SECONDARY –
- DRUGS
- CHEMICALS & TOXINS
- RADIATION
- INFECTION
- PNH
–MALNUTRITION

8. APPROACH TO PANCYTOPENIA
• HISTORY
• PHYSCICAL EXAMINATION
• INVESTIGATIONS

10. Family history of aplastic anemia, MDS, Leukemia or having congenital anomalies in
family members

18. APLASTIC ANEMIA

19. BACKGROUND
• Aplastic anemia, a hematologic disease, is the paradigm of bone marrow failure
syndromes.
• Almost universally fatal just a few decades ago, can now be cured or ameliorated by
stem-cell transplantation or immunosuppressive drug therapy.
• The pathophysiology is immune mediated in most cases, with activated type-1
cytotoxic-T cells implicated.
• The molecular basis of the aberrant immune response and deficiencies in
hematopoietic cells is now being defined genetically; examples are telomere repair
gene mutations in target cells and dysregulated T-cell activation pathways.

20. DEFINITION
• Aplastic anemia is a physiologic and anatomic failure of the bone
marrow characterized by a marked decrease or absence of blood-
forming elements in the marrow and resulting pancytopenia
(decreased red cells, white blood cells and platelets).
• NO ORGANOMEGALY - Splenomegaly, hepatomegaly and
lymphadenopathy do not occur in this condition.

21. APLASTIC ANEMIA - AETIOLOGY
• Bone Marrow Failure ------------- WHY??????????
• Stem cell defect (seed)
• Stromal cell defect (soil)
• Growth Factor defect (fertilizer)
• Evidence suggests that majority of cases of idiopathic AA are due to
immune suppression of the hematopoietic stem cell

22. Normal Bone marrow Hypoplastic Bone marrow

23. CAUSES
Aetiological Classification:
A-Primary:
• 1- congenital (as Fanconi's anemia).
• 2- Idiopathic acquired (67% of cases).
According to origin: B- Secondary:
• 1-chemicals (benzene, toluene, glue sniffing).
• 2-drugs (chemotherapeutics, antibiotics, gold,
phenytoin).
• 3-insecticides.
• 4-ionizing radiation.
• 5-infections (as hepatitis, influenza, MMR).
• 6-paraxysmal nocturnal haemoglobinuria.
• 7-Immunological disorder
• 8-Malnutrition
A. Inherited
B. Acquired: a. Idiopathic b. secondary

24. DRUGS CAN CAUSES ACQUIRED APLASTIC ANEMIA
Some of the medicines linked to aplastic anemia include:
• NSAIDs –indomethacin, piroxicam, diclofenac
• Amphetamines, including MDMA (Ectasy)
• Antibiotics: sulfonamides (“sulfa drugs”) and forms of penicillin, Chloramphenicol
• Anti-thyroid drugs: propylthiouracil and methimazole
• Carbonic anhydrase inhibitor: acetazolamide, methazolamide
• Diabetes medications: tolbutamide, carbutamide, and chlorpropamide
• Diuretics – furosemide, thiazides
• Anti-malarial drugs: quinacrine, chloroquine
• Phenothiazines (used for nausea, psychiatric problems)
• Allopurinol [gout]
• Ticlopidine -- used to prevent strokes and heart attacks
• Anti-seizure drugs like carbemazepine, phenytoin, and valproic acid
• Mesalazine [ IBS]

25. ASSOCIATION WITH APLASTIC ANEMIA
• Chemicals and pesticides eg Benzene,
organophosphates and carbamates
• Viral infection:
 Hepatitis (Viral Hepatitis, HBV)
 Viruses such as EBS, CMV, parvovirus B19, HIV.
• Pregnancy
• Auto-immune diseases like lupus (SLE) and RA
• Severe radiation poisoning (eg atomic bomb
explosion)

26. APLASTIC ANEMIA
PATHOPHYSIOLOGY

27. APLASTIC ANEMIA PATHOPHYSIOLOGY
Intrinsic abnormality of stem cells  which predisposes DNA damage and marrow
aplasia  Reduction in number of haemopoietic pluripotential stem cells ------->
defective production in the remaining stem cells.
OR
Immune reaction against stem cells. -------> suppression and killing of Haemopoietic
progenitors in bone marrow  unable to divide and differentiate sufficiently to
produce blood cells.
A reasonable theory suggests that exposure to an inciting antigen, cells and cytokines
of the immune system destroy stem cells in the marrow resulting in pancytopenia.
(Philip 6th)

28. APLASTIC ANEMIA PATHOPHYSIOLOGY
T-cells secrete γ-IFN (plays central
role) and TNF, act as potent inhibitors
of hematopoietic progenitor cells.
Cyt T-cells secrete IL-2, which causes
polyclonal expansion of T-cells.
Additionally, γ-IFN mediates its
hematopoietic suppressive activity
through IRF-1.
Direct cell-cell interactions between
effective lymphocytes and targeted
hematopoietic cells probably also
occur.
Oligoclonal expansion of CD41, CD81
T-cells supports an immune etiology.

29. • T-cells secrete Gamma-Interferon (γ-IFN) (plays central role) and TNF, act as potent inhibitors of
both early and late hematopoietic progenitor cells.
• They suppress hematopoiesis by their effects on mitotic cycle and, more importantly, by the
mechanism of cell killing (apoptosis).
• Cyt T-cells secrete IL-2, which causes polyclonal expansion of T-cells. Activation of Fas receptor
on hematopoietic stem cell by the Fas ligand (present on lymphocytes) leads to apoptosis of
targeted hematopoietic progenitor cells.
• Additionally, γ-IFN mediates its hematopoietic suppressive activity through IFN regulatory factor
1 (IRF-1), which inhibits transcription of cellular genes and their entry into cell cycle.
• γ-IFN induces production of nitric oxide, diffusion of which causes additional toxic effects on the
hematopoietic progenitor cells.
• Direct cell-cell interactions between effective lymphocytes and targeted hematopoietic cells
probably also occur.
• The oligoclonal expansion of CD41 and CD81 T-cells supports an immune etiology.
Philip 6th

30. EVALUATION OF APLASTIC ANEMIA
• Based on --Evaluation of pancytopenia
History
Physical examination
Investigation
 For diagnostic purpose
 For treatment purpose
 To exclude other disease

31. Pallor (anemia)
Anemia (low Hb)
Pallor with malaise,
weakness, fatigue,
anorexia and
palpitations.
Clinical Features

32. Thrombocytopenia (low platelet
counts), leading to
increased risk of hemorrhage.
Bleeding manifestations:
• Skin: bruising, petechiae, purpura,
echymoses.
• Mucosa: gum, nose & conjunctiva
• Internal organs: brain, kidneys, gut
etc.

33. Leukopenia (low WBC
count) --
Fevers and infections.
 Usually severe, recurrent
 Fungus, or unusual pathogens
(Septicemia, Meningitis,
Pneumonia, UTI, Mucositis,
Skin infection, Sinusitis )

34. LABORATORY INVESTIGATIONS
 CBC with PBF:
• 1. Hb: reduced; normocytic, normochromic or macrocytic.
• 2. Reticulocytopenia: absolute count more reliable.
• 3. Leukopenia: granulocytopenia often less than 1,500/mm3.
• 4. Thrombocytopenia: platelets often less than 30,000/mm3.
• 5. Fetal hemoglobin: may be slightly to moderately elevated.
 Bone marrow biopsy - with cellularity < 25%
Aplastic “anemia” misnomer (Anemia refers to Red Cell Lineage)
Pancytopenia

35. PARENTHESIS: (BONE MARROW EVALUATION)
• Aspiration:
• Morphology
• Lineage maturation
• Blasts
• Flow Cytometry
• Cytogenetics
• Molecular Genetics
• Biopsy:
• Cellularity
Fanconi Anemia

36. Normal Bone Marrow
Aplastic Anemia or
Bone Marrow Failure
BONE MARROW

37. INV CONTD.
Findings in Bone marrow analysis:
a. Morphology:
• Marked depression or absence of hematopoietic cells and replacement by fatty-tissue-containing
reticulum cells, lymphocytes, plasma cells and usually tissue mast cells.
• Megaloblastic changes & other features of dyserythropoiesis frequently seen in RBC precursors.
b. Biopsy: most essential (only way to assess cellularity) for diagnosis
- Exclude possibility of poor aspiration technique or poor bone marrow sampling;
- exclude infiltrative disease.
c. Chromosomal analysis including breakage assay & rearrangements of peripheral lymphocytes –
[normal]
- to rules out Fanconi anemia and MDS
d. Bone marrow cultures, antigen-based evaluation, and molecular testing for infectious agents
and/or DNA; when indicated.

38. INV. TO EXCLUDE OTHER DISEASE:
• Chromosome breakage assay: performed mainly on peripheral blood to rule out Fanconi anemia.
• Flow cytometry (CD59): to exclude paroxysmal nocturnal haemoglobinuria (PNH).
• Telomere length to screen - for Dyskeratosis congenita.
• Cytogenetic analysis of marrow particles to predict the subsequent development of leukemia.
• Viral serology: HAV, HBV, HCV, EBV antibody panel; parvovirus B19 IgG & IgM antibodies; varicella
antibody titer; CMV antibody titer, HIV antibody
• Quantitative immunoglobulins (Igs) - to rule out immunodeficiency
• Autoimmune disease:
- ANA, total hemolytic complement (CH50), DAT, C3, C4 and complement.
• Others:
- Liver function chemistries: to exclude hepatitis.
- Renal function chemistries: to exclude renal disease.
- HLA typing:
-to find out eligible family member for transplantation.

39. TYPES OF APLASTIC ANEMIA ON BASIS OF SEVERITY
Aplastic anemia Divided into 3 groups:
• •Moderate
• •Severe
• •Very severe
These categories were defined in 1975
by Dr. Bruce Camitta and his team.

40. MANAGEMENT
Aim: The main goal of aplastic anemia treatment is to increase the number of healthy cells in blood.
A. Supportive Care:
• The cause should be removed.
• Blood transfusion
• Iron chelation to treat iron overload if present
• Growth factors
• Prevention of infection- Antibiotics/Anti viral/anti fungal. Keep patients out of hospital as much as possible
• Drugs that impair platelet function, such as aspirin, should be avoided.
• Intramuscular injection should be given carefully, followed by ice pack.
• Antifibrinolytic agent, aminocaproic acid -to reduce mucosal bleeding except in Hematuria
• Good oral care - Teeth should be brushed with a cloth or soft toothbrush.

41. B. Definitive Therapy
Acquired aplastic anaemia can be treated with either
a. Hematopoietic stem cell transplantation (HSCT)
for children with severe or very severe aplastic anemia who have a matched
sibling donor.
b. Immunosuppressive therapy:
drugs used to treat aplastic anemia are:
• ATG (anti thymocyte globulin) or
• ALG (anti lymphocyte globulin)
• Cyclosporine
• Tacrolimus
BONE MARROW TRANSPLANTATION:
• Treatment of choice. gold standard treatment
• from HLA matched donor. Usually siblings.
• Long term survival rates: 60-70%.
It is effective, even only partially effective, and the blood counts may not
return to normal.
In this situation, many people can still lead a normal life and no need
treatment. Their blood counts, although not normal, will be high enough
for a normal life.
APLASTIC ANEMIA SPECIFIC TREATMENT

42. TREATMENT OF MODERATE APLASTIC ANEMIA
• The natural history of moderate aplastic anemia is uncertain and clinical experience varies
widely. For this reason, it is generally thought that these patients should be treated initially
with supportive therapy with very close follow-up.
• Indication of definitive treatment:
1. Progressive disease, to develop severe aplastic anemia (majority cases)
and/or
2. Significant and severe thrombocytopenia and bleeding,
3. Serious infections, or
4. Chronic red blood transfusion requirement
• Definitive treatment: same for severe aplastic anemia.

43. WHAT IS THE PROGNOSIS OF APLASTIC ANEMIA?
If left untreated AA can prove to be fatal.
• Outcomes for both IST and HSCT have improved considerably.
• Survival rates are >90% with either IST or HSCT.
• HSCT found curative for most patients, very successful in younger population.
• IST improves hematopoiesis and removes transfusion dependency, but time to
response is long, may be partial response and relapses are relatively common.
Mortality is usually due to—
• Bleeding and/or infection

45. • Thank you…………………..