Pediatric Acute Lymphoblastic Leukemia Workup

Upon initial evaluation, obtain a complete blood cell (CBC) count. A hematologist or hematopathologist must evaluate the peripheral smear for the presence and morphology of lymphoblasts. An elevated leukocyte count of more than 10 × 10⁹/L (>10 × 10³/µL) occurs in one half of patients with acute lymphoblastic leukemia (ALL). Neutropenia, anemia, and thrombocytopenia are often observed secondary to inhibition of normal hematopoiesis by leukemic infiltration. It is important to recognize that 20% of patients with ALL initially present with pancytopenia and no evidence of peripheral blasts.^[11]

Various metabolic abnormalities may include increased serum levels of uric acid, potassium, phosphorus, calcium, and lactate dehydrogenase (LDH). The degree of abnormality reflects the leukemic cell burden and destruction (lysis). Although not universally performed, coagulation studies can be helpful in patients with T-lineage ALL and should include tests of the prothrombin time (PT), activated partial thromboplastin time (aPTT), fibrinogen level, and D-dimer level to assess for disseminated intravascular coagulation (DIC).

No other imaging studies than chest radiography to evaluate for a mediastinal mass should be required. However, if the physical examination reveals enlarged testes, perform ultrasonography to evaluate for testicular infiltration. In addition, if anthracyclines are to be administered, obtain a baseline echocardiogram and an electrocardiogram (ECG).

To assess for central nervous system (CNS) involvement and to administer intrathecal chemotherapy, lumbar puncture with cytospin morphologic analysis is performed before systemic chemotherapy is administered.

Acute lymphoblastic leukemia (ALL) cells rearrange their immunoglobulin and T-cell receptor (TCR) genes and express antigen receptor molecules in ways that correspond to such processes in normal developing B and T lymphocytes, so that acute lymphoblastic leukemia can be classified as B-lineage or T-lineage ALL.

The diagnosis of mature B-cell leukemia, which accounts for only 1-3% of childhood ALL, depends on the detection of surface immunoglobulin on leukemic blasts. Lymphoblasts with this phenotype have a distinctive morphology, with deeply basophilic cytoplasm containing prominent vacuoles, designated L3 in the French-American-British (FAB) system (see Histologic Features). Mature B-cell ALL should be differentiated from other B-lineage ALL.

B-lineage ALL accounts for 80% of childhood ALL and involves lymphoblasts that have cell-surface expression of 2 or more B-lineage–associated antigens (ie, CD19, CD20, CD24, CD22, CD21, or CD79).^[2] CD10 is commonly expressed, which makes it a useful diagnostic marker, and the presence of aberrant myeloid markers (eg, CD7) is occasionally noted but has little prognostic impact. B-cell precursors of ALL can be further subclassified as early pre–B-cell, pre–B-cell, or transitional pre–B-cell, but distinguishing these subtypes is usually not clinically relevant.

T-lineage ALL is identified by the expression of T-cell–associated surface antigens, of which cytoplasmic CD3 is specific. T-cell acute lymphoblastic leukemia cases can be classified by early, mid, or late thymocytes. The prognosis of patients with T-cell ALL has historically been worse than that of patients with B-lineage ALL. However, the outlook for patients with T-cell leukemia is comparable to that of precursor B-cell ALL when intensive chemotherapy is used.

In more than 90% of pediatric acute lymphoblastic leukemia (ALL) cases, specific genetic alterations can be found in the leukemic blasts, which have important diagnostic, therapeutic, and prognostic implications. In addition, molecular techniques, including fluorescence in situ hybridization (FISH), reverse transcriptase-polymerase chain reaction (RT-PCR), and Southern blot analysis help identify translocations not detected on routine karyotype analysis and to distinguish lesions that appear cytogenetically identical but are molecularly different.

Of the many abnormalities described, t(12;21)(p13;q22) or ETV6-RUNX1 (formerly known as TEL-AML1) and hyperdiploidy (>50 chromosomes/cell) account for 50% of chromosomal abnormalities found and confer a favorable prognosis. Trisomy 4, trisomy 10, and trisomy 17 (”triple trisomy”) may be seen in some hyperdiploid cells and share the favorable outcome. Hypodiploidy (< 44 chromosomes/cell), t(4;11)(q21;q23) MLL-AF4 or MLL gene rearrangement, and t(9;22)(q34;q11), or Philadelphia chromosome positivity confer a poor prognosis.

Traditionally, the response to leukemia treatment has been assessed morphologically, which can be challenging when looking for small numbers of leukemic cells, especially in bone marrow specimens recovering from chemotherapy or after transplantation.

Molecular analysis plays a promising role in the diagnosis and treatment of acute lymphoblastic leukemia (ALL) and in monitoring patients' responses to therapy.

Studies of minimal residual disease (MRD) may be based on the detection of chimeric transcripts generated by fusion genes, the detection of clonal TCR or immunoglobulin heavy-chain (IgH) gene rearrangements, or the identification of a phenotype specific to the leukemic blasts.^[12]

The methods for detecting MRD have been shown to have a much higher sensitivity than that of morphology. All studies using MRD techniques have shown significant correlations between end-of-induction leukemia burden and outcome.^[13] As a result, current treatment protocols use MRD measurements for acute lymphoblastic leukemia risk assignment.^[14]

Perform testicular ultrasonography if the testes are enlarged upon physical examination.

Some clinicians use renal ultrasonography to evaluate for leukemic kidney involvement as an assessment of risk for tumor lysis syndrome.

Bone marrow aspirate and biopsy results confirm the diagnosis of acute lymphoblastic leukemia (ALL). In addition, special stains (immunohistochemistry), immunophenotyping, cytogenetic analysis, and molecular analysis help in classifying each case. See the images below for examples of bone marrow aspirate findings.

According to the French-American-British (FAB) classification system, acute lymphoblastic leukemia (ALL) is classified into 3 groups based on morphology, as follows:

• L1: The lymphoblast cells are usually small, with scant cytoplasm and inconspicuous nucleoli. L1 accounts for 85% of all cases of childhood acute lymphoblastic leukemia.
• L2: The lymphoblast cells are larger than in L1. The cells demonstrate considerable heterogeneity in size, with prominent nucleoli, and abundant cytoplasm. L2 accounts for 14% of all childhood acute lymphoblastic leukemia.
• L3: The lymphoblast cells are large and notable for their deep cytoplasmic basophilia. They frequently have prominent cytoplasmic vacuolation and are morphologically identical to Burkitt lymphoma cells. L3 accounts for 1% of childhood acute lymphoblastic leukemia cases.

Although the FAB system was used in the past, it is no longer useful (except for L3), because current standard diagnosis is based on immunophenotype and molecular techniques.