TCF3-HLF-Positive Acute Lymphoblastic Leukemia Resembling Burkitt Leukemia: Cell Morphologic and Immunophenotypic Findings

MLPA, multiplex ligation-dependent probe ampli ﬁ cation; PCR, polymerase chain reaction.


Introduction
Acute lymphoblastic leukemia (ALL), the most common malignancy in childhood and adolescence, is a heterogeneous disease with various subtypes on the basis of distinct cell morphology and genetic alterations. These distinctions are noted through cytogenetic or molecular analyses and represent prognostic outcomes. 1 By contrast, Burkitt leukemia (BL), a mature B lymphocyte neoplasm, is a highly aggressive hematologic malignancy characterized by uniformly medium-sized cells containing abundant basophilic cytoplasm, lipid vacuoles, and round nuclei with multiple nucleoli and clumped chromatin. 1 It is associated with the proto-oncogene MYC at chromosomal locus 8q24. 2 According to chromosomal analyses, approximately 80% of BL cases harbor a t(8;14) (q24;q32) translocation, which is typically used to diagnose the disease. 1 Nonetheless, t(2;8) (p12;q24) and t(8;22) (q24;q11) may also occur. 2 It is uncertain if true BL may be diagnosed without MYC translocations and hematologic cancers with BL morphology that lacked the typical BL immune phenotype or MYC rearrangement have been reported as BL-like ALL. 3 Therefore, patients with leukemia with BL morphology and no MYC rearrangement may be diagnosed as BL-like ALL, BL without MYC rearrangement, or BL with cryptic MYC rearrangement, on the basis of nextgeneration sequencing. 4 However, distinctions should be made because of differences in management and prognosis.
Because of front-line therapies derived from non-Hodgkin lymphoma Berlin-Frankfurt-Münster 95 (NHL-BFM95) protocol, pediatric BL is associated with promising outcomes. By contrast, some types of ALL with specific genetic alterations, such as t(17;19) TCF3-HLF fusion ALL, may harbor a dismal prognosis. 5,6 In this study, we reported a pediatric case of TCF3-HLF fusion ALL wherein BL was initially diagnosed according to morphologic or immunophenotypic characteristics. The case emphasizes the importance of detecting cryptic fusion genes, especially those associated with considerable prognostic effects.
The patient was initially treated with the Taiwan Pediatric Oncology Group (TPOG) 10 B-NHL protocol R4 arm (AA→ BB→ CC→ AA→ BB→ CC) with concurrent rituximab administration. 5,7 Detailed medications are listed in Appendix Table A1. The bone marrow study showed complete remission after course AA. The CSF study similarly showed the absence of blasts after four courses of triple intrathecal chemotherapy (methotrexate, hydrocortisone, and cytarabine). The patient underwent sequential treatment: course BB with rituximab, course CC with rituximab, and twice triple intrathecal chemotherapy.

Discussion
This study reports a rare case of ALL in which the blast cells expressed typical morphologic features of BL (ie, mature B lymphoid antigens) but harbored TCF3-HLF fusion genes. The discrepant results obtained from different diagnostic tests (eg, morphologic interpretations, immunophenotyping, and karyotype) emphasized the significance of molecular genetic analysis and its crucial role in subgroup classification and prognostication.
TCF3, located at 19p, is rearranged with several genes in childhood ALL. A t(1;19) (q23;p13) translocation and TCF3-PBX1 fusion account for 5% of childhood ALL. Among reported BL-like ALL cases, only t(1;19) is repeatedly found in gene analyses, indicating that it plays a critical role in ALL resembling BL. 3 The phenotype is also associated with approximately 85% event-free 5-year survival rates. 7,8 By contrast, childhood ALL harboring t(17;19) (q22;p13) translocation and TCF3-HLF fusion is extremely rare (, 0.5% of cases) but very aggressive and associated with a poor outcome. [8][9][10] Recent next-generation sequencing studies on the pathogenesis of BL have revealed that mutations in the transcription factor TCF3 or its negative regulator ID3 occurred in approximately 70% of sporadic and immunodeficiencyrelated BL cases and 40% of endemic BL cases. TCF3 promotes survival and proliferation in lymphoid cells by activating the B-cell receptor/phosphatidylinositol 3-kinase signaling pathways and modulating the expression of cyclin D3, which is also mutated in 30% of BL. 11,12 The Berlin-Frankfurt-Munich protocol for pediatric Burkitt lymphoma/leukemia is promising. Few patients (7.9%; 157/1,979) demonstrate disease progression or relapse. 13 Consistent with the World Health Organization consensus, which classified BL as a leukemic variant of Burkitt lymphoma, the TPOG treats patients with BL using the TPOG 10 B-NHL-R4 protocol. 5 By contrast, patients with other forms of B-cell ALL are treated using the ALL protocol. 14 However, TCF3-HLF-positive ALL is highly resistant to conventional chemotherapy and has a high rate of treatment failure despite treatment intensification and hematopoietic stem cell transplantation. 6 Molecular tests are becoming increasingly valuable in disease diagnosis, and genetic markers are widely used for subtype classification and prognostication. Detection of TCF3 rearrangement is crucial in diagnosing ALL, and distinguishing different TCF3 fusion genes is essential in risk stratification. However, conventional cytogenetic analysis may fail to detect t(8;14) or t(17;19) and underestimate cryptic fusion genes. The MLPA telomere kit circumvents this pitfall because of its higher sensitivity in detecting aneuploidy. Furthermore, it has a shorter turnaround time and higher sensitivity than conventional cytogenetics analysis, irrespective of the mitotic index. 15,16 This case report highlights the relevance of discrepant diagnostics, the importance of genetic marker identification, and the need for molecular test development and application. MLPA aids in the identification of copy-number variations and aneuploidy, which may be missed in conventional cytogenetic analysis. Thus, it may be a reasonable supplement to routine diagnostics in patients with childhood B-ALL, especially those with unknown subtypes of B-cell ALL and without metaphases or normal karyotype.