Elsevier

Cancer Genetics

Volume 240, January 2020, Pages 40-44
Cancer Genetics

Case Report
Molecular and phenotypic characterization of an early T-cell precursor acute lymphoblastic lymphoma harboring PICALM-MLLT10 fusion with aberrant expression of B-cell antigens

https://doi.org/10.1016/j.cancergen.2019.11.002Get rights and content

Summary

T-cell acute lymphoblastic leukemia/lymphoma (T-ALL/LBL) is usually diagnosed based on the presence of immature lymphoid marker terminal deoxynucleotidyl transferase (TdT), and T-cell specific markers, specifically CD3, by immunohistochemistry (IHC) staining on bone marrow and/or extramedullary tissue. We present a novel, TdT and CD3 negative, aggressive early T-cell precursor LBL (ETP-LBL) initially misdiagnosed as a high grade B-cell lymphoma due to expression of CD79a and the erroneous detection of BCL2/IGH fusion. The patient was eventually evaluated using molecular diagnostic techniques, including fluorescence in situ hybridization (FISH) and next generation sequencing (NGS) assays that demonstrated PICALM-MLLT10 fusion and a NOTCH1 mutation in the absence of BCL2/IGH fusion. The use of NGS, specifically mate-pair sequencing (MPseq), subsequently confirmed an in-frame PICALM-MLLT10 fusion. Our retrospective analysis showed that PICALM-MLLT10 fusion has no association with CD3/TdT negativity, as 6/49 T-ALL/LBL cases from Mayo Clinic database (01/1998–09/2018), including this case, were noted to have PICALM-MLLT10 fusion; however, none of the other cases were associated with CD3/TdT negativity. We emphasize the importance of a comprehensive hematopathologic evaluation including multiple molecular studies for the appropriate interrogation and classification of a difficult acute leukemia diagnosis, and to prevent potential diagnostic errors of clinical significance.

Introduction

Approximately 5,930 new cases and 1,500 deaths are expected from acute lymphoblastic leukemia/lymphoma (ALL/LBL) in 2019 [1]. Acute lymphoblastic lymphoma comprises approximately 1–2% of all non-Hodgkin lymphomas. Over 90% of LBLs are of T-cell lineage, and are often difficult to diagnose and treat in adults [2]. T- LBL is an aggressive neoplasm of immature T-cells, and characteristically presents as a large mediastinal mass [3, 4]. The WHO classification currently includes LBL under the same classification as ALL; the presence of >25% leukemic cells in the bone marrow distinguishes ALL from LBL [5]. Morphology and immunophenotyping are usually sufficient to differentiate T-LBL from other non-Hodgkin lymphoma and mediastinal tumors. Occasionally, combinations of morphology, immunophenotyping, cytogenetics, FISH, and/or molecular studies are required to confirm the diagnosis of this rapidly progressive disease. Our case required all these diagnostic approaches of which fluorescence in situ hybridization (FISH) and a next generation sequencing (NGS) strategy, mate-pair sequencing (MPseq), provided important results for a diagnosis of early T-cell precursor -LBL (ETP-LBL).

A 41-year-old, previously healthy male presented to the emergency room of an outside hospital with progressive fatigue, weight loss, chest discomfort, dry cough and two episodes of syncope over the preceding week. Computed tomography (CT) of the chest showed a large 3.4 × 4.6 × 5.9 cm infiltrating mass involving the anterior and middle mediastinum, with encasement of the great vessels, thickening of the pericardium and epicardium, moderate sized pericardial effusion, and small bilateral pleural effusions. [Fig. 1] CT guided core needle biopsy was obtained initially at the outside institution, and was later repeated at our institution for a definitive pathological diagnosis. The H&E sections showed diffusely infiltrating, small to medium sized, neoplastic lymphocytes with necrosis and marked crush artifact. The small foci of viable tissue showed neoplastic lymphocytes with high nucleus to cytoplasm ratio, open chromatin, and brisk mitosis. [Fig. 2A] IHC staining was positive for CD34, CD79a, CD43, CD33, CD7, [Fig. 2B–F] BCL2, and MYC with a high proliferative rate by Ki-67 (>90%). The malignant cells were negative for CD20, CD19, CD22, PAX-5, CD3, CD5, CD4, CD8, CD1a, CD2, CD279, CD45 RO, CD99, TdT, MPO, CD30, CD68 (KP1 and PGM1), CD117, CD13, CD11c, CD14, CD10, BCL6, MUM-1, CD123, CD163, CD138, and HHV8. EBER in situ hybridization for EBV was negative. At the outside institution, FISH was reported positive for BCL2/IGH fusion and the patient was subsequently diagnosed with a high-grade CD20-negative B-cell lymphoma.

The patient was referred to our institution for definitive treatment and the outside slides were reviewed and a repeat CT guided biopsy of the mediastinal mass was obtained. Morphology and immunophenotyping yielded the same results as previously reported and remained inconclusive. A presumptive diagnosis of TdT-negative B-LBL versus high-grade B-cell lymphoma was considered. However, B-LBL presenting as a large tumor confined to the mediastinum is unusual and pathologic findings did not fit with mediastinal large B-cell lymphoma. As such, additional diagnostic testing was performed. Molecular studies by polymerase chain reaction for gene rearrangements of immunoglobulin heavy chain and kappa light chain were negative and T-cell receptor gene rearrangements for beta and gamma genes were equivocal. [Table 1] Internal FISH studies using an IGH/BCL2 dual-color dual-fusion probe were negative, and did not confirm the IGH/BCL2 fusion reported by the other institution. FISH studies with break-apart and fusion probes for MYC, BCL2, BCL6, MYC/IGH, MYC/IGK, and MYC/IGL were also negative. The remaining FISH probes demonstrated gains with 3 − 4 copies of most probe sets [Table 1]. A T-ALL FISH panel identified PICALM-MLLT10 fusion in 100% of nuclei, [Fig. 3] and also identified a CDKN2A deletion (at 9p21) in 73% of nuclei. A comprehensive B-ALL FISH panel was normal. The combination of the clinical presentation, cell morphology, ambiguous immunophenotype, and PICALM-MLLT10 fusion identified by FISH resulted in a final diagnosis of ETP-LBL. [Table 1] At this time, there was no evidence of disease outside of the mediastinum or bone marrow involvement on biopsy. MRI of the brain and the cervical spine were negative and cerebrospinal fluid (CSF) cytology was negative for lymphoblasts.

The patient was started on hyper-CVAD (Cyclophosphamide, Vincristine, Adriamycin, Dexamethasone) with high dose intravenous methotrexate (MTX) and cytarabine, along with prophylactic intrathecal MTX and cytarabine. Treatment course was complicated by pulmonary embolism, pneumocystis (PCP) pneumonia, and neutropenic fever. The patient had stable disease after 4 cycles of chemotherapy. Following cycle 5, patient presented with dyspnea and was found to have superior vena cava syndrome (SVC), consistent with disease progression, and received urgent radiation treatment. [Fig. 1] A repeat bone marrow biopsy showed 90% T-lymphoblastic involvement with a similar immunophenotype to that seen on the initial mediastinal biopsy. A bone marrow chromosome study identified 3 normal and 17 abnormal metaphases with a complex, hypo-tetraploid karyotype, including two copies of a t(10;11)(p13;q21) and several additional abnormalities. FISH studies confirmed PICALM-MLLT10 fusion associated with the t(10;11)(p13;q21). To further characterize the PICALM-MLLT10 fusion, MPseq was performed and revealed a t(10;11)(p12.31;q14.2) with breakpoints located within intron 3 of MLLT10 (NM_004641) and intron 19 of PICALM (NM_007166), resulting in a predicted in-frame PICALM-MLLT10 fusion. [Fig. 3] NGS sequencing also showed activated NOTCH1 [(Chr9(GRCh37):g.139390621dup; NM_017617.3 (NOTCH1): c.7570dup; p.Ser2524Phefs*82 (19%)] and TP53 [Chr17(GRCh37):g.7577587T>A; NM_000546.4 (TP53): c.694A>T; p.Ile232Phe (44%)] mutations. The patient was given second line treatment with ifosfamide, carboplatin, and etoposide (ICE) for one cycle, and eventually started on the BCL-2 inhibitor venetoclax and remained on this treatment regimen for the remainder of the hospitalization. Unfortunately, the patient developed septic shock in the setting of neutropenia and passed away within a month of disease progression to leukemia.

To evaluate if PICALM-MLLT10 fusion has association with CD3/TdT negativity in patients with T-ALL/LBL, we retrospectively analyzed patients with T-ALL/LBL from the Mayo Clinic database (01/1998–09/2018) after getting an approval from the institutional review board (IRB). Six of the 49 cases (12%), including this case, were noted to have PICALM-MLLT10 fusion; however, none of the other cases were associated with CD3/TdT negativity.

Section snippets

Discussion

The differential diagnosis for an adult with a primary mediastinal mass includes multiple hematopoietic entities [3]. T-LBL should be strongly suspected in patients presenting as a lymphoma confined to the mediastinum in the absence of diagnostic features for primary mediastinal large B-cell lymphoma or classical Hodgkin lymphoma. B-LBL presenting as a mediastinum-confined lymphoma is rare as our literature review identified only a single patient case [6]. A comprehensive pathologic and

Conclusion

In conclusion, we report an aggressive case of a TdT, CD3 negative and CD79a, CD33, and CD34 positive, ETP-LBL with PICALM-MLLT10 fusion detected by FISH and confirmed by MPseq. Our retrospective analysis showed no association between PICALM-MLLT10 fusion and CD3/TdT negativity in T-ALL/LBL patients. This case emphasizes the importance of including a comprehensive genomic and molecular interrogation in the diagnostic evaluation of patients with an acute leukemia of ambiguous immunophenotype. In

Author contributions

SK, MEM, RPK, JFP and JJ wrote the manuscript, JJ, RPK, JFP, IML and GV conducted the pathologic diagnosis and genomic research, HWT, JMF and JJ critically reviewed and edited the manuscript. The final manuscript was reviewed and approved by all the authors.

Funding

None.

Declaration of Competing Interest

SK, MEM, LJ, RPK, JFP, IML, HWT, JMF: no financial disclosures. GV: Algorithms described in this manuscript for mate-pair sequencing are licensed to WholeGenome LLC owned by GV.

Acknowledgements

None.

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