Mohammad Hussaini

Myeloid/lymphoid neoplasms with FGFR1 rearrangement

Abstract Myeloid/lymphoid neoplasms with FGFR1 rearrangement are a rare entity. We present a multicenter experience of 17 patients with FISH-confirmed FGFR1 rearrangement. The clinical presentation at diagnosis included myeloproliferative neoplasm (MPN) in 4 (24%) patients, acute leukemia (AL) in 7 (41%), and concomitant MPN with AL in 6 (35%). The two most frequently observed cytogenetic abnormalities were t(8;13)(p11.2;q12)(partner gene ZMYM2) and t(8;22)(p11.2; q11.2)(BCR). Seventy-eight percent of tested patients had a RUNX1 mutation, of whom all had AL. Overall response rate to frontline therapy was 69%, and 76% of patients subsequently received allogeneic stem cell transplant (ASCT). After a median follow-up of 11 months, median progression-free survival was 15 months and median overall survival was not reached. In conclusion, FGFR1-rearranged hematologic malignancies present with features of MPN and/or AL. FGFR1 and RUNX1 are therapeutic targets for ongoing and future clinical trials.

Comparison of the Mutational Profiles of Primary Myelofibrosis, Polycythemia Vera, and Essential Thrombocytosis

Abstract Objectives: To compare the mutational profiles of patients with primary myelofibrosis (PMF), polycythemia vera (PV), and essential thrombocytosis (ET). Methods: Next-generation sequencing results of 75 cases of PMF, 33 cases of PV, and 27 cases of ET were compared. Results: Mutation rates of ASXL1 and SRSF2 were significantly higher in PMF than in PV or ET. ASXL1 mutations appeared to be more frequently associated with risk of transformation to acute myeloid leukemia than JAK2 or TET2 mutations. The most common mutation-cytogenetic combinations in myeloproliferative neoplasm (MPN) were mutations of JAK2 or ASXL1 with del(20q) and were more common in patients with PMF and PV than in patients with ET. Differences were also found between patients with PMF and PV. Conclusions: PMF, PV, and ET show different mutational profiles, which may be helpful in resolving the differential diagnosis between MPNs. Due to the relatively small number of cases and variable testing over time, larger controlled studies are necessary to confirm the findings.

Allogeneic hematopoietic cell transplantation in T-cell prolymphocytic leukemia: A single-center experience

BACKGROUND T- cell prolymphocytic leukemia (T- PLL) is a rare aggressive hematological malignancy. Alemtuzumab, an anti-CD52 humanized monoclonal antibody, is the treatment of choice for remission induction. Allogeneic hematopoietic cell transplantation (allo-HCT) has been described to induce durable remissions and improve survival, but data is limited. PATIENTS AND METHODS We evaluated clinical outcomes of 11 patients, median age of 56 (range, 43-71) years who underwent allo-HCT for T-PLL. The majority of cases were in the first complete remission (CR1 = 9, CR2 = 1, second partial response PR2 = 1) at time of allo-HCT. Myeloablative conditioning was the most commonly prescribed preparative regimen (n = 8, 73%) and tacrolimus plus sirolimus was most commonly prescribed regimen for graft-versus-host disease prophylaxis (n = 5, 46%). RESULTS The median follow-up for surviving patients was 48 (range, 6-123) months. The 4-year progression-free survival (PFS) and overall survival (OS) were 45% (95% confidence interval (CI) = 13-78%) and 56% (95% CI = 24-89%), respectively. Cumulative incidence of non-relapse mortality (NRM) at 4-year post-transplantation was 34% (95%CI = 14-85%). The 4-year cumulative incidence of relapse/progression was 21% (95% CI = 6-71%). CONCLUSION Allo-HCT is an effective treatment for T-PLL. Patients must be evaluated for their candidacy for allo-HCT as soon as the diagnosis is confirmed. Efforts are needed to decrease NRM and relapse.

Transformation of T-Cell Acute Lymphoblastic Lymphoma to Peripheral T-Cell Lymphoma: A Report of Two Cases

Nonhepatosplenic/noncutaneous γδ peripheral T-cell lymphoma (NHNCγδ PTCL) represents a miscellaneous group of unrelated T-cell lymphomas of which only isolated cases have been reported. We describe two cases of transformation from T-lymphoblastic leukemia/lymphoma to NHNCγδ PTCL. Transformation into more aggressive disease is a rare event in T-cell lineage-derived hematologic malignancies compared to B-cell neoplasms. Nevertheless, both of our cases involved relapse as PTCL manifested with skin involvement and an overt shift from blastic morphology to large granular leukemia-like mature T cells. Among other notable molecular characteristics, expression of immature markers such as TdT was lost in both cases. Based on cytogenetics, phenotype, and morphology, both patients represent a novel phenomenon of clonal transformation from T-ALL to PTCL which has rarely been reported in the literature. Such transformation may carry important diagnostic and biological implications.

EBV-positive Richter's syndrome with laboratory features of Burkitt's lymphoma, in Ibrutinib-treated chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is a malignant proliferation of small clonal B-lymphoid cells with distinctive morphologic and immunophenotypic features, including clumped chromatin, few nucleolated larger cells (prolymphocytes), and aberrant coexpression of CD5 and CD23.[1] A minority of patients with CLL (2–10%) develop an aggressive large B-cell neoplasm, which in most cases has morphologic features of diffuse large B-cell lymphoma (90%, a.k.a. Richter’s syndrome [RS]), but can also manifest as Hodgkin lymphoma (10%) or, rarely, prolymphocytic leukemia (<1%).[2] RS has been increasingly recognized as an important resistance mechanism to the Bruton tyrosine kinase inhibitor Ibrutinib, accounting for approximately half of cases of disease progression on therapy.[3] The occurrence of RS in Ibrutinib-treated CLL is likely not coincidental, as most cases occur early during therapy and have a much worse overall survival than preIbrutinib RS (3.5 months vs. 8–30 months, respectively).[3] Nevertheless, the mechanism of RS-associated Ibrutinib resistance remains to be fully elucidated, as the BTK and PLCG2 mutations responsible for Ibrutinib resistance in untransformed CLL are largely absent in this setting. We, herein, report a unique case of RS in Ibrutinib-treated CLL, exhibiting cytologic, immunophenotypic and cytogenetic features of Burkitt’s lymphoma. To our knowledge, this is the first case report of RS with features of CD10-positive/MYC-translocated Burkitt’s lymphoma in the modern literature. A 57-year-old man with a history of hypertension, dyslipidemia, and nephrolithiasis was found to have asymptomatic lymphocytosis of small mature lymphocytes with clumped chromatin (absolute lymphocyte count 1⁄413.1 10/L). Peripheral blood flow cytometry showed a predominance of CD5-positive small clonal B-cells, which were positive for CD5, CD20 (dim), CD23, and lambda light chain (dim), but negative for FMC-7. A bone marrow biopsy revealed extensive involvement by the same process. Karyotype showed deletion of chromosomes 13 and 17 (Table 1). A diagnosis of CLL was established and the patient was initially managed with watchful waiting. On a follow up visit 16 months later, the patient was found to have worsening thrombocytopenia, generalized lymphadenopathy, marked splenomegaly, and B symptoms. A bone marrow biopsy showed extensive involvement by CLL and clonal evolution by chromosomal analysis (Table 1, 16 months after diagnosis), but no evidence of large cell transformation. A month later, he was treated with one cycle of fludarabine, cyclophosphamide, and rituximab (FCR), without significant response. Peripheral blood work up at this point showed lymphocytosis of 17.3 10/L with persistent CLL, ZAP-70-positivity by flow cytometry, and unmutated IGVH. Six months after starting of FCR, the patient was enrolled in a clinical trial with Ofatumumab, high doses of methylprednisone and lenalidomide (HiLOG), resulting in significant clinical improvement and stabilization of disease. A follow up bone marrow biopsy at 2 months of HiLOG therapy showed persistent CLL and further clonal evolution by karyotypic analysis (Table 1, 25 months after diagnosis), in the absence of large cell transformation or clinical progression. After 9 months of HiLOG therapy, the patient was noted to have bulky axillary lymphadenopathy, worsening transfusion-dependent cytopenias, and severe B symptoms. Routine peripheral blood values showed a white blood cell count of 15.3 10/L, hemoglobin of 980 g/L, platelets of 73 10/L, and lactate dehydrogenase of 331U/L. A peripheral blood smear showed persistent CLL with 2% prolymphocytes. A needle core biopsy performed on the largest right axillary lymph node (7 cm) showed persistent CLL without evidence of large cell transformation (Figure 1(A)). Similarly, a bone marrow biopsy showed involvement by CLL with no large cells. In-situ hybridization for EBV-encoded small RNAs (EBER) on the bone marrow biopsy showed no Epstein-Barr virus (EBV)-positive cells. Chromosomal analysis on the bone marrow aspirate demonstrated a highly complex karyotype with clonal evolution (Table 1, 32 months

Detection of Novel t(12;17)(p12;p13) in Relapsed Refractory Acute Myeloid Leukemia by Anchored Multiplex PCR(AMP)-based Next-Generation Sequencing.

Although several technologies can be used to detect gene fusions, anchored multiplex PCR next-generation sequencing (AMP-NGS) offers the advantage of novel fusion detection and the ability to multiplex multitudinous genes. We applied AMP-NGS technology in the evaluation of a 56-year-old gentleman with myelodysplastic syndrome transformed acute myeloid leukemia (AML). Patient was initially diagnosed with low-risk myelodysplastic syndrome-refractory cytopenias and multilineage dysplasia (MDS-RCMD), progressed to AML after failing hypomethylating agent therapy. At progression patients had normal cytogenetics but NGS profiling showed ETV6 c.416_417del CT frame shift and U2AF1 S34F mutations. Patient attains brief remission of 2 months after induction chemotherapy and then he was refractory to 2 salvage chemotherapy regimens. Reassessment after failing second salvage, identified t(12;17)(p13;p13)[20] by karyotype. It was postulated that the 12p13 locus might represent a new rearrangement of ETV6. AMP-NGS confirmed involvement of the ETV6 with discovery of a novel fusion partner, HIC1. The detection of the novel fusion partners was supported by the breakpoints originally observed by karyotype. This discovery of ETV6-HIC1 gene fusion by AMP-NGS technology provided new insight into a leukemogenic pathway in AML. Future use of this technology can serve as an adjunct tool in workup of patients with AML and can also help in formulating therapeutic strategies.

Abstract 3984: Novel t(X;21)(q26;q22) detected in a case of acute unclassifiable leukemia by application of anchored multiplex PCR-based next-generation sequencing

In this study, we demonstrate real-time clinical application of novel anchored multiplex PCR (AMP) technology in hematologic disease. A 78-year-old man with a history of hairy cell leukemia treated with Cladribine (2013) and bladder cancer (2014) developed dyspnea in August 2015. Work up revealed bicytopenia and leukocytosis. Outside bone marrow biopsy was read as B lymphoblastic leukemia/lymphoma. Rebiopsy was performed at our NCI-designated cancer center with extensive work up resulted in reclassification as acute unclassifiable leukemia (rare) with deferral to molecular diagnostics for guidance regarding classification. MLL, BCR-ABL, and MDS FISH panel were negative. Next generation sequencing (NGS) interrogating more than 400 genes showed FLT3-ITD, truncation of RUNX1 exon 6, ASXL1 p.G646Wfs*12, MLL p.V297L, and EZH2 p.V674M and p.C443Y mutations. The molecular profile favored therapy-related AML. Interestingly, cytogenetics revealed 46,Y,t(X;21)(q26;q22)[20]. FISH confirmed RUNX1 rearrangement but the partner was unknown. The t(X;21)(q26;q22) appears to be novel. However, the rare t(3;21)(q26.2;q22) has been cited to segregate with therapy-related MDS/AML. Subsequent aberrant activation of RUNX1 and MECOM is postulated to be pivotal in pathogenesis. Therefore, it is of great interest to determine the translocation partner associated with RUNX1 in our case. This type of analysis is not easily amenable to either Sanger or conventional NGS approaches. Fortuitously, with the recent advent of AMP technology (Archer FusionPlex) novel fusion detection has become possible. AMP was key in the characterization of this sample and promotes discovery of novel fusions by targeting one (known) of the two genes involved in a translocation event. Application of this cutting edge technology lead to the discovery of the novel fusion RUNX1-G6PD in this case. The identity of the novel fusion partners was supported by the breakpoints originally observed by the karyotype analysis. G6PD has been previously reported to be upregulated in acute leukemia. This discovery raises the possibility convergent pathogenic pathways and carries potential biological, diagnostic, and therapeutic implications. Citation Format: Laura Johnson, Helen Wang, Katelyn Trifilo, Brian Kudlow, Peter R. Pappenhausen, Mohammad Hussaini. Novel t(X;21)(q26;q22) detected in a case of acute unclassifiable leukemia by application of anchored multiplex PCR-based next-generation sequencing. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3984.

Abstract 3988: Detection of novel t(12;17)(p12;p13) in treatment-refractory/relapsed acute myeloid leukemia by anchored multiplex PCR(AMP)-based next-generation sequencing

Gene fusions are an important class of mutations and have been shown to drive many genetic diseases. While several technologies can be used to detect fusions, anchored multiplex PCR (AMP) next generation sequencing (NGS)-based detection offers the advantages of novel fusion detection and the ability to multiplex multitudinous genes. Here we report application of this technology in the evaluation of a 56-year-old man diagnosed with AML (2013). He was treated initially with Vidaza and later with CLAG-M induction (2014) with complete response and consolidation CLA (2015). He relapsed (4/2015) and was reinduced with 7+3 (Ida). He relapsed again and was reinduced with FCT but bone marrow biopsy showed persistent disease (75% blasts). Karyotype showed 46, XY, t(12;17)(p12;p13)[20]. It was postulated that ETV6 oncogene was involved based on 12p13 locus. The partner gene was unknown in this patient with multiply relapsed/refractory disease. AMP technology (Archer FusionPlex) confirmed involvement of the expected ETV6. However, more importantly, employing this technology lead to the discovery of a novel fusion partner, HIC1. The identity of the novel fusion partners was supported by the breakpoints originally observed by karyotype analysis. AMP promotes discovery of novel fusions by targeting one (known) of two genes involved in the fusion event. HIC1 regulates cellular growth and serves as a tumor suppressor gene. Inactivation of HIC1 has been associated with aggressive disease and poor survival in other cancer types. Interestingly, in vitro restoration of gene function by 5-aza-dC resulted in decreased cell proliferation and tumor aggressiveness in pancreatic cancer and head and neck squamous cell carcinoma suggesting the prospect of a therapeutic target. This discovery provides new insight into a possible leukemogenic pathway in AML and potential for targeted therapy. Citation Format: Laura Johnson, Katelyn Trifilo, Helen Wang, Brian Kudlow, Eric Padron, Peter R. Pappenhausen, Mohammad Hussaini. Detection of novel t(12;17)(p12;p13) in treatment-refractory/relapsed acute myeloid leukemia by anchored multiplex PCR(AMP)-based next-generation sequencing. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3988.