t(14;19)(q32;q13) is found in a fraction of chronic lymphocytic leukemia (CLL) patients and creates the IGH::BCL3 fusion gene, but this translocation has been observed in B-cell lymphomas other than CLL (non-CLL BLs). Ultra-high molecular weight DNA from liquid nitrogen-frozen leukemia cells of one CLL patient and OCT compound-embedded cryopreserved biopsies from two non-CLL BL patients, all of whom carried cytogenetic t(14;19), were subjected to optical genome mapping. In the CLL patient, t(14;19) resulted in fusion between IGHA1 and BCL3, whereas in the non-CLL BL patients, breakpoints on 19q13.32 involved NECTIN2 and BCAM as the IGH partners, both of which were located telomeric to BCL3. On der(19)t(14;19), chromosome 19 sequences centromeric to the breakpoints were fused to the germline IGHVs, or IGHV-D-J rearrangement sequences followed by the 5' Eµ enhancer and IGHM/IGHD constant genes. In the non-CLL BL patients, BCL3 was retained on der(19)t(14;19) and potentially affected by the translocated IGH.

BACKGROUND Six cycles of R-CHOP (rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone) are the standard first-line treatment for diffuse large-B-cell lymphoma (DLBCL). Past studies showed that adding rituximab to CHOP not only improved survival outcome but also reduced central nervous system (CNS) relapse (Boehme et al. Blood. 2009, Villa et al. Ann Oncol. 2010). However, the optimal dose of rituximab has not been fully investigated. Results from a subset analysis from the PETAL trial (Hüttmann et al. Ann Hematol 2019) showed no superiority of six cycles of R-CHOP plus two doses of rituximab (R8-CHOP6) over six cycles of R-CHOP (R6-CHOP6) in event free survival and overall survival (OS), although the population analyzed in the study was limited to patients who were sensitive to first two courses of chemotherapy. To our knowledge, there have been no prior reports addressing the effect of increasing rituximab on the incidence of CNS relapse. We aim to compare progression free survival (PFS), OS, and cumulative incidence of CNS relapse between patients receiving R6-CHOP and R8-CHOP in an observational cohort of patients with newly diagnosed CD20-positive DLBCL. METHODS We performed a retrospective cohort review of patients with newly diagnosed CD20-positive DLBCL receiving R6-CHOP6 or R8-CHOP6 as first-line therapy at two institutions in Japan between May, 2006 and July, 2022. Patients with CNS involvement at the time of diagnosis were excluded from the study cohort. PFS and OS were estimated with the use of the Kaplan-Meier method starting from the date of diagnosis. Differences between groups were assessed by log-rank test. A Cox proportional-hazards regression model that included IPI was used to perform an adjusted analysis. To evaluate the heterogeneity of additional rituximab effect on OS, subgroup analyses according to variables based on disease characteristics at baseline were done. A Cox model was used for each subgroup. The presence of interaction was tested by including an interaction term between the treatment effect and the subgroup covariate of interest. The cumulative incidence of CNS relapse was estimated using the method proposed by Gray. Death without CNS relapse was defined as a competing event and comparisons between groups were done using the Fine and Gray model, adjusted for CNS IPI. RESULTS A total of 453 patients were analyzed, of whom 294 patients received R6-CHOP6 and 159 patients received R8-CHOP6. Median age was 70 years (range, 19-93). The median follow-up time was 52 months (range, 5-198). Baseline patient characteristics in the treatment groups slightly differed in Eastern Cooperative Oncology Group (ECOG) performance-status score ≥ 2 (R6-CHOP6 vs. R8-CHOP6; 26.1% vs. 15.1%) and LDH greater than upper limit of normal (R6-CHOP6 vs. R8-CHOP6; 65.6% vs. 50.3%). The complete response rates were similar between the groups (R6-CHOP6 vs. R8-CHOP6; 91.8% vs. 89.8%). Five-year PFS was 68.6% (95% CI, 62.1-74.2) in the R6-CHOP6 group and 67.8% (95% CI, 59.7-74.6) in the R8-CHOP6 group (HR, 0.96; 95% CI, 0.68-1.36; p=0.838, IPI-adjusted HR, 1.05; 95% CI, 0.74-1.50; p=0.506). Five-year OS was 78.6% (95% CI, 72.3-83.6) in the R6-CHOP6 group and 81.1% (95% CI, 73.8-86.6) in the R8-CHOP6 group (HR, 0.92; 95% CI, 0.90-1.41; p=0.705, IPI-adjusted HR, 1.00; 95% CI, 0.65-1.54; p=0.994). Five-year cumulative incidence of CNS relapse was 7.5% (95% CI, 4.8-11.8) in the R6-CHOP6 group and 6.6% (95% CI, 3.6-12.1) in the R8-CHOP6 group (HR, 0.79; 95% CI, 0.37-1.71; p=0.533, CNS-IPI-adjusted HR, 0.95; 95% CI, 0.44-2.08; p=0.902). R8-CHOP6 was not associated with better OS than R6-CHOP6 in subgroup analyses stratified according to key covariates, including high-risk features. CONCLUSIONS Our populatiion-based cohort study showed that additional two rituximab applications after six cycles of R-CHOP did not improve outcome including CNS relapse. Six cycles of rituximab seem to be optimal in the frontline treatment combined with CHOP for patients with DLBCL.

A woman in her 80 s presented with generalized lymphadenopathy, bone marrow (BM) involvement, and leukemic manifestation. Lymph node biopsy revealed typical histopathology of mantle cell lymphoma (MCL) and the CD5+ and immunoglobulin μ+δ+/λ+ immunophenotype, with unmutated IGHV. BM was infiltrated with not only MCL but also another B-cell tumor that was CD5- and μbright+δ+/κ+, being consistent with M proteins in the serum and urine, with mutated IGHV. As the latter lymphoma component carried the MYD88 L265P mutation, this case represented a composite of MCL and lymphoplasmacytic lymphoma. Next-generation sequencing revealed a cryptic insertion of IGK enhancer sequences into the CCND1-major translocation cluster, accounting for CCND1 expression in MCL cells recognized by immunohistochemistry. Composite lymphoma is rare, but a correct diagnosis is required because effective treatments for each component are now available.

Early disease progression, including histological transformation, can occur in follicular lymphoma, but tumor heterogeneity makes comparison difficult. We analyzed cell-free DNA from 30 patients with follicular lymphoma to assess cell-free DNA concentration and gene mutations. Mutational profiles were also examined in 16 cases using DNA from formalin-fixed, paraffin-embedded tissue samples. Cell-free DNA concentrations were higher in cases with histological transformation. Both the number of mutations per patient and the number of mutated genes were greater in cell-free DNA than in formalin-fixed, paraffin-embedded tissue-derived DNA. Notably, TP53 mutations were more frequently detected in cell-free DNA. CIITA mutations were more commonly observed in cases with a low frequency of somatic hypermutation in VH genes. Cell-free DNA analysis in follicular lymphoma may become a complementary approach, overcoming the limitations of assessing disease status from single biopsy sites. Further studies will examine the usefulness of cell-free DNA in predicting disease progression in follicular lymphoma.

We characterized 5 B -cell tumors carrying t(14;19)(q32;q13) that creates the IGH::BCL3 fusion gene. The patients' ages ranged between 55 and 88 years. Two patients presented with progression or recurrence of B -cell chronic lymphocytic leukemia (B-CLL)/small lymphocytic lymphoma (SLL), two with diffuse large B -cell lymphoma (DLBCL) of non -germinal center B -like phenotype, and the remaining one with composite angioimmunoblastic T -cell lymphoma and Epstein -Barr virus -positive DLBCL. The presence of t(14;19)(q32;q13) was confirmed by fluorescence in situ hybridization (FISH), showing colocalization of 3' IGH and 3' BCL3 probes on der(14)t(14;19) and 5' BCL3 and 5' IGH probes on der(19)t(14;19). One B-CLL case had t(2;14)(p13;q32)/IGH::BCL11A, and 2 DLBCL cases had t(8;14)(q24;q32) or t(8;11;14)(q24;q11;q32), both of which generated IGH::MYC by FISH, and showed nuclear expression of MYC and BCL3 by immunohistochemistry. The IGH::BCL3 fusion gene was amplified by long-distance polymerase chain reaction in 2 B-CLL/SLL cases and the breakpoints occurred immediately 5' of BCL3 exon 1 and within the switch region associated with IGHA1. The 5 cases shared IGHV preferentially used in B-CLL cells, but the genes were unmutated in 2 B-CLL/SLL cases and significantly mutated in the remaining 3. B -cell tumors with t(14;19)(q32;q13) can be divided into B-CLL/SLL and DLBCL groups, and the anatomy ofIGH::BCL3 in the latter may be different from that of the former.

We describe two types of IGH::BCL2 breakpoints involving the 5' region of BCL2 (5' BCL2). One was ins(14;18)(q32;q21q21) observed in 2 follicular lymphoma (FL) cases, in which IGH was cleaved at 3' of IGHD and 5' of IGHJ and BCL2 was cleaved at 5' BCL2 and downstream regions, and a 281- or 201-kilobase pair fragment containing the BCL2 protein-coding sequences was invertedly inserted into IGH. In another type observed in 2 FL and 2 chronic lymphocytic leukemia (CLL) cases, breakage and reunion occurred within the switch region associated with IGHM (Sµ) and 5' BCL2, creating IGH Sµ::5' BCL2 fusion sequences on der(18)t(14;18)(q32;q21). The former is considered to be mediated by VDJ-recombination, while the latter by the class switch recombination process. There were no particular features in FL or CLL cases with IGH::5' BCL2 breakpoints compared with those with t(14;18)(q32;q21)/IGH::BCL2 involving the 3' breakpoint cluster regions.

Objective Testing for the Janus activating kinase 2 ( JAK2 ) V617F mutation is important for diagnosing and treating myeloproliferative neoplasms (MPNs). Recently, urine cell-free DNA (ucfDNA) was reported to be useful for detecting tumor-specific gene mutations in several solid tumors. However, its utility in detecting such mutations in hematological malignancies has not yet been assessed. In this study, we assessed whether or not the JAK2 V617F mutation could be detected in ucfDNA and whether or not its positivity rate in ucfDNA was associated with the JAK2 V617F allele ratio of peripheral blood cells in patients with MPN. Methods The JAK2 V617F allele ratio of genomic DNA from peripheral blood cells was determined using quantitative polymerase chain reaction (qPCR) or droplet digital PCR (ddPCR). ucfDNA was subjected to ddPCR. The correlation between the JAK2 V617F mutation positivity rates of blood-derived DNA and those of ucfDNA was assessed. Materials Twelve patients with polycythemia vera and 12 patients with essential thrombocythemia were enrolled. Ethylenediaminetetraacetic acid-treated peripheral blood (100 mL) and 15-30 mL of fresh urine were used. Results The JAK2 V617F mutation was detected in the ucfDNA from all 20 JAK2 V617F mutation-positive patients. In addition, the JAK2 V617F mutation positivity rate of ucfDNA was correlated with the JAK2 V617F allele ratio of blood-derived DNA, including in both estimated glomerular filtration rate (eGFR) groups (patients with an eGFR >= 50 or <50 mL/min/1.73 m (2) ). Conclusion Our results indicate that ucfDNA is a valuable tool for diagnosing and monitoring MPN. Given these findings, other disease-specific gene mutations in hematological malignancies may also be detectable in ucfDNA.