Shestakova, Anna

Large B-cell lymphoma with IRF4 rearrangement (LBCL-IRF4) predominantly affects children and young adults (CAYA) and presents as localized disease with excellent prognosis. Although most tumors harbor IRF4 rearrangements (IRF4-R), the existence of cryptic rearrangements has been suggested. Whether IRF4-R LBCL in adults represents the same biological entity remains unclear. To address this question, 35 CAYA patients diagnosed with LBCL-IRF4 and 7 adult (>40-year-old) LBCL with IRF4-R were investigated by an integrative molecular approach. Targeted sequencing structural variant analysis (SV-NGS) confirmed the IRF4-R in 84% of investigated cases. In CAYA, an enrichment (55% vs 29% in adults) of translocations involving IGH, with a breakpoint cluster 3' to IRF4 (EXOC2) was observed, including a cryptic insertion of IGHM-IGHJ5 into IRF4 locus. Novel translocations, IRF4::MIR142 in two CAYA (7%) and IRF4::BATF in one adult, were discovered. Whole-exome sequencing analysis identified unreported mutations in LBCL-IRF4 as YY1 and ZC3H12A (11% each). The high incidence of mutations affecting the B-cell receptor/NF-KB pathways in LBCL-IRF4 was confirmed in cases with diffuse component only. None of the CAYA tumors were classified by LymphGen tool. Adult IRF4-R LBCL showed higher levels of genetic complexity and a distinct mutational profile with mutations on KMT2D and DTX1 (43% each), and MYD88-L265P (29%), and were predicted as EZB or MCD (29% each). These differential genetic features were in line with different clinical presentations (extranodal involvement and advanced stage). In conclusion, IRF4-R architecture and mutational profile differ according to age, supporting that not all cases should be classified as LBCL-IRF4.

ABSTRACT Background Chronic myelomonocytic leukemia (CMML) is largely a disease of individuals older than 50 years; its occurrence in patients aged 50 years and younger is rare. The authors sought to characterize the clinicopathologic features and genetic profile of young‐onset (age 50 years and younger) CMML (y‐CMML) compared with older onset (older than 50 years) CMML (o‐CMML). Methods The authors conducted a multi‐institutional study of 32 patients with y‐CMML and 119 patients with o‐CMML, analyzing clinical, morphologic, cytogenetic, and molecular data. Results Patients who had y‐CMML frequently presented with splenomegaly (59% vs. 11%; p < .001) and hepatomegaly (25% vs. 2%; p < .001) compared with those who had o‐CMML. Although the overall incidence of cytogenetic abnormalities was similar, patients who had y‐CMML exhibited distinct mutational patterns. PTPN11 mutations were significantly more common in y‐CMML (13% vs. 3%; p = .034) than in o‐CMML, whereas TET2 (6% vs. 57%; p < .001) and SRSF2 (16% vs. 46%; p = .002) mutations were significantly less frequent. In addition, multi‐hit TET2 (6% vs. 34%; p = .002) and TET2‐SRSF2 (3% vs. 29%; p = .003) co‐mutations were rare in y‐CMML. However, the molecular differences did not affect the CMML‐specific risk‐stratification categories because the clinical outcomes, including overall survival and leukemia‐free survival, did not differ between the two groups. Conclusions CMML in patients aged 50 years and younger is rare and presents with unique clinical and mutational features, suggesting a distinct pathogenesis.

ESCRT-III undergoes dynamic assembly and disassembly to facilitate membrane exvagination processes including multivesicular body (MVB) formation, enveloped virus budding, and membrane abscission during cytokinesis. The AAA-ATPase Vps4 is required for ESCRT-III disassembly, however the coordination of Vps4 ATP hydrolysis with ESCRT-III binding and disassembly is not understood. Vps4 ATP hydrolysis has been proposed to execute ESCRT-III disassembly as either a stable oligomer or an unstable oligomer whose dissociation drives ESCRT-III disassembly. An in vitro ESCRT-III disassembly assay was developed to analyze Vps4 function during this process. The studies presented here support a model in which Vps4 acts as a stable oligomer during ATP hydrolysis and ESCRT-III disassembly. Moreover, Vps4 oligomer binding to ESCRT-III induces coordination of ATP hydrolysis at the level of individual Vps4 subunits. These results suggest that Vps4 functions as a stable oligomer that acts upon individual ESCRT-III subunits to facilitate ESCRT-III disassembly.

Vps4 is a key enzyme that functions in endosomal protein trafficking, cytokinesis, and retroviral budding. Vps4 activity is regulated by its recruitment from the cytoplasm to ESCRT-III, where the protein oligomerizes into an active ATPase. The recruitment and oligomerization steps are mediated by a complex network of at least 12 distinct interactions between Vps4, ESCRT-III, Ist1, Vta1, and Did2. The order of events leading to active, ESCRT-III–associated Vps4 is poorly understood. In this study we present a systematic in vivo analysis of the Vps4 interaction network. The data demonstrated a high degree of redundancy in the network. Although no single interaction was found to be essential for the localization or activity of Vps4, certain interactions proved more important than others. The most significant among these were the binding of Vps4 to Vta1 and to the ESCRT-III subunits Vps2 and Snf7. In our model we propose the formation of a recruitment complex in the cytoplasm that is composed of Did2-Ist1-Vps4, which upon binding to ESCRT-III recruits Vta1. Vta1 in turn is predicted to cause a rearrangement of the Vps4 interactions that initiates the assembly of the active Vps4 oligomer.