Branko Zevnik

Caspase-3 (CASP3) and caspase-7 (CASP7) are the two major executioner caspases that are proteolytically activated by upstream initiator caspases. They possess almost indistinguishable activity, which has led to the overall view that these caspases have functionally redundant roles. Here, we generate knock-in mice expressing cleavage-resistant CASP3(D175A) or CASP7(D198A). Our results show that proteolytic activation of CASP3 and CASP7 is decisive for their activity in vivo and controls redundant processes during embryonic development as combined expression of both CASP3(D175A) and CASP7(D198A) causes embryonic lethality. In adult mice, however, activation of CASP3 and CASP7 controls different processes in different tissues, without the involvement of apoptosis. While CASP7 activation is required for male fertility by controlling spermatogenesis, CASP3 activation appears crucial for lymphoid tissue development by regulating interferon signalling. Our findings shed light on emerging roles of caspases in non-apoptotic processes and provide impetus for reconsidering their involvement in physiological and pathological conditions.

Table S4 contains the individual mutations of PPMBC and MBC lymphomas.

Table S6 contains clinical parameters and sequencing results of human DLBCL samples.

Table S3 contains B cell receptor repertoire sequencing data and lists the individual sublcones of the dominant clones of MBC, SMBC and PPMBC lymphomas.

Table S1 contains the p values of comparisons of the V gene usage in non-malignant GC B cells between the individual genotypes. These comparisons were done on the family, gene and allele level. V usage was compared for all BCR sequences as well as for isotype subsets.

This file contains the Supplementary Figures S1 to S12.

Table S2 contains a summary of the B cell receptor sequencing data of lymphoma samples. It includes 1) an overview of the investigated samples, 2) a list of characteristics of the dominant clone of each lymphoma sample, 3) a list comparing the frequencies of V family usage in WT samples to the lymphoma frequency, 4) the frequencies of FR and CDR mutations in samples analyzed by full-length BCR sequencing.

Table S5 contains statistical values of gene set enrichment analyses of the KEGG, BIOCARTA and HALLMARK gene set collections.

Genomic profiling revealed the identity of at least 5 subtypes of diffuse large B-cell lymphoma (DLBCL), including the MCD/C5 cluster characterized by aberrations in MYD88, BCL2, PRDM1, and/or SPIB. We generated mouse models harboring B cell–specific Prdm1 or Spib aberrations on the background of oncogenic Myd88 and Bcl2 lesions. We deployed whole-exome sequencing, transcriptome, flow-cytometry, and mass cytometry analyses to demonstrate that Prdm1- or Spib-altered lymphomas display molecular features consistent with prememory B cells and light-zone B cells, whereas lymphomas lacking these alterations were enriched for late light-zone and plasmablast-associated gene sets. Consistent with the phenotypic evidence for increased B cell receptor signaling activity in Prdm1-altered lymphomas, we demonstrate that combined BTK/BCL2 inhibition displays therapeutic activity in mice and in five of six relapsed/refractory DLBCL patients. Moreover, Prdm1-altered lymphomas were immunogenic upon transplantation into immuno-competent hosts, displayed an actionable PD-L1 surface expression, and were sensitive to antimurine-CD19-CAR-T cell therapy, in vivo.Significance:Relapsed/refractory DLBCL remains a major medical challenge, and most of these patients succumb to their disease. Here, we generated mouse models, faithfully recapitulating the biology of MYD88-driven human DLBCL. These models revealed robust preclinical activity of combined BTK/BCL2 inhibition. We confirmed activity of this regimen in pretreated non–GCB-DLBCL patients.See related commentary by Leveille et al., p. 8.This article is highlighted in the In This Issue feature, p. 1