Multiple myeloma (MM), a tumor of germinal center (GC)-experienced plasma cells, comprises distinct genetic subgroups, such as the t(11;14)/CCND1 and the t(4;14)/MMSET subtype. We have generated genetically defined, subgroup-specific MM models by the GC B cell-specific coactivation of mouse Ccnd1 or MMSET with a constitutively active Ikk2 mutant, mimicking the secondary NF-κB activation frequently seen in human MM. Ccnd1/Ikk2ca and MMSET/Ikk2ca mice developed a pronounced, clonally restricted plasma cell outgrowth with age, accompanied by serum M spikes, bone marrow insufficiency, and bone lesions. The transgenic plasma cells could be propagated in vivo and showed distinct transcriptional profiles, resembling their human MM counterparts. Thus, we show that targeting the expression of genes involved in MM subgroup-specific chromosomal translocations into mouse GC B cells translates into distinct MM-like diseases that recapitulate key features of the human tumors, opening the way to a better understanding of the pathogenesis and therapeutic vulnerabilities of different MM subgroups.

Biomechanical alterations contribute to the decreased regenerative capacity of hematopoietic stem cells (HSCs) upon aging. RhoA is a key regulator of mechanosignaling, but its role in mechanotransduction in stem cell aging remains unclear. Here we show that murine HSCs respond to increased nuclear envelope (NE) tension by inducing NE translocation of P-cPLA2, which cell-intrinsically activates RhoA. Aged HSCs experience physiologically higher intrinsic NE tension, but reducing RhoA activity lowers NE tension in aged HSCs. Feature image analysis of HSC nuclei reveals that chromatin remodeling is associated with RhoA inhibition, including restoration of youthful levels of the heterochromatin marker H3K9me2 and a decrease in chromatin accessibility and transcription at retrotransposons. Finally, we demonstrate that RhoA inhibition upregulates Klf4 expression and transcriptional activity, improving aged HSC regenerative capacity and lympho/myeloid skewing in vivo. Together, our data outline an intrinsic RhoA-dependent mechanosignaling axis, which can be pharmacologically targeted to restore aged stem cell function.

Background/Objectives: Genetic and epigenetic alterations accumulate throughout life in hematopoietic stem and progenitor cells (HSPCs), leading to an increased risk of age-related hematological malignancies, such as acute myeloid leukemia (AML). While a few epigenetic drugs have entered clinical practice, studies focusing on histone post-translational modifications (PTMs) altered in AML remain limited. Methods: Here, we used murine HSPCs, murine AML mouse models, human leukemic cell lines, and leukemic patient samples to investigate whether targeting the methylation of histone 3 at lysine 9 (H3K9 methylation) affects HSPCs upon aging and leukemogenesis and might represent a possible target. Results: Our data show that H3K9 methylation changes upon aging in HSPCs and is linked to a pre-malignant phenotype. Low levels of H3K9 methylation in leukemic cells are required to maintain proliferative capacities in vitro. Increasing H3K9 methylation reduces the leukemogenesis of both young and aged murine and patient-derived leukemic cells. Conclusions: Thus, H3K9 methylation might be a potential and selective therapeutic target in AML patients.

Glioblastoma are incurable primary tumors of the central nervous system. The standard of care for newly diagnosed glioblastoma outside clinical trials includes a combination of radiation therapy, alkylating chemotherapy and tumor-treating fields. Yet, for progressive glioblastoma, only lomustine is registered in Europe. The REGOMA trial (Lombardi et al., Lancet Oncol 2019) had investigated regorafenib versus lomustine in a phase 2-trial. Furthermore, the use of regorafenib in a compassionate-use-program was recently reported (Tzaridis et al., Neuro Oncol 2019) Yet, the regorafenib arm in the Glioblastoma Adaptive Global Innovative Learning Environment (GBM Agile) master protocol was stopped after an interim analysis that showed a low probability of improvement of overall survival compared with control (press release 21 September 2022). We investigated here a functional genomics-based approach to discover and validate potential modulators of response to regorafenib treatment in experimental glioma in vitro, in vivo and ex vivo. We performed CRISPR/Cas9 activation (Calabrese P65-HSF activation library) and knockout (genome-wide knockout library Brunello) screens during treatment with regorafenib, sequenced and analyzed the genomic DNA of the remaining cells. We discovered functionally-instructed molecular hits, designed a drug library for further validation by functional assays in vitro and combination treatments in murine glioma models in vivo. Taken together, we discovered and validated functionally-instructed combination treatment options involving regorafenib. Our finalized results will be presented.