Mubin, N. | bioRxiv Science

bioRxiv2026

Synthetic lethality of p97 inhibition and PSMC2-overexpressionovercomes bortezomib resistance in multiple myeloma

The aim of the present study was to analyze genomic profiles obtained from multiple myeloma (MM) patient tumors to identify prognostic classifiers of clinical response and survival following bortezomib monotherapy and then target key actionable classifiers that underly bortezomib resistance. We discovered higher levels of proteasome PSMC2 messenger RNA in myeloma cells from patients that did not respond to bortezomib and that higher PSMC2 expression correlated with inferior progression-free and overall survival. Pharmacologic screening revealed that CB-5339, an ATP-competitive inhibitor of the molecular chaperone p97/VCP that shuttles ubiquitin-conjugated substrates to proteasomes, was synthetic lethal in MM cells engineered to overexpress PSMC2. p97 co-immunoprecipitated with 26S proteasomes affinity-purified from PSMC2-overexpressing cells which exhibited greater activity and reduced bortezomib sensitivity relative to PSMC2-KO cells. Bortezomib treatment followed by in-situ cross-linking and western blotting revealed higher levels of K48- and K63-linkage-specific ubiquitin chains covalently bound to 26S proteasomes isolated from PSMC2-KO cells compared to proteasomes affinity-purified from PSMC2-overexpressing cells. Western blotting demonstrated higher levels of 19S RPs, 26S proteasomes, and a PSMC2-PSMC1-PSMD2 subcomplex in PSMC2-overexpressing cells relative to PSMC2-KO cells. CB-5339 and bortezomib co-treatment synergistically targeted tumor cells obtained from MM patients previously exposed to bortezomib therapy and reduced the in vivo growth of PSMC2-overexpressing myeloma xenografts in mice. Taken together, we have discovered a genetic vulnerability that provokes proteasome-intrinsic bortezomib resistance and validated the potential of synthetic lethality to guide therapeutic interventions. Our study reaffirms that transcriptomic heterogeneity in MM is a primary driver of prognostic variability.

cancer biology↗

bioRxiv2022

Removal of pomt1 in zebrafish leads to loss of α-dystroglycan glycosylation and dystroglycanopathy phenotypes

Biallelic mutations in Protein O-mannosyltransferase 1 (POMT1) are among the most common causes of a severe group of congenital muscular dystrophies (CMDs) known as dystroglycanopathies. POMT1 is a glycosyltransferase responsible for the attachment of a functional glycan mediating interactions between the transmembrane glycoprotein dystroglycan and its binding partners in the extracellular matrix (ECM). Disruptions in these cell-ECM interactions lead to multiple developmental defects causing brain and eye malformations in addition to CMD. Removing Pomt1 in the mouse leads to early embryonic death due to the essential role of dystroglycan during placental formation in rodents. Here, we characterized and validated a model of pomt1 loss of function in the zebrafish showing that developmental defects found in individuals affected by dystroglycanopathies can be recapitulated in the fish. We also discovered that pomt1 mRNA provided by the mother in the oocyte supports dystroglycan glycosylation during the first few weeks of development. Muscle disease, retinal synapse formation deficits, and axon guidance defects can only be uncovered during the first week post fertilization by generating knock-out embryos from knock-out mothers. Conversely, maternal pomt1 from heterozygous mothers was sufficient to sustain muscle, eye, and brain development only leading to loss of photoreceptor synapses at 30 days post fertilization. Our findings show that it is important to define the contribution of maternal mRNA while developing zebrafish models of dystroglycanopathies and that offspring generated from heterozygous and knock-out mothers can be used to differentiate the role of dystroglycan glycosylation in tissue formation and maintenance.

neuroscience↗