Inhibition of proteasome deubiquitinase activity: a strategy to overcome resistance to conventional proteasome inhibitors?
Submitted 5 Inactivating PSMB5 mutations and P-glyco-protein (MDR1/ ABCB1) mediate resistance to proteasome inhibitors: ex vivo efficacy of (immuno) proteasome inhibitors in mononuclear blood cells from rheumatoid arthritis patients 106 Abstract Bortezomib (BTZ), a registered proteasome inhibitor (PI) for multiple myeloma, has also been proposed as a potential anti-rheumatic agent. Its reported side effects however, make it unappealing for long-term administration and resistance may also develop. To overcome this, second generation PIs became available. Here, we investigated whether a novel class of peptide epoxyketone-based PIs including carfilzomib, ONX 0912 and ONX 0914, might escape two established BTZ-resistance mechanisms: (a) mutation(s) in the proteasome β5 subunit (PSMB5) targeted by these PIs, and (b) drug efflux mediated by ABC transporters. THP1 myeloid sublines with acquired resistance to BTZ (54-235 fold) due to mutations in PSMB5, displayed marked cross-resistance but less pronounced to carfilzomib (9-32 fold), ONX 0912 (39-62 fold) and ONX 0914 (27-97 fold). As for ABC transporter mediated efflux: lymphoid CEM/VLB cells with P-glycoprotein (Pgp/MDR1) overexpression exhibited substantial resistance to carfilzomib (114-fold), ONX 0912 (23-fold), ONX 0914 (162-fold) whereas less resistance to BTZ (4.5-fold) was observed. Consistently, the β5 subunit-associated chymo-trypsin-like proteasome activity was significantly less inhibited in these CEM/ VLB cells. Ex vivo analysis of peripheral blood mononuclear cells (PBMCs) from therapy naive rheumatoid arthritis patients revealed that, although basal Pgp levels were low, it compromised the inhibitory effect of carfilzomib and ONX 0914. However, the use of P121, a Pgp transport inhibitor, restored parental cell inhibitory levels both in CEM/VLB cells as well as in PBMCs. These results indicate that the pharmacologic activity of these PIs may be hindered by drug resistance mechanisms involving PSMB5 mutations and PI extrusion via Pgp.