Testing sensitivity to cd30 targeting via brentuximab vedotin in multiple myeloma

Abstract

Introduction Multiple myeloma (MM) is a hematological malignancy characterized by terminally differentiated plasma cells (PCs), immunosuppression and end-organ damage. Despite the therapeutic progress achieved with the introduction of the proteasome inhibitors, immunomodulators and monoclonal antibodies, for most patients MM remains a non-curable disease due to intrinsic or acquired drug resistance. To improve the treatment of PCs malignancies new antigens have been identified and, among them, CD30 could be a therapeutic target. Currently, the only targeted therapy anti-CD30 is the antibody-drug conjugate brentuximab vedotin (BV), approved in US and Europe for use in Hodgkin Lymphoma, and never tested in the setting of MM. Methods Combining in vitro assays, immunofluorescence and gene expression profile we evaluated the response to BV treatment alone or in combination with bendamustine (BDM) or bortezomib (BTZ) of three human MM cell lines (NCI-H929, U266, OPM2) and primary plasma cells from 15 newly diagnosed (ND) or relapsed/refractory (RR) MM patients. Results In primary samples, we found a strong expression of CD30 in about 30% of cases, all of them primary refractory to btz. Thus, we explored if CD30 targeting could overcome btz refractoriness. In human MM cell lines, there was a gradual increase in CD30 expression from OPM2 to U266 (p<0.05). BV rapidly led to proliferative arrest of OPM2 within 24 hours, while higher concentrations 50 µg/mL and 100 µg/mL and longer time (48 hours) were required for U266, even if there was not a linear effect between the quantity of baseline CD30 and percentage of cell death. Moreover, we tested if BDM could induce CD30 expression to increase BV-sensitivity, as previously shown in Hodgkin Lymphoma. However, exposure to BDM (25-100 micromol/L up to 48 hours) did not lead to upregulation of CD30 nor inhibition of proliferation, confirming to be a non-toxic agent for MM cells. In addition, combination BV-BDM did not produce a significant killing effect on MM cell lines over BV alone. Then, we tested if acting on NFkB pathway, BV could increase BTZ sensitivity. Surprisingly, in all human MM cell lines NCI-H929, U266, OPM2, increasing BTZ-BV doses (both in the range 0-100 nM for up to 48 hours) showed protective role of BV against BTZ, thus BV could not overcome BTZ refractoriness. Our previous studies showed that btz refractoriness is associated to mitochondrial dysfunction (Tibullo, 2020). In BTZ resistant cell lines OPM2 and U266, long-term exposure to BV (48 hours) induced mitochondrial depolarization, as assessed by mitotracker and DioC2 essays, without affecting the expression profile of key-genes involved in mitochondrial fission and fusion (Fis1, OPA1, MNF2, Drp1), suggesting that mechanisms other than mitochondrial dysfunction are involved in the protective role of BV when used in combination with BTZ. Conclusions Taken together, our data suggest lack of efficacy of BV in MM, despite high basal expression of CD30, and open a new scenario about resistance signaling to BTZ independent from NFkB signalling and mitochondrial dysfunction.