Bortezomib Targets Multiple Myeloma Endothelial Cells.

Introduction: Bone marrow (BM) angiogenesis is an important hallmark of multiple myeloma (MM) which correlates with progression. Although MM remains incurable despite conventional and high-dose chemotherapy, the proteasome inhibitor Bortezomib (Velcade, formerly PS-341), can overcome conventional drug resistance in vitro and in vivo and it has recently been FDA approved for treatment of relapsed and refractory multiple myeloma. Here we evaluated whether anti-angiogenesis may contribute to the anti-MM activity of PS-341. We examined the effect of PS-341 on the angiogenic phenotype of endothelial cells (ECs) isolated from BM of patients with MM.

Methods: MMECs were extracted from BM of patients with active MM using a lectin-based method. The MMEC population contained >95% factor VIII-related antigen (FVIII-RA)⁺ and CD31⁺ cells, as assessed by fluorescence activated cell sorting (FACS). Contamination by macrophages and plasma cells was <5%, evaluated by FACS for CD14 and CD38 positivity, respectively, as well as by RT-PCR and Western blot for CD38. Viability, assessed by trypan blue was >90%. MTT assay and [³H] thymidine uptake were used to evaluate the effects of PS-341 on survival and proliferation, respectively, of MMECs. Proliferation of MM.1S cells cocoltured with MMECs was measured by [³H] thymidine uptake. Cytokine (IL-6, VEGF) levels were quantitated by ELISA. Other in vitro angiogenesis functions examined included chemotaxis, spreading on fibronectin (FN), and morphogenesis on Matrigel. Ongoing work is looking at the effect of PS-341 on angiogenesis in vivo by using a chick embryo chorioallantoic membrane (CAM) model.

Results: PS-341, at concentrations achievable in the plasma of patients, inhibited in vitro MMEC and HUVEC functions related to angiogenesis, including proliferation, chemotaxis, spreading on FN, and capillary formation on Matrigel. All these functions were affected in a dose-dependent fashion. A significant concentration-dependent reduction of VEGF and IL-6 production was observed in the presence of PS-341, as demonstrated by ELISA. Importantly, binding of MM.1S cells to MMECs triggers tumor cell proliferation, and PS-341 inhibits proliferation of adherent MM.1S cells in a dose-dependent fashion. Similar data were demonstrated in HUVECs.

Conclusions: These data therefore demonstrate that PS-341 acts both directly and indirectly against MMECs, another mechanism which may contribute to the anti-MM activity of PS-341.