Efficacy of combination of rituximab (R), obatoclax (O), and bortezomib (B) against rituximab-sensitive (RSCL) and rituximab-resistant B-cell lymphoma cell lines (RRCL).

Abstract

8094 Background: The interactions between the members of the Bcl-2 like proteins play an important role in the development, progression and prognosis in various subtypes of B-cell lymphomas. R is an anti-CD20 chimeric monoclonal antibody, O (obatoclax) is a novel pan bcl-2 inhibitor, and B is a proteasome inhibitor with multiple mechanisms of action. These agents can alter the balance/interaction of Bcl-2 family members to induce cell death. Methods: In an effort to expand the targeted therapeutic options for B-cell lymphoma patients, we evaluated the biological activity of combining R, B and O against RSCL (Raji, RL, SU-DHL6, OCI-LY3 and OCI-LY19) and RRCL (Raji-4RH and RL-4RH). We developed RRCL by exposing R-naive cells to escalating doses of R ± human serum (HS). Lymphoma cells were incubated with either RPMI or O (2.5 μ M) with or without B (10nM) for 42 hours. Subsequently, cells were exposed to either R or Isotype control (1μ g/ml) + HS (12.5%). Following an additional 6 hours period of incubation, changes in mitochondrial potential and cell viability were measured by alamar blue reduction. Lymphoma cells were exposed for 48 hrs to R, O, B or various combinations (BO, RB, RBO, etc), and changes in Bcl-2 family members were determined by Western blotting. Results: Significant anti-tumor activity was observed with each agent alone in RSCL and to a lesser degree in RRCL. O was equally effective in both RSCL and RRCL. Enhanced anti-tumor activity was observed by combining R+O or O+B. Furthermore, the triple combination of R+B+O resulted in maximum cell kill (80%) in both RSCL and RRCL. In vitro exposure of lymphoma cells to B or O altered the levels of pro-apoptotic (Bak, PUMA, Noxa) and anti-apoptotic (Mcl-1, Bcl-XL) proteins. Conclusions: Our data suggest that deregulation of apoptosis by combining B, R and O is a potent strategy in both RSCL and RRCL. Ongoing in vivo studies will further define the therapeutic role of combining these three targeted agents in B-cell lymphoma. No significant financial relationships to disclose.