Abstract 2579: Bax345/BLyS: A novel, completely human fusion construct targeting malignant B cells and delivering a unique mitochondrial toxin

Abstract

Abstract B lymphocyte stimulator (BLyS) is crucial for B-cell survival and the biological effects of BLyS are mediated by three cell surface receptors BAFF-R, TACI, and BCMA. Increased expression of BLyS and its receptors has been identified in diffuse large B cell lymphoma (DLBCL) and mantle cell lymphoma (MCL). DLBCL and MCL are an aggressive sub-type of B-cell, non-Hodgkin's lymphoma (NHL). Despite recent advances in the understanding of the molecular and cellular basis for their pathogenesis, these tumors are still associated with poor response to treatment and a fatal outcome. Bcl-2 associated x protein (Bax) is a proapoptotic protein of the Bcl-2 family and is a gateway protein to mitochondria dependent apoptosis. We generated a highly cytotoxic fusion protein designated Bax345/BLyS for receptor-mediated delivery to malignant B cells. The Bax345/BLyS fusion protein has the BH3 and BH1 domains of Bax and contains full-length BLyS. This fusion protein was generated in bacteria and purified by IMAC. The final material was shown to be highly cytotoxic to activated B cell (ABC)-subtype DLBCL (IC50 ∼ 0.1 nM) and mantle cell lymphoma lines (IC50 ∼ 0.1-1 nM). The cytotoxic effects of Bax345/BLyS were inhibited by pretreatment with free BLyS demonstrating that the effects of the fusion construct are mediated through interaction with the BLyS receptors. Treatment of cells with Bax345/BLyS induced down-regulation of Akt, p-Akt, Mcl-1, and survivn and triggered accumulation of cells in the sub G1 phase of the cell cycle. In addition, treatment of cells with Bax345/BLyS induced up-regulation of Bax and apoptosis through caspase-3 activation, PARP cleavage, and DNA fragmentation. Our results suggest that BLyS has the potential to serve as an excellent targeting ligand for the specific delivery of the pro-apoptotic molecule Bax345 to neoplastic B cells expressing the BLyS receptors, and Bax345/BLyS may be an excellent candidate for the treatment of the NHLs which are resistant to conventional chemotherapeutic regimens. This work was supported in part by the Clayton Foundation for Research and by LLS 6234-07 Translational Research Award from the Leukemia and Lymphoma Society. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2579.