Preclinical studies of TW-37, a new nonpeptidic small-molecule inhibitor of Bcl-2, in diffuse large cell lymphoma xenograft model reveal drug action on both Bcl-2 and Mcl-1

Purpose: Overexpression of Bcl-2 protein has been observed in more than 80% of B-cell lymphomas, including diffuse large cell lymphoma (DLCL), the most common subtype of non-Hodgkin's lymphoma. We have previously employed the natural product (-) -gossypol to test its therapeutic potential as a small-molecule inhibitor of Bcl-2 for the treatment of B-cell lymphomas. Experimental Design: Recently, we have used a structure-based strategy to design a new class of potent small-molecule inhibitor acting on Bcl-2. One such lead compound is the benzenesulfonyl derivative TW-37, which was designed to target the BH3-binding groove in Bcl-2 where proapoptotic Bcl-2 proteins, such as Bak, Bax, Bid, and Bim bind. Results: In our fluorescence polarization - based binding assays using recombinant Bcl-2, Bcl-X-L, and Mcl-1 proteins,TW-37 binds to Bcl-2, Bcl-X-L, and Mcl-1 with K-i values of 290, 1,110 and 260 nmol/L, respectively. Hence,TW-37 is a potent inhibitor of Bcl-2 and has >3-fold selectivity over Bcl-X-L. In vitro,TW-37 showed significant antiproliferative effect in a de novo chemoresistant-WSU-DLCL2 lymphoma cell line and primary cells obtained from a lymphoma patient with no effect on normal peripheral blood lymphocytes. Coimmunoprecipitation experiments showed that TW-37 disrupted heterodimer formation between Bax or truncated-Bid and antiapoptotic proteins in the order Mcl-1 > Bcl-2 >> Bcl-X-L. As expected, TW-37 caused apoptotic death. Pre-exposure of lymphoma cells to TW-37 significantly enhanced the killing effect of cyclophosphamide-doxorubicin-vincristine-prednisone (CHOP) regimen. The maximum tolerated dose of TW-37 in severe combined immunodeficient (SCID) mice was 40 mg/kg for three i.v. injections when given alone and 20 mg/kg, x3 when given in combination with CHOP. Using WSU-DLCL2 SCID mouse xenograft model, the addition of TW-37 to CHOP resulted in more complete tumor inhibition compared with either CHOP or TW-37 alone. Conclusions: We conclude that the administration of TW-37, as a potent Bcl-2 and Mcl-1 inhibitor, to standard chemotherapy may prove an effective strategy in the treatment of B-cell lymphoma.