Expression and pharmacologic inhibition of the anti-apoptotic Bcl-2 family members in feline tumor cell lines

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Spontaneous tumors in companion animals offer a promising opportunity as models for human cancer biology and translational therapeutics. Feline oral squamous cell carcinomas (OSCC) and soft tissue sarcomas (STS) are excellent examples of spontaneous tumors that arise in companion animals that share similar biologic behavior and clinical outcome when compared to their human counterparts. Here, we report that the anti-apoptotic proteins Bcl-2, Bcl-xL and Bcl-w may play an important role in the survival of feline OSCC and STS cells and that inhibition of this family of proteins by ABT-737, a small molecule inhibitor that targets the Bcl-2 family, leads to cell death in vitro. More importantly, we show that inhibition of these Bcl-2 family members by ABT-737 leads to chemosensitization of OSCC and STS cells and enhances the effects of certain chemotherapies. In this study, the feline OSCC cell line SCCF-1 and four feline STS cell lines (Tietz, Ela-1, Kai and Ham) were examined for expression of Bcl-2, Bcl-xL and Bcl-w via western blot analysis. All three of the Bcl-2 family members were expressed in all of the STS cell lines tested. Interestingly, the OSCC cell line SCCF-1 had undetectable levels of Bcl-2 and low expression of Bcl-w when compared to the panel of STS cell lines. OSCC did however have significantly higher expression of the Bcl-xL compared to STS cell lines. After determining that at least one of the Bcl-2 family members was expressed in the OSCC and STS cell lines, we examined the effect of ABT-737 on the cell lines in vitro. Cells were incubated with increasing amounts of ABT-737 for 72 hours and analyzed using a colorimetric proliferation assay. The results showed that ABT-737 was able to inhibit cell growth in OSCC cells and 3 of the 4 STS cell lines by greater that 75% when concentrations reached 20 μM. SCCF-1, Ela-1, Ham, and Kai showed greater than 50% growth inhibition with concentrations between 1 μM and 5 μM ABT-737 in minimal growth media containing 1% fetal bovine serum (MEM-1%). Staining of the cells with annexin-FITC and propidium iodide indicated that the inhibitory effect of ABT-737 is a result of apoptosis and not growth arrest. SCCF-1 (OSCC) and Ela-1 (STS) cells were then incubated in MEM-1% with 1 μM ABT-737 and increasing levels of the chemotherapeutic drugs doxorubicin (DOX) and carboplatin (CPT). The results showed that ABT-737 was able to sensitize the cells to the effects of both DOX and CPT. The information gained from these studies leads us to believe that the use of ABT-737 in combination with conventional chemotherapeutic agents may improve the clinical outcome of cats with OSCC or STS.