Activity of dolastatin 10 against small-cell lung cancer in vitro and in vivo: induction of apoptosis and bcl-2 modification

Purpose: Dolastatin 10 is a natural cytotoxic peptide which acts through the inhibition of microtubule assembly. Studies have suggested that such agents can induce apoptosis in association with bcl-2 phosphorylation. Since bcl-2 overexpression is common in small-cell lung cancer (SCLC), we evaluated the activity of dolastatin 10 in SCLC cell lines and xenografts. Methods: In vitro growth inhibition was evaluated with a standardized MTT assay and apoptosis with fluorescent microscopy and a TUNEL assay. Immunoblot analysis and phosphatase digestion were used to determine bcl-2 modification. In vivo activity was evaluated in subcutaneous and metastatic SCLC xenograft models in SCID mice. Results: Dolastatin 10 had growth inhibitory activity against four SCLC cell lines (NCI-H69, -H82, H446, H510) with IC50 values ranging from 0.032 to 0.184 nM. All four cell lines exhibited evidence of apoptosis after 48 h of exposure to 1.3 nM dolastatin 10. Immunoblot analysis revealed that 1.3 nM dolastatin 10 altered the electrophoretic mobility of bcl-2 in NCI-H69 and -H510 cells within 16 h of treatment. Incubation of protein extract from dolastatin 10-treated NCI-H69 and -H510 cells with calcineurin resulted in the disappearance of the altered mobility species, suggesting dolastatin 10-induced bcl-2 phosphorylation. In in vivo studies, 450 mu g/kg of dolastatin 10 IV x 2 given after intravenous injection of NCI-H446 cells completely inhibited tumor formation. In established subcutaneous NCI-H446 xenografts, 450 mu g/kg of dolastatin 10 IV induced apoptosis in the majority of tumor cells within 96 h, resulting in a log(10) cell kill of 5.2 and an increase in median survival from 42 to 91 days. Conclusions: These findings suggest that dolastatin 10 has potent activity against SCLC and that the modulation of apoptotic pathways deserves further evaluation as an anticancer strategy.