The ubiquitin-proteasome pathway plays a critical role in the degradation of cellular proteins and cell cycle control. As such, dysregulating the degradation of such proteins should have profound effects on tumor growth and should cause cells to undergo apoptosis. Proteasome inhibitors display marked anticancer tendencies in hematopoietic cells; however, this activity is much less potent against solid tumor malignant cells. In addition, it can mediate toxicities such as peripheral neuropathy and can induce primary/secondary resistances. These observations prompted us to investigate whether sodium butyrate (NaB), a natural product for inducing tumor cell apoptosis that exhibits a low degree of clinical toxicity, can enhance the anticancer effects of the proteasome inhibitors MG115, MG132, PSI-1, PSI-2 and epoxomicin in human colorectal cancer. Combinations of the proteasome inhibitors and NaB markedly inhibited the proliferation of human colorectal cancer cells in a dose- and time-dependent manner, particularly compared to single treatment with NaB and the proteasome inhibitors. A combination of the proteasome inhibitors and NaB produced additive and synergistic growth inhibitory effects. The human colorectal cancer cells treated with NaB, proteasome inhibitors, and their combinations exhibited growth arrest at the G(1), G(2)-M/S, and G(1) or S/G(2)-M phases of the cell cycle, respectively. NaB stimulated the conversion of procaspase-3 into caspase-3 and induced the cleavage of poly(ADP-ribose) polymerase and lamin-B, which are two hallmarks of apoptosis. NaB, the proteasome inhibitor epoxomicin, and their combination stimulated the release of cytochrome-c from mitochondria, which was most likely responsible for the activation of caspase-3. Combined treatment with NaB and the proteasome inhibitors caused marked induction of apoptosis compared to single treatments of the proteasome inhibitors and NaB; the apoptotic signals were abolished by the inhibitors of caspase-3 activity. The proteasome inhibitors and their combinations with NaB counteracted the suressive effect that was induced by NaB on the levels of p53. NaB, the proteasome inhibitors and their combinations decreased the levels of BcL2, but increased the levels of Bax, p21(waf1) and p27(Kip1). Taken together, these results indicate a synergistic interaction between proteasome inhibitors and sodium butyrate that could present a new approach for the treatment of colorectal cancer.
