RNAi-mediated inhibition of cyclin D1 decreases colon cancer cell proliferation and MMP protein expression

Abstract Introduction: Cyclin D1, a downstream protein of the PI3K/GSK-3 pathway, regulates cell cycle progression and proliferation. Overexpression of cyclin D1 is associated with a poor prognosis in various human cancers (eg, colorectal cancers). The purpose of this study was to determine whether targeted inhibition of cyclin D1, using small interfering RNA (siRNA), alters proliferation and expression of the matrix metalloproteinase proteins (MMPs) in the colon cancer cell line KM20. Methods: KM20 cells were treated with cyclin D1 siRNA or scrambled sequences (control); cell proliferation and viability was assessed. Proliferating cell nuclear antigen (PCNA) expression was measured by ELISA. Western blots were performed to determine expression of cyclin D1, PI3K subunits p85alpha and p110 and GSK-3alpha/beta (upstream regulators of cyclin D1), and MMP-2 and MMP-9 (tumor invasion proteins regulated by cyclin D1). Results: Cyclin D1 expression was effectively blocked by siRNA treatment. Expression of MMP-2 and MMP-9 was decreased in KM20 cells by cyclin D1 inhibition; PI3K subunit and GSK-3 expression was not affected. Compared to control, decreased KM20 cell proliferation and PCNA translocation was noted with cyclin D1 siRNA treatment; no significant changes were noted in apoptosis. Conclusions: We show that selective inhibition of cyclin D1 inhibits MMP expression and KM20 cell proliferation, most likely due to cell cycle arrest. Selective targeting of cyclin D1 may enhance the effects of standard chemotherapeutic agents and provide novel adjunct treatment for selected colorectal cancers.