Selective Activation of M 1 Muscarinic Receptors Attenuates Human Colon Cancer Cell Proliferation.

M 3 muscarinic receptor (M 3 R) activation stimulates colon cancer cell proliferation, migration, and invasion; M 3 R expression is augmented in colon cancer and ablating M 3 R expression in mice attenuates colon neoplasia. Several lines of investigation suggest that in contrast to these pro-neoplastic effects of M 3 R, M 1 R plays an opposite role, protecting colon epithelial cells against neoplastic transformation. To pursue these intriguing findings, we examined the relative expression of M 1 R versus M 3 R in progressive stages of colon neoplasia and the effect of treating colon cancer cells with selective M 1 R agonists. We detected divergent expression of M 1 R and M 3 R in progressive colon neoplasia, from aberrant crypt foci to adenomas, primary colon cancers, and colon cancer metastases. Treating three human colon cancer cell lines with two selective M 1 R agonists, we found that in contrast to the effects of M 3 R activation, selective activation of M 1 R reversibly inhibited cell proliferation. Moreover, these effects were diminished by pre-incubating cells with a selective M 1 R inhibitor. Mechanistic insights were gained using selective chemical inhibitors of post-muscarinic receptor signaling molecules and immunoblotting to demonstrate M 1 R-dependent changes in the activation (phosphorylation) of key downstream kinases, EGFR, ERK1/2, and p38 MAPK. We did not detect a role for drug toxicity, cellular senescence, or apoptosis in mediating M 1 R agonist-induced attenuated cell proliferation. Lastly, adding M 1 R-selective agonists to colon cancer cells augmented the anti-proliferative effects of conventional chemotherapeutic agents. Collectively, these results suggest that selective M 1 R agonism for advanced colon cancer, alone or in combination with conventional chemotherapy, is a therapeutic strategy worth exploring.