Interacting post-muscarinic receptor signaling pathways potentiate matrix metalloproteinase-1 expression and invasion of human colon cancer cells

Abstract

M3 muscarinic receptor (M3R) expression is increased in colon cancer; M3R activation stimulates colon cancer cell invasion via cross-talk with epidermal growth factor receptors (EGFR), post-EGFR activation of mitogen-activated protein kinase (MAPK) ERK1/2, and induction of matrix metalloproteinase-1 (MMP1) expression. MMP1 expression is strongly associated with tumor metastasis and adverse outcomes. Here, we asked whether other MAPKs regulate M3R agonist-induced MMP1 expression. In addition to activating ERK1/2, we found that treating colon cancer cells with acetylcholine (ACh) stimulated robust time- and dose-dependent phosphorylation of p38 MAPK. Unlike ERK1/2 activation, ACh-induced p38 phosphorylation was EGFR-independent and blocked by inhibiting protein kinase C-α (PKC-α). Inhibiting activation of PKC-α, EGFR, ERK1/2, or p38-α/β alone attenuated but did not abolish ACh-induced MMP1 expression, a finding that predicted potentiating interactions between these pathways. Indeed, ACh-induced MMP1 expression was abolished by incubating cells with either an EGFR or MEK/ERK1/2 inhibitor combined with a p38-α/β inhibitor. Activating PKC-α and EGFR directly with the combination of phorbol 12-myristate 13-acetate (PMA) and EGF potentiated MMP1 gene and protein expression, and cell invasion. PMA- and ACh-induced MMP1 expression were strongly diminished by inhibiting Src and abolished by concurrently inhibiting both p38-α/β and Src, indicating that Src mediates the cross-talk between PKC-α and EGFR signaling. Using siRNA knockdown, we identified p38-α as the relevant p38 isoform. Collectively, these studies uncover novel functional interactions between post-muscarinic receptor signaling pathways that augment MMP1 expression and drive colon cancer cell invasion; targeting these potentiating interactions has therapeutic potential.

M3 receptor modulates extracellular matrix synthesis via ERK1/2 signaling pathway in human bladder smooth muscle cells

“…Many studies have shown that ERK1/2 is a postal M 3 receptor signaling molecule. Research on colon cancer cells suggested that stimulating M 3 receptor could induce the activation of ERK1/2, thereby promoting the proliferation of cancer cells 24,25 . In gastric cancer cells, Yu et al 26 found that acetylcholine could activate M3 receptor to promote phosphorylation of ERK1/2 and AKT to increase cell proliferation.…”

Targeting M3 Muscarinic Receptors for Colon Cancer Therapy

“…In the United States, colorectal cancer is the fourth most prevalent neoplasm and the second most frequent cause of cancer-related death [1]. While there have been recent advances in early detection and prevention of colon cancer, treatment options remain limited, especially for patients with advanced disease [2, 3]. In addition to identifying genetic mutations that initiate colon cancer, to develop new treatment modalities it is essential to have a better understanding of the mechanisms that govern cancer progression [2, 3].…”

Prediction of novel target genes and pathways involved in bevacizumab-resistant colorectal cancer

“…Although this study is the first to demonstrate the aforementioned relationship in mCRC, MMPs have received attention in terms of their role in the mechanism underlying resistance to antiangiogenic therapy, because increased MMP2 and MMP9 expression levels have been associated with resistance to the anti-VEGF and antiplacental growth factor drug aflibercept and with poor OS [ 53 , 54 ]. Furthermore, MMP1 expression has been strongly associated with tumor metastasis and adverse outcomes in mCRC and has been suggested as a potential prognostic and therapeutic target [ 55 – 59 ]. A previous study reported that BRCA1 is associated with early onset CRC and functions as a DNA repair gene to cytotoxic drugs [ 60 ].…”