Breast cancer is the most commonly diagnosed cancer in women worldwide, and an unmet clinical need exists for novel approaches to target invasive and metastatic breast cancers refractory to current treatments. The Rho-associated kinase (ROCK) pathway is progressively activated in human breast cancer and is highest in invasive forms. However, whether ROCK activity has a causal role in promoting mammary tumourigenesis has remained enigmatic. We have discovered, by conditionally activating ROCK in the PyMT mouse model of breast cancer, that ROCK activity significantly enhances mammary tumour burden compared to that observed in mice expressing a kinase-dead (KD) version of ROCK. Similarly, blocking ROCK activity by administering pharmacologic inhibitors, or knocking down ROCK via an siRNA approach, significantly reduces tumour burden.
Intriguingly, ROCK activity in tumours also resulted in an increased number of cancer-associated fibroblasts (CAFs) in ROCK-PyMT tumours compared to KD-PyMT tumours of comparable size. CAFs derived from ROCK-PyMT tumours, or those exposed to culture medium conditioned by ROCK-PyMT primary tumour cells, exhibit enhanced migration and upon co-engraftment promote tumour growth from cancer cells when compared to CAFs derived from KD-PyMT control tumours. Our findings strongly suggest that one mechanism by which ROCK regulates breast cancer progression is by enhancing the cancer-promoting properties of CAFs via fibroblast-educating paracrine signalling mediators. Proteomic analysis revealed that that the ER-stress protein Creld2 is enriched in the secretome of ROCK-activated cells and can enhance the cancer-promoting functions of CAFs. Deeper analysis revealed a novel mechanism whereby ROCK selectively activates the PERK arm of the unfolded protein response that transcriptionally activates Creld2 resulting in increased secretion of the Creld2 protein, which educates fibroblasts to a tumour-promoting form.
Our work suggests novel therapeutic strategies against invasive breast cancer, based on inhibiting Creld2 to abolish the tumour-promoting education of cancer-associated fibroblasts.