by ecancer reporter Clare Sansom

The molecular events involved in the development of colorectal cancer are generally well understood, and include the inactivation of the tumour suppressor gene Apc (Adenomatous polyposis coli).

Inactivation of this gene is also involved in the normal regeneration of the intestinal epithelium, and this process is often used as a model for early stage colorectal cancer development.

The regeneration process is known to be driven by the Wnt signalling pathway.

The protein complex mTORC1, or mechanistic target of rapamycin complex 1, which controls protein synthesis, is a mediator of cell growth and proliferation and researchers have suggested that this complex is also important in regeneration of the intestinal epithelium and colorectal cancer development.

Increased phosphorylation of mTORC1 effector proteins is observed after Apc deletion and Wnt activation.

A group of researchers led by Owen Sansom of the Cancer Research UK Beatson Institute, Glasgow, Scotland have investigated the role of mTORC1 in this cellular proliferation pathway and, thus, its potential as a target for chemoprevention or early stage colorectal cancer therapy.

Firstly, they observed that rapamycin, which inhibits the serine/threonine kinase mTOR that is a core part of this complex, inhibits epithelial regeneration but not normal cellular proliferation or apoptosis in the intestine.

This confirmed that the mTORC1 signalling pathway might be a valid therapeutic target during the early development of colorectal cancer.

Sansom and his co-workers deleted the gene Rptor, another essential component of this complex, in rat intestinal epithelium, and showed that this prevented the proliferation associated with regeneration and Apc deletion but had no effect on normal gut homeostasis.

Rapamycin was found to prevent the growth of tumours in the intestines of transgenic mice following the induction of Apc deletion.

Similarly, giving rapamycin to mice from the same strain with established adenomas caused those tumours to regress, and the mice survived longer than similar mice that had not received rapamycin.

Examination of tumours taken from mice that had been treated with rapamycin showed an initial loss of proliferation and a gain in the number of lysozyme-secreting Paneth cells, which – like progenitor cells – show high levels of Wnt signalling.

After 30 days, however, the tumours had shrunk into small, non-proliferative lesions without Paneth cells.

These results suggested that rapamycin had caused the progenitor cells to differentiate into Paneth-like cells.

There was no increase in proteins involved in the classic cell cycle arrest pathway, such as p53, which suggested that this pathway had not been engaged and that the tumours would regrow if rapamycin treatment was discontinued.

Intestinal tumours did, indeed, regrow in these mice starting 40-60 days after the end of rapamycin treatment, suggesting that tumour stem cells were still present.

The researchers next investigated the role of mTORC1 in regulating protein synthesis following loss of Apc.

They measured the number of polysomes in wild type, Apc-deficient and Apc/Rptor deficient intestinal epithelial cells.

The number of polysomes in the cells decreased in the Apc deficient cells but not in those in which both genes had been deleted.

This suggests that Apc deletion causes either reduced translation initiation or faster translational elongation in these cells, and that the defect can be reversed by removing Rptor.

Further experiments confirmed that translational elongation is the rate limiting step of protein synthesis following Apc deletion (and thus Wnt activation) and suggested that mTORC1 is activated to overcome this.

Other inhibitors of protein elongation, such as cyclohexamide, were found to limit cellular proliferation associated with Apc deletion in a similar way to rapamycin.

Dissection of the signalling pathways downstream of mTORC1 using further strains of transgenic mice showed that inhibition of the kinase eEF2 by this complex is required for the proliferation of Apc-deleted cells.

The researchers tested mRNA and protein levels of cell cycle regulators in Apc-deleted cells and found that while several cyclins and CDKs showed increases in both RNA and protein, cyclin D3 only showed increased protein levels without any increase in mRNA.

This indicated that the regulation of cyclin D3 translation occurs at the level of elongation; however, the contribution of this cyclin to the proliferative phenotype of Apc-deficient cells is still unknown.

Taken together, these results show that translational elongation mediated by the mTORC1 complex is essential for the proliferation of Apc-deficient cells and that this can be inhibited by mTOR inhibitors such as rapamycin.

Rapamycin and its analogues, which are already in clinical use, may therefore be useful drugs for treating early stage colorectal cancer, or as a chemopreventive in those at high risk of developing the disease.

Reference

Faller, W.J., Jackson, T.J., Knight, J.R.P. and 20 others (2014). mTORC1-mediated translational elongation limits intestinal tumour initiation and growth. Nature, published online ahead of print 5 November 2014.