New molecular mechanism for colorectal cancer

Jun 4 2008

Scientists in Portugal just found a new molecular mechanism behind colorectal cancer in which a mutated and a normal, but over-expressed, gene cooperate and are both needed to create the disease.

The research, published in the journal Gastroenteroloy, also reveals how a technique called RNA interference can - by inactivating both genes - kill, in just 48 hours, as much as 80% of cancer cells. These are extremely promising results if transferred into new therapies for humans against a disease, which is still one of the most common cancers in the western world.

Colorectal cancer affects the colon, rectum and appendix and is not only the third most common form of cancer, but also the second cancer-related cause of death in the Western world, according to the World Health Organization. The disease kills about 655,000 people per year worldwide, with 16,000 only in the UK, even if it has a high cure rate if early detected and treated.

It is known that about 30 to 40 percent of colorectal cancer cases result from a mutated KRAS gene, which affects cell division. When this gene is mutated it becomes hyper-activated, leading to uncontrolled cell multiplication, which, together with resistance to death, are the hallmarks of all cancerous cells. And in fact, the capabilities of KRAS mutations to induce cancer depend on another molecule - Rac1 - that complements its effect on cell division by inhibiting cell death and further stimulating cell division. Together they create immortal and abnormally growing cells, the exact definition of cancer cells.

More recently, among colorectal cancers negative for the KRAS mutation, a related abnormality has been identified, this time on a gene called BRAF, which, like KRAS, is also involved in cell growth and division. When studied in laboratory, however, BRAF mutations were not enough by themselves to produce cancer, suggesting that a second event was necessary for malignant transformation. The fact that therapies targeting BRAF have a low success rate in these tumours , further supported the existence of a second event and highlighted the urgency to further investigate the mechanism behind BRAF-mutated cancers, which, after all, comprise as much as 10% of all colorectal cancers cases.

Paulo Matos, Raquel Seruca, Peter Jordan and colleagues at the Centre of Human Genetics in the National Health Institute Dr. Ricardo Jorge in Lisbon and the Institute of Molecular Pathology and Immunology of the University of Porto (IPATIMUP), Porto, Portugal have previously found abnormally high quantities of a variant of Rac1 - called Rac1b - among some colorectal cancers. This - together with the fact that Rac1 is crucial for K-Ras-induced tumours - led the researchers to hypothesise that maybe Rac1b was the (mysterious) partner of B-Raf V600 in colorectal tumours.

To test this possibility Matos, Seruca , Jordan and colleagues analysed cells from 61 different colorectal cancer patients, together with normal mucosa cells from 13 patients. Confirming their hypothesis a strong association between the most common Raf mutation - B-Raf V600 - and Rac1b was found, with 82% of B-Raf V600 positive tumours showing Rac1b over-expression. In contrast, K-Ras mutated tumours and normal mucosal tissue had almost no Rac1b.

The next question to Matos, Seruca , Jordan and colleagues was to see if the two molecules did in fact cooperate in the formation of the tumour, since B-Raf V600 was known to be incapable, by itself, of producing cancer. For this, the researchers inhibited the gene for B-Raf V600 or the one for Rac1b, or both at the same time, and analysed the resulting tumour cells.

Gene inhibition was done using a method called RNA interference (or RNAi). The first step during gene expression is to pass the information, contained in the gene (the piece of DNA) to be expressed, into a molecule of RNA called messenger RNA. The RNAi method consists in introducing into the cells a small double molecule of RNA with the same sequence of the messenger RNA corresponding to the gene we want to inactivate. Because double RNA molecules do not occur naturally the cell will destroy it, triggering too the destruction of the messenger RNA with the same sequence and effectively silencing the gene, as its expression is interrupted. The big advantage of this method is its specificity, since, contrary to other cancer treatments like radio- or even chemo-therapy, it will only result in the death of the target cells.

Matos, Seruca, Jordan and colleagues' RNAi experiment revealed that when the genes for B-Raf V600 or Rac1b were inactivated there was a reduction or in cell viability and/or division but, most striking, was the result of their combined inactivation. In fact, combined "inhibition" of the two genes resulted in 80% of the colorectal cancer cells dying in the period of 48 hours. This confirmed that the two molecules functionally cooperate in the development of some K-Ras negative colorectal tumours and explains why B-Raf mutations alone are not sufficient to achieve cancer, while also suggesting a promising specific molecular target for therapy against this type of cancer. Supporting the specific importance of Rac1b in BRAF-mutated colorectal cancer, its inactivation on KRAS-mutated colorectal cancers had no effect on cells' survival or division.

Matos, Seruca, Jordan and colleagues' results are extremely promising as they reveal that the relatively simple and very specific (so with less secondary effects for the patient) technique of RNAi can, when targeting both B-Raf V600 and Rac1b, kill almost all the colorectal tumour. This, if translated into a therapy for humans could be the difference between patients' life or death and, as such, prompts the urgent need for further clinic-oriented investigation. But, as Peter Jordan one of the leaders of the project emphasizes "It is important to remember though, that despite these novel findings, it is still most crucial to move forwards with cancer prevention through changes in diet and life style"

The discovery of a new molecule involved in the pathogenesis of this cancer also raises the question of its suitability as a marker in order to identify and follow closely those individuals with propensity for the disease since the disease has such a high cure rate if detected early. In fact, although screening is already done, at the moment this is only done in individuals from families with the hereditary form of colorectal cancer.

Finally, high quantities of Rac1b have already been detected in some breast cancers what, with the new results, raise the possibility that the molecule can have a role in this (and others?) epithelial cancer and, as such, also needs to be further investigated.