Cancer is the second-most common cause of death in developed countries. The statistics suggest that, in the next 20 years, the number of global cancer cases will increase more than 50%, and the deaths caused by this disease will increase more than 60%, with respect to the current figures.
Cancer appears when a group of cells from a tissue begin to divide uncontrollably and acquire the ability to migrate and spread throughout the body. In the fight against cancer, four major strategies are used: surgery, radiotherapy, chemotherapy, and biological therapies. Most of the drugs used in cancer chemotherapy exhibit a strong anti-proliferative action by interfering with cell division process or mitosis.
A key protein in mitosis process is tubulin. The assembly of tubulin gives rise to microtubules that are biopolymeric structures responsible for the separation of chromosomes in mitosis. Many natural products, such as colchicine, and the ones used in chemotherapy, taxanes or vinca alkaloids, target tubulin or microtubules.
The main problem of this biological action is the lack of selectivity towards cancer cells, which explains the dangerous side effects it may cause. Fortunately, since it has been proven that cancer cells have characteristics that are common to all of them, regardless of the tissue in which they are found, the emphasis in anticancer drug development has been shifting from antimitotic nonselective agents to compounds that target one or more of these common hallmarks of cancer cells. Among the ten hallmarks of cancer cells described to date, their ability to avoid senescence and to induce angiogenesis should be highlighted.
Senescence is strongly related to telomeres, which are the terminal zones of chromosomes. Telomeres progressively shorten because of cellular replication, leading, after several cell generations, to a permanent cell cycle arrest, known as senescence. Telomerase is a ribonucleoprotein enzyme that maintains the length of the telomeres but it is not expressed in somatic normal cells while it is over-expressed in cancer cells. Telomerase expression has been detected in about 90% of all malignant tumors and is associated with high levels of hTERT, the gene that encodes for the hTERT protein which is an essential constituent of telomerase. Drugs able to inhibit telomerase are potentially useful weapons in the fight against cancer.
On the other hand, as has been noted above, cancer cells are able to induce angiogenesis, which is the process that leads to the formation of new blood vessels towards the tumor and consequently, allows it to grow and spread. Tumor-induced angiogenesis is mainly sustained by the over-secretion of some proteins from cancer cells, among which the vascular endothelial growth factor (VEGF) stands out. When secreted VEGF binds to a membrane receptor in endothelial cells, VEGFR-2 activates these cells to proliferate and differentiate into microvessels towards the tumor cells. Further, expression of both hTERT and VEGF genes shows a positive association with c-Myc, a transcription factor that regulates cell proliferation, differentiation, and promotes changes in the tumor microenvironment.
Colchicine is an alkaloid isolated from the poisonous meadow saffron Colchicum autumnale L. and was the first tubulin destabilizing agent to be discovered. Colchicine was approved by the FDA in 2009 to treat familial Mediterranean fever and acute gout flares. Although colchicine is one of the most potent antimitotic drugs, it is not used in chemotherapy due to its narrow toxicity window, which explains the efforts to get colchicine-like compounds approved for cancer treatment. As a matter of fact, some derivatives of colchicine, such as thiocolchicoside (Neoflax™, Muscoril™), are being used as anti-inflammatory and analgesic drugs. Recent studies have suggested that the anti-inflammatory effect of colchicine may be due to its activity at the transcriptional level.
A few years ago we designed, synthesized, and studied some hybrid molecules bearing a colchicine moiety and a pironetin analogue fragment. We found that, in addition to binding to tubulin, these compounds were able to downregulate the expression of some oncogenes such as c-MYC, hTERT, and VEGF. These three genes are of paramount importance in the cancer-generation process. Our results pointed to the colchicine fragment being responsible for the observed oncogene downregulation, though high doses were required (50 nM-15 ÎĽM). Taking into account that colchicine is not only able to target tubulin but also to downregulate some oncogene expression, we decided to design new colchicine derivatives with reduced cytotoxicity and enhanced selectivity.
One of our strategies was to replace the N-acetyl residue of the parent molecule that was replaced by haloacetyl, cyclohexylacetyl, phenylacetyl, and various aroyl moieties. These synthetic compounds showed antiproliferative action at the nanomolar level (measured as IC50 values for three tumor cell lines HT-29, MCF-7 and A549 and one non-cancer cell line, HEK-293), in many cases with a higher potency than colchicine itself. The proportion between the IC50 of a compound for a non-cancerous line and the IC50 of that compound in a tumor line is called the selectivity index. A high selectivity index may be related to a good therapeutic margin. o-Chlorobenzoylcolchicine along with m– and p-bromobenzoylcolchicine, were the compounds that showed selectivity indexes greater than 1, much higher than that of the natural product.
Some of the colchicine derivatives, particularly the haloacetyl ones, show full inhibition of tubulin polymerization in a similar manner to colchicine. A characteristic feature of the antimitotic compounds is the arrest of the cell cycle in the G2/M phase due to their binding to tubulin. The most active of our colchicine derivatives, as regards the cell cycle, were again the chlorobenzoyl and bromobenzyl derivatives, which caused cell cycle arrest at the G2/M phase when tested at 20 nM, along with the bromoacetyl derivative, which arrests the cell cycle at 15 nM.
In addition, these colchicine derivatives have been shown to be fairly active downregulating at very low concentrations the expression of the c-Myc, hTERT and VEGF genes, as well as VEGF protein secretion. Thus, the enhanced oncogene downregulation effect, shown mainly by o-chlorobenzoylcolchicine, which was achieved at concentrations lower than its IC50 values, might broaden the therapeutic window of these colchicine derivatives. They appear to be effective anti-cancer drug candidates as they are able to decrease the expression of oncogenes involved in tumor aggressiveness at concentrations in which there is no antimitotic effect.
In the same way that the glass slipper transformed ragged Cinderella into a beautiful princess, the haloaroyl unit might transform the toxic colchicine into a potential therapeutically useful anticancer agent.
These findings are described in the article entitled Effects on tubulin polymerization and down-regulation of c-Myc, hTERT and VEGF genes by colchicine haloacetyl and haloaroyl derivatives, recently published in the European Journal of Medicinal Chemistry.  This work was conducted by Eva Falomir, Juan Murga, Ana Marzo-Mas, and Miguel Carda from Jaume I University, and J. Alberto Marco from the University of Valencia.