EXECUTIVE SUMMARY

Novel Inducers of Apoptosis

SUMMARY:  Siuta Consulting has been retained by the Arizona Biomedical Research Commission to identify partners for a novel strategy to advantageously alter the cancer cell proteome to kill cancer cells by inducing a natural cell death process as well as fighting drug resistance that has been developed by Dr. Seth D. Rose in the Department of Chemistry and Biochemistry at Arizona State University.  Dr. Rose’s strategy is twofold: (1) irreversibly inhibiting enzymes involved in cell division, and (2) preventing drug efflux pumps from removing the drug from the cell to restore enzyme activity.  This strategy offers a truly new approach for the development of chemotherapeutic agents for effective inhibition of cancer cell growth.

Several novel compounds have been synthesized and tested in vitro against more than 60 human cancer cell lines.  The lead compound, RG-66, was found to be active against several cell lines (pancreatic, leukemia, colon, central nervous system, ovarian, and breast) at concentrations below 10 nanomolar.  Xenograft testing is in progress and results should be available shortly.

BACKGROUND:  Human cancers rely on many proteins to activate the cells to divide in an unrestrained manner.  Many of these proteins are involved in the cell division process of mitosis.  Tubulin, for example, is needed for partitioning chromosomes into daughter cells.  Other proteins are involved in passing the cell through various checkpoints in the cell cycle designed to arrest a cell that is abnormal.  Others are involved in the process of self-destruction of abnormal cells (apoptosis).  Successful anticancer strategies can deplete the cell of proteins essential for cell division or to remove restraints on proteins that induce apoptosis.  Cancer cells respond by attempting to pump the drug out of the cell to prevent necrosis or self-destruction of the cancer cell.

TECHNOLOGY:  The investigators used the principle of covalent bonding of potential irreversible inhibitors to various biomolecular targets in the cancer cell.  After covalent bonding to its molecular target, the drug efflux pumps in principle cannot reverse the action of the drug by removing unbound drug from the cell.  The action of the drug cannot be undone because a covalent bond links the drug to the enzyme.  These compounds are easily synthesized by processes that could readily be scaled up.

To learn more about the mode of action of this series of compounds in cancer cells, cell cycle analysis was carried out on BxPc-3 pancreatic cancer cells.  Exposure of the pancreatic cancer cells to one of the compounds at 100 µM for 6 and 12 hours resulted in accumulation of cells at G2/M, followed by cell death due to apoptosis.  Apoptosis was confirmed by the 5-fold increase in caspase-3 activity relative to the untreated control.  This suggests that the compound interacted with proteins involved in mitosis with the result that the cells were unable to complete mitosis.  Also, time-dependent decrease in b-tubulin upon treatment with this agent was observed, as well as a profound synergism with paclitaxel. 

A second, related class of compounds showed micromolar activity against cancer cells and was also very efficient at inducing G2/M block and apoptosis.  Further studies were performed to elucidate the mechanism of action in cancer cells.  In these experiments, the mode by which the cells were dying was explored to ascertain whether the cells were undergoing apoptosis (programmed cell death) or necrosis.  It was found that the compounds did indeed induce apoptosis, and a larger effect was found at a lower concentration (50 micromolar) than was found with the clinically used, broad-spectrum anticancer drug etoposide (100 micromolar) as a positive control.  

PUBLICATIONS:  Some of this research has been published:

K. J. Okolotowicz, W. J. Lee, R. F. Hartman, A. Y. Kim, S. R. Ottersberg, D. E. Robinson, Jr., S. R. Lefler and S. D. Rose, Inactivation of Protein Farnesyltransferase by Active-Site-Targeted Dicarbonyl Compounds, Arch. Pharm. Pharm. Med. Chem., 334 (6), 194-202 (2001) and is available upon request.

Additional unpublished data are also available upon request.

PATENT STATUS:  Some of the compounds of this invention are covered in the following patents:

1. United States Patent 6,576,436 entitled "Anticancer Agents Based on Prevention of Protein Prenylation" issued on June 10, 2003.

2. United States Patent 7,012,097 entitled "Anticancer Agents Based on Prevention of Protein Prenylation" issued on March 14, 2006.

3. United States Patent 7,019,031 entitled "Anticancer Agents Based on Regulation of Protein Prenylation" issued on March 28, 2006.

The more recently synthesized and most active compounds are covered in United States Patent Applications that were filed in May 2006 and May 2007.

LICENSE TERMS:  The present compounds provide a novel approach to the treatment of cancer and are available for worldwide licensing.