Mechanism of Action of Novel Antiproliferative Oligonucleotides

Principal Investigator: BATES, PAULA
Institution Receiving Award: LOUISVILLE, UNIVERSITY OF
Program: PCRP
Proposal Number: PC001482
Award Number: DAMD17-01-1-0067
Funding Mechanism: New Investigator Award
Partnering Awards:
Award Amount: $323,324.00


Prostate cancer is the second leading cause of cancer death among American men and is responsible for around 40,000 deaths per year. This disease is more prevalent in older men, and with increasing life expectancy, prostate cancer is an escalating health problem. Localized prostate cancer can be treated by surgical removal of the prostate, or radiotherapy. However, for many men, the disease has spread beyond the prostate by the time of diagnosis. For these individuals, treatment with compounds that block the action of male hormones (androgens) is generally effective, but rarely curative. Unfortunately, in almost all cases, patients eventually become resistant to hormonal therapies, usually within a few years. Many chemotherapy agents have been tested in prostate cancer clinical trials, but none have produced any significant increase in survival rate. Therefore, new therapeutic agents are much needed for the treatment of hormone-independent prostate cancer.

Prostate cancer, like all cancers, develops when cells grow without the normal control mechanisms. This is usually a result of inappropriate or increased expression of proteins involved in cell proliferation, or because of inactivation of proteins that would normally ensure regulation of cell growth. Ideally, a therapeutic agent would target only these dysregulated proteins (or the genes that encode them) to inhibit the growth of cancer cells while leaving healthy cells undamaged.

Oligonucleotides are short fragments of DNA that can be synthesized chemically. They have been tested as agents that can target the dysregulated proteins by binding to the specific DNA or RNA molecules that encode them. One of these `antisense┬┐ oligonucleotides is now approved by the Food and Drug Administration for use in humans, and many others are in clinical trials. These trials have shown that oligonucleotides are safe and have fewer side effects than most cancer chemotherapy drugs. However, these antisense oligonucleotides have not achieved optimal activity, partly because they bind to molecules other than their target DNA or RNA.

We have discovered that certain types of oligonucleotide, called G-rich oligonucleotides or GROs, can completely inhibit the growth of prostate cancer cells growing in cell culture. Because of the DNA sequence of these GROs, we have postulated that they work by a different mechanism from the antisense oligonucleotides described above. We have now shown that the GROs form unusual folded structures that are very stable under biological conditions. We have also shown that the growth-inhibiting effects of the GROs are somehow related to their binding to a specific cellular protein (in contrast to antisense oligonucleotides, which bind to RNA). Most importantly, we have identified the GRO-binding protein as nucleolin, a protein involved in cell growth. This protein is present at much higher levels in cancer cells than in most normal cells and is thus a good target for therapeutic intervention. The novel GROs are highly active in prostate cancer cells and some other types of cancer cells (for example, breast cancer cells), but are less active at inhibiting the growth of noncancer cells. Because of their high activity, potential selectivity for cancer cells over normal cells, and novel mechanism of action, these novel G-rich oligonucleotides represent promising new agents in the fight against prostate cancer.

The goal of this proposal is to further explore the mechanism by which the novel GROs can inhibit cell growth. We aim to determine which functions of the putative target protein (nucleolin) are inhibited and how binding to the GROs causes this. We anticipate that the proposed studies will validate our hypothesis that GROs work by binding to nucleolin, and therefore confirm that this protein is a new target for cancer therapy. We will then determine which parts of the nucleolin protein interact with the GROs, and this information will be useful for future studies to design nucleolin inhibitors that bind to the same region. We will also use a new technology known as proteomics that can simultaneously analyze the levels and characteristics of many thousands of proteins to examine changes between untreated prostate cancer cells and cells which have stopped growing as a result of treatment with GROs. The information obtained from these experiments will not only clarify exactly how the novel oligonucleotides work, but may identify other altered proteins which could be new markers or targets for prostate cancer.

In summary, GROs are novel oligonucleotides with enormous potential as new therapeutic agents for prostate cancer. They are highly active against prostate cancer cells and will be potentially tumor-selective with few unpleasant side effects. Because they work by an entirely new mechanism, they may offer significant advantages over existing treatments. The primary aim of our proposed research is to understand this new mechanism, which could also lead to further novel markers or therapies for prostate cancer.