Recombination and Mutation in Breast Cancer

Institution: Pangene Corporation
Investigator(s): David Zarling, Ph.D. -
Award Cycle: 1997 (Cycle III) Grant #: 3IB-0052 Award: $260,988
Award Type: IDEA
Research Priorities
Biology of the Breast Cell>Pathogenesis: understanding the disease



Initial Award Abstract (1997)
Breast cancer is associated with numerous genetic alterations that can be detected directly in the chromosomal DNA by methods that visualize the attachment of probes labeled with fluorescent markers. This standard method of fluorescent in situ hybridization (FISH) is time consuming, insensitive and is not allele specific. Our lab has recently developed an enhanced FISH method for the detection of a tumor suppressor gene (p53) and an oncogene (ERBB2) in human breast cancer. Our method is rapid (30 min), sensitive, and appears to have a very low background. We plan to adapt our recombinase-enhanced hybridization method with novel ‘cyclizable’ DNA probes to develop an allele-specific DNA hybridization method for the specific and rapid detection of mutant breast cancer genes. To develop and apply this novel technology to breast cancer, we will employ a collaborative multidisciplinary research program combining the expertise of three laboratories: Pangene Corporation, the University of California at San Francisco, and Uppsala University in Sweden.

A second objective of this grant is to study the status of recombination (exchange of strands of homologous chromosomal DNA) genes and proteins in breast cancer cells. We have made preliminary observations that one of the key genes involved, RAD51, is abnormally regulated in breast cancer cells. We plan to expand on this observation and understand the molecular basis for the abnormal regulation of RAD51 and its potential role in the onset and progression of breast cancer. In related experiments, we will evaluate the possibility of using RAD51 as a marker for breast cancer diagnosis. Recently, the p53 tumor suppressor protein was reported to interact with the RAD51 protein and control its biochemical functions, including DNA strand exchange. In contrast, mutant p53 fails both to interact with RAD51 protein and to control DNA strand exchange. These related observations, together with the fact that breast cancer cells often have mutant p53 and abnormal rates of recombination, strongly suggests that abnormal regulation of recombination enzymes exist in breast cancer.

Thus, the goals of our research are to (i) provide a rapid method of evaluating breast cancer biopsy samples to detect genetic abnormalities, and (ii) explore novel mechanisms by which already known mutations are associated with DNA repair and recombination defects. These novel techniques offer the prospect of tailoring therapy options to the particular cellular and genetic defects of individual patients.


Final Report (1999)
Current breast cancer therapies include radiation therapy and/or treatment with DNA damaging chemotherapeutic agents. Many tumors are highly resistant to radiation and chemotherapy. One of the major reasons for radio- and chemo-resistance is enhanced cellular ability to repair and recombine radiation or chemotherapy damaged DNA due to overexpression of Rad51 recombinase in tumors. Hence, radiation therapy and chemotherapy can be improved by combining them with inhibitors of Rad51 activity.

The overall objectives of our study were to 1) measure Rad51 expression in breast cancer and to assess its value for breast cancer diagnostics and therapeutics, 2) determine the interactions between Rad51 protein and normal or mutant tumor suppressor proteins and oncogenes, and 3) characterize the expression of recombinases involved in homologous recombination, particularly Rad51 and its overexpression in breast cancer cells.

We determined the status of the human Rad51 protein in breast cancer cells. Rad51 is overexpressed in all 90 of the breast tumors studied. Aberrant overexpression of the Rad51 gene is directly associated with radiation resistance and chemotherapeutic drug resistance. The aberrant expression of Rad51 in tumor cells also appears to be consistent with the observations of enhanced recombination and genomic instabilities in breast tumor cells. The molecular basis for expression of Rad51 in breast tumor cells is under investigation. We are continuing studies on the therapeutic consequences of inhibiting the overexpression of Rad51 in breast tumor cells. We and our collaborators have shown that experimental downregulation of Rad51 with specific Rad51 inhibitor formulations causes increased sensitivity of human breast tumor cells to chemotherapy in vitro and enables long term survival in tumor-bearing mice treated with Rad51 inhibitors and radiation in vivo.

Rad51 inhibitors appear to be highly effective therapeutic agents for both breast tumor therapy and diagnosis. Our findings indicate that Rad51 is an important new target for breast cancer therapy. Rad51 inhibitors are well positioned to be conveniently integrated into current breast cancer protocols.