Efficient Deletion of Normal Brca2-Deficient Intestinal Epithelium by Poly(ADP-Ribose) Polymerase Inhibition Models Potential Prophylactic Therapy

Abstract

The genes encoding the BRCA1 and BRCA2 tumor suppressors are the most commonly mutated in human familial breast cancers. Both have separate roles in the maintenance of genomic stability through involvement in homologous recombination, an error-free process enabling cells to repair DNA double-strand breaks. We have previously shown that cremediated conditional deletion of Brca2 within the mouse small intestine sensitizes the tissue to DNA damage. Eventually, the tissue repopulates via stem cells in which recombination at the floxed Brca2 allele has not taken place. In this study, we have treated Brca2-deficient small intestine with a potent small-molecule inhibitor of poly(ADP-ribose) polymerase 1 (PARP1), an enzyme predominantly involved in the recognition of DNA single-strand breaks. Brca2 deficiency rendered otherwise normal cells exquisitely sensitive to PARP inhibition, resulting in very high levels of apoptosis as early as 6 hours after treatment, with evidence for repopulation of the tissue at 12 hours. Furthermore, the intestines of animals treated with serial injections of the inhibitor repopulated very rapidly in comparison with those from untreated mice. Our results represent the first in vivo demonstration that inhibition of PARP1 activity confers exquisite sensitivity to death in physiologically normal Brca2-deficient cells, suggesting that such a regimen may be extremely potent prophylactically in women heterozygous for the BRCA2 gene, as well as against established tumors lacking functional BRCA2. (Cancer Res 2005; 65(22): 10145-8)

DNA Repair, Cancer and Cancer Therapy

“…Inhibition of this protein leads to severe and highly selective toxicity in BRCA-deficient cells. Similar results were obtained on xenografts and in animal models of spontaneous BRCA2 loss of function Farmer et al, 2005;Hay et al, 2005). PARP inhibitors have been previously used as chemosensitizing and radiosensitizing agents.…”

Inhibition of poly(ADP-ribose) polymerase down-regulates BRCA1 and RAD51 in a pathway mediated by E2F4 and p130

“…However, our unexpected finding that PARP inhibition, itself, can suppress BRCA1 and RAD51 expression identifies a previously uncharacterized mechanism of action for PARP inhibitors and may provide the basis for new therapeutic strategies that combine PARP inhibition with agents that exploit decreased HDR. It also suggests that proposed strategies to use PARP inhibitors for cancer chemoprevention may be counterproductive (8 Clonogenic Survival Assays. Clonogenic survival was determined by colony formation, as previously described (30).…”

“…PARP-1 is thought to play a key role in DNA repair, primarily by modifying chromatin factors at sites of DNA damage and thereby recruiting repair factors. Inhibitors of PARP have attracted interest for cancer therapy because cancer cells deficient in BRCA1 or BRCA2 due to inactivating mutations are sensitive to PARP inhibition (5)(6)(7)(8). This has been attributed to the role of PARP in recruiting base excision repair (BER) factors that remove damaged bases and fix single-strand breaks (SSBs) (1).…”

Mouse models of BRCA1 and BRCA2 deficiency: past lessons, current understanding and future prospects

“…To further validate this approach to targeting BRCAdeficient cells, a small intestine-specific inducible Cre line (Ah-Cre) was crossed to mice carrying conditional Brca2 F9-10 alleles as well as a reporter for Cre-mediated recombination, ROSA26R (Soriano, 1999;Hay et al, 2005b). Subsequent treatment of mice with PARP1 inhibitors resulted in depletion of reporter-positive cells specifically in Ah-Cre;Brca2 F9-10/F9-10 ;Rosa26R intestines but not in Ah-Cre;Brca2 F9-10/ þ ;Rosa26R intestines, suggesting specific depletion of Brca2-deficient cells (Hay et al, 2005a). These findings pave the way for extending the application of PARP1 inhibitors from therapeutic towards prophylactic settings.…”