Abstracts: Abstracts of Papers Submitted to the 36th Annual Meeting of the American Pancreatic Association, November 3-4, 2005
Depts of 1Surgery, 2Physiology, and 3Radiation Oncology, University of Michigan, Ann Arbor, MI; 4Depts of Cancer Biology and Medical Oncology, 5M.D. Anderson Cancer. Center, Houston, TX
We have recently determined that Ataxia-Telangectasia Group D Associated gene (ATDC) is overexpressed in pancreatic cancer, however the cellular function of ATDC is unknown. Based on preliminary studies, we hypothesize that ATDC is a novel DNA repair gene.
We performed experiments in Panc-1 cells (high ATDC expression) and Panc-1 cells stably expressing shRNA targeting ATDC or control shRNA constructs. Silencing of ATDC in shRNA-expressing cells was confirmed by western blotting. The effect of down-regulating ATDC on phosphorylation of histone H2AX (measure of DS-DNA damage) by radiation (10 Gy for 1-24 hrs) was assessed by Western blotting. The impact of ATDC silencing on the DNA damage-dependent S-phase checkpoint was evaluated by measuring radio-resistant DNA synthesis (RDS). Cell survival 14 days following radiation exposure (0-10 Gy) was determined by a colonigenic assays.
Silencing of ATDC in Panc-1 cells resulted in marked upregulation of DS DNA damage (measured by phosphorylation of H2AX) following exposure to radiation (10 Gy), with little evidence of DS DNA damage in control shRNA cells. In RDS assays, 1 hr after exposure to 10Gy radiation, ATDC silencing significantly increased RDS, from 45 ± 3% (control shRNA) to 75 ± 8% (ATDC shRNA) (p < 0.05). In clonagenic assays, ATDC silencing decreased the surviving cell fraction at all radiation levels, with 17% of ATDC-silenced cells surviving at a dose of 2 Gy compared to 50% of control shRNA cells (p < 0.05).
Silencing of ATDC renders pancreatic cancer cells more sensitive to DS DNA damage induced by cellular insult (ionizing radiation). ATDC may function as a novel DNA repair gene in protecting pancreatic cancer cells from normal apoptotic mechanisms that function in damaged cells. These findings may also explain the relative resistance of pancreatic cancer to radiation and offer a molecular target for treatment.