Rectal cancer was diagnosed in 40,340 Americans in 2005. In spite of multimodality therapy, the rate of recurrence remains high and 50% of patients die of their disease. In order to understand the factors leading to ionizing radiation (IR) resistance to apoptosis, we submitted primary colon cancer cells (stage II) SW480 and lymph node metastatic (stage III) SW620 cells to DNA fragmentation studies following IR treatments at 0, 2, 4, and 6 Gy. Our studies demonstrated that SW620 cells are more resistant to apoptosis after ionizing radiation (IR) compared to SW480 cells (32.9 ± 5.9 vs. 71.0 ± 11.5%; p = 0.041) at 4 Gy and (41.7 ± 8.7 vs. 97.0 ± 3.0 %; p = 0.004) at 6 Gy. We hypothesized that the apoptotic signaling pathway in SW620 might have been genetically modified resulting in either the down-regulation of

Smac/DIABLO or the up-regulation of the inhibitors of apoptosis (IAPs). Because survivin is up-regulated in several malignancies and because XIAP is the most potent mammalian caspase inhibitor, we elected to study these IAPs in radioresistance to apoptosis in SW620 cells. Immunocytochemistry was performed in cell treated with IR at 0, 2, 4, and 6 Gy with antibodies specific for Smac/DIABLO in SW480 and SW620 cells. Following the same treatment, Western blot analysis was performed with antibodies specific for survivin and XIAP. Our results demonstrated a substantial increase in translocation of Smac/DIABLO in SW480 cells compared to SW620 cells at 2 and 4 Gy (80.9 ± 3.7 % vs. 36.6 ± 5.6; p = 0.001 and 100 vs. 82.1%; p = 0.03, respectively). Survivin demonstrated a moderate increase in protein levels in both cell lines. However, XIAP protein levels were markedly increased in SW620 cells in an IR-dose dependent manner compared to SW480 cells. This data suggest that metastatic colorectal cancer cells resistant to IR-mediated apoptosis have a substantially increased XIAP/Smac ratio. Thus, factors targeting inhibition of NFκB or pro-apoptotic mediators resulting in an increase mitochondrial permeability to Smac/DIABLO (or Smac-mimetics) are potential targets for radiosensitization of rectal cancers resistant to conventional radio-therapeutic modalities.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA