Cytosine Β-D Arabino-Furanoside (Ara-C) and CPT-11 inhibit cell proliferation in human colorectal cancer with microsatellite instability

Abstract

2061 Background: Recent studies suggest 5-Fluorouracil-based adjuvant chemotherapy does not improve survival in colorectal cancer (CRC) patients with microsatellite instability (MSI) (NEJM, 349 July: 247–257, 2003). We have reported decreased expression of p12DOC-1 is associated with increased proliferation and apoptosis arrest in human CRC with MSI (Oncogene, 22: 6304–10, 2003). The purpose of this study was to measure differential expression of p12DOC-1 using a Cancer Cell Line Profiling (CCLP) Array (Clontech) and to assess the efficacy of CPT-11 and oxaliplatin, in human MSI CRC cell lines. Methods: A CCLP array containing 702 complex cDNA samples from 26 different human cancer cell lines was treated with a range of chemotherapeutic agents. The array was hybridized with a p32-dCTP labeled p12DOC-1 probe. Array results were confirmed using real-time RT-PCR. In addition, the toxicity of CPT-11 and oxaliplatin (neither agent tested on array) against CRC cell lines, RKO (MSI) and SW480 (microsatellite stable), was measured pre and post transfection with pcDNA3-p12DOC-1. Cell survival was assessed by MTT assay and expressed as IC50 values (50% inhibitory concentration). Results: Ara-C increased p12DOC-1 expression in 2 of 3 MSI CRC cell lines by CCLP array and was confirmed by real-time RT-PCR. Significantly decreased S phase and increased apoptosis was also observed. RT-PCR confirmed an 8-fold increase in p12DOC-1 expression in MSI CRC cell lines after transfection with pcDNA3-p12DOC-1. IC50 values for CPT-11 were significantly higher in all cell lines with increased p12DOC-1 expression (mean IC50= 37 μM), compared to the RKO wild type control (minimal DOC-1 expression) (IC50 =1 μM) (p< 0.05). No differential IC50 values were observed for oxaliplatin. Conclusions: Ara-C induces p12DOC-1 expression, inhibits cell proliferation and increases apoptosis in MSI CRC. In addition, MSI CRC is extremely sensitive to CPT-11. These results suggest that differential gene expression analysis may assist selection of optimal chemotherapeutic agents specifically for the treatment of 5FU resistant MSI CRC. No significant financial relationships to disclose.