Microarray-based detection and expression analysis of drug resistance in an animal model of peritoneal metastasis from colon cancer

Abstract

Chemotherapy drugs efficiently eradicate rapidly dividing differentiated cells by inducing cell death, but poorly target slowly dividing cells, including cancer stem cells and dormant cancer cells, in the later course of treatment. Prolonged exposure to chemotherapy results in a decrease in the proportion of apoptotic cells in the tumour mass. To investigate and characterize the molecular basis of this phenomenon, microarray-based expression analysis was performed to compare tHcred²-DEVD-EGFP-caspase 3-sensor transfected C-26 tumour cells that were harvested after engraftment into mice treated with or without 5-FU. Peritoneal metastasis was induced by intraperitoneal injection of C-26 cells, which were subsequently reisolated from omental metastatic tumours after the mice were sacrificed by the end of the 10th day after tumour injection. The purity of reisolated tHcred2-DEVD-EGFP-caspase 3-sensor-expressing C-26 cells was confirmed using FLIM, and total RNA was extracted for gene expression profiling. The validation of relative transcript levels was carried out via real-time semiquantitative RT‒PCR assays. Our results demonstrated that chemotherapy induced the differential expression of mediators of cancer cell dormancy and cell survival-related genes and downregulation of both intrinsic and extrinsic apoptotic signalling pathways. Despite the fact that some differentially expressed genes, such as BMP7 and Prss11, have not been thoroughly studied in the context of chemoresistance thus far, they might be potential candidates for future studies on overcoming drug resistance.

Keywords: Apoptosis; BMP7; Cell survival; Chemoresistance; Chemotherapy; Prss11.

Fig. 1

FLIM of caspase-3 sensor transfected C26 cells in metastatic omental tissue and following their reisolation. The top-left panel depicts a representative image of the normal peritoneal cavity in a mouse. The top-right panel presents an image of peritoneal metastasis generated by the injection of caspase-3 sensor-transfected C26 cells into the peritoneal cavity. The lower-left panel shows a FLIM image of metastatic omental tissue, while the lower-right panel shows a FLIM image after the C26 cells were reisolated and seeded onto a culture dish

Fig. 2

Comparison of mice teated with or without 5-FU for 120 h. The relative expression values for the two mouse experimental groups (mice treated with 5-FU for 120 h compared to untreated mice) are plotted, significantly differentially expressed genes are shown (P < 0.05)