Cancer cell adaptation to chemotherapy

Abstract

Background: Tumor resistance to chemotherapy may be present at the beginning of treatment, develop during treatment, or become apparent on re-treatment of the patient. The mechanisms involved are usually inferred from experiments with cell lines, as studies in tumor-derived cells are difficult. Studies of human tumors show that cells adapt to chemotherapy, but it has been largely assumed that clonal selection leads to the resistance of recurrent tumors.

Methods: Cells derived from 47 tumors of breast, ovarian, esophageal, and colorectal origin and 16 paired esophageal biopsies were exposed to anticancer agents (cisplatin; 5-fluorouracil; epirubicin; doxorubicin; paclitaxel; irinotecan and topotecan) in short-term cell culture (6 days). Real-time quantitative PCR was used to measure up- or down-regulation of 16 different resistance/target genes, and when tissue was available, immunohistochemistry was used to assess the protein levels.

Results: In 8/16 paired esophageal biopsies, there was an increase in the expression of multi-drug resistance gene 1 (MDR1) following epirubicin + cisplatin + 5-fluorouracil (ECF) chemotherapy and this was accompanied by increased expression of the MDR-1 encoded protein, P-gp. Following exposure to doxorubicin in vitro, 13/14 breast carcinomas and 9/12 ovarian carcinomas showed >2-fold down-regulation of topoisomerase IIalpha (TOPOIIalpha). Exposure to topotecan in vitro, resulted in >4-fold down-regulation of TOPOIIalpha in 6/7 colorectal tumors and 8/10 ovarian tumors.

Conclusion: This study suggests that up-regulation of resistance genes or down-regulation in target genes may occur rapidly in human solid tumors, within days of the start of treatment, and that similar changes are present in pre- and post-chemotherapy biopsy material. The molecular processes used by each tumor appear to be linked to the drug used, but there is also heterogeneity between individual tumors, even those with the same histological type, in the pattern and magnitude of response to the same drugs. Adaptation to chemotherapy may explain why prediction of resistance mechanisms is difficult on the basis of tumor type alone or individual markers, and suggests that more complex predictive methods are required to improve the response rates to chemotherapy.

Figure 1

Changes due to chemotherapy in biopsies taken before and after chemotherapy. Cytotoxicity of a) doxorubicin; b) mitoxantrone; c) topotecan; d) paclitaxel and e) cisplatin (expressed as IC50 μM) in paired samples obtained from ovarian cancer patients before and after they were treated with an anthracycline containing regimen (paclitaxel+mitoxantrone n = 7; liposomal doxorubicin n = 2). Each line represents an individual patient. (f) Gene expression changes pre- and post-treatment (paclitaxel plus mitoxantrone chemotherapy) in one patient were analysed by qRT-PCR following in vitro exposure to doxorubicin.

Figure 2

Changes in relative gene expression in 6 paired esophageal tumor biopsies. a) Changes in MDR1 relative gene expression and b) changes in TS relative gene expression. Each dot represents the relative mRNA level for an individual tumor, measured before and after ECF chemotherapy.

Figure 3

Changes in relative expression of putative chemoresistance genes in tumor-derived cells. Each dot represents the relative mRNA level for an individual sample, measured after in vitro drug exposure compared with untreated control cells. (a) TOPO IIα expression in breast (black lines) and ovarian (red lines) tumor cells after doxorubicin exposure. (b) TOPO I expression in breast (black lines) and ovarian (red lines) tumor cells after doxorubicin exposure. (c) MRP1 expression in breast (black lines) and ovarian (red lines) tumor cells after doxorubicin exposure. (d) TOPO I expression in ovarian tumor cells after topotecan exposure. (e) TOPO IIα expression in ovarian tumor cells after topotecan exposure. (f) TS expression in breast (red lines) and colorectal (black lines) tumor cells after 5-FU exposure. (g) ERCC1 expression in breast (black lines) and ovarian (red lines) tumor cells after cisplatin exposure. (h) TS expression in esophageal tumor cells after ECF exposure. The numerical data for these graphs is summarized in Tables 5-8.