Search

The combination of the topoisomerase I (Topo I) inhibitor CPT-11 with the anti-epidermal growth factor receptor (EGFR) agent Gefitinib (Iressa™, ZD1839) represents a promising medical approach for colorectal cancer patients. In this report, we provide pre-clinical evidences for their optimal combination schedule in HT-29 and LoVo human colon cancer cell lines. We analyzed the different effects that three different combination schedules of SN-38 (the active CPT-11 metabolite) and Gefitinib (Gefitinib before; Gefitinib simultaneously; Gefitinib after SN-38) have on cell growth, cell cycle, apoptosis, and expression/phosphorylation of EGFR, Topo I and some steps of the signal transduction pathway. We first determined the IC50 of each drug choosing the 5 days exposure for Gefitinib (0.6 and 3.8 μM for LoVo and HT-29 cells, respectively) and 1 day exposure for SN-38 (0.31 and 0.5 μM for LoVo and HT-29 cells, respectively). The different drug combination schedules were tested in various concentrations by using equiactive concentrations of the two drugs. The cytotoxicity of Gefitinib and SN-38 combination was schedule- and concentration-dependent but not cell line-specific. The most synergistic schedule was Gefitinib given after SN-38, with combination indexes (CI) of 0.007 and 0.454 in HT-29 and LoVo, respectively. Analysis of bio-molecular targets showed that Gefitinib was able to modulate SN-38 ability to inhibit Topo I, to accumulate cells in S-phase, and to induce apoptosis. Interestingly, SN-38 was able to activate EGFR and its signal transduction pathway. Confirming preliminary clinical experience of Gefitinib with other cytotoxic drugs, it seems that Gefitinib after SN-38 represents the best cytotoxic combination schedule but the biomolecular basis for this synergism remain to be completely elucidated. [Copyright &y& Elsevier]

ZD1839 (‘Iressa’), a selective epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), is currently undergoing preclinical and clinical evaluation in several solid tumors. The present study aimed to assess the effect of ZD1839 in combination with oxaliplatin in the colon cancer cell lines HT-29 and LoVo. For in vitro chemosensitivity testing, cells were treated with serial dilutions of each drug sequentially at a fixed ratio of doses that corresponded to 1/20, 1/10, 1/5, 1/2, 1, 1.5 and 2 times the individual ic50 values. Oxaliplatin followed by ZD1839 produced a synergistic effect. In contrast, oxaliplatin following ZD1839 exhibited an additive effect at best. Mass spectrometry examination revealed that ZD1839 modestly enhanced cellular oxaliplatin accumulation and platinum-DNA (Pt-DNA) adducts (P>0.05). In additional studies, high-performance liquid chromatography revealed that oxaliplatin had no effect on ZD1839 accumulation. In contrast, ZD1839 markedly inhibited removal of Pt-DNA adducts (P<0.05). With oxaliplatin treatment (1 day) followed by ZD1839 (1 day), then incubation with drug-free medium (1 day), 90% of Pt-DNA adducts remained. Apoptosis examination revealed that oxaliplatin-induced apoptosis was prolonged by sequential oxaliplatin followed by ZD1839 treatment compared with oxaliplatin alone. Further experiments revealed that ZD1839 decreased cellular γ-glutamyltransferase activity.Conclusions: The above observations provide a mechanistic explanation for the synergy of oxaliplatin followed by ZD1839, and suggest that this treatment combination warrants further preclinical and clinical investigation. [Copyright &y& Elsevier]