Synergistic cytotoxic activity of thiazolo[5,4-b]quinoline derivative D3CLP in combination with cisplatin against human cervical cancer cells

I. González-Sánchez, A. Navarrete-Castro, A. Lira-Rocha, M. A. Loza-Mejía, C. A. Mendoza-Rodríguez, C. Coronel-Cruz, M. A. Cerbón-Cervantes. Faculty of Chemistry, National Autonomous University of Mexico, M é xico, D. F., Mexico

BACKGROUND

In different types of cancer, the combined chemotherapy has been shown better results than the use of single antineoplastic drug. These drugs are diverse, and a great number becomes both from synthetic or natural sources. D3CLP is a cytotoxic thiazolo[5,4-b]quinoline synthetic derivative which has been shown that significantly activate effector caspases in leukemia cells, while it display low toxicity against non-tumoral cells. The aim of this study was to determine if D3CLP could enhance the cytotoxicity in combination with other anti-neoplastic drugs against different tumoral cell lines.

METHODS

Different human cancer cells lines from cervix (HeLa), leukemia (K-562) and breast (MCF-7) were cultured and administrated with D3CLP, imatinib, tamoxifen or cisplatin, respectively. From single cytotoxic activity data, new doses were proposed for evaluating the combined effect of D3CLP with chemotherapeutic drugs in current clinical use in three proportions based on its IC50. The cell viability induced after 48 h of treatment was evaluated by MTT assay. Data from combined treatments were analyzed by using both the combination index and isobologram method to determine synergism, additive or antagonism. DNA fragmentation drug induced was evaluated by TUNEL assay for D3CLP, cisplatin and its combination in a ratio 3:1 to explore the cell death mechanism. The results are the mean of three independent experiments±SEM and were analyzed by student’s t-test and a p<0.05 was considered significant.

RESULTS

The concentrations that induce 50% of cell viability (IC50) after treatment of D3CLP were 8.55±1.29, 11.22±1.45 and 8.81±1.17 µM in K-562, MCF-7 and HeLa cells, respectively. Additionally, the IC50 for imatinib, tamoxifen and cisplatin were 2.37±0.12, 22.71±1.59 and 15.62±0.48 µM in K-562, MCF-7 and HeLa cells, respectively. The combination analysis indicate that D3CLP with anti-neoplastic drugs induced a synergism by combination index for both D3CLP plus imatinib in K-562 leukemia cells in 3:1 and 1:1 proportions, and for D3CLP with cisplatin in a 3:1 ratio in HeLa cells (Table 1). Furthermore, isobolographic analysis indicates a statistically significant synergism for the combination of D3CLP with cisplatin in a 3:1 ratio in HeLa cells.

Table 1. Combination index for the combined treatment.

CI<1, CI=1 and CI>1; represents synergism, additive or antagonism, respectively.

The treatment of HeLa cells with D3CLP induces features suggestive of apoptosis since 12 h, while cisplatin has shown at 36 h. This indicates that, the cellular target by which D3CLP induces cell death is different from that of cisplatin. Furthermore, differences in the mechanisms of action of these drugs validates the use of ecuation for the mixture of mutually exclusive drugs in the CI described by Chou and Talay.

CONCLUSION

This study demonstrated the in vitro synergism of D3CLP and cisplatin in HeLa cells, which was confirmed by both CI and isobolographic analysis, and suggest that this compound is a good candidate to be evaluated in preclinical studies.