Drug targeting to decrease cardiotoxicity - determination of the cytotoxic effect of GnRH-based conjugates containing doxorubicin, daunorubicin and methotrexate on human cardiomyocytes and endothelial cells.

Abstract

Background: Cardiomyopathy induced by the chemotherapeutic agents doxorubicin and daunorubicin is a major limiting factor for their application in cancer therapy. Chemotactic drug targeting potentially increases the tumor selectivity of drugs and decreases their cardiotoxicity. Increased expression of gonadotropin-releasing hormone (GnRH) receptors on the surface of tumor cells has been reported. Thus, the attachment of the aforementioned chemotherapeutic drugs to GnRH-based peptides may result in compounds with increased therapeutic efficacy. The objective of the present study was to examine the cytotoxic effect of anticancer drug–GnRH-conjugates against two essential cardiovascular cell types, such as cardiomyocytes and endothelial cells. Sixteen different previously developed GnRH-conjugates containing doxorubicin, daunorubicin and methotrexate were investigated in this study. Their cytotoxicity was determined on primary human cardiac myocytes (HCM) and human umbilical vein endothelial cells (HUVEC) using the xCELLigence SP system, which measures impedance changes caused by adhering cells on golden electrode arrays placed at the bottom of the wells. Slopes of impedance–time curves were calculated and for the quantitative determination of cytotoxicity, the difference to the control was analysed.Results: Doxorubicin and daunorubicin exhibited a cytotoxic effect on both cell types, at the highest concentrations tested. Doxorubicin-based conjugates (AN-152, GnRH-III(Dox-O-glut), GnRH-III(Dox-glut-GFLG) and GnRH-III(Dox=Aoa-GFLG) showed the same cytotoxic effect on cardiomyocytes. Among the daunorubicin-based conjugates, [4Lys(Ac)]-GnRH-III(Dau=Aoa), GnRH-III(Dau=Aoa-YRRL), {GnRH-III(Dau=Aoa-YRRL-C)}2 and {[4N-MeSer]-GnRH-III(Dau-C)}2 had a significant but decreased cytotoxic effect, while the other conjugates – GnRH-III(Dau=Aoa), GnRH-III(Dau=Aoa-K(Dau=Aoa)), [4Lys(Dau=Aoa)]-GnRH-III(Dau=Aoa), GnRH-III(Dau=Aoa-GFLG), {GnRH-III(Dau-C)}2 and [4N-MeSer]-GnRH-III(Dau=Aoa) – exerted no cytotoxic effect on cardiomyocytes. Mixed conjugates containing methotrexate and daunorubicin – GnRH-III(Mtx-K(Dau=Aoa)) and [4Lys(Mtx)]-GnRH-III(Dau=Aoa) – showed no cytotoxic effect on cardiomyocytes, as well.Conclusion: Based on these results, anticancer drug–GnRH-based conjugates with no cytotoxic effect on cardiomyocytes were identified. In the future, these compounds could provide a more targeted antitumor therapy with no cardiotoxic adverse effects. Moreover, impedimetric cytotoxicity analysis could be a valuable technique to determine the effect of drugs on cardiomyocytes.