Novel Doxorubicin Derivatives: Synthesis and Cytotoxicity Study in 2D and 3D in Vitro Models.

Roman Akasov,Maria Drozdova,Daria Zaytseva-Zotova,Maria Leko,Natalia Klyachko,Thierry Vandamme,Pavel Chelushkin,Elena Markvicheva,Isabelle Chevalot,Sergey Burov,Annie Marc

doi:10.15171/apb.2017.071

Abstract

Purpose: Multidrug resistance (MDR) of tumors to chemotherapeutics often leads to failure of cancer treatment. The aim of the study was to prepare novel MDR-overcoming chemotherapeutics based on doxorubicin (DOX) derivatives and to evaluate their efficacy in 2D and 3D in vitro models.Methods: To overcome MDR, we synthesized five DOX derivatives, and then obtained non-covalent complexes with human serum albumin (HSA). Drug efficacy was evaluated for two tumor cell lines, namely human breast adenocarcinoma MCF-7 cells and DOX resistant MCF-7/ADR cells. Additionally, MCF-7 cells were entrapped in alginate-oligochitosan microcapsules, and generated tumor spheroids were used as a 3D in vitro model to study cytotoxicity of the DOX derivatives.Results: Due to 3D structure, the tumor spheroids were more resistant to chemotherapy compared to monolayer culture. DOX covalently attached to palmitic acid through hydrazone linkage (DOX-N2H-Palm conjugate) was found to be the most promising derivative. Its accumulation levels within MCF-7/ADR cells was 4- and 10-fold higher than those of native DOX when the conjugate was added to cultivation medium without serum and to medium supplemented with 10% fetal bovine serum, respectively. Non-covalent complex of the conjugate with HSA was found to reduce the IC50 value from 32.9 µM (for free DOX-N2H-Palm) to 16.8 µM (for HSA-DOX-N2H-Palm) after 72 h incubation with MCF-7/ADR cells.Conclusion: Palm-N2H-DOX conjugate was found to be the most promising DOX derivative in this research. The formation of non-covalent complex of Palm-N2H-DOX conjugate with HSA allowed improving its anti-proliferative activity against both MCF-7 and MCF-7/ADR cells.

Highlights

Doxorubicin (DOX) is an anthracycline antibiotic which is widely used to treat hematological malignancies, carcinomas, and soft tissue sarcomas since early 1980th
In MCF-7 cells free DOX was found to accumulate in the cell nuclei, while all the obtained derivatives were observed outside the nuclei
Similar tendency was revealed for DOX conjugate with the hydrazide of 1-carboxy-5fluorouracil (DOX-5FU), DOX conjugate with aminoguanidine (DOX-AMG) and Palm-N2HDOX derivatives, while DOX conjugate with (4-carboxybutyl)triphenylphosphonium bromide (DOX-TPP) and N-Palm-DOX were localized outside the nucleus

Summary

Introduction

Doxorubicin (DOX) is an anthracycline antibiotic which is widely used to treat hematological malignancies, carcinomas, and soft tissue sarcomas since early 1980th. The major molecular mechanisms responsible for direct anticancer DOX effects include inhibition of topoisomerase II, nuclear DNA damage, and induction of reactive oxygen species.[1,2,3] there are some limitations of DOX cancer therapy, including lack of solubility, rather poor biodistribution, and non-specific action leading to cardiac and renal toxicity.[4]. In response to anticancer DOX therapy, multidrug resistance could be developed.[5,6] The resistance of tumor cells to DOX could be mediated through various pathways, including physiological factors (e.g. interstitial fluid pressure in tumors, diffusion limitations, hypoxia, etc.) and cellular factors which are generally associated with overexpression of ATP-binding cassette efflux transporters in cancer cells.[7,8] the physiological characteristics of tumor tissue, such as hypoxia, low nutrient supply, and low pH have been

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