Abstract
Tissue architecture and cell–extracellular matrix (cell–ECM) interaction determine the organ specificity; however, the influences of these factors on anticancer drugs preclinical studies are highly neglected. For considering such aspects, three-dimensional (3D) cell culture models are relevant tools for accurate analysis of cellular responses to chemotherapy. Here we compared the MCF-7 breast cancer cells responses to cisplatin in traditional two-dimensional (2D) and in 3D-reconstituted basement membrane (3D-rBM) cell culture models. The results showed a substantial increase of cisplatin resistance mediated by 3D microenvironment. This phenotype was independent of p53 status and autophagy activity and was also observed for other cellular models, including lung cancer cells. Such strong decrease on cellular sensitivity was not due to differences on drug-induced DNA damage, since similar levels of γ-H2AX and cisplatin–DNA adducts were detected under both conditions. However, the processing of these cisplatin-induced DNA lesions was very different in 2D and 3D cultures. Unlike cells in monolayer, cisplatin-induced DNA damage is persistent in 3D-cultured cells, which, consequently, led to high senescence induction. Moreover, only 3D-cultured cells were able to progress through S cell cycle phase, with unaffected replication fork progression, due to the upregulation of translesion (TLS) DNA polymerase expression and activation of the ATR-Chk1 pathway. Co-treatment with VE-821, a pharmacological inhibitor of ATR, blocked the 3D-mediated changes on cisplatin response, including low sensitivity and high TLS capacity. In addition, ATR inhibition also reverted induction of REV3L by cisplatin treatment. By using REV3L-deficient cells, we showed that this TLS DNA polymerase is essential for the cisplatin sensitization effect mediated by VE-821. Altogether, our results demonstrate that 3D-cell architecture-associated resistance to cisplatin is due to an efficient induction of REV3L and TLS, dependent of ATR. Thus co-treatment with ATR inhibitors might be a promising strategy for enhancement of cisplatin treatment efficiency in breast cancer patients.
Highlights
Breast cancer is a major public health problem due to its high incidence and mortality[1]
3D-cultured breast cancer cells are much more resistant to cisplatin than 2D-cultured ones, subject to similar levels of DNA damage To assess changes on cellular sensitivity to cisplatin potentially mediated by cell architecture and cell–extracellular matrix (ECM) interaction, we evaluated the behavior of cisplatin-treated cells under distinct cell culture conditions: the standard monolayer culture on plastic (2D culture) versus in 3Dreconstituted basement membrane gels (3D-rBM), in which cells form micro-tissue-like structures[14]
This remarkable increase in cisplatin resistance observed in 3D-reconstituted basement membrane (3D-rBM) cultures could be as much a result of the cell–ECM contact as due to morphological changes and increased cell-to-cell contact induced by growth in 3D
Summary
Breast cancer is a major public health problem due to its high incidence and mortality[1]. Cells are embedded in a complex threedimensional (3D) microenvironment, whose cellular and chemical composition is tissue specific. Gomes et al Cell Death and Disease (2019)10:459 cellular polarity and tissue architecture. It defines the organ specificity and function[2]. In pathological conditions, such as cancer, cellular microenvironment displays a central role. By mimicking fundamental aspects of the in vivo microenvironment, 3D cell culture models might provide more assertive conclusion regarding the effectiveness of chemotherapy and emerge as more robust methodologies for drug screening analysis[5]
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