The widespread clinical application of paclitaxel (PTX) in cancer treatment has been significantly limited by the emergence of drug resistance and the presence of drug-tolerant persister cells. To systematically identify key regulators of this resistance, we conducted a genome-wide CRISPR/Cas9 knockout screen, which revealed that cell division cycle 6 (CDC6) is a critical determinant of cell adhesion-mediated PTX resistance. Furthermore, our results illustrate that CDC6, an essential DNA replication licensing factor, functions through a pathway distinct from previously well-characterized resistance mechanisms. Genetic depletion of CDC6 considerably sensitizes cells, markedly increasing PTX-induced cell death. In addition to its established role in chromosome stability, CDC6 physically interacts with tropomodulin-3 (Tmod3) in the cytoplasmic compartment. This interaction enhances CDC6 protein stability and drives drug resistance phenotypes through the regulation of actin cytoskeleton remodeling and facilitating focal adhesion assembly. In addition, combination treatment with PTX and actin filament inhibitors synergistically enhanced the antitumor efficacy both in vitro and in vivo. Overall, our studies elucidate the mechanisms through which CDC6 functions as a key regulator of PTX resistance and provide a potential therapeutic strategy to increase PTX efficacy through the modulation of the cytoskeletal-adhesion axis.
Hang Yin,Xiaoling Yuan,Lihua Luo,Yichao Lu,Bing Qin,Junlei Zhang,Yingying Shi,Chunqi Zhu,Jie Yang,Xiang Li,Mengshi Jiang,Zhenyu Luo,Xinyu Shan,Dawei Chen,Jian You
The development of gene delivery has attracted increasing attention, especially when the introduction and application of the CRISPR/Cas9 gene editing system appears promising for gene therapy. However, ensuring biosafety and high gene editing efficiency at the same time poses a great challenge for its in vivo applications. Herein, a pardaxin peptide (PAR)-modified cationic liposome (PAR-Lipo) is developed. The results are indicative that significantly enhanced gene editing efficiency can be obtained through the mediation of PAR-Lipos compared to non-Lipos (non-PAR-modified liposomes) and Lipofectamine 2000, owing to its protection toward carried nucleotide by the prevention of lysosomal capture, prolongation of retention time in cells through the accumulation in the endoplasmic reticulum (ER), and more importantly, facilitation of the nuclear access via an ER-nucleus route. Accumulation of PAR-Lipos in the ER may improve the binding of Cas9 and sgRNA, thus further contributing to the eventually enhanced gene editing efficiency. Given their high biosafety, PAR-Lipos are used to mediate the knockout of the oncogene CDC6 in vivo, which results in significant tumor growth inhibition. This work may provide a useful reference for enhancing the delivery of gene editing systems, thus improving the potential for their future clinical applications.
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