Hwang Hyemin, Kim Jimin, Kim Tae-Hun, Han Yeonju, Choi Dayoung, Cho Sua, Kim Seunghwan, Park Sanghee, Park Taehyun, Piccinini Filippo, Rhee Won Jong, Pyun Jae-Chul, Lee Misu
Division of Life Sciences, College of Life Science and Bioengineering, Incheon National University, Incheon, 22012, Republic of Korea.
Department of Materials Science and Engineering, Yonsei University, 50 Yonsei-Ro, Seodaemun-Gu, Seoul, 03722, Republic of Korea.
BMC Cancer. 2024 Dec 20;24(1):1557. doi: 10.1186/s12885-024-13342-y.
Hepatocellular carcinoma (HCC) stands as the sixth most prevalent cancer globally, presenting a substantial health challenge, particularly due to late-stage diagnoses that limit treatment effectiveness. Sorafenib, a multi-kinase inhibitor, is the primary chemotherapeutic agent for advanced HCC, but it only extends survival by 2-3 months. However, drug resistance remains a major clinical challenge, necessitating the exploration of new molecular mechanisms, including the role of microRNAs (miRNAs) in sorafenib resistance. In this study, we aimed to identify miRNAs within exosomes derived from sorafenib-resistant HCC cells to elucidate the molecular mechanisms underlying resistance.
Sorafenib-resistant cells were generated by culturing the human HCC cell line Huh7 in a medium containing 20 µM sorafenib for six months. Exosomes were isolated from the conditioned medium 24 h before cell harvest using exosome-depleted serum medium. miRNA sequencing and western blotting were used to analyze the expression profiles of exosomal miRNAs and proteins, respectively. pH measurement was performed to assess pH changes in response to sorafenib treatment and miRNA modulation.
A total of 180 exosomal miRNAs were found to be dysregulated between sorafenib-treated control Huh7 (Huh7S) and sorafenib-resistant Huh7 (Huh7RS) cells, as well as between untreated control Huh7 and Huh7RS cells. Among these, miR-6126 was significantly downregulated in Huh7RS cells compared to Huh7S cells. Functional studies using 2-dimensional (D) and 3D cell culture systems revealed that miR-6126 overexpression reduced sorafenib resistance in Huh7RS cells, while its inhibition increased resistance in Huh7 cells. miR-6126 downregulated key proteins involved in cancer stem cell maintenance, such as CD44 and HK2. Furthermore, the pH level was elevated in cells overexpressing miR-6126 following sorafenib treatment, whereas inhibiting miR-6126 resulted in a lower pH.
Exosomal miR-6126 plays a pivotal role in sorafenib resistance and tumorigenesis, highlighting its potential as a novel therapeutic target for overcoming drug resistance in HCC.
肝细胞癌（HCC）是全球第六大常见癌症，构成了重大的健康挑战，尤其是因为晚期诊断限制了治疗效果。索拉非尼是一种多激酶抑制剂，是晚期HCC的主要化疗药物，但它仅能将生存期延长2至3个月。然而，耐药性仍然是一个主要的临床挑战，因此有必要探索新的分子机制，包括微小RNA（miRNA）在索拉非尼耐药中的作用。在本研究中，我们旨在鉴定源自索拉非尼耐药HCC细胞的外泌体中的miRNA，以阐明耐药的分子机制。
通过在含有20 μM索拉非尼的培养基中培养人HCC细胞系Huh7六个月来产生索拉非尼耐药细胞。在收获细胞前24小时，使用不含外泌体的血清培养基从条件培养基中分离外泌体。分别使用miRNA测序和蛋白质印迹分析外泌体miRNA和蛋白质的表达谱。进行pH测量以评估对索拉非尼治疗和miRNA调节的pH变化。
在索拉非尼处理的对照Huh7（Huh7S）和索拉非尼耐药的Huh7（Huh7RS）细胞之间，以及未处理的对照Huh7和Huh7RS细胞之间，共发现180种外泌体miRNA表达失调。其中，与Huh7S细胞相比，miR-6126在Huh7RS细胞中显著下调。使用二维（2D）和三维（3D）细胞培养系统进行的功能研究表明，miR-6126过表达降低了Huh7RS细胞中的索拉非尼耐药性，而其抑制则增加了Huh7细胞中的耐药性。miR-6126下调了参与癌症干细胞维持的关键蛋白，如CD44和HK2。此外，索拉非尼处理后，过表达miR-6126的细胞中的pH水平升高，而抑制miR-6126则导致较低的pH。
外泌体miR-6126在索拉非尼耐药和肿瘤发生中起关键作用，突出了其作为克服HCC耐药性的新型治疗靶点的潜力。
