Targeting the <i>MIR34C-5p</i>-ATG4B-autophagy axis enhances the sensitivity of cervical cancer cells to pirarubicin

Pirarubicin (THP) is a newer generation anthracycline anticancer drug. In the clinic, THP and THP-based combination therapies have been demonstrated to be effective against various tumors without severe side effects. However, previous clinical studies have shown that most patients with cervical cancer are not sensitive to THP treatment, and the associated mechanisms are not clear. Consistent with the clinical study, we confirmed that cervical cancer cells were resistant to THP in vitro and in vivo. Our data demonstrated that THP induced a protective macroautophagy/autophagy response in cervical cancer cells, and suppression of this autophagy dramatically enhanced the cytotoxicity of THP. By scanning the mRNA level change of autophagy-related genes, we found that the upregulation of <i>ATG4B</i> (autophagy-related 4B cysteine peptidase) plays an important role in THP-induced autophagy. Moreover, THP increased the mRNA level of <i>ATG4B</i> in cervical cancer cells by promoting mRNA stability without influencing its transcription. Furthermore, THP triggered a downregulation of <i>MIR34C-5p</i>, which was associated with the upregulation of <i>ATG4B</i> and autophagy induction. Overexpression of <i>MIR34C-5p</i> significantly decreased the level of ATG4B and attenuated autophagy, accompanied by enhanced cell death and apoptosis in THP-treated cervical cancer cells. These results for the first time reveal the presence of a <i>MIR34C-5p</i>-ATG4B-autophagy signaling axis in THP-treated cervical cancer cells in vitro and in vivo, and the axis, at least partially, accounts for the THP nonsensitivity in cervical cancer patients. This study may provide a new insight for improving the chemotherapeutic effect of THP, which may be beneficial to the further clinical application of THP in cervical cancer treatment.吡柔比星（Pirarubicin，THP）是新一代蒽环类抗肿瘤药物。临床研究表明，THP及其联合疗法对多种肿瘤均具有确切疗效，且未引发严重不良反应。然而既往临床研究显示，多数宫颈癌患者对THP治疗不敏感，其具体作用机制尚未阐明。与临床研究结果一致，我们在体内及体外实验中均证实宫颈癌细胞对THP产生耐药性。本研究数据显示，THP可在宫颈癌细胞中诱导保护性巨自噬（macroautophagy，以下简称自噬）反应，抑制该自噬过程可显著增强THP的细胞毒性。通过筛查自噬相关基因的mRNA水平变化，我们发现自噬相关4B半胱氨酸肽酶（autophagy-related 4B cysteine peptidase，ATG4B）的上调在THP诱导的自噬过程中发挥关键作用。此外，THP可通过促进mRNA稳定性而非影响转录过程，上调宫颈癌细胞中ATG4B的mRNA水平。进一步研究发现，THP会触发MIR34C-5p的表达下调，这与ATG4B的上调及自噬诱导密切相关。过表达MIR34C-5p可显著降低ATG4B的蛋白水平并减弱自噬活性，同时增强THP处理后宫颈癌细胞的死亡与凋亡程度。本研究首次在THP处理的宫颈癌细胞体内外模型中，揭示了MIR34C-5p-ATG4B-自噬信号轴的存在，该信号轴可部分解释宫颈癌患者对THP的不敏感性。本研究可为改善THP的化疗效果提供全新研究视角，有望推动THP在宫颈癌治疗中的进一步临床应用。