非编码小核仁RNA SNORD33在三阴性乳腺癌铂类化疗抵抗中的作用及其分子机制

Triple-negative breast cancer (TNBC) represents a continuing challenge due to limited approaches of treatments. Platinum has a favorable efficacy in the treatment of TNBC. However, many patients relapse despite having received platinum therapy, and resistance inevitably develops. ..Small nucleolar RNAs (snoRNAs), a subclass of small non-coding RNAs with the function of rRNA modification and RNA slicing, play a crucial role in controlling cell behavior and carcinogenesis. snoRNAs can also be detected in plasma and is considered as potential non-invasive biomarkers for early cancer diagnosis and prognosis prediction. ..In our preliminary experiments, we found that small nucleolar RNA 33 (SNORD33) was significantly decreased in cisplatin-resistant TNBC cells. Patients with low plasma SNORD33 level showed worse efficacy of cisplatin. Downregulation of SNORD33 promoted the resistance of cisplatin in TNBC cells, and inhibited cell apoptosis. Despite no changes have been found in resistance of cisplatin due to interfering host gene or slicing to miRNA-like fragments, we found that SNORD33 could bind to methyl-CpG binding protein 2 (MeCP2).These data suggested that SNORD33 might interact with MeCP2 and regulate the transcription of target genes involving apoptosis pathway. ..In this proposal, we plan to investigate the interaction between SNORD33 and MeCP2 and interference of platinum. We will employ multidiscipline approaches encompassing biochemistry and molecular biology, breast tumor in situ model and patient-derived xenograft model to pursue our aims above. The proposed studies in this application will not only allow us to advance the knowledge of molecular mechanisms in snoRNA research, but may also produce new leads to predict efficacy and treat TNBC.

三阴性乳腺癌是乳腺癌领域的治疗难点。铂类是三阴性乳腺癌治疗的一线药物，但多数患者治疗后仍出现耐药。小核仁RNA（snoRNA）参与肿瘤发生发展并可通过血浆检测。课题组前期研究发现，顺铂耐药的三阴性乳腺癌细胞SNORD33低表达，血浆SNORD33低水平的患者铂类治疗疗效不佳；下调SNORD33的细胞表现为顺铂耐药，凋亡减少；未见其宿主基因及miRNA样作用参与调控铂类敏感性，而发现SNORD33可与甲基CpG结合蛋白2（MeCP2）结合。提示SNORD33可能通过与MeCP2结合，调控下游凋亡相关靶基因转录，影响铂类敏感性。本项目拟结合使用分子生物学技术、小鼠原位肿瘤及PDX模型等手段，深入研究SNORD33通过与MeCP2相互作用、干扰铂类药物作用等机制影响铂类耐药，并探索其作为预测疗效及提高铂类敏感性靶点的可行性。以期丰富对snoRNA这种非编码RNA的认知，拓展其临床应用价值。

三阴性乳腺癌是具有高侵袭性及转移性的乳腺癌亚型，患者复发比例高，总体生存率低，是临床治疗难点。铂类药物是治疗三阴性乳腺癌的有效药物，然而，耐药仍不可避免。既往包括BRCA突变在内的标志物对铂类疗效的预测效力在不同研究中的结果并不一致。. snoRNA是一类小分子非编码RNA，参与核糖体的生物合成以及剪接体的剪切组装，且在血浆中含量丰富，可作为液体活检的潜在生物标志物。研究团队发现，顺铂耐药的三阴性乳腺癌细胞SNORD33表达显著降低，下调SNORD33促进顺铂耐药。进而，在既往一线接受含铂和非铂方案治疗的转移性三阴性乳腺癌患者的血浆样本中发现，血浆SNORD33低水平的患者无进展生存时间（PFS）和总生存时间（OS）显著缩短，血浆SNORD33特异性预测铂类疗效，是使用含铂方案治疗患者PFS的独立预测因子。研究基于肝转移状态、转移灶数目的临床特征及血浆SNORD33水平绘制了预测含铂方案PFS时间的列线阵图，用于预测患者铂类药物治疗不同时间临床缓解可能性，以供临床医生/患者参考，选择治疗方案。. 研究发现SNORD33存在与甲基化结合蛋白MeCP2的结合，SNORD33下调增加MeCP2与下游靶基因启动子甲基化区域，释放MeCP2的转录抑制活性，进一步抑制包括GADD45α、MYOD1、FOXF1、CDKL5、CLDN6在内的凋亡相关的基因的表达，细胞凋亡减少，使得三阴性乳腺癌细胞对铂类的抵抗增加。. 本研究结果提供了高准确性、检测便利、非侵入性的新的分子标记物和治疗的特异性靶标。采集0.2毫升血浆检测SNORD33水平，结合临床特征，即可实现预测转移性三阴性乳腺癌、肺腺癌及膀胱癌患者铂类药物疗效，为肿瘤患者带来更多获益。