School of Basic Medical Sciences, Capital Medical University, Beijing, 100069, China.
Laboratory for Clinical Medicine, Capital Medical University, Beijing, 100069, China.
Mol Cancer. 2024 Oct 31;23(1):243. doi: 10.1186/s12943-024-02145-1.
Small cell lung cancer (SCLC) stands as one of the most lethal malignancies, characterized by a grim diagnosis and prognosis. The emergence of multi-drug resistance poses a significant hurdle to effective therapy. Although previous studies have implicated the long noncoding RNA LYPLAL1-DT in the tumorigenesis of SCLC, the precise role of the highly expressed LYPLAL1-DT in SCLC chemoresistance and the underlying mechanism remain inadequately understood.
cDDP-, VP-16- and PTX-resistant SCLC cells lines were established. The viabilities of SCLC cells were assessed by CCK-8 assay in vitro and xenograft tumor formation assay in vivo. Apoptosis was evaluated by FACS, Western blot and JC-1 fluorescence staining, while autophagy was explored via autophagic flux detection under confocal microscopy and autophagic vacuole investigation under transmission electron microscopy (TEM). The functional role and mechanism of LYPLAL1-DT were further investigated by gain- and loss-of-function assays in vitro. Furthermore, the therapeutic efficacy of the combination of venetoclax and HCQ with cDDP, VP-16 or PTX was evaluated by cell line, cell-derived xenograft (CDX) and patient-derived xenograft (PDX) mice model.
Our findings revealed that LYPLAL1-DT is upregulated in chemoresistant SCLC cell lines. Gain- and loss-of-function assays demonstrated that LYPLAL1-DT impairs sensitivity to cDDP, VP-16, or PTX both in vitro and in vivo. Overexpression of LYPLAL1-DT significantly enhanced autophagy and inhibited apoptosis in SCLC cells. Further analyses, including RIP and RNA pull-down assays, revealed that LYPLAL1-DT promotes the expression of BCL2 by sponging miR-204-5p and is implicated in the assembly of the autophagy-specific complex (BECN1/PtdIns3K complex). Combining venetoclax and HCQ with cDDP, VP-16, or PTX effectively mitigated chemoresistance in SCLC cells and suppressed tumor growth in CDX and PDX models without inducing obvious toxic effects.
Our findings demonstrate that upregulation of LYPLAL1-DT sequesters apoptosis through the LYPLAL1-DT/miR-204-5p/BCL2 axis and promotes autophagy by facilitating the assembly of the BECN1/PtdIns3K complex, thereby mediating multi-drug resistance of SCLC. The triple combination of venetoclax, HCQ, in conjunction with cDDP, VP-16 or PTX overcomes refractory SCLC, shedding light on a potential therapeutic target for combating SCLC chemoresistance.
小细胞肺癌（SCLC）是最致命的恶性肿瘤之一，其诊断和预后均十分严峻。多药耐药的出现对有效治疗构成了重大障碍。尽管先前的研究表明长链非编码 RNA LYPLAL1-DT 参与了 SCLC 的肿瘤发生，但高表达的 LYPLAL1-DT 在 SCLC 化疗耐药中的确切作用及其潜在机制仍了解不足。
建立了顺铂（cDDP）、依托泊苷（VP-16）和紫杉醇（PTX）耐药的 SCLC 细胞系。通过体外 CCK-8 测定和体内异种移植肿瘤形成测定评估 SCLC 细胞的活力。通过流式细胞术、Western blot 和 JC-1 荧光染色评估细胞凋亡，通过共聚焦显微镜下自噬流检测和透射电子显微镜（TEM）下自噬小体观察评估自噬。通过体外的 gain-和 loss-of-function 测定进一步研究 LYPLAL1-DT 的功能作用和机制。此外，通过细胞系、细胞衍生的异种移植（CDX）和患者衍生的异种移植（PDX）小鼠模型评估 venetoclax 和 HCQ 联合 cDDP、VP-16 或 PTX 的治疗效果。
我们的研究结果表明，LYPLAL1-DT 在耐药性 SCLC 细胞系中上调。gain-和 loss-of-function 测定表明，LYPLAL1-DT 在体外和体内均降低了 SCLC 细胞对 cDDP、VP-16 或 PTX 的敏感性。LYPLAL1-DT 的过表达显著增强了 SCLC 细胞的自噬并抑制了细胞凋亡。进一步的分析，包括 RIP 和 RNA 下拉测定，表明 LYPLAL1-DT 通过海绵吸附 miR-204-5p 促进 BCL2 的表达，并参与了自噬特异性复合物（BECN1/PtdIns3K 复合物）的组装。venetoclax 和 HCQ 联合 cDDP、VP-16 或 PTX 可有效减轻 SCLC 细胞的化疗耐药性，并抑制 CDX 和 PDX 模型中的肿瘤生长，而没有引起明显的毒性作用。
我们的研究结果表明，上调的 LYPLAL1-DT 通过 LYPLAL1-DT/miR-204-5p/BCL2 轴抑制细胞凋亡，并通过促进 BECN1/PtdIns3K 复合物的组装来促进自噬，从而介导 SCLC 的多药耐药性。venetoclax、HCQ 与 cDDP、VP-16 或 PTX 的三联组合克服了难治性 SCLC，为治疗 SCLC 化疗耐药提供了新的潜在靶点。
