Study on the role and mechanism of LAPTM4B in promoting malignant proliferation of renal cell carcinoma via the PI3K/AKT signaling pathway
LAPTM4B通过PI3K/AKT信号通路促进肾细胞癌恶性增殖的作用及机制研究

摘要

目的 研究LAPTM4B在肾细胞癌（RCC）中表达情况以及其激活PI3K/AKT信号传导机制促使肿瘤恶变增殖的作用机理。方法 收集自2019年6月至2023年6月份在我院接受根治性肾切除手术的RCC患者的癌组织与癌旁正常组织样本各64份，以免疫组化染色法、Western blot及荧光定量PCR的方法检测LAPTM4B的表达量；建立LAPTM4B持续性下调的786-O细胞株，应用CCK-8法、克隆形成法以及流式细胞仪分析细胞增殖情况及其周期变化情况；运用Western blot检测PI3K/AKT信号通路主要蛋白的磷酸化状态；借助PI3K激活剂740Y-P进行救援实验以探索LAPTM4B影响肾细胞癌生长时是否经过PI3K-AKT途径。结果 LAPTM4B在肾癌组织中的表达量明显高于邻近正常肾组织（P<0.001）。而且较高的表达量也与肿瘤的临床分期、Fuhrman分级以及淋巴结转移密切相关（P<0.05）。敲低LAPTM4B之后，786-O细胞的生长活力显著减弱，G1相延迟增多，p-PI3K以及p-AKT蛋白水平同时减少，在加入740Y-P之后，这种抑制作用得到了一定的缓解，再次证明了LAPTM4B是通过活化PI3K/AKT信号网络来促进肾脏癌细胞的恶性增殖。结论 LAPTM4B在肾细胞癌高度过量表达，通过活化PI3K/AKT信号通路使得癌细胞发生恶性增殖，具有作为肾细胞癌治疗相关药物靶标的前景。

Abstract

Objective To investigate the expression of LAPTM4B in renal cell carcinoma (RCC) and the mechanism by which it activates the PI3K/AKT signaling pathway to promote tumor malignancy and proliferation.
Methods Sixty-four pairs of tumor tissue and adjacent normal tissue samples from RCC patients who underwent radical nephrectomy in our hospital from June 2019 to June 2023 were collected. The expression level of LAPTM4B was detected by immunohistochemical staining, Western blot, and fluorescence quantitative PCR. A 786-O cell line with continuous down-regulation of LAPTM4B was established. The cell proliferation and its cycle changes were analyzed by CCK-8 assay, colony formation assay, and flow cytometry. The phosphorylation status of the main proteins of the PI3K/AKT signaling pathway was detected by Western blot. Rescue experiments with the PI3K activator 740Y-P were conducted to explore whether LAPTM4B affects the growth of renal cell carcinoma through the PI3K-AKT pathway.
Results The expression level of LAPTM4B in renal cancer tissues was significantly higher than that in adjacent normal renal tissues (P < 0.001). Moreover, the higher expression level was also closely related to the clinical stage, Fuhrman grade, and lymph node metastasis of the tumor (P < 0.05). After knockdown of LAPTM4B, the growth vitality of 786-O cells was significantly weakened, the G1 phase was delayed, and the levels of p-PI3K and p-AKT proteins were simultaneously reduced. After adding 740Y-P, this inhibitory effect was alleviated, further confirming that LAPTM4B promotes the malignant proliferation of renal cell carcinoma cells by activating the PI3K/AKT signaling network.
Conclusion   LAPTM4B is highly overexpressed in renal cell carcinoma, and its activation of the PI3K/AKT signaling pathway leads to malignant proliferation of cancer cells, presenting a prospect as a target drug for renal cell carcinoma treatment.