PIK3CA mutation and PTEN suppression lead to tumorigenesis and drug resistance in colorectal cancer (CRC). There is no research on the role of circular RNAs (circRNAs) in regulating PIK3CA mutation and MEK inhibitor resistance in CRC. The expression of circLHFPL2 in PIK3CA-mutant and wild-type cells and tissues was quantified by RNA-sequencing and qRT-PCR. CCK-8 assay and colony formation assay were used to evaluate cell viability. Annexin V/PI staining was implemented to assess cell apoptosis. Luciferase assay, biotin-coupled microRNA capture, and RIP assay were used to validate the interaction among potential targets. Western blotting and qRT-PCR assays were used to evaluate the expression of involved targets. Xenograft tumor in a nude mouse model was used to explore the role of circRNAs in vivo. RNA sequencing defined downregulated expression of circLHFPL2 in both PIK3CAH1047R (HCT116) and PIK3CAE545K (DLD1) cells. CircLHFPL2 was also downregulated in PIK3CA-mutant CRC primary cells and tissues, which was correlated with poor prognosis. CircLHFPL2 was mainly localized in the cytoplasm and its downregulation was attributed to the PI3K/AKT signaling pathway activated by phosphorylating Foxo3a. CircLHFPL2 inhibited PI3KCA-Mut CRC progression both in vitro and in vivo. Furthermore, our work indicated that circLHFPL2 acts as a ceRNA to sponge miR-556-5p and miR-1322 in CRC cells and in turn modulate the expression of PTEN. Importantly, circLHFPL2 was able to overcome PIK3CA-mediated MEK inhibitor resistance in CRC cells. Downregulation of circLHFPL2 sustains the activation of the PI3K/AKT signaling pathway via a positive feedback loop in PIK3CA-mutant CRC. In addition, downregulation of circLHFPL2 leads to MEK inhibitor resistance in CRC. Therefore, targeting circLHFPL2 could be an effective approach for the treatment of CRC patients harboring oncogenic PIK3CA mutations.

中文翻译：

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PIK3CA突变介导的circLHFPL2下调通过上调PTEN抑制结直肠癌进展

PIK3CA 突变和 PTEN 抑制导致结直肠癌 (CRC) 的肿瘤发生和耐药性。目前尚无关于环状 RNA（circRNAs）在调节 CRC 中 PIK3CA 突变和 MEK 抑制剂耐药中的作用的研究。circLHFPL2 在 PIK3CA 突变体和野生型细胞和组织中的表达通过 RNA 测序和 qRT-PCR 进行量化。CCK-8测定和集落形成测定用于评估细胞活力。实施膜联蛋白 V/PI 染色以评估细胞凋亡。荧光素酶测定、生物素偶联 microRNA 捕获和 RIP 测定用于验证潜在靶标之间的相互作用。Western印迹和qRT-PCR测定用于评估相关靶标的表达。使用裸鼠模型中的异种移植肿瘤来探索 circRNA 在体内的作用。RNA 测序确定了 circLHFPL2 在 PIK3CAH1047R (HCT116) 和 PIK3CAE545K (DLD1) 细胞中的下调表达。CircLHFPL2 在 PIK3CA 突变的 CRC 原代细胞和组织中也下调，这与不良预后相关。CircLHFPL2 主要定位于细胞质，其下调归因于磷酸化 Foxo3a 激活的 PI3K/AKT 信号通路。CircLHFPL2 在体外和体内均抑制 PI3KCA-Mut CRC 进展。此外，我们的工作表明，circLHFPL2 作为 ceRNA 在 CRC 细胞中海绵 miR-556-5p 和 miR-1322，进而调节 PTEN 的表达。重要的是，circLHFPL2 能够克服 CRC 细胞中 PIK3CA 介导的 MEK 抑制剂耐药性。circLHFPL2 的下调通过 PIK3CA 突变 CRC 中的正反馈回路维持 PI3K/AKT 信号通路的激活。此外，circLHFPL2 的下调导致 CRC 中的 MEK 抑制剂耐药性。因此，靶向circLHFPL2可能是治疗携带致癌PIK3CA突变的CRC患者的有效方法。