Table_1_PLEKHA4 is a novel prognostic biomarker that reshapes the tumor microenvironment in lower-grade glioma.pdf

BackgroundLower-grade glioma (LGG) is a primary intracranial tumor that carry a high risk of malignant transformation and limited therapeutic options. Emerging evidence indicates that the tumor microenvironment (TME) is a superior predictor for tumor progression and therapy response. PLEKHA4 has been demonstrated to be a biomarker for LGG that correlate with immune infiltration. However, the fundamental mechanism by which PLEKHA4 contributes to LGG is still poorly understood.
MethodsMultiple bioinformatic tools, including Tumor Immune Estimation Resource (TIMER), Gene Expression Profiling Interactive Analysis (GEPIA2), Shiny Methylation Analysis Resource Tool (SMART), etc., were incorporated to analyze the PLEKHA4. ESTIMATE, ssGSEA, CIBERSORT, TIDE and CellMiner algorithms were employed to determine the association of PLEKHA4 with TME, immunotherapy response and drug sensitivities. Immunohistochemistry (IHC)-based tissue microarrays and M2 macrophage infiltration assay were conducted to verify their associations.
ResultsPLEKHA4 expression was found to be dramatically upregulated and strongly associated with unfavorable overall survival (OS) and disease-specific survival (DSS) in LGG patients, as well as their poor clinicopathological characteristics. Cox regression analysis identified that PLEKHA4 was an independent prognostic factor. Methylation analysis revealed that DNA methylation correlates with PLEKHA4 expression and indicates a better outcome in LGG. Moreover, PLEKHA4 was remarkably correlated with immune responses and TME remodeling, as evidenced by its positive correlation with particular immune marker subsets and the putative infiltration of immune cells. Surprisingly, the proportion of M2 macrophages in TME was strikingly higher than others, inferring that PLEKHA4 may regulate the infiltration and polarization of M2 macrophages. Evidence provided by IHC-based tissue microarrays and M2 macrophage infiltration assay further validated our findings. Moreover, PLEKHA4 expression was found to be significantly correlated with chemokines, interleukins, and their receptors, further supporting the critical role of PLEKHA4 in reshaping the TME. Additionally, we found that PLEKHA4 expression was closely associated with drug sensitivities and immunotherapy responses, indicating that PLEKHA4 expression also had potential clinical significance in guiding immunotherapy and chemotherapy in LGG.
ConclusionPLEKHA4 plays a pivotal role in reshaping the TME of LGG patients, and may serve as a potential predictor for LGG prognosis and therapy.

背景：低级别胶质瘤（Lower-grade glioma, LGG）是一类原发性颅内肿瘤，具有较高的恶变风险且治疗手段有限。越来越多的研究证据表明，肿瘤微环境（tumor microenvironment, TME）是预测肿瘤进展与治疗响应的更优指标。已有研究证实PLEKHA4可作为与免疫浸润相关的LGG生物标志物，但PLEKHA4促进LGG发生发展的核心机制仍不甚明确。
方法：本研究纳入多种生物信息学工具，包括肿瘤免疫估算资源（Tumor Immune Estimation Resource, TIMER）、基因表达谱交互分析（Gene Expression Profiling Interactive Analysis, GEPIA2）、Shiny甲基化分析资源工具（Shiny Methylation Analysis Resource Tool, SMART）等，用于分析PLEKHA4的相关特征。采用ESTIMATE、ssGSEA、CIBERSORT、TIDE及CellMiner算法，探究PLEKHA4与肿瘤微环境、免疫治疗响应及药物敏感性的关联。通过基于免疫组化（immunohistochemistry, IHC）的组织芯片实验与M2巨噬细胞浸润实验，验证上述关联。
结果：研究发现，LGG患者体内PLEKHA4的表达显著上调，且与不良总生存期（overall survival, OS）、疾病特异性生存期（disease-specific survival, DSS）及较差的临床病理特征密切相关。Cox回归分析显示，PLEKHA4是一项独立的预后因素。甲基化分析结果表明，DNA甲基化水平与PLEKHA4的表达存在关联，且可提示LGG患者的良好预后。此外，PLEKHA4与免疫应答及肿瘤微环境重塑显著相关，具体表现为其与特定免疫标记子集及推定的免疫细胞浸润水平呈正相关。值得注意的是，肿瘤微环境中M2巨噬细胞的占比显著高于其他免疫细胞类型，提示PLEKHA4可能调控M2巨噬细胞的浸润与极化。基于免疫组化的组织芯片实验与M2巨噬细胞浸润实验所得结果进一步验证了本研究的发现。同时，PLEKHA4的表达与趋化因子、白细胞介素及其受体的表达显著相关，进一步佐证了其在重塑肿瘤微环境中的关键作用。此外，本研究还发现PLEKHA4的表达与药物敏感性及免疫治疗响应密切相关，表明PLEKHA4在指导LGG患者的免疫治疗与化疗方案选择中具有潜在临床应用价值。
结论：PLEKHA4在重塑LGG患者的肿瘤微环境中发挥关键作用，有望成为预测LGG患者预后与治疗方案的潜在生物标志物。