Background:Colorectal cancer is the third most diagnosed cancer and a leading cause of cancer-related deaths worldwide. Early detection significantly improves patient outcomes, yet many cases are identified only at late stages. The high molecular and genetic heterogeneity of colorectal cancer presents major challenges in accurate diagnosis, prognosis, and therapeutic stratification. Recent advances in gene expression profiling offer new opportunities to discover genes that play a role in colorectal cancer carcinogenesis and may contribute to early diagnosis, prognosis prediction, and the identification of novel therapeutic targets.Methods:This study involved 142 samples: 84 primary tumor samples, 27 liver metastases, and 31 adjacent non-tumor tissues serving as controls. RNA sequencing was performed on a subset of tissues (12 liver metastases and 3 adjacent non-tumor tissues) using a targeted RNA panel covering 395 cancer-related genes. Data processing and differential gene expression analysis were carried out using the DRAGEN RNA and DRAGEN Differential Expression tools. The expression of six genes involved in hypoxia and epithelial-to-mesenchymal transition (EMT) pathways (SLC16A3,ANXA2,P4HA1,SPP1,KRT19, andLGALS3) identified as significantly differentially expressed was validated across the whole cohort via quantitative real-time PCR. The relative expression levels were determined using the ΔΔct method and log2FC, and compared between different groups based on the sample type; clinical parameters; and mutational status of the genesKRAS,PIK3CA,APC,SMAD4, andTP53.Results:Our results suggest that the expression of all the validated genes is significantly altered in metastases compared to non-tumor control samples (p< 0.05). The most pronounced change occurred for the genesP4HA1andSPP1, whose expression was significantly increased in metastases compared to non-tumor and primary tumor samples, as well as between clinical stages of CRC (p< 0.001). Furthermore, all genes, except forLGALS3, exhibited significantly altered expression between non-tumor samples and samples in stage I of the disease, suggesting that they play a role in the early stages of carcinogenesis (p< 0.05). Additionally, the results suggest the mutational status of theKRASgene did not significantly affect the expression of any of the validated genes, indicating that these genes are not involved in the carcinogenesis ofKRAS-mutated CRC.Conclusions:Based on our results, the genesP4HA1andSPP1appear to play a role in the progression and metastasis of colorectal cancer and are candidate genes for further investigation as potential biomarkers in CRC.
背景：结直肠癌是全球第三大常见癌症，也是癌症相关死亡的主要原因。早期检测能显著改善患者预后，但许多病例仅在晚期才被发现。结直肠癌的高度分子和遗传异质性给准确诊断、预后判断和治疗分层带来了重大挑战。基因表达谱分析的最新进展为发现参与结直肠癌发生发展的基因提供了新机遇，这些基因可能有助于早期诊断、预后预测及新型治疗靶点的识别。 方法：本研究纳入142例样本，包括84例原发肿瘤样本、27例肝转移样本及31例作为对照的癌旁非肿瘤组织。使用覆盖395个癌症相关基因的靶向RNA panel对部分组织（12例肝转移和3例癌旁非肿瘤组织）进行RNA测序。数据处理和差异基因表达分析采用DRAGEN RNA和DRAGEN差异表达工具完成。通过定量实时PCR，在全部队列中验证了被确定为显著差异表达的六个涉及缺氧和上皮-间质转化（EMT）通路的基因（SLC16A3、ANXA2、P4HA1、SPP1、KRT19和LGALS3）。采用ΔΔct法和log2FC计算相对表达水平，并根据样本类型、临床参数以及KRAS、PIK3CA、APC、SMAD4和TP53基因的突变状态在不同组间进行比较。 结果：我们的结果表明，与非肿瘤对照样本相比，所有验证基因在转移灶中的表达均发生显著改变（p < 0.05）。变化最显著的是P4HA1和SPP1基因，与非肿瘤样本和原发肿瘤样本相比，其在转移灶中的表达显著增加，且在结直肠癌不同临床分期之间也存在显著差异（p < 0.001）。此外，除LGALS3外，所有基因在非肿瘤样本与疾病I期样本间的表达均呈现显著差异，提示这些基因在癌变早期阶段发挥作用（p < 0.05）。另外，结果表明KRAS基因的突变状态对任何验证基因的表达均无显著影响，说明这些基因不参与KRAS突变型结直肠癌的癌变过程。 结论：基于我们的研究结果，P4HA1和SPP1基因似乎在结直肠癌的进展和转移中发挥作用，是作为结直肠癌潜在生物标志物进一步研究的候选基因。
SPP1as a Potential Stage-Specific Marker of Colorectal Cancer
肿瘤(癌症)患者之家