biostudies-literatureUnknown13Lukacova E<h4>Introduction</h4>Colorectal cancer (CRC) can develop through several dysregulated molecular pathways, including the serrated pathway, characterized by CpG island methylator (CIMP) phenotype. Although the tumor tissue is a commonly tested material, sample types such as stool or plasma, bring a new, non-invasive approach. Several cancer-related methylated genes have been identified in CRC patients, including gene <i>GRIA4</i>, showing promising diagnostic potential. The aim of our study was to develop a sensitive droplet digital PCR (ddPCR) assay to examine <i>GRIA4</i> hypermethylation status in CRC patients and evaluate its diagnostic potential in tissue and liquid biopsy samples.<h4>Methods</h4>In total, 23 patients participated in this study, 7 patients with primary CRC and 16 patients with liver metastasis of clinically known CRC. We obtained tumor and non-tumor tissues (N=17), blood samples pre- and post-surgery (N=22), and blood of five volunteers without a personal cancer history. We have developed and optimized a ddPCR assay for <i>GRIA4</i> hypermethylation detection, from tissue and plasma samples.<h4>Results</h4>We detected significantly increased <i>GRIA4</i> methylation in tumor tissues compared to their adjacent non-tumor tissue, p<0.0001. Receiver operating characteristic (ROC) analysis defined cutoff values to separate primary tumors and metastases from non-tumor colon/rectum, specifically 36.85% for primary tumors and 34.81% for metastases. All primary tumors were above this threshold. When comparing the methylation levels of metastatic vs. non-tumor tissue, a smaller increase was observed in liver metastasis versus colon tissue (3.6× gain; p=0.001), then in liver metastasis versus adjacent liver tissue (17.4× gain; p<0.0001). On average, <i>GRIA4</i> hypermethylation in primary tumor plasma was 2.8-fold higher (p=0.39), and in metastatic plasma, 16.4-fold higher (p=0.0011) compared to healthy individuals. Hypermethylation in metastatic plasma was on average 5.9 times higher (p=0.051) than in primary tumor plasma. After tumor removal surgery, average hypermethylation decrease in plasma was 1.6× for primary (p=0.037) and 4.5× for metastatic patients (p=0.023).<h4>Discussion</h4>Based on our data, it can be inferred that <i>GRIA4</i> serves as a tissue specific biomarker for the colon/rectum tissue, thus is suitable for cancer classification. This biomarker showed the potential to be an attractive target for early non-invasive detection of metastases of clinically known CRC, although additional analysis has to be performed.Frontiers in oncology1205791https://www.ebi.ac.uk/biostudies/studies/S-EPMC10354553biostudies-literatureHypermethylated <i>GRIA4</i>, a potential biomarker for an early non-invasive detection of metastasis of clinically known colorectal cancer.PMC10354553Mersakova SPodlesniy PTuryova ELukacova ELaca LKudelova EBurjanivova TVanochova AGrendar MMiklusica JLasabova ZKasubova IMikolajcik PfalseHypermethylated <i>GRIA4</i>, a potential biomarker for an early non-invasive detection of metastasis of clinically known colorectal cancer.<h4>Introduction</h4>Colorectal cancer (CRC) can develop through several dysregulated molecular pathways, including the serrated pathway, characterized by CpG island methylator (CIMP) phenotype. Although the tumor tissue is a commonly tested material, sample types such as stool or plasma, bring a new, non-invasive approach. Several cancer-related methylated genes have been identified in CRC patients, including gene <i>GRIA4</i>, showing promising diagnostic potential. The aim of our study was to develop a sensitive droplet digital PCR (ddPCR) assay to examine <i>GRIA4</i> hypermethylation status in CRC patients and evaluate its diagnostic potential in tissue and liquid biopsy samples.<h4>Methods</h4>In total, 23 patients participated in this study, 7 patients with primary CRC and 16 patients with liver metastasis of clinically known CRC. We obtained tumor and non-tumor tissues (N=17), blood samples pre- and post-surgery (N=22), and blood of five volunteers without a personal cancer history. We have developed and optimized a ddPCR assay for <i>GRIA4</i> hypermethylation detection, from tissue and plasma samples.<h4>Results</h4>We detected significantly increased <i>GRIA4</i> methylation in tumor tissues compared to their adjacent non-tumor tissue, p<0.0001. Receiver operating characteristic (ROC) analysis defined cutoff values to separate primary tumors and metastases from non-tumor colon/rectum, specifically 36.85% for primary tumors and 34.81% for metastases. All primary tumors were above this threshold. When comparing the methylation levels of metastatic vs. non-tumor tissue, a smaller increase was observed in liver metastasis versus colon tissue (3.6× gain; p=0.001), then in liver metastasis versus adjacent liver tissue (17.4× gain; p<0.0001). On average, <i>GRIA4</i> hypermethylation in primary tumor plasma was 2.8-fold higher (p=0.39), and in metastatic plasma, 16.4-fold higher (p=0.0011) compared to healthy individuals. Hypermethylation in metastatic plasma was on average 5.9 times higher (p=0.051) than in primary tumor plasma. After tumor removal surgery, average hypermethylation decrease in plasma was 1.6× for primary (p=0.037) and 4.5× for metastatic patients (p=0.023).<h4>Discussion</h4>Based on our data, it can be inferred that <i>GRIA4</i> serves as a tissue specific biomarker for the colon/rectum tissue, thus is suitable for cancer classification. This biomarker showed the potential to be an attractive target for early non-invasive detection of metastases of clinically known CRC, although additional analysis has to be performed.2023-01-01T00:00:00Z20232026-06-02T12:02:46.256Z2026-04-18T03:11:17.637ZS-EPMC103545533747638210.3389/fonc.2023.1205791