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Kildusiene 1,2, *, Ryte Rynkeviciene 3, Auguste Kaceniene 2, Rima Miknaite 1,3, Kestutis Suziedelis 1,3 and Giedre Smailyte 2,4  \nAcademic Editors: C˘alin C˘ainap and Nicolae Crisan  \nReceived: 31 May 2025  \nRevised: 5 July 2025  \nAccepted: 7 July 2025  \nPublished: 10 July 2025  \nCitation: Kildusiene, I.;  \nRynkeviciene, R.; Kaceniene, A.; Miknaite, R.; Suziedelis, K.; Smailyte, G. Analysis of LINE-1 DNA Methylation in Colorectal Cancer, Precancerous Lesions, and Adjacent Normal Mucosa. Medicina 2025, 61, 1243. [https://doi.org/10.3390/](https://doi.org/10.3390/)[ ](https://doi.org/10.3390/)medicina61071243  \nCopyright: © 2025 by the authors. Published by MDPI on behalf of the Lithuanian University of Health Sciences. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ([https://creativecommons.org/](https://creativecommons.org/)[ ](https://creativecommons.org/)[licenses/by/4.0/](licenses/by/4.0/)) .  \n1 Institute of Biosciences, Life Sciences Center, Vilnius University, LT-03101 Vilnius, Lithuania; [kestutis.suziedelis@gf.vu.lt](kestutis.suziedelis@gf.vu.lt) (K.S.)  \n2 Laboratory of Cancer Epidemiology, National Cancer Institute, P. Baublio 3B, LT-08406 Vilnius, Lithuania; [auguste.kaceniene@nvi.lt](auguste.kaceniene@nvi.lt) (A.K.); [giedre.smailyte@nvi.lt](giedre.smailyte@nvi.lt) (G.S.)  \n3 Laboratory of Molecular Oncology, National Cancer Institute, Santariskiu 1, LT-08406 Vilnius, Lithuania; [ryte.rynkeviciene@nvi.lt](ryte.rynkeviciene@nvi.lt)  \n4 Institute of Health Sciences, Faculty of Medicine, Vilnius University, LT-03101 Vilnius, Lithuania  \n* Correspondence: [inga.kildusiene@nvc.santa.lt](inga.kildusiene@nvc.santa.lt)  \nAbstract  \nBackground and Objectives: Colorectal cancer (CRC) is a major cause of cancer morbidity and mortality worldwide. Genetic and epigenetic changes, especially DNA methylation alterations, are key in CRC development. LINE-1 hypomethylation marks global DNA methylation loss and genomic instability, making it a potential early CRC biomarker. This study investigates the methylation status of LINE-1 in colorectal adenocarcinoma, precancerous lesions (tubular and serrated adenomas), and the surrounding normal mucosa, aiming to elucidate its role as an epigenetic marker in early colorectal tumorigenesis. Materials and Methods: Paired lesion and normal tissue samples from 66 patients were analyzed for LINE-1 methylation at three CpG sites using bisulfite pyrosequencing. Results: Adenocarcinomas and tubular adenomas showed significant hypomethylation, especially at loci A and B, while serrated adenomas exhibited no significant differences. Conclusions: LINE-1 hypomethylation is associated with colorectal tumorigenesis, with distinct patterns observed between tubular and serrated adenomas, indicating distinct pathways forming and progressing specific adenomas. These findings support the potential of LINE-1 methylation as an early epigenetic biomarker for CRC risk stratification and highlight the need for further research into its clinical utility.  \nKeywords: LINE-1; colorectal cancer; methylation  \n1. Introduction  \nColorectal cancer (CRC) is the third most common cancer worldwide, with over 1.15 million new cases and 576,000 deaths annually [1] . CRC develops through a multistep process driven by genetic and epigenetic alterations, primarily via three pathways: chromosomal instability (CIN, ~85%), microsatellite instability (MSI, ~15%), and CpG island methylator phenotype (CIMP, ~25%) [2,3] .  \nEpigenetic mechanisms—DNA methylation, histone modifications, and non-coding RNAs—regulate gene expression without altering the DNA sequence [4–7] . DNA methylation, especially at CpG islands in gene promoters, is the most studied in CRC and is linked to gene silencing, carci","cbCaiozBe920V7z5","https://ap.wps.com/l/cbCaiozBe920V7z5","pdf",749545,"English","# Abstract\n# 1. Introduction\n# 2. Materials and Methods\n## 2.1. Study Population\n## 2.2. Tissue Collection and Classification\n## 2.3. LINE-1 Methylation Analysis","[{\"question\":\"What is the main biomarker examined in this study?\",\"answer\":\"The study focuses on LINE-1 DNA methylation, assessed as an indicator of global DNA methylation status and genomic instability relevant to CRC development.\"},{\"question\":\"Which patient and tissue groups were analyzed?\",\"answer\":\"Paired lesion and adjacent normal tissue samples were analyzed from patients with colorectal adenocarcinoma, tubular adenomas, and serrated adenomas.\"},{\"question\":\"How was LINE-1 methylation measured and compared?\",\"answer\":\"LINE-1 methylation was measured at three CpG sites using bisulfite pyrosequencing, then compared across lesion types and their paired normal mucosa.\"}]","Analysis of LINE-1 DNA Methylation in Colorectal Cancer, Precancerous Lesions, and Adjacent Normal Mucosa | PDF",1790172173,18]