Hipermetylacja promotora genu CDH1 w DNA komórek jako biomarker progresji raka jamy ustnej
Authors:   Klinika Chirurgii Szczękowo-Twarzowej Uniwersytecki Szpital Kliniczny im. F. Chopina w Rzeszowie, Brodowski Robert 
Published: June 2023

Malignant neoplasms of the head and neck region are the sixth most common tumors in the world wide population, with 650 000 new cases diagnosed every year and 350 000 deaths. In this group of cancers, the most common is oral squamous cell carcinoma (OSCC), which is recognized in 90% of malignant tumors in the mouth and 38% of all head and neck cancers. Due to the large number of newly diagnosed cases, especially in relation with high mortality and low 5-year survival rate, oral cancer is a serious medical problem with a significant impact on various aspects of public health as well as having a social and economic impact. The latest epidemiological data were presented by the National Cancer Registry concerning 2019 year. There were 2,168 new cases of oral cancer, including 1,726 cases in men and 442 women and 1,428 deaths caused by oral cancer, in this group there were 1,165 men and 263 women. According to this register, in the decade 2010–2020, in the Podkarpackie Province, a total of 1,044 patients suffered from malignant oral tumors, including 819 men and 225 women. During this period, a total of 753 patients died of oral cancer, in this group there were 636 men and 117 women. Despite significant advances in preventive activities and a large development of oral cancer therapeutic tools and techniques, the percentage of 5-year survival rates of OSCC patients has remained unchanged for over 20 years. Bad results and prognosis for the patients are closely related to the late diagnosis of cancer and the initiation of the therapeutic process, often in advanced stage of malignant disease, tumor infiltration of surrounding tissues, metastases to regional lymph nodes and rarely distant metastases to other organs. Long-term results of oral cancer treatment are still unsatisfactory and epidemiological studies have not improved for many years. Due to these reasons, many researchers aimed at finding new biomarkers, allowing precise monitoring of the cancer process and starting treatment when a patient has not clinical symptoms. One of the research methods aimed at developing a tumor biomarker is epigenetics. The process of carcinogenesis occurs as a result of simultaneous genetic and epigenetic changes. The mechanisms of epigenetic regulation of gene expression (from the Greek epi – above the genome), including hereditary but potentially reversible gene expression changes which are encoded by genome modifications and chromatin components and are important in the process of tumorigenesis but without changes in the DNA sequence. One of the epigenetic mechanisms of changes in the normal DNA chain is the methylation process. Epigenetic changes in the DNA methylation pattern have been observed in many types of tumors and are considered to be essential in carcinogenesis. Epigenetic changes observed in the early stages of tumor development are considered as abnormalities necessary for tissue neoplasia and progression of cancer development. Different types of tumors express characteristic methylation patterns, but they also have common features of epigenetic changes that are used in early detection of tumors, they allow tracking their progression and have some significance in assessing prognosis. In the world there is rapid technological progress in detection and monitoring of epigenetic changes in human tissues, which is used in clinical practice as early biomarkers of cancer changes and markers of cancer progression. The methylation process consists in attaching a methyl group to the fifth carbon of the cytosine ring, however only 3-4% of all cytosines in the DNA undergo this process. Changes in methylation of DNA are recognized as the first epigenetic biomarker associated with the development of cancer, changing the proper functioning of the gene and they include hypermethylation, hypomethylation and blocking imprinting. Epigenetic methylation disorders develop gradually in the ongoing process of carcinogenesis. Hypermethylation of CpG islands appears early in this process and the number of methylated CpG is successively increasing with the development of cancer and the progression of cancer malignancy. The process of the methylation of the new CpG islands begins and spreads, takes up more and more areas of the genome. This is probably due to the aging of the genome and the result of mediators of inflammation and oxidative stress in the cell. The CDH1 gene is located on the long arm of chromosome 16, in the 16q22.1 locus and encodes the E-cadherin protein (epithelial cadherin). E-cadherin is a 120 kD transmembrane homodimer composed of 882 amino acid residues. Scientific studies evaluating the expression of E-cadherin in various types of cancer, have demonstrated the key role of this protein during cancer progression and invasion. A low level of activity or lack of E-cadherin is closely related to cell changes and motility, resulting in more cancer metastases to the tissues. The low expression or lack of E-cadherin protein results of blocking the CDH1 suppressor gene encoding this protein. The most important mechanism of weakening the expression or completely blocking the formation of E-cadherin is the hypermethylation of the promoter of the CDH1 gene. Reduced expression of E-cadherin is the main cause of dysfunction of intercellular adhesion, resulting in a partial or complete reduction of progeny cell adherence in tumor tissue and consequently an increased ability to infiltrate surrounding tissues as well as early ability to create metastases. The aim of the study was to evaluate the effect of epigenetic disorders in the hypermethylation mechanism of the CDH1 gene promoter in the DNA of tumor cells and in normal cells, on the clinical course, dynamics and prognosis of oral cancer. This was retrospective study. In the group of patients treated for oral cancer in the Clinic of Maxillofacial Surgery University Clinical Hospital in Rzeszów, Oncology Clinic of the Podkarpackie Center of Oncology and histological preparations obtained from the Department of Clinical Pathology of University Clinical Hospital in Rzeszów there were 58 men and 21 women. The Bioethical Commission of the University of Rzeszów, resolution No. 14/01/2019, agreed to conduct a methylation study of the CDH1 suppressor gene as part of the research project: “Molecular disorders in various types of cancer and autoimmune diseases”. In this study I looked at the impact of hypermethylation of the CDH1 suppressor gene in the DNA of oral cancer cells and normal cells from the surgical margin after cancer excision, on the clinical course and the treatment results of patients operated on oral cancer. The main studies concerned the correlation between CDH1 hipermethylation and the time of onset of tumor recurrence, the time of metastases to regional lymph nodes, distant metastases and time of patients’ survival. The research was conducted in the context of searching for an answer to the question whether hypermethylation of the CDH1 gene could be a biomarker used to patients monitoring after oral cancer surgery. Analysis of the results obtained in the study allowed to draw conclusions that this knowledge can be used in the treatment and follow-up of oral cancer. A statistically significant correlation was found between hypermethylation of the CDH1 suppressor gene in the DNA of oral cancer cells and the incidence of metastases to the neck lymph nodes after the primary cancer surgery. This knowledge should be used for a more frequent examination of patients after oral cancer surgery, in whom hypermethylation was found and in case of the metastases to the regional lymph nodes are found, appropriate treatment can be started as soon as possible. Similar observations were made regarding the time of metastases to the neck lymph nodes, which was 4 months shorter in the group of patients with confirmed hypermethylation of the CDH1 gene. In addition, the studies showed a positive and statistically significant correlation between DNA hypermethylation in the CDH1 gene and the group of patients abusing alcohol, as well as for smokers who smoked tobacco for many years. In my studies, there was no statistical correlation between hypermethylation of the CDH1 gene in the DNA of oral cancer cells and normal cells of operative margin, and the frequency of cancer recurrence in the oral cavity, time of local recurrence, the frequency of distant metastases, time of occurrence of distant metastases and metastases to regional lymph nodes. This research has expanded our knowledge on the molecular basis of oncogenesis of oral squamous cell carcinoma, combining the expression of the CDH1 suppressor gene with determination of the methylation of the promoter regions of the tested gene and their effect on the clinical course of cancer and treatment results. Continuing epigenetic research may in the future contribute to the development of a clinically useful biomarker panel, support the diagnosis of oral cancer and become the foundation for the development of new therapeutic methods and oncological drugs based on epigenetic mechanisms.