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Background: CPUK02 (15-Oxosteviol benzyl ester) is a new ent-kaurenoid derivative of stevioside and exhibits strong anti-cancer activity. Nowadays, the pattern of epigenetic in cancer has been topic of many studies and DNA methylation targeting represents a relevant strategy for cancer treatment. Since, no study conducted to this mechanism, we attempt to evaluate whether CPUK02 induce its anti-cancer effects via alteration the level of mRNA DNMT3B, DNMT3A expression and ESR1 methylation pattern in breast cancer cells line. Methods: MCF-7 (ER +) and MDA-MB231 (ER-) cell lines were treated for 24, 48 hours with 1 μM CPUK02 and 5-AZA-CdR (DNA methyltransferase inhibitor). Quantitative expression of DNMT3B and DNMT3A genes and ESR1 promoter methylation was assessed by Real-Time PCR and MS-PCR, respectively. Results: CPUK02 restored ESR1 promoter unmethylated allele in MDA-MB 231 cells. Also treatment with CPUK02 decreased the expression of both DNMT3A and DNMT3B genes like 5-AZA. The expression of DNMT genes were diminished by half compared with control cells. Conclusions: These results showed that CPUK02 has an anticancer effect on MDA-MB 231 cells which this effect can be done through several pathways., (Creative Commons Attribution License)

Introduction: Colorectal cancer (CRC) is a leading cause of cancer deaths worldwide but current molecular targeted therapy is not providing major success in CRC treatment, so early detection by non-invasive methods continues to be vital. Aberrant methylation of CpG islands in promoter regions is associated with inactivation of various tumor suppressor genes. O6-methyguanine-DNA methyltransferase (MGMT) is a DNA repair enzyme that removes mutagenic and cytotoxic adducts from O6-guanine in DNA. Aberrant hypermethylation of the MGMT promoter has been associated with lack of mRNA expression, with concomitant loss of protein content and enzyme activity. AIM: Our aim was to determine whether MGMT promoter methylation might be detectable in circulating free DNA in the serum of CRC patients and normal individuals using a methylation specific (MSP) polymerase chain reaction (PCR) method. Methods: A total of 70 subjects were enrolled in the study. Of these, 30 patients who were diagnosed previously as untreated colon adenocarcinoma by a gastroenterologist and the other 40 were nearly age-matched individuals who had a normal colonoscopic evaluation (except for hemorrhoids or fissures) and normal pathologic reports. After bisulphite modification of DNA, serum samples were examined for MGMT promoter methylation using MSP. Results: Ninety percent of CRC patients had MGMT promoter hypermethylation as compared to no methylation in normal subjects’ serum. Most of the cancers were stage П and moderately differentiated adenocarcinomas; nearly 60% were found in the left colon. No statistically significant correlation was found between the promoter methylation status and gender and age. Discussion and Conclusions: MGMT hypermethylation can be detected in free circulating DNA in serum of CRC patients and can be used “as a clinical biomarker” for early diagnosis and prognostic assessment of the disease. Our data confirm previous studies indicating utility for free circulating DNA as a serum biomarker for early detection, diagnosis and monitoring of CRC patients., (Creative Commons Attribution License)

Background: The pathogenesis of sporadic colorectal cancer (CRC) is influenced by the patient genetic background and environmental factors. Based on prior understanding, these are classified in two major pathways of genetic instability. Microsatellite instability (MSI) and CPG island methylator phenotype (CIMP) are categorized as features of the hypermethylated prototype, and chromosomal instability (CIN) is known to be indicative of the non-hypermethylated category. Secreted frizzled related protein 2 (SFRP2), APC1A in WNT signaling pathway and the DNA repair gene, O6-methylguanine-DNA methyltransferase (MGMT), are frequently hypermethylated in colorectal cancer. Detection of methylated DNA as a biomarker by easy and inexpensive methods might improve the quality of life of patients with CRC via early detection of cancer or a precancerous condition., Aim: To evaluate the rate of SFRP2 and MGMT hypermethylation in both polyp tissue and serum of patients in south Iran as compared with matched control normal population corresponding samples., Materials and Methods: Methylation-specific PCR was used to detect hypermethylation in DNA extracted from 48 polypoid tissue samples and 25 healthy individuals., Results: Of total polyp samples, 89.5% had at least one promoter gene hypermethylation. The most frequent methylated locus was SFRP2 followed by MGMT-B (81.2 and 66.6 percent respectively). Serologic detection of hypermethylation was 95% sensitive as compared with polyp tissue. No hypermethylation was detected in normal tissue and serum and its detection in patients with polyps, especially of serrated type, was specific., Conclusions: Serologic investigation for detection of MGMT-B, SFRP2 hypermethylation could facilitate prioritization of high risk patients for colonoscopic polyp detection and excision.

Background: The ubiquitin-proteasome system (UPS) degrades a variety of proteins which attach to specific signals. The ubiquitination pathway facilitates degradation of damaged proteins and regulates growth and stress responses. This pathway is altered in various cancers, including acute lymphoblastic leukemia, head and neck squamous cell carcinoma and breast cancer. Recently it has been reported that expression of newly characterized human genes, UBE2Q1 and UBE2Q2, putative members of ubiquitin-conjugating enzyme family (E2), has been also changed in colorectal cancer. Epigenetics is one of the fastest-growing areas of science and nowadays has become a central issue in biological studies of diseases. According to the lack of information about the role of epigenetic changes on gene expression profiling of UBE2Q1 and UBE2Q2, and the presence of CpG islands in the promoter of these two human genes, we decided to evaluate the promoter methylation status of these genes as a first step., Materials and Methods: The promoter methylation status of UBE2Q1 and UBE2Q2 was studied by methylation-specific PCR (MSP) in tumor samples of 60 colorectal cancer patients compared to adjacent normal tissues and 20 non-malignant controls. The frequency of the methylation for each gene was analyzed by chi-square method., Results: MSP results revealed that UBE2Q2 gene promoter were more unmethylated, while a higher level of methylated allele was observed for UBE2Q1 in tumor tissues compared to the adjacent normal tissues and the non malignant controls., Conclusions: UBE2Q1 and UBE2Q2 genes show different methylation profiles in CRC cases.

Inflammatory bowel disease (IBD) is a disease strongly associated with colorectal cancer (CRC) as a well-known precancerous condition. Alterations in DNA methylation and mutation in K-ras are believed to play an early etiopathogenic role in CRC and may also an initiating event through deregulation of molecular signaling. Epigenetic silencing of APC and SFRP2 in the WNT signaling pathway may also be involved in IBD-CRC. The role of aberrant DNA methylation in precancerous state of colorectal cancer (CRC) is under intensive investigation worldwide. The aim of this study was to investigate the status of promoter methylation of MGMT-B, APC1A and SFRP2 genes, in inflamed and normal colon tissues of patients with IBD compared with control normal tissues. A total of 52 IBD tissues as well as corresponding normal tissues and 30 samples from healthy participants were obtained. We determined promoter methylation status of MGMT-B, SFRP2 and APC1A genes by chemical treatment with sodium bisulfite and subsequent MSP. The most frequently methylated locus was MGMT-B (71%; 34 of 48), followed by SFRP2 (66.6 %; 32 of 48), and APC1A (43.7%; 21 of 48). Our study demonstrated for the first time that hypermethylation of the MGMT-B and the SFRP2 gene promoter regions might be involved in IBD development. Methylation of MGMT-B and SFRP2 in IBD patients may provide a method for early detection of IBD-associated neoplasia.