Coexistence of K-ras mutations and HPV infection in colon cancer

BACKGROUND: Activation of the ras genes or association with human papillomavirus infection have been extensively studied in colorectal cancer. However, the correlation between K-ras mutations and HPV in colorectal cancer has not been investigated yet. In this study we aimed to investigate the presence of K-ras mutations and their correlation with HPV infection in colon cancer. METHODS: K-ras mutations were analyzed by a mutagenic PCR assay and digestion with specific restriction enzymes to distinguish the wild-type and mutant codons. HPV infection was analyzed by PCR amplification and hybridization with specific probes by Southern blotting. Stattistical analyses were performed by the chi-square and Fisher's exact tests RESULTS: HPV gene fragments were detected in 43 tumors and 17 normal tissue samples. HPV 18 was the prevalent type in the tumor tissue. A mutation at codon 12 of the K-ras gene was present in 31 patients. 56% of the HPV-positive tumors also harbored a K-ras mutation. Codon 13 mutations were not observed. These data indicate that infection with high risk HPV types and mutational activation of the K-ras gene are frequent events in colorectal carcinogenesis. CONCLUSION: Our findings suggest that mutational activation of the K-ras gene is a common event in colon carcinogenesis and that HPV infection may represent an important factor in the development of the premalignant lesions leading to the neoplastic phenotype

Promoter DNA Methylation of Oncostatin M receptor-β as a Novel Diagnostic and Therapeutic Marker in Colon Cancer

In addition to genetic changes, the occurrence of epigenetic alterations is associated with accumulation of both genetic and epigenetic events that promote the development and progression of human cancer. Previously, we reported a set of candidate genes that comprise part of the emerging “cancer methylome”. In the present study, we first tested 23 candidate genes for promoter methylation in a small number of primary colon tumor tissues and controls. Based on these results, we then examined the methylation frequency of Oncostatin M receptor-β (OSMR) in a larger number of tissue and stool DNA samples collected from colon cancer patients and controls. We found that OSMR was frequently methylated in primary colon cancer tissues (80%, 80/100), but not in normal tissues (4%, 4/100). Methylation of OSMR was also detected in stool DNA from colorectal cancer patients (38%, 26/69) (cut-off in TaqMan-MSP, 4). Detection of other methylated markers in stool DNA improved sensitivity with little effect on specificity. Promoter methylation mediated silencing of OSMR in cell lines, and CRC cells with low OSMR expression were resistant to growth inhibition by Oncostatin M. Our data provide a biologic rationale for silencing of OSMR in colon cancer progression and highlight a new therapeutic target in this disease. Moreover, detection and quantification of OSMR promoter methylation in fecal DNA is a highly specific diagnostic biomarker for CRC

Detection of RB1 Gene Copy Number Variations Using a Multiplex Ligation-Dependent Probe Amplification Method

Multiplex ligation-dependent probe amplification (MLPA) is based on simultaneous multiplex PCR of specific probes that hybridize to multiple different target DNA regions. The method can identify copy number changes, gross gene rearrangements, methylation patterns or even point mutations. MLPA has been a reliable approach to identify copy number changes in the clinical and research settings and is widely used for the screening and investigation of copy number variations and genomic aberrations of interest in various diseases. In this chapter the analysis of the copy number changes in the RB1 gene locus by MLPA is described

Role of the IncRNAs in malignant melanoma and their involvement in metastasis

Malignant melanoma is an aggressive disease and its incidence is still rising. Despite available targeted therapies the prognosis of patients with advanced disease is relatively poor. Therefore, detailed understanding of the mechanisms that lead to melanoma development and characterization of the underlying molecular events associated with the outcome is essential for a more effective therapy. The molecular and cellular biology of melanomas involves a complex network of multiple factors interacting with different signaling pathways and disrupting the gene regulatory mechanisms. Recently, the long non-coding RNAs (IncRNAs) were identified as new transcriptional regulators modulating gene expression at various levels and playing an important role in diverse biological processes including carcinogenesis, tumor development and progression. Several IncRNAs have been shown to provide potential prognostic markers and represent novel therapeutic targets in different cancers. Aberrant expression of IncRNAs are frequently observed in various cancers including melanomas. However, studies investigating IncRNAs in malignant melanoma are limited and their potential or functional role in driving metastatic progression in particular is largely unknown. A recent report has revealed a mechanism by which the IncRNAs might mediate metastasis in melanoma. The study provided evidence that the IncRNA growth-arrest specific transcript 5 (GAS5) can modulate the metastatic capacity of melanoma cells by suppressing expression of the matrix metalloproteases MMP2 and MMP9. It was shown that knocking down GAS5 resulted in upregulation of the MMPs which may facilitate new therapeutic implications. In this article the current understanding on the role of IncRNAs and the associated functional mechanisms in melanoma pathogenesis and their involvement in promoting metastases are evaluated