Absence of oncogenic mutations of RAS family genes in soft tissue sarcomas of 100 Japanese patients

Background: Activating point mutations of genes of the RAS family (KRAS, HRAS and NRAS genes) are frequently found in carcinomas, but their prevalence in sarcomas varies considerably among ethnic groups. No extensive studies in Japanese patients have been performed. Materials and Methods: Mutation analyses of three RAS genes (KRAS, HRAS and NRAS) were performed using polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) analyses and PCR direct sequencing in one hundred cases of soft tissue sarcoma (STS) as well as six STS cell lines from Japanese patients. Results: No mutations were found in two hot spot regions (codon 12- 13 and 61) of the three RAS genes. Conclusion: Activating mutations of the RAS gene family are uncommon events in soft tissue sarcomas in Japanese patients. The detection of oncogene mutations in tumors has not only led to a better understanding of tumorigenesis, but also aided in diagnosis and treatment. RAS genes are the most frequently found activating oncogenes in human carcinomas. They gain their oncogenic activity via a single amino acid substitution in codon 12, 13 or 61 of the KRAS-, HRAS- or NRAS-encoded p21 protein (1). Point mutations occurring in RAS genes give rise to proteins with reduced intrinsic GTPase activity associated with oncogenesis (2). Some reports have indicated that RAS oncogenes have no ability to transform primary cells and mutant Ras proteins can only transform cells that have undergone predisposing changes such as immortalization (3, 4). We have previously reported that an activated HRAS gene transformed mesenchymal stem cells only when they were immortalized beforehand by the introduction of a human telomerase catalytic subunit (hTERT) and BMI1 gene (5). Activated RAS genes have frequently been found in adenocarcinomas of the pancreas (90%), colon (50%), thyroid (50%) and lung (30%) (6). Soft tissue sarcomas (STSs) account for fewer than 1% of human malignancies and fewer than 2% of cancer deaths (7, 8). They present a heterogeneous group of tumors with respect to origin and morphological features. Previous studies have provided inconsistent results, with the frequency of RAS mutations in STS ranging from 0% to 44% (9-17). The inconsistency may be due to small sample numbers, incomplete sensitivity of the detection methods or the patients' ethnic origin. Moreover, no reports have investigated all codons 12, 13 and 61 of KRAS, HRAS and NRAS genes in a large number of samples. In the present study, extensive analyses in codons 12, 13 and 61 of the KRAS, HRAS and NRAS genes were performed in one hundred STS samples and six STS cell lines from Japanese patients. To increase sensitivity, two methods were used to detect mutations: polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) and PCR direct sequencing.