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N- and KRAS mutations in primary testicular germ cell tumors: incidence and possible biological implications
R A Olie1, L H Looijenga, L Boerrigter
1Laboratory of Experimental Patho-Oncology, Dr. Daniel den Hoed Cancer Center, Rotterdam, The Netherlands.
Abstract:
Recently, conflicting results have been reported on the incidence of RAS mutations in primary testicular germ cell tumors of adults (TGCTs). In four studies a low incidence of mutations (less than 15%) in a variety of TGCTs or derived cell lines was found, whereas in two other studies a high incidence of N- or KRAS mutations (over 40%) was shown. A total of 62 testicular seminomas (SE) and 34 nonseminomatous TGCTs (NS) were studied thus far. The largest series consisted of 42 TGCTs, studied on paraffin embedded tissue. We present the results of analysis for the presence of N- and KRAS mutations, in codons 12, 13, and 61, in snap frozen samples of 100 primary TGCTs, comprising 40 SE and 60 NS. Using the polymerase chain reaction (PCR) and allele specific oligonucleotide hybridization (ASO), mutations were found in five SE (three in NRAS and two in KRAS, all codon 12), and in one NS (KRAS, codon 12). To exclude underestimation of the incidence of RAS mutations in TGCTs due to the presence of an excess of wild type alleles in the analyzed sample, a PCR technique preferentially amplifying KRAS alleles with a mutation in codon 12 was applied to all SE. This approach, allowing a 250 times more sensitive assay, resulted in the detection of only one additional SE with a mutation. Based on a critical analysis of published data and on our results from the largest series of frozen samples investigated thus far, we conclude that N- or KRAS mutations are rare and apparently not essential for initiation or progression of TGCTs.
Insights
RAS mutations are rare in adult testicular germ cell tumors (TGCTs), with N- or KRAS mutations not appearing essential for tumor development. This study analyzed 100 frozen TGCT samples, finding a low incidence of these specific mutations.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Conflicting reports exist regarding the incidence of RAS mutations in adult testicular germ cell tumors (TGCTs).
- Previous studies showed a low (<15%) or high (>40%) incidence of N- or KRAS mutations in TGCTs and derived cell lines.
- A total of 62 testicular seminomas (SE) and 34 nonseminomatous TGCTs (NS) have been analyzed in prior research.
Purpose of the Study:
- To investigate the incidence of N- and KRAS mutations in codons 12, 13, and 61 in primary TGCTs.
- To clarify the role of RAS mutations in the initiation and progression of TGCTs.
- To analyze a large series of frozen TGCT samples for RAS mutations.
Main Methods:
- Analysis of 100 primary TGCT samples (40 SE, 60 NS) using snap-frozen tissue.
- Utilized polymerase chain reaction (PCR) and allele-specific oligonucleotide hybridization (ASO) to detect N- and KRAS mutations.
- Employed a highly sensitive PCR technique to preferentially amplify mutated KRAS alleles in codon 12 to avoid underestimation.
Main Results:
- Mutations were detected in five SE (three NRAS, two KRAS, all codon 12) and one NS (KRAS, codon 12) in the initial analysis.
- A sensitive PCR assay detected only one additional SE with a mutation, indicating a low overall incidence.
- RAS mutations were found to be rare in the analyzed cohort of TGCTs.
Conclusions:
- N- or KRAS mutations are rare in primary testicular germ cell tumors (TGCTs).
- These mutations do not appear to be essential for the initiation or progression of TGCTs.
- The findings provide a clearer understanding of the genetic landscape of TGCTs.