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Analysis of N-ras gene mutations in medulloblastomas by polymerase chain reaction and oligonucleotide probes in
A Iolascon1, A Lania, M Badiali
1Department of Pediatrics, University of Naples, Italy.
Abstract:
Precise data on the incidence of transforming ras oncogenes in pediatric tumors and the correlations with the histopathological properties of the tumors are very limited. Additionally the presence of ras activation in medulloblastomas has not been investigated so far. Using a combination of techniques including in vitro gene amplification by polymerase chain reaction (PCR) and detection of single base mutations by sequence-specific oligonucleotides we studied N-ras activation (mutations at codon 12, 13, and 61) in 32 medulloblastomas. DNA was isolated from 20 microns sections of formalin-fixed paraffin-embedded tissue. Mutations were found in 3 out of 32 examined medulloblastomas. In all cases only mutations of codon 61 were found: two of three mutations were C to A mutations at position 1 of the codon 61 (leading to a substitution of a glutamine residue for a lysine) and one was A to T mutation at position 3 in the same codon (glutamine-histidine). Our results indicate 10% incidence N-ras mutation in medulloblastoma, higher than in other CNS tumors studied so far. The main advantages of the procedure described are its greatly improved sensitivity, the increased speed with which tumor samples can be analyzed, and the possibility of using paraffin-embedded sections to analyze various rare tumors in retrospect.
Insights
This study investigated N-ras oncogene activation in pediatric medulloblastomas. We found a 10% incidence of N-ras mutations, primarily at codon 61, offering insights into pediatric tumor genetics.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Limited data exists on ras oncogene incidence in pediatric tumors.
- Ras activation in medulloblastomas has not been previously studied.
Purpose of the Study:
- To investigate N-ras gene activation in pediatric medulloblastomas.
- To correlate ras activation with tumor histopathology.
Main Methods:
- Analyzed 32 medulloblastomas using polymerase chain reaction (PCR) and sequence-specific oligonucleotides.
- Utilized DNA from formalin-fixed paraffin-embedded tissue sections.
- Focused on mutations at codons 12, 13, and 61 of the N-ras gene.
Main Results:
- N-ras mutations were detected in 3 out of 32 medulloblastomas (10% incidence).
- All identified mutations occurred at codon 61.
- Specific mutations included C to A and A to T at codon 61, altering the glutamine residue.
Conclusions:
- N-ras mutation incidence in medulloblastoma is higher than in other CNS tumors.
- The described PCR and oligonucleotide method is sensitive, rapid, and applicable to paraffin-embedded tissues for retrospective analysis of rare tumors.