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Genetic alterations in rat colon tumors induced by heterocyclic amines
M Toyota1, T Ushijima, H Kakiuchi
1National Cancer Center Research Institute, Tokyo, Japan.
Background:
In rat colon tumors induced by the cooked food mutagens 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), ras and p53 alterations are rarely detected. To investigate the roles of the APC gene and microsatellite instability (MI) in PhIP-induced colon carcinogenesis, mutations of the APC gene and alterations of microsatellites were examined.
Methods:
Complimentary DNA sequence of the rat APC gene were determined by polymerase chain reaction (PCR) using primers based on the human APC sequence. PCR-single strand conformation polymorphism (SSCP) analysis was performed using primers based on sequences of flanking introns and exon 15. Microsatellite alterations were also analyzed using 85 microsatellite sequences dispersed through most of the rat chromosomes.
Results:
Five mutations in the APC gene were detected in four of eight PhIP-induced rat colon tumors. All five mutations involved deletion of a guanine base in a 5'-GGGA-3' sequence. Only 2 of 13 IQ-induced colon tumors had mutations of the APC gene and these were base substitution mutations. Seven of eight PhIP-induced colon tumors had microsatellite alterations in at least one locus, whereas no alterations were observed in the IQ-induced colon tumors.
Conclusions:
The specific 5'-GGGA-3' to 5'-GGA-3' mutation and MI demonstrated in this study are strong evidence of a mutational fingerprint of PhIP.
Insights
Mutations in the APC gene and microsatellite instability (MI) were analyzed in rat colon tumors. PhIP exposure specifically induced APC gene mutations and MI, suggesting a PhIP mutational fingerprint.
Area of Science:
- Oncology
- Molecular Biology
- Carcinogenesis
Background:
- Colon tumors induced by cooked food mutagens 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) rarely show ras and p53 alterations.
- The roles of the Adenomatous Polyposis Coli (APC) gene and microsatellite instability (MI) in PhIP-induced colon carcinogenesis require investigation.
Purpose of the Study:
- To investigate the role of the APC gene in PhIP-induced colon carcinogenesis.
- To examine microsatellite alterations in PhIP- and IQ-induced rat colon tumors.
Main Methods:
- Rat APC gene complementary DNA (cDNA) sequences were determined using polymerase chain reaction (PCR) with human APC primers.
- PCR-single strand conformation polymorphism (SSCP) analysis was performed on exon 15 and flanking introns of the APC gene.
- Microsatellite alterations were analyzed across 85 microsatellite loci throughout the rat genome.
Main Results:
- Five distinct mutations in the APC gene were identified in 4 of 8 PhIP-induced colon tumors, all involving a guanine deletion in a 5'-GGGA-3' sequence.
- In contrast, only 2 of 13 IQ-induced tumors exhibited APC gene mutations, characterized by base substitutions.
- Seven of eight PhIP-induced tumors displayed microsatellite alterations, while none were observed in IQ-induced tumors.
Conclusions:
- The specific 5'-GGGA-3' to 5'-GGA-3' mutation in the APC gene is a hallmark of PhIP exposure.
- The observed microsatellite instability (MI) in PhIP-induced tumors further supports a unique mutational fingerprint associated with PhIP.
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