Additional characterization of a hexanucleotide polymorphic site in the first intron of human H-ras gene: comparative study of its alterations in non-small cell lung carcinomas and sporadic invasive breast carcinomas

Intron 1 of the human I-I-l ns gene possesses a polymorphism consisting of repetitions of the GGGCCT consensus. Three alleles have been reported at th is locus. We confirmed that two, pi and PZ, display four and two repeats, r espectively, with their internal sequence structure similar to that previou sly described. The third, P3, previously assigned as a three-unit repetitio n allele according to its electrophoretic mobility and with no other inform ation regarding its internal structure, was also found. Sequence analysis o f the P3 allele revealed that it consists of three perfect repeats of the G GGCCT consensus. This polymorphism is present only in human c-H-ras gene, a lthough single hexanucleotide repeats are found scattered within intron 1 o f this gene in rodents. Analysis of this locus in matched tumor/distant nor mal samples from. (i) 38 patients with nonsmall-cell lung carcinoma (NSCLC) , and (ii) 35 patients with sporadic invasive breast carcinoma, revealed: ( 1) 6.6% and 19% loss of heterozygosity (LOH) respectively, and (2) 10.5% an d 2.9% hexanucleotide instability (III) respectively, detected by the prese nce of shifted in length alleles. Shifted alleles exhibited altered interna l sequence structure in comparison to normal ones, suggesting complex mutat ional events. The same pattern of alterations was also detected in tissues adjacent to lung adenocarcinomas and dysplasias adjacent to squamous cell c arcinomas (7.7% LOH, 5.9% III), implying that abnormalities at this locus m ay be early events in lung carcinogenesis. The frequency of alterations (LO H vs. HI) was significantly different among NSCLC and breast cancer (P=.005 ), probably due to the different tumor biology of each system. Finally, alt ered mRNA expression of H-ras gene was detected in all cases with HI, but t his finding was also observed in samples without I-II. In view of reports s howing that elements in intron 1 of H-ras gene potentially influence its tr anscriptional regulation, from our results we cannot exclude that the hexan ucleotide locus could be an element with possible involvement in expression al regulation of this gene. (C) 2001 Elsevier Science Tnc. All rights reser ved.