Demonstration of a polymorphism in the gene ERCC1 by two DNA sequencing methods.

New methodology is important to the advancement of biomedical science. We recently described a polymorphism within exon IV of the ERCC1 gene, which is associated with an approximate 50% decrease in codon usage in some biological systems. In this report we show that this polymorphism can be readily demonstrated by standard manual DNA sequencing, and by a recently developed methodology which relies on spectrophotometric principles. The major advantage of this new methodology is that several-fold more samples can be assessed per unit time, at reduced cost.

Platinum-sensitive and platinum-resistant ovarian cancer tissues show differences in the relationships between mRNA levels of p53, ERCC1 and XPA.

We investigated the association between p53 mRNA expression and clinically relevant surrogates of nucleotide excision repair (ERCC1 and XPA) in 28 ovarian cancer specimens. We observed that platinum-resistant tumors showed higher mRNA levels of p53, ERCC1, and XPA than platinum-sensitive tumors; mRNA expression patterns in responders differed substantially from nonresponders; and p53 expression showed a strong correlation with the expression of ERCC1, and of XPA in platinum-sensitive tumor tissues, but not with platinum-resistant tumors. 47% of the mutations from p53 sequence analysis were not related to clinical response to chemotherapy. We conclude that the p53 influence on DNA repair in human malignancy may vary substantially from tumor to tumor, and that such differences are not necessarily related to the mutational status of p53.

Expression of an alternatively spliced ercc1 messenger-RNA species, is related to reduced DNA-repair efficiency in human T-lymphocytes.

We have previously shown that in non-drug-selected human T lymphocytes, DNA repair is the primary determinant of cellular resistance to cisplatin (1). In this system, we have assessed mRNA levels of expression of the nucleotide excision repair (NER) genes ERCC1 and XPA, as well as the alternatively spliced species of ERCC1 which lacks exon VIII. The focus of these studies, was to try to identify the possible relative roles of normal XPA, full-length ERCC1, and alternatively spliced ERCC1, in a system where DNA repair is a clear determinant of cisplatin resistance. ERCC1 expression was directly related to cisplatin-DNA adduct repair capability, as well as directly related to cisplatin resistance, suggesting a primary role for ERCC1 in effecting DNA repair. XPA expression was approximately equivalent in each cell line, regardless of the level of DNA repair activity, suggesting a helper role for the product of this gene. The mRNA levels of the alternatively spliced species of ERCC1 were strongly inversely related to DNA repair activity, suggesting a possible inhibitory influence on the DNA repair process. This interpretation is consistent with alternative splicing of several known oncogenes, where the alternatively spliced species has an inhibitory effect on the full-length gene product. The NER pathway appears to be vitally important in effecting cisplatin resistance in non-drug-selected T lymphocytes. Further, it appears that NER may have at least one inhibitory regulatory component.