Screening de alterações na região hotspot do gene BRCA2 em tumores de mama caninos

Abstract

Breast tumors are common tumors in female dogs. Representing 25 to 50% of canine tumors, and 50% are considered malignant. They are considered special for researchers due to similarities to human breast cancer. The identification of genetic risk factors is crucial to improving the prevention, diagnosis and treatment. The main genetic factor involved in mammary carcinogenesis is the alteration of tumor suppressor genes, including BRCA2. When mutated is great risk to develop breast tumor. Thus, this study aims to identify changes in the BRCA2 gene hotspot region related to the development of canine mammary tumors, with the analysis of exons 11, 12, 24 and 27, which are the most studied in dogs, for that gene and which shows changes already described in literature in order to identify genetic alterations in dogs that have mammary tumor correlating the changes found with the histopathological features, and identify potential markers of tumor process for the construction of diagnostic kits and prognosis disease. Samples were used, of neoplastic breast tissue and non-neoplastic canine treated at HOVET / UFRA. The samples were stored in microtubes with RNA Later® and stored at -20 ° C. DNA extraction was performed with phenol-chloroform method, and then performed PCR. The identification of changes took place in BioEdit v.5.0.6 program, by aligning the sequences obtained with the corresponding exons derived from GenBank. Statistical analysis was performed using the chi-square test, G test and Fisher exact test, odds ratio by GraphPad Prism 6.0. 40 samples of 23 animals were used, of which only one was male, aged between 4-15 years, several races with the Poodle being the most affected after the mongrel dogs with tumor histogenesis sarcoma type is more frequent. In the analysis of the changes found it was observed that the exon 11 had the highest number of polymorphisms in the hotspot region analyzed: A355G, CCC1954AAA, C2121A, A2414G and A511C; and only the change A511C was not correlated with increased risk of developing tumors compared with the clinical and pathological characteristics of the patients. The exon 12 had a G / A polymorphism (rs23255542), and allele frequencies of this were not in Hardy-Weinberg equilibrium in tumor groups and non-tumor analyzed and that the presence of the mutant allele in individuals who used contraceptive increased 13 times the risk of developing tumors, suggesting that this alteration may influence even without causing amino acid change in canine mammary tumorigenesis. The only change observed in exon 24 (A / G, position 7,747,634) was found only in heterozygous at a low frequency in the population, while the polymorphism of indel type observed in exon 27 (10204 ins / del AAA) showed allelic frequencies They are not in Hardy-Weinberg equilibrium. This event, despite being observed at a high frequency in the population studied, was not correlated with any clinical or histopathological characteristics analyzed. Thus, it may be suggested that polymorphisms of exons 11:12 have potential as biomarkers of canine mammary tumorigenesis in this population.