Predisposition gene identification in thyroid and breast carcinomas syndrome by exome sequencing

Abstract

Hereditary cancer syndromes are estimated to account for 10% of all tumors, while 15% of cases are associated with familial aggregation of cancer. The etiology of solid tumors, including breast carcinomas (BC) and thyroid carcinomas (TC), is multifactorial, dependent on environmental and genetic factors playing a variable role in the disease etiology. However, a significant incidence of these tumors is associated with a hereditary component. Thus, the identification of predisposition factors and characterization of the genetic risk are essential for diagnosis, prognosis and genetic counseling of affected members of these families. Within this context, the Exome Sequence has been applied for searching the disease-related variations, mainly in monogenic inherited disorders. About 85% of the large effect mutations are found at protein coding sequence, mainly in exonic regions and splicing sites. The aim of the study is identify predisposition genes associated with breast and thyroid carcinomasby exoma sequencing analysis in patients with at least one of these tumors and familial cancer history as well. Patients were selected according to stricted and well-defined inclusion and exclusion criteria. Inclusion criteria consisted of proband with non-medullary TC and/or BC with familial cancer history of these tumors AND at least two first- or second-degree relatives with one of these tumors developed before 45 years of age OR one first-degree relative with one of these tumors and proband diagnosed before 40 years of age AND history of other tumors. Exclusion criteria comprised female patients with BC treated by radiotherapy followed by TC, patients with medullary thyroid carcinomas and probands affected with common mutations in major hereditary predisposition genes. All patients were followed at Oncogenetic Department of AC Camargo Cancer Center. Twenty-four blood samples and 4 tumor samples from patients whose fulfill these criteria were collected for exome analysis. Paired-end libraries were prepared following the manufacturer's protocols and whole exome capture was carried out using the protocol for Nextera Exome Enrichment kit (Illumina), targeting 62 Mb of sequence. Sequencing was carried out for the captured libraries with the HiSeq 1000 using 100 bp paired-end reads following Illumina's standard protocol using the Illumina cluster station cBot and HiSeq. To achieve high level sensitivity and accuracy for detecting all the mutations in the whole exome, a 140x coverage of depth was estimated. We alredy generated 164 Gb of sequence for all sample and, after cleaning, > 67% of reads were kept. A mean of 99,89% of these reads were mapped against reference genome. The next step is identify the variants mutations. The detection of novel genetic alterations associated with thyroid and breast carcinomas could extend the spectrum of hereditary cancer syndromes, allowing the identification of patients at risk of developing these tumors, as well as contributing to diagnosis and genetic counseling of these families. (AU)