Germline mutations in the BRCA1 gene have been reported to increase the lifetime risk of developing breast and/or ovarian cancer (BOC). By new sequencing technologies, numerous variants of uncertain ...significance (VUS) are identified. It is mandatory to develop new tools to evaluate their functional impact and pathogenicity. As the expression of pathogenic BRCA1 variants in
increases the frequency of intra- and inter-chromosomal homologous recombination (HR), and gene reversion (GR), we validated the two HR and the GR assays by testing 23 benign and 23 pathogenic variants and compared the results with those that were obtained in the small colony phenotype (SCP) assay, an additional yeast-based assay, that was validated previously. We demonstrated that they scored high accuracy, sensitivity, and sensibility. By using a classifier that was based on majority of voting, we have integrated data from HR, GR, and SCP assays and developed a reliable method, named yBRCA1, with high sensitivity to obtain an accurate VUS functional classification (benign or pathogenic). The classification of BRCA1 variants, important for assessing the risk of developing BOC, is often difficult to establish with genetic methods because they occur rarely in the population. This study provides a new tool to get insights on the functional impact of the BRCA1 variants.
Approximately 30–50% of hereditary breast and ovarian cancer (HBOC) is due to the presence of germline pathogenic variants in the BRCA1 (OMIM 113705) and BRCA2 (OMIM 600185) onco-suppressor genes, ...which are involved in DNA damage response. Women who carry pathogenic BRCA1 variants are particularly likely to develop breast cancer (BC) and ovarian cancer (OC), with a 45–79 percent and 39–48 percent chance, respectively. The BRCA1 c.4096+1G>A variant has been frequently ascertained in Tuscany, Italy, and it has also been detected in other Italian regions and other countries. Its pathogenetic status has been repeatedly changed from a variant of uncertain significance, to pathogenic, to likely pathogenic. In our study, 48 subjects (38 of whom are carriers) from 27 families were genotyped with the Illumina OncoArray Infinium platform (533,531 SNPs); a 20 Mb region (24.6 cM) around BRCA1, including 4130 SNPs (21 inside BRCA1) was selected for haplotype analysis. We used a phylogenetic method to estimate the time to the most recent common ancestor (MRCA) of BRCA1 c.4096+1G>A founder pathogenic variant. This analysis suggests that the MRCA lived about 155 generations ago—around 3000 years ago.
Multiple Endocrine Neoplasia type 1 is a rare genetic syndrome mainly caused by mutations of
gene and characterized by a combination of several endocrine and non-endocrine manifestations. The ...objective of this study was to describe cutaneous lesions and other non-endocrine manifestations of
in a cohort of patients with familial (F) and sporadic (S)
, compare the prevalence of these manifestations between the two cohorts, and investigate the correlation with
mutation status.
We collected phenotypic and genotypic data of 185 patients with F-
and S-
followed from 1997 to 2022. The associations between F-
and S-
or
mutation-positive and mutation-negative patients and non-endocrine manifestations were determined using chi-square or Fisher's exact tests or multivariate exact logistic regression analyses.
The prevalence of angiofibromas was significantly higher in F-
than in S-
in both the whole (p < 0.001) and index case (p = 0.003) cohorts. The prevalence of lipomas was also significantly higher in F-
than in S-
(p = 0.009) and in
mutation-positive than in
mutation-negative (p = 0.01) index cases. In the whole cohort, the prevalence of lipomas was significantly higher in
mutation-positive compared to
mutation-negative patients (OR = 2.7, p = 0.02) and in F-
than in S-
(p = 0.03), only after adjustment for age. No significant differences were observed for the other non-endocrine manifestations between the two cohorts. Hibernoma and collagenoma were each present in one patient (0.5%) and meningioma and neuroblastoma in 2.7% and 0.5%, respectively. Gastric leiomyoma was present in 1.1% of the patients and uterine leiomyoma in 14% of women. Thyroid cancer, breast cancer, lung cancer, basal cell carcinoma, melanoma, and colorectal cancer were present in 4.9%, 2.7%, 1.6%, 1.6%, 2.2%, and 0.5% of the whole series, respectively.
We found a significantly higher prevalence of angiofibromas and lipomas in F-
compared with S-
and in
mutation-positive compared to
mutation-negative patients. In patients with one major endocrine manifestation of
, the presence of cutaneous lesions might suggest the diagnosis of
and a possible indication for genetic screening.
Primary ciliary dyskinesia (PCD) is a highly heterogeneous genetic disorder caused by defects in motile cilia. The hallmark features of PCD are the chronic infections of the respiratory tract, ...moreover, clinical manifestations include also laterality defects and risk of male infertility. Clinical phenotypes of PCD are the result of mutations in genes encoding components of axonema or factors involved in axonemal assembly. Recent studies have identified over 45 PCD-associated genes, therefore, molecular analysis represents a powerful diagnostic tool to confirm and uncover new genetic causes of this rare disease.
Here, we describe a female infant of Moroccan origin with normal pressure hydrocephalus (NPH) in addition to most common PCD symptoms. Transmission Electron Microscopy (TEM) and molecular tests, such as a Next generation Sequencing panel and a custom array CGH, were performed for diagnosis of PCD. TEM revealed outer dynein arm (ODA) defects, whilst molecular analyses detected a novel 6,9 kb microdeletion in DNAI2 gene.
Since DNAI2 mutations are very rare, this case report contributes to better delineate the important role of DNAI2 as causative of PCD phenotype, suggesting, furthermore, that the variations in DNAI2 may be as a new genetic risk factor for NPH. Indeed, although the association of hydrocephalus with PCD has been well documented, however, only a small number of human patients show this defect. Furthermore, this study highlights the importance of high-throughput technologies in advancing our understanding of heterogeneous genetic disorders.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
With the progress of sequencing technologies, an ever-increasing number of variants of unknown functional and clinical significance (VUS) have been identified in both coding and non-coding regions of ...the main Breast Cancer (BC) predisposition genes. The aim of this study is to identify a mutational profile of coding and intron-exon junction regions of 12 moderate penetrance genes (ATM, BRIP1, CDH1, CHEK2, NBN, PALB2, PTEN, RAD50, RAD51C, RAD51D, STK11, TP53) in a cohort of 450 Italian patients with Hereditary Breast/Ovarian Cancer Syndrome, wild type for germline mutation in BRCA1/2 genes. The analysis was extended to 5′UTR and 3′UTR of all the genes listed above and to the BRCA1 and BRCA2 known regulatory regions in a subset of 120 patients. The screening was performed through NGS target resequencing on the Illumina platform MiSeq. 8.7% of the patients analyzed is carriers of class 5/4 coding variants in the ATM (3.6%), BRIP1 (1.6%), CHEK2 (1.8%), PALB2 (0.7%), RAD51C (0.4%), RAD51D (0.4%), and TP53 (0.2%) genes, while variants of uncertain pathological significance (VUSs)/class 3 were identified in 9.1% of the samples. In intron-exon junctions and in regulatory regions, variants were detected respectively in 5.1% and in 32.5% of the cases analyzed. The average age of disease onset of 44.4 in non-coding variant carriers is absolutely similar to the average age of disease onset in coding variant carriers for each proband’s group with the same cancer type. Furthermore, there is not a statistically significant difference in the proportion of cases with a tumor onset under age of 40 between the two groups, but the presence of multiple non-coding variants in the same patient may affect the aggressiveness of the tumor and it is worth underlining that 25% of patients with an aggressive tumor are carriers of a PTEN 3′UTR-variant. This data provides initial information on how important it might be to extend mutational screening to the regulatory regions in clinical practice.
Congenital adrenal hyperplasia (CAH) is a heterogeneous group of autosomal recessive disorders due to defects in adrenal steroid biosynthesis. In about 90% of patients, CAH is caused by pathogenetic ...variants in CYP21A2 gene, impairing the function of 21-hydroxylase (21-OH) enzyme. CAH can present as classical form (simple virilizing or salt wasting) or as non-classical form (NC-CAH). NC-CAH is due to pathogenetic variants in the CYP21A2 gene that result in 20–70% residual activity of 21-hydroxylase. Early diagnosis may be missed, mainly in childhood, jeopardizing long-term outcome. This paper will review some information on clinical findings, symptoms, diagnostic approaches, and treatments of NC-CAH in childhood, allowing better management and long-term outcome.
Disorders of sexual development (DSDs) encompass a group of congenital conditions associated with atypical development of internal and external genital structures. Among those with DSDs are 46,XX ...males, whose condition mainly arises due to the translocation of SRY onto an X chromosome or an autosome. In the few SRY-negative 46,XX males, overexpression of other pro-testis genes or failure of pro-ovarian/anti-testis genes may be involved, even if a non-negligible number of cases remain unexplained. A three-year-old boy with an SRY-negative 46,XX karyotype showed a normal male phenotype and normal prepubertal values for testicular hormones. A heterozygous de novo in tandem duplication of 50,221 bp, which encompassed exons 2 and 3 of the Doublesex and Mab-3-related transcription factor 1 (DMRT1) gene, was detected using MPLA, CGH-array analysis, and Sanger sequencing. Both breakpoints were in the intronic regions, and this duplication did not stop or shift the coding frame. Additional pathogenic or uncertain variants were not found in a known pro-testis/anti-ovary gene cascade using a custom NGS panel and whole genome sequencing. The duplication may have allowed DMRT1 to escape the transcriptional repression that normally occurs in 46,XX fetal gonads and thus permitted the testicular determination cascade to switch on. So far, no case of SRY-negative 46,XX DSD with alterations in DMRT1 has been described.
Leigh Syndrome (LS, OMIM 256000) is an early-onset, progressive neurodegenerative disorder characterized by broad clinical and genetic heterogeneity; it is the most frequent disorder of mitochondrial ...energy production in children. LS inheritance is complex because patients may present mutations in mitochondrial DNA (mtDNA) or in nuclear genes, which predominantly encode proteins involved in respiratory chain structure and assembly or in coenzyme Q10 biogenesis. However, during the last 15 years, the discovery of several genetic mutations and improved knowledge of the natural history of LS has significantly increased our understanding of this mitochondrial disorder.
Here we describe a 19-year-old male with clinical and neuroimaging LS diagnosed at 3 years of age. Genetic analyses of the whole mtDNA for maternally inherited LS (MILS) and neuropathy ataxia retinitis pigmentosa (NARP) syndrome failed to reveal any pathogenic mutations.
Recently, a missense mutation in ECHS1 and a ~ 35 kb deletion in 10q26.3 involving the region including the gene were identified by WES (whole exome sequencing), uncovering the genetic diagnosis clinically hypothesized for 15 years. We also report the long-term follow-up of this patient, showing a comparison with classical LS or other Leigh-like pictures.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
In this study, we determined if BRCA1 partners involved in DNA double-strand break (DSB) and mismatch repair (MMR) may contribute to breast and ovarian cancer development. Taking advantage the ...functional conservation of DNA repair pathways between yeast and human, we expressed several BRCA1 missense variants in DNA repair yeast mutants to identify functional interaction between BRCA1 and DNA repair in BRCA1-induced genome instability. The pathogenic p.C61G, pA1708E, p.M775R, and p.I1766S, and the neutral pS1512I BRCA1 variants increased intra-chromosomal recombination in the DNA-repair proficient strain RSY6. In the
, and
deletion strains, the BRCA1 variants p.C61G, pA1708E, p.M775R, p.I1766S, and pS1215I did not increase intra-chromosomal recombination suggesting that a functional DNA repair pathway is necessary for BRCA1 variants to determine genome instability. The pathogenic p.C61G and p.I1766S and the neutral p.N132K, p.Y179C, and p.N550H variants induced a significant increase of reversion in the
Δ strain; the neutral p.Y179C and the pathogenic p.I1766S variant induced gene reversion also, in the
Δ strain. These results imply a functional interaction between MMR and BRCA1 in modulating genome instability. We also performed a somatic mutational screening of
, and
genes in tumor samples from 34 patients and identified eight pathogenic or predicted pathogenic rare missense variants: four in
, one in
, one in
, and two in
. Although we found no correlation between BRCA1 status and these somatic DNA repair variants, this study suggests that somatic missense variants in DNA repair genes may contribute to breast and ovarian tumor development.
Breast Cancer 2 gene (BRCA2) mutation carriers have a 45% chance of developing breast cancer and a 11% risk of developing ovarian cancer by the age of 70. While hundreds of BRCA2-truncating mutations ...have been associated with an increased cancer risk in carriers, the contribution of unclassified variants (UCVs) to cancer risk remains largely undefined. BRCA2-defective cells show a high degree of chromosome instability. Although a functional assay based on the BRCA2 capability to stimulate DSB-induced homologous recombination (HR) as a way to classify UCVs has been proposed, so far no data are available concerning the effect of BRCA2 UCVs on spontaneous HR. In this study, we proposed a novel functional HR-based assay that determines the effect of the transient overexpression of the BRCA2 variant on spontaneous HR. This assay will help one in the difficult task of classifying UCVs, and it will give more information on how BRCA2 may induce genome instability and on the basic mechanism of BRCA2-induced tumourigenesis. We chose 11 BRCA2 UCVs not previously described or classified in other articles, and distributed along the entire BRCA2-coding region. They are as follows: G173V, D191V, S286P, M927V, T1011R, L1019V, N1878K, S2006R, R2108C, G2353R and V3091I. Basically, because the expression of BRCA2wt and the neutral variants did not increase spontaneous HR, we classified the variants G173V, S286P, M927V, T1011R and L1019V as HR-negative and presumed that they were not pathogenic. The HR-positive variants, D191V, N1878K, S2006R, R2108C, G2353R, and V3091I, which increased HR as much as the cancer-associated variant G2748D, could probably be classified as pathogenic. We observed that all our variants in the C-terminus of the protein behaved differently from the wt, suggesting a role for this protein region in spontaneous HR.
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