Heterozygous Pathogenic Nonsense Variant in the ATM Gene in a Family with Unusually High Gastric Cancer Susceptibility.

Germline pathogenic variants (PVs) in the Ataxia Telangiectasia mutated (ATM) gene (MIM* 607585) increase the risk for breast, pancreatic, gastric, and prostatic cancer and, to a reduced extent, ovarian and colon cancer and melanoma, with moderate penetrance and variable expressivity. We describe a family presenting early-onset gastric cancer and harboring a heterozygous pathogenic ATM variant. The proband had gastric cancer (age 45) and reported a sister deceased due to diffuse gastric cancer (age 30) and another sister who developed diffuse gastric cancer (age 52) and ovarian serous cancer. Next generation sequencing for cancer susceptibility genes (APC, ATM, BRD1, BRIP1, CDH1, CDK4, CDKN2A, CHEK2, EPCAM, MLH1, MRE11, MSH2, MSH6, MUTYH, NBN, PALB2, PMS2, PTEN, RAD50, RAD51C, RAD51D, RECQL1, SMAD4, STK11, and TP53) was performed. Molecular analysis identified the truncating c.5944C>T, p.(Gln1982*) variant in the ATM (NM_000051.3; NP_000042.3) in the proband. The variant had segregated in the living affected sister and in the unaffected daughter of the deceased affected sister. Familial early-onset gastric cancer is an unusual presentation for ATM-related malignancies. Individual variants may result in different specific risks. Genotype-phenotype correlations are challenging given the low penetrance and variable expressivity. Careful family history assessments are pivotal for prevention planning and are strengthened by the availability of molecular diagnoses.

Long QTc in hypertrophic cardiomyopathy: A consequence of structural myocardial damage or a distinct genetic disease?

Hypertrophic cardiomyopathy (HCM) is an autosomal dominant disease, characterized by the presence of unexplained left ventricular hypertrophy. This condition is often associated with electrocardiographic abnormalities including QTc prolongation occurring in 13% of patients. The main explanation for prolonged QTc in HCM is myocardial hypertrophy and the related structural damage. However, other mechanisms, including long QT syndrome (LQTS) genes mutations, may be involved. In the present study we explored the hypothesis of a distinct genetic basis underlying QTc prolongation in HCM by investigating the potential co-inheritance of pathogenic gene variants associated with LQTS and HCM. For this purpose, starting from a cohort of 150 HCM patients carrying pathogenic variants in sarcomere genes, we selected 25 patients carrying a QTc prolongation unexplained by any other cause. The QTc was considered prolonged if greater than 450 ms in males and greater than 470 ms in females. The NGS analysis was performed with Illumina TrueSight Cardio panel genes on Illumina MiniSeq platform. We identified pathogenic/likely pathogenic variants in the KCNQ1 in two patients (c.1781G > A, p. Arg594Gln; c.532G > A, p. Ala178Thr) (8%). Variants of uncertain significance were identified in SCN5A, KCNJ5, AKAP9 and ANK2 in four patients (16%). Although the results are limited by the small number of patients included in the study, they highlight a minor contribution of LQTS genes for QTc prolongation in HCM patients. The screening for ion channel genes mutations may be considered in HCM patients with prolonged QTc unexplained by any other cause. This in-depth molecular diagnosis may contribute to improve risk stratification and treatment planning.

A New SMAD4 Splice Site Variant in a Three-Generation Italian Family with Juvenile Polyposis Syndrome.

Juvenile polyposis syndrome (JPS) is an autosomal dominant disorder characterized by hyperplastic polyps in the upper and lower gastrointestinal (GI) tract with a high risk of developing GI cancers. We have described a three-generation Italian family with all the spectrum of SMAD4 phenotype. A multigene panel test was performed on the genomic DNA of the proband by next-generation sequencing, including genes related to hereditary GI tumor syndromes. Molecular analysis revealed the presence of the c.1140-2A>G substitution in the SMAD4 gene, a novel splice variant that has never been described before. Our family is remarkable in that it illustrates the variable expressivity of the SMAD4 phenotype within the same family. The possibility of phenotype variability should also be considered within family members carrying the same mutation. In JPS, a timely genetic diagnosis allows clinicians to better manage patients and to provide early surveillance and intervention for their asymptomatic mutated relatives in the early decades of life.

Beyond BRCA1 and BRCA2: Deleterious Variants in DNA Repair Pathway Genes in Italian Families with Breast/Ovarian and Pancreatic Cancers.

The 5-10% of breast/ovarian cancers (BC and OC) are inherited, and germline pathogenic (P) variants in DNA damage repair (DDR) genes BRCA1 and BRCA2 explain only 10-20% of these cases. Currently, new DDR genes have been related to BC/OC and to pancreatic (PC) cancers, but the prevalence of P variants remains to be explored. The purpose of this study was to investigate the spectrum and the prevalence of pathogenic variants in DDR pathway genes other than BRCA1/2 and to correlate the genotype with the clinical phenotype. A cohort of 113 non-BRCA patients was analyzed by next-generation sequencing using a multigene panel of the 25 DDR pathways genes related to BC, OC, and PC. We found 43 unique variants in 18 of 25 analyzed genes, 14 classified as P/likely pathogenic (LP) and 28 as variants of uncertain significance (VUS). Deleterious variants were identified in 14% of index cases, whereas a VUS was identified in 20% of the probands. We observed a high incidence of deleterious variants in the CHEK2 gene, and a new pathogenic variant was detected in the RECQL gene. These results supported the clinical utility of multigene panel to increase the detection of P/LP carriers and to identify new actionable pathogenic gene variants useful for preventive and therapeutic approaches.

Rapid detection of copy number variations and point mutations in BRCA1/2 genes using a single workflow by ion semiconductor sequencing pipeline.

Molecular analysis of BRCA1 (MIM# 604370) and BRCA2 (MIM #600185) genes is essential for familial breast and ovarian cancer prevention and treatment. An efficient, rapid, cost-effective accurate strategy for the detection of pathogenic variants is crucial. Mutations detection of BRCA1/2 genes includes screening for single nucleotide variants (SNVs), small insertions or deletions (indels), and Copy Number Variations (CNVs). Sanger sequencing is unable to identify CNVs and therefore Multiplex Ligation Probe amplification (MLPA) or Multiplex Amplicon Quantification (MAQ) is used to complete the BRCA1/2 genes analysis. The rapid evolution of Next Generation Sequencing (NGS) technologies allows the search for point mutations and CNVs with a single platform and workflow. In this study we test the possibilities of NGS technology to simultaneously detect point mutations and CNVs in BRCA1/2 genes, using the OncomineTM BRCA Research Assay on Personal Genome Machine (PGM) Platform with Ion Reporter Software for sequencing data analysis (Thermo Fisher Scientific). Comparison between the NGS-CNVs, MLPA and MAQ results shows how the NGS approach is the most complete and fast method for the simultaneous detection of all BRCA mutations, avoiding the usual time consuming multistep approach in the routine diagnostic testing of hereditary breast and ovarian cancers.

Downregulation of the major histocompatibility complex class I molecules by human herpesvirus type 8 and impaired natural killer cell activity in primary effusion lymphoma development.

Primary effusion lymphomas (PELs) are invariably infected by human herpesvirus type 8 (HHV8) and often co-infected by Epstein-Barr virus (EBV). We found that expression of major histocompatibility complex class I (MHC-I) surface molecules was significantly decreased in PEL cells when compared with HHV8 negative lymphomas, irrespective of EBV infection. MHC-I downregulation rendered PEL cells sensitive to recognition and killing by natural killer (NK) cells. Intriguingly, analysis of MHC-I non-restricted cytotoxicity in two PEL patients indicated a reduced NK cell activity when compared with healthy individuals. These data suggest that PEL outgrowth may require an impaired NK cell function.

NK cell activity controls human herpesvirus 8 latent infection and is restored upon highly active antiretroviral therapy in AIDS patients with regressing Kaposi's sarcoma.

Kaposi's sarcoma (KS) develops upon reactivation of human herpesvirus 8 (HHV8) infection and virus dissemination to blood and tissue cells, including endothelial and KS spindle cells where the virus is mostly present in a latent form. However, this may likely require the presence of compromised host immune responses and/or the evasion of infected cells from the host immune response. In this regard, mechanisms of evasion of productively infected cells from both CTL and NK cell responses, and resistance of latently infected cells from specific CTL, have already been shown. Here we show that cells which are latently infected by HHV8 are indeed efficiently lysed by NK cells from individuals with a normal immune response. Notably, NK cell-mediated immunity was found to be significantly reduced in AIDS patients with progressing KS as compared to both HIV-negative patients with indolent classic KS or normal blood donors. However, it was restored after treatment with the highly active antiretroviral therapy (HAART) in AIDS-KS patients, that showed regression and clearance of HHV8 from PBMC. By contrast, AIDS-KS patients with a more aggressive disease and no clinical response had persistent HHV8 viremia associated with reduced NK cell cytotoxicity. These results suggest a key role for NK cells in the control of HHV8 latent infection, KS development, and in disease remission upon HAART.