NBS1 Research Articles

Clinical measurement of cellular DNA damage hypersensitivity in patients with DNA repair defects

BackgroundDNA repair deficiency disorders are rare inherited diseases arising from pathogenic (disease-causing) variants in genes involved in DNA repair. There are no standardized diagnostic assays for the investigation of pathological significance of unknown variants in DNA repair genes. We hypothesized that our assays for measuring in vitro patient blood cell hypersensitivity to DNA-damaging agents can be used to establish the pathological significance of unknown variants in DNA repair genes. Six patients with variants in the DNA repair genes PRKDC (two siblings), DCLRE1C (two siblings), NBN, and MSH6 were included. Here, we used the cell division assay (CDA) and the γ-H2AX assay, which were both developed and clinically validated by us, to measure patient cell hypersensitivity in response to ionizing radiation, mitomycin C, cytarabine and doxorubicin.ResultsRadiation hypersensitivity was detected in the two patients with variants in the PRKDC gene (p < 0.0001 for both at 3.5 Gy), and the two patients with DCLRE1C variants (p < 0.0001 at 3.5 Gy for sibling 1 and p < 0.0001 at 1 Gy for sibling 2). The cells from the patients with the PRKDC variant were also deficient in removing γ-H2AX (p < 0.001). The cells from the patient with variants in the NBN gene were hypersensitive to mitomycin C (p = 0.0008) and deficient in both induction and removal of γ-H2AX in response to radiation.ConclusionsThe combination of the CDA and the γ-H2AX assay is useful in investigating the significance of unknown variants in some DNA repair genes.

Importance of Diagnosis in Breast Cancer with Non-BRCA Pathogenic Germline Variants of Cancer Susceptibility Genes using High-Throughput Sequencing Analysis

Objectives: The aim was to point out the importance of the diagnosis rate of breast cancer (BC) by analyzing the cancer predisposition genes except BRCA1/2 with multigene testing. Methods: In this study, 232 non-BRCA cases with BC and/or BC family history (FH) were analyzed using the next-generation sequencing method. Results: Twenty-two different pathogenic/likely pathogenic variants were determined in 24 (10.34%) of cases, and these variants were detected in the CHEK2 (7/24, 29.1%), ATM (5/24, 20.8%), MUTYH (3/24, 12.5%), BLM (2/24, 8.3%), WRN (2/24, 8.3%), TP53 (1/24, 4.1%), BRIP1 (1/24, 4.1%), MSH2 (1/24, 4.1%), NBN (1/24, 4.1%), and PTEN (1/24, 4.1%) genes including three novel variants which were identified in the BLM, ATM, and MSH2 (3/22, 13.6%) genes. Fourteen of 24 (58.3%) cases had BC diagnosis, and 10 of 24 (41.6%) cases had a FH of BC. Conclusion: Among non-BRCA BC and/or BC FH cases, cancer susceptibility gene frequency was 10.34% in this study. CHEK2 and ATM genes had relatively high mutation rates. Keywords: Breast cancer, Cancer susceptibility, Non-BRCA1/2, Targeted gene analysis

Effects of probiotic feed additives (biosol and Zemos) on growth and related genes in broiler chickens

The goal of this study was to look into the effect of two probiotics as feed additives: Biosol (lipotropic factor containing probiotics) and/or Zemos, and to determine the best regimen for using both probiotics to improve growth performance, carcass traits, blood parameters, and transcription of some growth and immunity-related genes in broiler chickens. A commercial hatchery provided 400 one-day-old Cobb broiler chicks. The chicks were divided into four equal groups at random, each with 100 birds. Group 1 served as a negative control. Group 2 was given Probiotic 1(Biosol) in drinking water at a dose of 120 g/10,000 chick each day. Group 3 received probiotic 2 (Zemos) at a dose of 0.25 mL/L of drinking water for 3 days/week. Group 4 was given combined probiotics (Biosol + Zemos), with the same weekly dose of each probiotic alternatively. During the experiment, all chickens were fed and hydrated ad libitum and exposed to 24 h of light. Our findings revealed that chicks fed on diets supplemented with Biosol and Biosol + Zemos showed an increase in body weight gain and lower feed conversion rates. Total protein levels were elevated while cholesterol and triglyceride levels dropped, whereas, no effect was detected in the uric acid levels among those groups. Furthermore, the carcass traits of Biosol and Biosol + Zemos increased as a result of the dietary supplementations, however, the weight of the drumstick showed no difference among the four groups. Both probiotics influenced the cecal count of lactobacilli, total aerobes, E. coli, and Enterococci in, as well as the transcript levels of the MTOR, SMYD, TLR-4, and NBN genes. It is possible to conclude that supplementing broilers with probiotics and Biosol, in particular, can increase their growth performance, and improve the biochemical characteristics of the blood and transcript levels of the genes under investigation. Highlights Supplementation of the broiler diet with probiotics is essential Biosol and Zemos increased the growth performance Both probiotics improved the biochemical blood parameters Biosol up-regulated growth and immunity related genes

The Effect of Gene Mutations on Metastasis and Overall Survival in Metastatic and Nonmetastatic Colon Cancers.

Objective:It is known that many genes are associated with colon cancer. We aimed to investigate the effect of gene mutations on metastasis and overall survival in metastatic and non metastatic colon cancers. Methods:A total of 50 patients with metastatic (n=25) and non metastatic (n=25) diagnosed with colon cancer between 2010 and 2018 were included in the study. APC, MUTYH, RAD50, MEN1, ATM, PALB2, NSH2, BRCA1, BRCA2, MLH1, BRIP1, TP53, PTEN, BARD1, MSH6, PMS2, NBN, and FAM175A gene mutations were evaluated using the next generation sequencing method. The effect of gene mutations on metastasis and overall survival were evaluated. Results:The mean age of patients with colon cancer without distant metastasis was 48.64±14.72 years and for patients with distance metases was 56.68±11.65. The mean survival time of colon cancer patients with distant organ metastasis after the metastasis date was 104.36±58.59 weeks. The presence of APC, MUTYH, and TP53 genetic mutations was observed with a higher rate in metastatic colon cancer (p<0.05). Conclusion:We showed that APC, MUTYH, and TP53 mutations are associated with distant organ metastasis.

Exonic sequencing and MLH3 gene expression analysis of breast cancer patients.

Breast cancer is the most common cancer in women worldwide. Detection of breast cancer susceptibility genes is an important issue. Also, MLH3 is a DNA mismatch repair gene and mutation in this gene is harmful in different cancers. This study aimed to use exome sequencing to uncover previously undetected breast cancer-predisposing variants. Also, we investigated the MLH3 gene expression of breast cancer patients which can be a breast cancer susceptibility gene. A total of 80 samples including 40 paired normal and cancer tissue samples were collected at Zheen International Hospital, Erbil, Iraq. Exome sequencing was used to identify mutations. Different in silico tools were used to predict the effect of mutation on the structural features or protein function. Real-time PCR was used for assessing the expression of MLH3 in breast cancer patients. We identified 26 variants in breast cancer patients, 22 inherited variants were found in MLH3, CHECK2, BRCA1, BRCA2, BLM, TP53, MSH6, NBN and PTEN genes and 4 somatic variants were found in PALB2, RAD50 and RBM10 genes. It was found that the expression of the MLH3 gene in tumor samples was significantly down-regulated compared with normal tissues. Statistically, high significance was found. The decreased expression of MLH3 was significant in all ranges of ages and all breast cancer types. Also, the expression of MLH3 decreased significantly in patients with breast cancer grades of II and III. In conclusion, MLH3 can be used as a susceptibility gene especially in grades II and III of breast cancer.

Diagnostic and therapeutic approach to children with Nijmegen breakage syndrome in relation to development of lymphoid malignancies.

Nijmegen breakage syndrome (NBS) is a rare chromosomal instability disorder. The majority of patients carry founder mutation in the NBN gene (c.657_661del5). Characteristic features of the NBS include progressive microcephaly, dysmorphic facial features, immunodeficiency, and high predisposition to malignancy with cumulative cancer incidence by the age of 20 years, and amounted to over 70%. The aim of study is to present the latest methods of diagnosis, potential cancer risk factors and treatment of lymphoid malignancies in children with NBS. To review the evidence using PubMed and Google Scholar search which included articles published between 2009-2021, focusing on articles published between 2013-2021. The average delay in diagnosis of NBS ranges from 4-5 years. Neonatal screening of T-cell excision circles (TRECs) and kappa-deleting recombination excision circles (KRECs) seems favourable in NBS. There are no specific protocols for the treatment of lymphoid malignancies in children with NBS, and full- dose chemotherapy is the most frequently applied method. Reducing the doses of chemotherapy does not significantly reduce the toxicity. Main cause of death is cancer progression and treatment-related mortality mostly associated with infectious complications. Patients with diagnosed cancer who received haematopoietic stem cell transplantation (HSCT) had significantly higher 20-year OS than those who did not (42.7% vs. 30.3%). Further meta-analysis is essential to establish the best monitoring and treatment regimen in patients with NBS and lymphoid malignancies.

Neuroprotective Effect of Moringa Oleifera Extract on Acetamiprid Induced Neurotoxicity and Apoptosis in Albino Rats

Acetamiprid (ACMP) can cause neurotoxicity and induces neuronal apoptosis. The ameliorative effects ofMoringa Oleifera (MO) seed extract against the neurotoxicity induced by ACMP in adult male albino ratswere investigated. Forty male albino rats were divided into 4 equal groups. The first group served as a control,the second group administered ACMP 26.8mg/kg b.w. (representing 1/10 LD50) by gavage. The third groupwas given combination of ACMP 26.8 mg/kg b.w. and MO seed extract 150 mg/kg b.w. the fourth group wasgiven MO extract only 150 mg/kg b.w. for 28 consecutive days. Results proved the efficacy of MO extractas a neuroprotective agent through the reduction of the oxidant parameter malondialdhyde (MDA) contentby 26.9%, significant increase of reduced glutathione content (GSH) by 21.4%, and elevation of catalase(CAT) and superoxide dismutase (SOD) activities by (62.2% and 40.8%) respectively, compared to ACMPintoxicated group. Consequently, gene expression analysis for Bax and NBN genes was downregulatedby 54.5% and 36.3% respectively in MO treated rats (group III) compared with ACMP intoxicated group.Moreover, restoration of the pathological tissue injuries was noticed. In conclusion, MO proved to be aneffective neuroprotective and anti-apoptotic agent against ACMP-induced toxicity.

New Approach for Detection of Normal Alternative Splicing Events and Aberrant Spliceogenic Transcripts with Long-Range PCR and Deep RNA Sequencing.

Simple SummaryRNA splicing defects, caused by genetic variants, are a common molecular mechanism of disease. To detect variants that cause splicing impairment, mRNA-based studies must be performed. Classical mRNA assays are time-consuming, which is why we have validated a new reliable straightforward approach to detect normal alternative splicing events and also splicing aberrations. Using our approach, we were able to reclassify three variants of uncertain significance in NBN and STK11 genes, which is of great importance for a proper clinical management of the patients.RNA sequencing is a promising technique for detecting normal and aberrant RNA isoforms. Here, we present a new single-gene, straightforward 1-day hands-on protocol for detection of splicing alterations with deep RNA sequencing from blood. We have validated our method’s accuracy by detecting previously published normal splicing isoforms of STK11 gene. Additionally, the same technique was used to provide the first comprehensive catalogue of naturally occurring alternative splicing events of the NBN gene in blood. Furthermore, we demonstrate that our approach can be used for detection of splicing impairment caused by genetic variants. Therefore, we were able to reclassify three variants of uncertain significance: NBN:c.584G>A, STK11:c.863-5_863-3delCTC and STK11:c.615G>A. Due to the simplicity of our approach, it can be incorporated into any molecular diagnostics laboratory for determination of variant’s impact on splicing.

Expression Levels of the TERT Gene Can be Used to Risk Stratify Aplastic Anemia Patients: Study in an Indian Cohort

Introduction: Acquired Aplastic anemia (AA) is a bone marrow failure syndrome characterized by pancytopenia and marrow hypoplasia, and is mediated by immune destruction of hematopoietic stem cells. Mutations in several genes including telomerase, a ribonucleoprotein enzyme complex, consisting of a reverse transcriptase enzyme (TERT), an RNA template (TERC), and several stabilizing proteins, and the associated shelterin complex have been found in cases of both congenital and idiopathic AA. In particular, several TERT and TER C mutations reduce telomerase activity in vitro and accelerate telomere attrition in vivo . Shortened telomeres have been observed in a third of idiopathic AA patients, but only 10% of these patients have mutations in genes of the telomerase complex. We have previously demonstrated that in addition to keeping telomeres from shortening, telomerase directly regulates transcriptional programs of developmentally relevant genes (Ghosh et al, Nat Cell Biol, 2012). We postulate that changes in expression of telomerase associated genes, specifically TERT, contribute to the etiology of aplastic anemia.Patient cohort and Methods: In an effort to better understand the molecular and clinical correlates of this disease, 24 idiopathic AA patient blood samples were collected at a tertiary medical center in Bangalore, India. After obtaining informed consent, high throughput exome panel sequencing on an Illumina HiSeq 2500 platform of the 24 samples was performed, followed by variant calling, annotation and filtration of the generated data to reveal rare pathogenic variants. Additionally, RT-PCR analysis of harvested RNA from each patient was carried out and expression of levels of TERT measured in comparison to samples from normal individuals.Results: Mutational profiling of 24 AA patients by exome sequencing revealed a total number of 2837 non- synonymous mutations (MAF < 0.01%). These included nonsense, frame shift, splice site, missense and in-frame mutations. Recurrent gene mutations were identified in 129 genes. However, pathway analysis of these gene mutations was uninformative with respect to disease etiology. Since DNA mutational data from the AA patients was unrevealing, and given that AA is a telomeropathy, we next chose to measure RNA expression levels of the TERT gene, an integral component of the Telomerase complex. RT-PCR of TERT expression compared to that of normal samples (n=6) revealed an 8 fold reduction in TERT expression 17/24 patients, while 7/24 patients maintained normal TERT expression.Examination of the average number of gene mutations in the TERT-normal versus the TERT-Low patients revealed that the mutational burden was significantly higher in TERT-normal (166 mutations) compared to TERT-low (112 mutations) expressers (p=0.04). High throughput sequencing data also revealed that while TERT-normal patients had no mutations in genes associated with the telomerase/shelterin complex, TERT-low patients had mutations in TERT, TEP1, TINF2, NBN, TPP1, HSP90A and POT1 genes: all associated with the telomerase complex, most of which were predicted to be pathogenic variants. TERT-low patients also showed more evidence of clonal evolution (multiple patients with mutations in TET2, and in BCORL1, FLT-3, MLP, BRAF genes) compared to TERT-normal patients.Clinically TERT-low patients were younger in age (average age 29) compared with the TERT-normal patients (average age 40). TERT-low patients were more likely to have severe aplastic anemia (SAA) leading to higher mortality and poorer response to therapy, with 6/17 patients dying and 4/17 not responding to ATG therapy.Summary and Conclusions: Our observations suggest that expression of TERT can serve as a risk stratification marker in patients with AA. In spite of the higher mutational burden in TERT-normal patients (probably due to their comparative advanced age) TERT-low patients had poorer clinical outcomes which correlated with mutations in the TERT/Shelterin complex and with mutations in genes associated with clonal evolution. While these findings need to be validated in larger cohorts of patients, we suggest that evaluation of TERT expression levels in newly diagnosed AA patients could serve to risk stratify these patients and influence the choice of therapy administered. DisclosuresNo relevant conflicts of interest to declare.

Genomic Correlates of DNA Damage in Breast Cancer Subtypes.

Simple SummaryBreast cancer (BC) is the most common invasive tumor in women and the second leading cause of cancer-related death. Therefore, identification of druggable targets to improve current therapies and overcome resistance is a major goal. In this work, we performed an in silico analysis of transcriptomic datasets in breast cancer, and focused on those involved in DNA damage, as were clearly upregulated using gene set enrichment analyses (GSEA), particular the following pathways: ATM/ATR, BARD1 and Fanconi Anemia. BHLHE40, RFWD2, BRIP1, PRKDC, NBN, RNF8, FANCD2, RAD1, BLM, DCLRE1C, UBE2T, CSTF1, MCM7, RFC4, YWHAB, YWHAZ, CDC6, CCNE1, and FANCI genes were amplified/overexpressed in BC, and correlated with detrimental prognosis. Finally, we selected the best transcriptomic signature of genes within this function that associated with clinical outcome to identify functional genomic correlates of outcome.Among the described druggable vulnerabilities, acting on the DNA repair mechanism has gained momentum, with the approval of PARP inhibitors in several indications, including breast cancer. However, beyond the mere presence of BRCA1/BRCA2 mutations, the identification of additional biomarkers that would help to select tumors with an extreme dependence on DNA repair machinery would help to stratify therapeutic decisions. Gene set enrichment analyses (GSEA) using public datasets evaluating expression values between normal breast tissue and breast cancer identified a set of upregulated genes. Genes included in different pathways, such as ATM/ATR, BARD1, and Fanconi Anemia, which are involved in the DNA damage response, were selected and confirmed using molecular alterations data contained at cBioportal. Nineteen genes from these gene sets were identified to be amplified and upregulated in breast cancer but only five of them NBN, PRKDC, RFWD2, UBE2T, and YWHAZ meet criteria in all breast cancer molecular subtypes. Correlation of the selected genes with prognosis (relapse free survival, RFS, and overall survival, OS) was performed using the KM Plotter Online Tool. In last place, we selected the best signature of genes within this process whose upregulation can be indicative of a more aggressive phenotype and linked with worse outcome. In summary, we identify genomic correlates within DNA damage pathway associated with prognosis in breast cancer.

Geographical Distribution, Incidence, Malignancies, and Outcome of 136 Eastern Slavic Patients With Nijmegen Breakage Syndrome and NBN Founder Variant c.657_661del5

Nijmegen breakage syndrome (NBS) is a DNA repair disorder characterized by combined immunodeficiency and a high predisposition to lymphoid malignancies. The majority of NBS patients are identified with a homozygous five base pair deletion in the Nibrin (NBN) gene (c.657_661del5, p.K219fsX19) with a founder effect observed in Caucasian European populations, especially of Slavic origin. We present here an analysis of a cohort of 136 NBS patients of Eastern Slav origin across Belarus, Ukraine, Russia, and Latvia with a focus on understanding the geographic distribution, incidence of malignancy, and treatment outcomes of this cohort. Our analysis shows that Belarus had the highest prevalence of NBS (2.3 per 1,000,000), followed by Ukraine (1.3 per 1,000,000), and Russia (0.7 per 1,000,000). Of note, the highest concentration of NBS cases was observed in the western regions of Belarus and Ukraine, where NBS prevalence exceeds 20 cases per 1,000,000 people, suggesting the presence of an “Eastern Slavic NBS hot spot.” The median age at diagnosis of this cohort ranged from 4 to 5 years, and delay in diagnosis was more pervasive in smaller cities and rural regions. A total of 62 (45%) patients developed malignancies, more commonly in males than females (55.2 vs. 34.2%; p=0.017). In 27 patients, NBS was diagnosed following the onset of malignancies (mean age: 8 years). Malignancies were mostly of lymphoid origin and predominantly non-Hodgkin lymphoma (NHL) (n=42, 68%); 38% of patients had diffuse large B-cell lymphoma. The 20-year overall survival rate of patients with malignancy was 24%. However, females with cancer experienced poorer event-free survival rates than males (16.6% vs. 46.8%, p=0.036). Of 136 NBS patients, 13 underwent hematopoietic stem cell transplantation (HSCT) with an overall survival of 3.5 years following treatment (range: 1 to 14 years). Indications for HSCT included malignancy (n=7) and immunodeficiency (n=6). Overall, 9% of patients in this cohort reached adulthood. Adult survivors reported diminished quality of life with significant physical and cognitive impairments. Our study highlights the need to improve timely diagnosis and clinical management of NBS among Eastern Slavs. Genetic counseling and screening should be offered to individuals with a family history of NBS, especially in hot spot regions.

Mutational landscape of primary and recurrent Ewing sarcoma

IntroductionEwing sarcoma (ES) is a highly aggressive malignancy of bone and soft tissues characterized by the presence of a genetic fusion involving the EWSR1 gene. More than one-third of patients develop distant metastases, which are associated with unfavorable prognosis. Knowledge about the disease’s genetic landscape may help foster progress in using targeted therapies in the treatment of ES.Aim of the studyThe objective is to assess the mutational landscape of ES in pretreatment samples, tumor samples after neoadjuvant chemotherapy, and in metastatic/recurrent tumors in children and adultsMaterial and methodsDNA from 39 formalin-fixed paraffin-embedded tumor samples of 22 patients (17 adults, 5 children) were analyzed by targeted next generation sequencing (NGS) using the Oncomine Comprehensive Assay v3gene panel. Additional functional analyses were performed between patient subgroups.ResultsAll samples were characterized by low tumor mutation burden (< 10 mut/Mb). The most commonly mutated genes were PIK3R1 (59%) and POLE (50%). The most widely detected variants in biopsy samples were PIK3R1 T369I (50%), FGFR1 E159K, and TP53 at codon 72 (both in 27.3%). Additionally, the ATR,BRCA1, RAD50,ATM,CHEK1, and NBN genes showed a significantly higher number of mutations in ES. Mutations in PIK3R1 were significantly more frequent in adults, while mutations in the pathways responsible for cell cycle control, DNA repair, and transcriptional regulation were more frequent in children.ConclusionsBesides EWSR1 fusion, ES is characterized by numerous point mutations that are potential targets for precision medicine. There is high genomic heterogeneity that may explain differences in outcomes between patient subgroups.

Liver Cirrhosis in Chronic Hepatitis B Patients Is Associated with Genetic Variations in DNA Repair Pathway Genes.

Simple SummaryAs DNA repair enzymes affect dynamics of liver damage and are involved in HBV viral replication, this study focused on the role of genetic variations within genes representing key DNA-repair pathways in HBV-induced liver cirrhosis. The obtained results have demonstrated that SNPs within XRCC1, ERCC2 genes may confer susceptibility to liver cirrhosis in chronic hepatitis B patients. Liver cirrhosis (LC), contributing to more than 1 million of deaths annually, is a major healthcare concern worldwide. Hepatitis B virus (HBV) is a major LC etiological factor, and 15% of patients with chronic HBV infection (CHB) develop LC within 5 years. Recently, novel host genetic determinants were shown to influence HBV lifecycle and CHB course. DNA repair enzymes can affect dynamics of liver damage and are involved in HBV covalently closed circular DNA (cccDNA) formation, an essential step for viral replication. This study aimed to evaluate the possible role of genes representing key DNA-repair pathways in HBV-induced liver damage. MALDI-TOF MS genotyping platform was applied to evaluate variations within XRCC1, XRCC4, ERCC2, ERCC5, RAD52, Mre11, and NBN genes. Apart from older age (p < 0.001), female sex (p = 0.021), portal hypertension (p < 0.001), thrombocytopenia (p < 0.001), high HBV DNA (p = 0.001), and high aspartate aminotransferase (AST) (p < 0.001), we found that G allele at rs238406 (ERCC2, p = 0.025), T allele at rs25487 (XRCC1, p = 0.012), rs13181 GG genotype (ERCC2, p = 0.034), and C allele at rs2735383 (NBN, p = 0.042) were also LC risk factors. The multivariate logistic regression model showed that rs25487 CC (p = 0.005) and rs238406 TT (p = 0.027) were independently associated with lower risk of LC. This study provides evidence for the impact of functional and potentially functional variations in key DNA-repair genes XRCC1 and ERCC2 in HBV-induced liver damage in a Caucasian population.

Mutations in ATM, NBN and BRCA2 predispose to aggressive prostate cancer in Poland

In designing national strategies for genetic testing, it is important to define the full spectrum of pathogenic mutations in prostate cancer (PCa) susceptibility genes. To investigate the frequency of mutations in PCa susceptibility genes in Polish familial PCa cases and to estimate gene-related PCa risks and probability of aggressive disease, we analyzed the coding regions of 14 genes by exome sequencing in 390 men with familial prostate cancer and 308 cancer-free controls. We compared the mutation frequencies between PCa cases and controls. We also compared clinical characteristics of prostate cancers between mutation carriers and noncarriers. Of the 390 PCa cases, 76 men (19.5%) carried a mutation in BRCA1, BRCA2, NBN, ATM, CHEK2, HOXB13, MSH2 or MSH6 genes. No mutations were found in BRIP1, PTEN, TP53, MLH1, PMS2 and SPOP. Significant associations with familial PCa risk were observed for CHEK2, NBN, ATM, and HOXB13. High-grade (Gleason 8-10) tumors were seen in 56% of BRCA2, NBN or ATM carriers, compared to 21% of patients who tested negative for mutations in these genes (OR = 4.7, 95% CI 2.0-10.7, P = .0003). In summary, approximately 20% of familial prostate cancer cases in Poland can be attributed to mutations in eight susceptibility genes. Carriers of mutations in BRCA2, NBN and ATM develop aggressive disease and may benefit from intensified screening and/or chemotherapy.

Successful pregnancy after complete resection of leiomyomatosis peritonealis disseminate without recurrence:a case report with next-generation sequencing analysis and literature review

BackgroundPeritoneal leiomyomatosis disseminate (LPD) is a rare disease characterized by widespread dissemination of leiomyomas nodules throughout the peritoneal and omental surfaces. Reports of pregnancy with LPD are even rarer. Therefore, there is no clear consensus on the treatment of LPD on pregnancy, and the pathogenesis is still unclear.Case presentationWe reported a case of LPD patient who developed during pregnancy. The patient underwent a cesarean section at 32 weeks of gestation while removing all visible tumors, and no LPD lesions were seen in the subsequent cesarean section at full term. NGS of LPD lesions detected 4 mutations with focal high-level amplifications of CDK4 (cyclin-dependent kinases 4), NBN (Nibrin), DAXX (death domain associated protein), and MYC (myelocytomatosis oncogene). Immunohistochemistry staining analysis among benign leiomyoma, LPD, and leiomyosarcoma verified that LPD was an unusual intermediate between benign and malignant uterine smooth muscle tumors. Besides, LPD is a hormonal-dependent leiomyoma. After a detailed literature search, we summarized the detailed clinical features and follow-up information of patients with LPD during pregnancy.ConclusionsThis is the first reported LPD case of successful term pregnancy without recurrence, following resection of all visible lesions in a prior pregnancy. LPD is an unusual intermediate between benign and malignant uterine smooth muscle tumors.

A case of Nijmegen syndrome in a child with frequent relapses of a respiratory infection

The article provides clinical dynamic observation of a child with a primary immunodeficiency state, presents the stages of the differential diagnostic search for congenital genetic pathology. Nijmegen syndrome is an autosomal recessive disease caused by a mutation of the NBN gene, characterized by microcephaly, Bird facial features, delayed physical development, immunodeficiency, congenital malformations, increased sensitivity to x-ray radiation and a high susceptibility to cancer. This syndrome is relevant for Russian pediatricians due to the high frequency of carriage of the Slavic NBN gene mutation among the population of the Russian Federation. The presented clinical observation describes a patient with characteristic phenotypic features, frequent episodes of acute respiratory viral infections (up to 8 times a year) with the addition of a bacterial infection, fever up to febrile numbers. The duration of verification of the immunodeficiency state was 5 years. Despite the complicated course of infectious diseases, the general well-being of the child did not suffer significantly, the level of neuropsychic development corresponded to age. The diagnosis of Nijmegens syndrome was made on the basis of characteristic phenotypic features, anamnesis, and laboratory data, instrumental studies and confirmed by the results of molecular genetic analysis.

Pathogenic Variants in Cancer Predisposition Genes and Prostate Cancer Risk in Men of African Ancestry.

In studies of men of European ancestry, rare pathogenic variants in DNA repair pathway genes have been shown to be associated with risk of aggressive prostate cancer. The contribution of rare coding variation to prostate cancer risk in men of African ancestry has not been established. We sequenced a panel of 19 DNA repair and cancer predisposition genes in 2,453 African American and 1,151 Ugandan prostate cancer cases and controls. Rare variants were classified as pathogenic or putatively functionally disruptive and examined in association with prostate cancer risk and disease aggressiveness in gene and pathway-level association analyses. Pathogenic variants were found in 75 out of 2,098 cases (3.6%) and 31 out of 1,481 controls (2.1%) (OR=1.82, 95% CI=1.19 to 2.79, P=0.0044) with the association being stronger for more aggressive disease phenotypes (OR=3.10, 95% CI=1.54 to 6.23, P=0.0022). The highest risks for aggressive disease were observed with pathogenic variants in the ATM, BRCA2, PALB2 and NBN genes, with odds ratios ranging from ~4 to 15 in the combined study sample of African American and Ugandan men. Rare, non-pathogenic, non-synonymous variants did not have a major impact on risk of overall prostate cancer or disease aggressiveness. Rare pathogenic variants in DNA repair genes have appreciable effects on risk of aggressive prostate cancer in men of African ancestry. These findings have potential implications for panel testing and risk stratification in this high-risk population.

Radiographic Surveillance of Patients with Non-BRCA1/2 Pathogenic Variants.

The National Comprehensive Cancer Network (NCCN) developed clinical practice guidelines for germline pathogenic variants in highly penetrant genes, such asTP53andPTEN, and in moderately penetrant genes, such asCHEK2, ATM andPALB2. Whether the practice of radiographic surveillance of patients with pathogenic variants in genes other than BRCA1/2 complies with current NCCN guidelines remains unclear. Retrospective review of patients identified with pathogenic variants in genes other than BRCA1/2 from 2007 through 2017 to determine if radiographic surveillance was in accordance with NCCN guidelines for mammography and consideration of magnetic resonance imaging (MRI). Exclusions included variants of unknown significance, pathogenic variants not associated with an increased risk of breast cancer, and previous breast cancer diagnosis. After exclusions, 35 patients with pathogenic variants in ATM, CDH1, CHEK2, NBN, PALB2, PTEN, and STK11 genes were reviewed to assess whether radiographic surveillance was in accordance with NCCN guidelines. Guidelines for those with variants in ATM, CHEK2 and NBN includes annual mammography with tomosynthesis and consideration of breast MRI at age 40, variants in CDH1 and PALB2 at age 30, variants in PTEN at age 30-35 or 5-10 years before the earliest family breast cancer, and variants in STK11 at age 25. Of these 35 patients, 11 (31%) received mammography only; 11 (31%) received mammography and MRI, and 13 (37%) received no radiographic surveillance. Two of the 35 (6%) patients who received radiographic surveillance were diagnosed with ductal carcinoma in situ or invasive breast cancer. Thirty-one percent of patients with pathogenic variants in genes other than BRCA1/2 received both mammography and MRI. Thirty-seven percent of patients with these highly penetrant and moderately penetrant genes received no radiographic follow-up, clearly demonstrating an opportunity for improvement.

Challenges in reporting pathogenic/potentially pathogenic variants in 94 cancer predisposing genes - in pediatric patients screened with NGS panels

The benefit of reporting unsolicited findings in Next Generation Sequencing (NGS) related to cancer genes in children may have implications for family members, nevertheless, could also cause distress. We aimed to retrospectively investigate germline variants in 94 genes implicated in oncogenesis, in patients referred to NGS testing for various rare genetic diseases and reevaluate the utility of reporting different classes of pathogenicity. We used in silico prediction software to classify variants and conducted manual review to examine unsolicited findings frequencies in 145 children with rare diseases, that underwent sequencing - using a 4813 gene panel. The anonymized reanalysis revealed 18250 variants, of which 126 were considered after filtering. Six pathogenic variants (in BRCA1,BMPR1A,FANCA,FANCC,NBN genes) with cancer related phenotype and three unsolicited variants (in BRCA2,PALB2,RAD50 genes) were reported to patients. Additionally, three unsolicited variants in ATR, BLM (in two individuals), and FANCB genes presented potential cancer susceptibility, were not reported to patients. In retrospect, 4.8% (7/145) of individuals in our cohort had unsolicited NGS findings related to cancer. More efforts are needed to create an updatable consensus in reporting variants in cancer predisposing genes, especially for children. Consent process is crucial to inform of both value and risk of additional genetic information.

Nijmegen breakage syndrome in two half sibs with peripheral T-cell lymphoma and cortical T-cell acute lymphoid leukemia.

Nijmegen breakage syndrome (NBS) is a rare autosomal recessive disorder, characterized by spontaneous chromosomal instability with predisposition to immunodeficiency and cancer.We present a repeated NBS in two sons from one woman after two marriages. We describe the clinical data, cytogenetic, and molecular findings of a prenatally diagnosed fetus, and his brothers with NBS. The first patient developed peripheral T-cell lymphoma at the age of 16 years and died 5 months after the protocol start. The diagnosis of NBS was established after his death. The second patient was born after the fifth pregnancy, third delivery in the second marriage; he developed cortical T-cell leukemia at the age of 3 years, received hematopoietic stem cells transplantation (HSCT) and he is alive now. In a year after repeated NBS case in this family, mother became pregnant again and the mutation was detected in the male fetus after the prenatal diagnosis; the pregnancy was aborted. At the age of 41 years, mother’s seventh pregnancy finished by miscarriage. In three months, she was pregnant again, only one mutation in NBN gene was detected during the prenatal diagnostics in the female fetus; healthy female was born at term.To our knowledge, this is the first time to describe the repeated cases of two patients born with Nijmegen breakage syndrome from one mother and two different fathers. This case highlights the value of checking NBN carrier in Belarusian families during genetic counselling.