Mutational screening of affected cardiac tissues and peripheral blood cells identified novel somatic mutations in GATA4 in patients with ventricular septal defect

The aim of this study was to examine how somatic mutations of the GATA4 gene contributed to the genesis of ventricular septal defect （VSD）. The coding and intron-exon boundary regions of GATA4 were sequenced of DNA samples from peripheral blood cells and cardiac tissues of twenty surgically treated probands with VSD. Seven novel heterozygous variants were detected in cardiac tissues from VSD patients, but they were not detected in the peripheral blood cells of VSD patients or in 500 healthy control samples. We replicated 14 single nucleotide polymorphisms （SNPs） reported in NCBI. Bioinformatics analysis was performed to analyze the possible mechanism by which mutations were linked to VSD. Among those variants, c. 1004C〉A （p.S335X） occurred in the highly conserved domain of GATA4 and generated a termination codon, which led to the production of truncated GATA4. The seven novel heterozygous GATA4 mutations were only identified in cardiac tissues with VSD, suggesting that they are of somatic origin. A higher mutation rate in cardiac tissues than in peripheral blood cells implies that the genetic contribution to VSD may have been underestimated.

Failure to Identify Somatic Mutations in Monozygotic Twins Discordant for Schizophrenia by Whole Exome Sequencing

Background： Schizophrenia （SCZ） is a severe, debilitating, and complex psychiatric disorder with multiple causative factors. An increasing number of studies have determined that rare variations play an important role in its etiology. A somatic mutation is a rare form of genetic variation that occurs at an early stage of embryonic development and is thought to contribute substantially to the development of SCZ. The aim of the study was to explore the novel pathogenic somatic single nucleotide variations （SNVs） and somatic insertions and deletions （indels） of SCZ. Methods： One Chinese family with a monozygotic （MZ） twin pair discordant for SCZ was included. Whole exome sequencing was performed in the co-twin and their parents. Rigorous filtering processes were conducted to prioritize pathogenic somatic variations, and all identified SNVs and indels were further confirmed by Sanger sequencing. Results： One somatic SNV and two somatic indels were identified after rigorous selection processes. However, none was validated by Sanger sequencing. Conclusions： This study is not alone in the failure to identify pathogenic somatic variations in MZ twins, suggesting that exonic somatic variations are extremely rare. Further efforts are warranted to explore the potential genetic mechanism of SCZ.

Mutational separation and clinical outcomes of TP53 and CDH1 in gastric cancer

BACKGROUND Gastric cancer(GC)is a deadly tumor with the fifth highest occurrence and highest global mortality rates.Owing to its heterogeneity,the underlying mechanism of GC remains unclear,and chemotherapy offers little benefit to individuals.AIM To investigate the clinical outcomes of TP53 and CDH1 mutations in GC.METHODS In this study,202 gastric adenocarcinoma tumor tissues and their corresponding normal tissues were collected.A total of 490 genes were identified using target capture.Through t-test and Wilcoxon rank-sum test,somatic mutations,microsatellite instability,and clinical statistics,including overall survival,were detected,compared,and calculated.RESULTS The mutation rates of 32 genes,including TP53,SPEN,FAT1,and CDH1 exceeded 10%.TP53 mutations had a slightly lower overall occurrence rate(33%).The TP53 mutation rate was significantly higher in advanced stages(stage Ⅲ/Ⅳ)than that in early stages(stage Ⅰ/Ⅱ)(P<0.05).In contrast,CDH1 mutations were significantly associated with diffuse GC.TP53 is related to poor prognosis of advanced-stage tumors;nevertheless,CDH1 corresponds to a diffuse type of cancer.TP53 is exclusively mutated in CDH1 and is primarily affected by two distinct GC mechanisms.CONCLUSION Different somatic mutation patterns in TP53 and CDH1 indicate two major mechanisms of GC.

Somatic TP53 mutations and comparison of different TP53 functional domains in human cancers:data analysis from the IARC TP53 database and the National Cancer Institute GDC data portal

P53 gene mutations have been known to be highly related to the majority of human cancers.The colocation of biologists and bioinformaticians have constructed many databases for cancer research.Although the relationship between the presence of TP53 mutation and cancers has been reported in various studies,few reports TP53 mutation distribution in different functional domains.Hence,we use 2 databases(The TP53 Mutation Database of the International Agency for the Research on Cancer and The Genomic Data Commons data portal)to compare survival rate with and without TP53 mutations in a certain cancer,as well as to find most frequent mutation sites in different functional domains of the TP53 protein.Our study shows that most somatic mutations of TP53 and high mutation rate sites are concentrated in the DNA-binding domain,and the survival of certain cancers varies with and without P53.

The emerging role of somatic tumor sequencing in the treatment of urothelial cancer

The development of rapid genome sequencing has greatly enhanced our understanding of the molecular biology underlying many malignancies.Whole exome sequencing has highlighted the individualistic nature of malignancies on a patient-to-patient basis and begun to revolutionize therapeutic approaches.In recent years,whole genome sequencing of urothelial malignancies has identified a host of somatic mutations which contribute to growth,progression,and metastasis of urothelial carcinoma of the bladder and upper tract urothelial carcinoma.As genetic sequencing continues,additional targets will be identified,allowing development of novel therapeutic agents targeting cancer on a molecular level,with the goal of delivering highly individualized care based on the underlying mutational profile of the patient’s malignancy.In this review,we aim to discuss known genetic alterations of urothelial malignancy and the implications these mutations carry in terms of prognostication and development of targeted therapeutic agents.We will focus on RNA-expression profiling and genomic DNA profiling,with a focus on comprehensive whole exome and whole genome sequencing relative to selected urothelial carcinoma-associated genes and circulating tumor DNA analysis.

A new era of mutation rate analyses: Concepts and methods

The mutation rate is a pivotal biological characteristic,intricately governed by natural selection and historically garnering considerable attention.Recent advances in high-throughput sequencing and analytical methodologies have profoundly transformed our understanding in this domain,ushering in an unprecedented era of mutation rate research.This paper aims to provide a comprehensive overview of the key concepts and methodologies frequently employed in the study of mutation rates.It examines various types of mutations,explores the evolutionary dynamics and associated theories,and synthesizes both classical and contemporary hypotheses.Furthermore,this review comprehensively explores recent advances in understanding germline and somatic mutations in animals and offers an overview of experimental methodologies,mutational patterns,molecular mechanisms,and driving forces influencing variations in mutation rates across species and tissues.Finally,it proposes several potential research directions and pressing questions for future investigations.

Exome analysis for Cronkhite-Canada syndrome: A case report

BACKGROUND Cronkhite-Canada syndrome(CCS)is a rare,non-genetic disorder characterized by multiple gastrointestinal polyps,and ectodermal lesions such as alopecia,fingernail atrophy,and skin mucosal pigmentation.Unfortunately,the pathogenesis of CCS is currently unknown.CASE SUMMARY Here,we describe the case of an elderly female with diarrhea,fatigue,and hair loss,who experienced abdominal pain for over half a year and was found to have multiple gastrointestinal polyps.She was diagnosed with CCS and was treated with albumin supplementation and prednisone,and her electrolyte imbalance was corrected.Following treatment,her symptoms significantly improved.To elucidate the role of potential genetic events in the pathogenesis of CCS,we performed exome sequencing using an extract of her colorectal adenoma.CONCLUSION Our data revealed multiple somatic mutations and copy number variations.Our findings provide a novel insight into the potential mechanisms of CCS etiology.

Tumor-to-tumor metastasis of clear cell renal cell carcinoma to contralateral synchronous pheochromocytoma:A case report

BACKGROUND Tumor-to-tumor metastasis(TTM)is an uncommon condition.Only a few cases of renal cell carcinoma(RCC)as donor tumor of TTM have been reported in literature,and none of these studies have described RCC metastasizing to synchronous pheochromocytoma(PCC).CASE SUMMARY The patient was a 54-year-old woman who presented with recurrent dull abdominal pain for six months,which was further aggravated for one more month.Enhanced computed tomography revealed a tumor mass in the right kidney and another mass in the left retroperitoneum/adrenal gland.Histopathology and immunochemistry of resected specimens confirmed the diagnosis of clear cell renal cell carcinoma(CCRCC)of the right kidney,and the left retroperitoneum revealed a typical PCC with CCRCC metastasis.Whole exome sequencing revealed the presence of a c.529A>T somatic mutation of the Von Hippel Lindau(VHL)gene in the metastasized CCRCC,which was also present in the primary right kidney CCRCC,as confirmed by Sanger sequencing.No VHL mutation was detected in the PCC or in normal right kidney tissue.Fluorescence in situ hybridization revealed loss of chromosome 3p in both the primary right kidney CCRCC and CCRCC metastasized to PCC in the left kidney.CONCLUSION This is the first case showing metastasis of CCRCC to PCC,thus leading to tumorto-tumor metastasis.

Genotoxicity of Some Essential Oils Frequently Used in Aromatherapy

Genotoxic properties of the essential oils extracted from Artemisia dracunculus (tarragon), Ocimum basilicum (basil), Cinnamomum loureirii (cinnamon), Laurus nobilis (laurel), Satureja montana (savory) and Rosmarinus officinallis (rosemary) are studied by Drosophila melanogaster Somatic Mutation and Recombination Test (SMART). The high bioactivation crossed with a high cytochrome P450-dependent bioactivation capacity is used. This assay is principally based on the loss of heterozygosity of the suitable recessive markers’ multiple wing hairs (mwh) and flare-3 (flr3) which can lead to the formation of mutant clones of larval cells, and which are then going to be expressed as spots on the wings of adult flies. Third-instar larvae are treated for 48 hr with different concentrations of the essential oils dissolved in Tween-80 at 0.2% or 2%. The wings of the emerging adults are analyzed for the occurrence of different types of mutant spots. No statistically significant differences in spot frequencies between negative controls and treated series are observed. These results suggest that the six essential oils at concentrations tested are not genotoxic towards somatic cells of D. melanogaster.

Myeloproliferative neoplasms complicated withβ-thalassemia:Two case report

BACKGROUND BCR-ABL-negative myeloproliferative neoplasms(MPNs)are clonal hematopoietic stem cell disorders characterized by the proliferation of one or more myeloid lineages and by mutually exclusive JAK2 V617F,CALR,and MPL[A1]mutations.The combination of MPN and thalassemia is extremely unusual.Several cases with myeloproliferative neoplasms andβ-thalassemia have been reported.However,these have not been extensively reviewed.The present report describes two cases of myeloproliferative neoplasms complicated withβ-thalassemia and reviews all similar cases reported in the literature.CASE SUMMARY We report two patients who were diagnosed with myeloproliferative neoplasms complicated withβ-thalassemia.Both patients had abnormal increases in platelet counts.Based on bone marrow pathology and molecular biology assessment,we made the diagnosis of myeloproliferative neoplasms complicated withβ-thalassemia.The female patient was given hydroxyurea and interferon,which enabled good control of her blood counts;the male patient was given ruxolitinib tablets,thalidomide tablets,and interferon to control the condition,but the patient poorly responded to drug treatment and died of gastrointestinal bleeding six months later.CONCLUSION Given the findings of our cases and the literature review,we hypothesize that myeloproliferative neoplasms complicated withβ-thalassemia can lead to rapid disease progression and a poor prognosis.

Tumor gene mutations and messenger RNA expression： correlation with clinical response to icotinib hydrochloride in non-small cell lung cancer

Background Molecular targeted drugs is now widely used in non-small cell lung cancer （NSCLC） clinical treatment. Icotinib hydrochloride is a new type of oral epidermal growth factor receptor （EGFR） tyrosine kinase inhibitors （EGFR-TKIs）. In this study, we examined the role of EGFR, K-RAS, B-RAF somatic mutations and EGFR mRNA expression in tumor specimens from advanced NSCLC patients as predicators of the efficacy of icotinib hydrochloride. Methods We analyzed tumor paraffin-embedded specimens, which were obtained from 14 of 40 patients with advanced NSCLC who enrolled in the stage I clinical trial of icotinib hydrochloride. Somatic mutations were evaluated by mutant-enriched liquidchip （MEL） technology, and EGFR mRNA expression was measured by branched DNA liquidchip （MBL） technology. Results In the 14 specimens, seven patients showed EGFR mutations, exon 19 deletion （3/7） and exon 21 point mutation （4/7）; and two patients showed K-RAS mutation. No mutations in EGFR exon 20. or B-RAF were detected. In patients with EGFR mutation, one patient developed progress disease （PD）, three patients had stable disease （SD）, two patients had partial responses （PR） and one patient had a complete response （CR）. In patients with wild-type EGFR, four patients had PD, three patients acquired SD, and none had PR/CR （P=-0.0407）. EGFR mutations were associated with better progress-free survival （PFS） （141 days vs. 61 days） but without a statistically significant difference （P=0.8597）, and median overall survival （OS） （-〉449 days vs. 140 days）. EGFR mRNA expression levels were evaluated （three high, eight moderate, one low, and two that can not be measured due to insufficient tumor tissue） and no statistically significant relationships was observed with response, PFS or OS. Conclusions The EGFR mutation rate was consistent with that reported in the Asian population, so the MEL technology is reliable for measuring EGFR mutation with high throughput and rapidity. EGFR exon 19 deletions and exon 21 point mutation are predictive biomarkers for response to icotinib hvdrochloride as second line treatment or above.

A simple and efficient method of DNA extraction from formalin-fixed and paraffin-embedded tissues for screening EGFR gene mutation by gene sequencing

Epidermal growth factor receptor （EGFR） is frequently overexpressed in non-small-cell lung cancer （NSCLC） and plays a key role in tumorigenesis.1 Small molecule tyrosine kinase inhibitors （TKIs）,such as gefitinib,erlotinib,and icotinib,which can inhibit receptor tyrosine kinase activity of EGFR become clinically available for the treatment of non-small-cell lung cancer （NSCLC）.2 NSCLC patients with EGFR mutation have experienced a marked response to EGFR-TKIs therapy.3 Detection of mutations of the EGFR gene is critical for predicting the response to therapy with TKIs.4

Mutation spectrum in GNAQ and GNA11 in Chinese uveal melanoma

Uveal melanoma is the most common intraocular cancer in the adult eye.R183 and Q209 were found to be mutational hotspots in exon 4 and exon 5 of GNAQ and GNA11 in Caucasians.However,only a few studies have reported somaticmutations in GNAQ or GNA11 in uveal melanoma in Chinese.We extracted somatic DNA from paraffin-embedded biopsies of 63 Chinese uveal melanoma samples and sequenced the entire coding regions of exons 4 and 5 in GNAQ and GNA11.The results showed that 33%of Chinese uveal melanoma samples carried Q209 mutations while none had R183 mutation in GNAQ or GNA11.In addition,seven novel missense somatic mutations in GNAQ(Y192C,F194L,P170S,D236N,L232F,V230A,and M227I)and four novel missense somatic mutations in GNA11(R166C,I200T,S225F,and V206M)were found in our study.The high mutation frequency of Q209 and the novel missense mutations detected in this study suggest that GNAQ and GNA11 are common targets for somatic mutations in Chinese uveal melanoma.

Characteristics of homologous recombination repair pathway genes mutation in ovarian cancers

Background:Few studies have investigated the characteristics of non‐BRCA homologous recombination repair(HRR)pathway somatic mutations,and the impact of these mutations on efficacy of treatment in ovarian cancer patients is not clear.Therefore,we conducted this study to analyze the frequency and spectrum of somatic mutations in HRR pathway genes in patients with ovarian cancer and to examine the relationships between somatic mutations in HRR pathway genes and their effects on the efficacy of platinum‐based chemotherapy.Methods:We performed targeted sequencing of 688 genes related to the occurrence,development,treatment,and prognosis of solid tumors.Somatic mutations were identified by paired analysis of tumor tissue and germline DNA in blood cells.Results:A total of 38 patients with ovarian cancer were included in the study,and 35(92.1%)patients were diagnosed with high‐grade serous carcinoma.All patients exhibited somatic mutations in the tumor tissue samples.The commonly mutated genes were TP53(73.7%),BRCA2(55.3%),NF1(52.6%),BRCA1(47.4%),and CDH1(47.4%).Overall,71.1%of the patients exhibited mutation in at least one HRR pathway gene.The most frequently altered HRR genes were BRCA2(55.3%),followed by BRCA1(47.4%),ATM(44.7%),BARD1(42.1%),and CHEK1(36.8%).The median progression‐free survival(PFS)in patients with HRR pathway mutation was 36.0 months compared with 13.6 months in patients with no HRR pathway mutation(hazard ratio[HR],0.25;95%confidence interval[CI],0.08–0.77;p=0.016).Patients harboring BRCA1/2 and/or CDK12 mutations displayed a longer PFS(median,36.0 months)compared with patients with no BRCA1/2 or CDK12 mutation(median,13.6 months;HR,0.21;95%CI,0.07–0.61;p=0.004).In multivariate analysis Cox proportional hazards models,after adjustment for tumor stage at diagnosis and histology of initial diagnosis,patients with HRR pathway mutation had a longer PFS than patients with HRR wild‐type genes(p=0.006).Conclusions:HRR pathway somatic mutations are common in Chinese patients with ovarian cancer.HRR pathway somatic mutations were associated with improved sensitivity to platinum-based chemotherapy.Large-scale prospective studies are needed to verify our findings.

Whole-exome mutational landscape of metastasis in patient-derived hepatocellular carcinoma cells

In order to explore the genomic basis for liver cancer metastasis,whole-exome sequencing(WES)was performed on patient-derived hepatocellular carcinoma(HCC)cell lines with differential metastatic potentials and analyzed their clonal evolution relationships.An evolutionary tree based on genomic single nucleotide polymorphism(SNP)was constructed in MegaX software.The WES data showed that the average percentage of heterogeneous mutations in each HCC cell lines was 16.55%(range,15.38%e18.17%).C:G>T:A and T:A>C:G somatic transitions were the two most frequent substitutions.In these metastatic HCC cell lines,non-silent gene mutations were found in 21.88%of known driver genes and 10 classical signaling pathways.The protein interaction network was constructed by STRING,and hub genes were found in the shared trunk mutation genes and the heterogeneous branch mutations respectively.In cBioPortal database,some of the selected hub genes were found to be associated with poor overall survival(OS)of HCC patients.Among the mutated HCC driver genes,a novel KEAP1 mutation with a homozygous frameshift truncation at the c-terminal Nrf2 binding region was detected and verified in MHCC97-H and HCC97LM3 cells.In conclusion,WES data demonstrate that HCC cell lines from tumor biopsy specimens of the same patient have obtained different metastatic potentials through repeated selection in rodents in vivo,and they do indeed have a genetic relationship at the genomic level.

A human antibody potently neutralizes RSV by targeting the conserved hydrophobic region of prefusion F

Respiratory syncytial virus(RSV) continues to pose serious threats to pediatric populations due to the lack of a vaccine and effective antiviral drugs. RSV fusion(F) glycoprotein mediates viral-host membrane fusion and is a key target for neutralizing antibodies. We generated 23 full-human monoclonal antibodies(hm Abs) against prefusion F protein(pre-F) from a healthy adult with natural RSV infection by single B cell cloning technique. A highly potent RSV-neutralizing hm Ab, named as 25-20, is selected, which targets a new site Φ-specific epitope. Site-directed mutagenesis and structural modelling analysis demonstrated that 25-20 mainly targets a highly conserved hydrophobic region located at the a4 helix and a1 helix of pre-F, indicating a site of vulnerability for drug and vaccine design. It is worth noting that 25-20 uses an unreported inferred germline(i GL) that binds very poorly to pre-F, thus high levels of somatic mutations are needed to gain high binding affinity with pre-F. Our observation helps to understand the evolution of RSV antibody during natural infection. Furthermore, by in silico prediction and experimental verification, we optimized 25-20 with KD values as low as picomolar range. Therefore, the optimized 25-20 represents an excellent candidate for passive protection against RSV infection.

TSNAdb v2.0:The Updated Version of Tumor-specific Neoantigen Database

In recent years,neoantigens have been recognized as ideal targets for tumor immunotherapy.With the development of neoantigen-based tumor immunotherapy,comprehensive neoantigen databases are urgently needed to meet the growing demand for clinical studies.We have built the tumor-specific neoantigen database(TSNAdb)previously,which has attracted much attention.In this study,we provide TSNAdb v2.0,an updated version of the TSNAdb.TSNAdb v2.0 offers several new features,including(1)adopting more stringent criteria for neoantigen identification,(2)providing predicted neoantigens derived from three types of somatic mutations,and(3)collecting experimentally validated neoantigens and dividing them according to the experimental level.

Utility of circulating tumor DNA in patients undergoing hepatectomy for colorectal liver metastases

In this study recently published in Annals of Surgery,Newhook et al.studied the influence of circulating tumor DNA(ctDNA)in the management of patients undergoing hepatic resection for colorectal liver metastases(CRLM)(1).The primary objective was to study the association between the dynamic of ctDNA and somatic mutations with survival after resection of CRLM.The secondary objectives were to evaluate the impact of surgery on perioperative ctDNA dynamics and the impact of its detection on survival.

Comparative genomic analysis of esophageal squamous cell carcinoma and adenocarcinoma:New opportunities towards molecularly targeted therapy

Esophageal cancer is one of the most lethal cancers worldwide because of its rapid progression and poor prognosis.Esophageal squamous cell carcinoma(ESCC)and esophageal adenocarcinoma(EAC)are two major subtypes of esophageal cancer.ESCC predominantly affects African and Asian populations,which is closely related to chronic smoking and alcohol consumption.EAC typically arises in Barrett’s esophagus with a predilection for Western countries.While surgical operation and chemoradiotherapy have been applied to combat this deadly cancer,molecularly targeted therapy is still at the early stages.With the development of large-scale next-generation sequencing,various genomic alterations in ESCC and EAC have been revealed and their potential roles in the initiation and progression of esophageal cancer have been studied.Potential therapeutic targets have been identified and novel approaches have been developed to combat esophageal cancer.In this review,we comprehensively analyze the genomic alterations in EAC and ESCC and summarize the potential role of the genetic alterations in the development of esophageal cancer.Progresses in the therapeutics based on the different tissue types and molecular signatures have also been reviewed and discussed.

Personalized treatment based on mini patient-derived xenografts and WES/RNA sequencing in a patient with metastatic duodenal adenocarcinoma

Background:Treatment guidelines for a variety of cancers have been increasingly used in clinical practice,and have resulted in major improvement in patient outcomes.However,recommended regimens(even first-line treat-ments)are clearly not ideal for every patients.In the present study,we used mini patient-derived xenograft(mini-PDX)and next-generation sequencing to develop personalized treatment in a patient with metastatic duodenal adenocarcinoma.Methods:Resected metachronous metastatic tumor tissues were implanted into SCID mice to determine the sensitivity to a variety of drug regimens.Mutation profiles were assessed using both DNA whole-exome sequencing(DNA-WES)and RNA sequencing.The results of the analyses were used to select optimal treatment for the patient with metastatic duodenal adenocarcinoma.Results:Assessment with mini-PDX models took only 7 days.The results showed high sensitivity to S-1 plus cis-platin,gemcitabine plus cisplatin and everolimus alone.The patient received gemcitabine plus cisplatin initially,but the treatment was terminated due to toxicity.The patient was then switched to treatment with S-1 alone.The overall disease-free survival was 34 months.DNA-WES and RNA sequencing identified KRAS mutation(A146T),TP53(C229Yfs*10)and RICTOR amplification in the metastatic duodenal adenocarcinoma.These findings provided further support to the results of the mini-PDX,and suggest mTOR inhibitors should be used if and when relapse eventually occurs in this patient.Conclusions:Mini-PDX model combined with WES/RNA sequencing can rapidly assess drug sensitivity in cancer patients and reveal key genetic alterations.Further research on this technology for personalized therapy in patients with refractory malignant tumors is warranted.