Fat-1 Gene Research Articles

Fatty acid synthase (Fasn) inhibits the expression levels of immune response genes via alteration of alternative splicing in islet cells

BackgroundIncreasing evidence has shown that fatty acid synthase (Fasn) is associated with diabetes mellitus (DM) and insulin resistance, however, it remains unclear how Fasn upregulation leads to dysregulation of energy homeostasis in islet cells. Consequently, uncovering the function of Fasn in islet cells. Consequently, uncovering the function of FASN in islet cells is immensely important for finding a treatment target. AimIn this study, we elucidated the biological function of Fasn on the target genes in a rat insulinoma INS-1 cell line. MethodsWe created a Fasn overexpressing rat insulinoma cell line (Fasn-OE), and performed bulk RNA-sequencing (RNA-seq) experiments on Fasn-OE and INS-1 (control) cells. We first identified differentially expressed genes (DEGs) using Bioconductor package edgeR, and then discovered enriched gene ontology terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways using the KEGG Orthology Based Annotation System (KOBAS) 2.0 web server. Furthermore, we identified alternative splicing events (ASEs) and regulated alternative splicing events (RASEs) by applying the ABLas pipeline. The reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used for validation of selected differentially expressed genes (DEGs) and Fasn-regulated alternative splicing genes (RASGs). ResultsIn this study we found that Fasn overexpression led to significant changes of gene expression profiles, including downregulations of mRNA levels of immune related genes, including Bst2, Ddit3, Isg15, Mx2, Oas1a, Oasl, and RT1-S3 in INS-1 cell line. Furthermore, Fasn positively regulated the expression of transcription factors such as Fat1 and Ncl diabetes-related genes. Importantly, Fasn overexpression to result in alternative splicing events including in a metabolism-associated ATP binding protein mRNA Abcc5. In Gene Ontology analysis, the downregulated genes in Fasn-OE cells were mainly enriched in inflammatory response and innate immune response. In Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis, the downregulated genes were mainly enriched in TNF signaling pathway and cytokine-mediated signaling pathways. ConclusionsOur findings showed that upregulation of Fasn may play a critical role in islet cell immunmetabolism via modifications of immune/inflammatory related genes on transcription and alternative splicing level, which provide novel insights into characterizing the function of Fasn in islet cell immunity and for the development of chemo/immune therapies.

Molecular profiling to identify potential therapeutic targets in hepatocellular carcinoma.

471 Background: Hepatocellular carcinoma (HCC) is the most common type of primary liver cancer and is often diagnosed at an advanced stage with limited effective therapies. The advent of comprehensive genomic molecular profiling has the potential to identify novel therapeutic options in the treatment of HCC. In this descriptive clinical utility study, we evaluated the utility of the Oncotype MAP Pan-Cancer Tissue (MAP) test to identify potential therapeutic options, including clinical trials. Methods: Between October 2018 and October 2020, we identified 41 samples from patients with HCC submitted for MAP testing. The MAP test uses tumor tissue to identify single nucleotide variants, indels, copy number alterations, and select structural variants and fusions by next-generation sequencing (NGS) of a 257 gene panel. Tumor mutational burden and microsatellite instability are also determined. A custom immunohistochemical (IHC) panel is also performed. Results from NGS and IHC are run against a proprietary knowledgebase of genomic and proteomic data that includes FDA approvals, NCCN Recommendations and published biomarker data on potentially targetable alterations. Eligibility for ongoing clinical trials is determined using the NCI database. Results: Of the 41 samples, 37 (90%) met minimum tissue requirements (3mm2 and 15% tumor cellularity) for DNA sequencing. Of these 37 samples, 36 (97%) were successfully sequenced for NGS testing. Median turnaround time from the date of accessioning to the date of laboratory report was 5 days (interquartile range (IQR) 4-6 days). The most frequently identified genomic variants characterized as pathogenic, or unknown significance, were TP53, CTNNB1 and FAT1 genes. Thirty-two samples (86%) had a genomic alteration that could potentially direct treatment. Six samples (16%) had findings directing to an approved treatment for HCC, 4 (11%) had findings for an approved treatment of a different tumor type, and 22 (59%) had findings for one or more clinical trials. Conclusions: The Oncotype MAP Pan-Cancer Tissue test identified molecular variants that could be used to inform treatment decisions in 86% of HCC samples, including 16% to an approved HCC therapy. The Oncotype MAP test may be considered as another tool in the management of advanced HCC patients.

Fat-1 expression alleviates atherosclerosis in transgenic rabbits.

Atherosclerosis is the main cause of cardiovascular diseases. The Fat‐1 gene can express the n‐3 fatty acid desaturase, which converts n‐6 polyunsaturated fatty acids (PUFA) to n‐3 PUFAs. The role of n‐3 PUFAs in atherosclerosis is widely debated. This study explored the effect of n‐3 PUFAs on atherosclerosis in rabbits. In this study, atherosclerosis was induced in Fat‐1 transgenic rabbits and their littermate (WT) rabbits by feeding a high‐cholesterol diet containing 0.3% cholesterol and 3% soybean oil for 16 weeks. Plasma lipid, fatty acid and pathological analyses of atherosclerotic lesions were conducted. Fatty acid composition in the liver and muscle showed that n‐3 PUFAs increased and n‐6 PUFAs decreased in the Fat‐1 group. Plasma high‐density lipoprotein cholesterol (HDL‐C) levels were significantly increased in the Fat‐1 group, and the atherosclerotic lesion area of the aortic arch in Fat‐1 transgenic rabbits was significantly reduced. Histological analysis showed that smooth muscle cells (SMCs) in atherosclerotic lesions decreased significantly. In conclusion, n‐3 PUFAs improve atherosclerosis in Fat‐1 transgenic rabbits, and this process may depend on the increase in plasma HDL‐C and the decrease in the amount of SMCs in atherosclerotic plaques.

Clinical significance of FAT1 gene mutation and mRNA expression in patients with head and neck squamous cell carcinoma.

The FAT1 gene functions as a tumor suppressor or promoter and remains incompletely understood. We examined the clinical significance of FAT1 in head and neck squamous cell carcinoma (HNSCC) using four publicly available HNSCC cohorts and one HNSCC cohort enrolled at a tertiary medical center. We developed FAT1 signatures reflecting FAT1 mutations and mRNA expression using one cohort. Patients with HNSCC were classified into FAT1‐associated low risk (FAT1‐LR; n = 195) and FAT1‐associated high risk (FAT1‐HR; n = 371) subgroups. The five‐year overall survival and recurrence‐free survival rates were significantly lower in the FAT1‐HR subgroup than in the FAT1‐LR subgroup (P = 0.01 and 0.003, respectively). The clinical significance of FAT1 was validated using four independent cohorts. Cox proportional hazards models showed that the FAT1 signature was an independent prognostic factor for HNSCC patients. In addition, FAT1 signature was associated with the response to radiotherapy, advanced stage, and human papilloma virus (HPV) status in HNSCC patients. In conclusion, the FAT1 gene signature was associated with prognosis of HNSCC and may help to provide personalized treatments for HNSCC patients.

Cardioprotective Effects of n-3 Polyunsaturated Fatty Acids: Orchestration of mRNA Expression, Protein Phosphorylation, and Lipid Metabolism in Pressure Overload Hearts.

Background: Pressure overload can result in dilated cardiomyopathy. The beneficial effects of n-3 polyunsaturated fatty acids (n-3 PUFAs) on heart disorders have been widely recognized. However, the molecular mechanisms underlying their protective effects against cardiomyopathy remain unclear.Methods: Pressure overload in mice induced by 8 weeks of transverse aortic constriction was used to induce dilated cardiomyopathy. A transgenic fat-1 mouse model carrying the n-3 fatty acid desaturase gene fat-1 gene from Caenorhabditis elegans was used to evaluate the mechanism of n-3 PUFAs in this disease. Echocardiography, transmission electron microscopy, and histopathological analyses were used to evaluate the structural integrity and function in pressure overloaded fat-1 hearts. mRNA sequencing, label-free phosphoprotein quantification, lipidomics, Western blotting, RT-qPCR, and ATP detection were performed to examine the effects of n-3 PUFAs in the heart.Results: Compared with wild-type hearts, left ventricular ejection fraction was significantly improved (C57BL/6J [32%] vs. fat-1 [53%]), while the internal diameters of the left ventricle at systole and diastole were reduced in the fat-1 pressure overload hearts. mRNA expression, protein phosphorylation and lipid metabolism were remodeled by pressure overload in wild-type and fat-1 hearts. Specifically, elevation of endogenous n-3 PUFAs maintained the phosphorylation states of proteins in the subcellular compartments of sarcomeres, cytoplasm, membranes, sarcoplasmic reticulum, and mitochondria. Moreover, transcriptomic analysis predicted that endogenous n-3 PUFAs restored mitochondrial respiratory chain function that was lost in the dilated hearts, and this was supported by reductions in detrimental oxylipins and protection of mitochondrial structure, oxidative phosphorylation, and ATP production.Conclusions: Endogenous n-3 PUFAs prevents dilated cardiomyopathy via orchestrating gene expression, protein phosphorylation, and lipid metabolism. This is the first study provides mechanistic insights into the cardioprotective effects of n-3 PUFAs in dilated cardiomyopathy through integrated multi-omics data analysis.

A novel frameshift homozygous mutation in FAT1 gene causes ptosis, nephropathy, and syndactyly in an Emirati family: case report and literature review

Background: Single gene mutations are important causes of glomerular disease in children. Of these genes, mutations in the FAT1 gene have been recently described in the literature as a cause of nephropathy in isolated form or multisystem involvement. The spectrum of renal disease associated with FAT1 gene mutations varies from asymptomatic proteinuria and hematuria to severe nephrotic syndrome and end-stage renal disease. Case Presentation: In this case report, we describe a 3-year-old child and two other family members with a novel frameshift homozygous mutation in the FAT1 gene consistent with the diagnosis of autosomal recessive colobomatous-microphthalmia, ptosis, nephropathy, and syndactyly syndrome with variable expression of the phenotype. Conclusion: This report adds to the genotype-phenotype correlation, highlighting the clinical importance of considering FAT1 gene defects as part of the differential diagnosis for congenital ptosis, syndactyly and nephropathy, especially with multiple affected family members.

Systematic Review and Meta-analysis of the Most Common Genetic Mutations in Esophageal Squamous Cell Carcinoma.

Esophageal cancer is the second most common cancer among men and women. There is a need to systematically assess the current evidence to map out the contribution of genetic factors in the development of esophageal squamous cell carcinoma (ESCC). A literature search was carried out on published and unpublished studies up to August 2021 in Medline (PubMed), Embase (Ovid), Scopus, Proquest, Web of Science, and Google scholar. Studies that have reported the frequency of genetic mutations in ESCC were included in this study. A total of 1238 titles were retrieved through searches, and finally, 56 articles, including 8114 samples, met our predefined inclusion criteria. Of the included studies, 31 were conducted in China, 12 in Japan, and the remaining were conducted in various nations, including Brazil, Korea, and Iran. Most of our included studies evaluated the TP53 (n = 37 studies) and PIK3CA (n = 30 studies) gene mutations. TP53 (68.6%; 95% CI: 61.6-74.9), CCND1 (39.3%; 95% CI: 26.2-54.1), MDM2 (24.9%; 95% CI: 9.5-51.0), NOTCH1/2/3 (17.9%; 95% CI: 15.0-21.2), KMT2D (17.4%; 95% CI: 12.4-23.8), CDKN2A (15.0%; 95% CI: 8.1-26.1), PIK3CA (13.8%; 95% CI: 10.3-18.1), FAT1 (13.3%; 95% CI: 11.7-15.0), and EGFR (9.9%; 95% CI: 5.6-17.0) were the most common involved genetic factors in developing ESCC. This systematic review and meta-analysis revealed that more than 10% of ESCC patients had changes in TP53, CCND1, MDM2, NOTCH1/2/3, KMT2D, CDKN2A, PIK3CA, and FAT1 genes, which can highlight their role in developing ESCC. TP53, CCND1, and MDM2 are the most prevalent, demonstrating 68.6%, 39.3%, and 24.9% of the mutations in ESCC patients.

Germline variants in DNA repair genes are associated with young-onset head and neck cancer

The genetic predisposition to head and neck carcinomas (HNSCC) and how the known risk factors (papillomavirus infection, alcohol, and tobacco consumption) contribute to the early-onset disease are barely explored. Although HNSCC at early onset is rare, its frequency is increasing in recent years. Germline and somatic variants were assessed to build a comprehensive genetic influence pattern in HNSCC predisposition and patient outcome. Whole-exome sequencing was performed in 45 oral and oropharynx carcinomas paired with normal samples of young adults (≤49 years). We found FANCG, CDKN2A, and TPP germline variants previously associated with HNSCC risk. At least one germline variant in DNA repair pathway genes was detected in 67% of cases. Germline and somatic variants (including copy number variations) in FAT1 gene were identified in 9 patients (20%) and 12 tumors (30%), respectively. Somatic variants were found in HNSCC associated genes, such as TP53, CDKN2A, and PIK3CA. To date, 55 of 521 cases from the large cohort of TCGA presented < 49 years old. A comparison between the somatic alterations of TCGA-HNSCC at early onset and our dataset revealed strong similarities. Protein-protein interaction analysis between somatic and germline altered genes revealed a central role of TP53. Altogether, germline alterations in DNA repair genes potentially contribute to an increased risk of developing HNSCC at early-onset, while FAT1 could impact the prognosis.

Targeted Next-Generation Sequencing of Cancer-Related Genes in a Norwegian Patient Cohort With Head and Neck Squamous Cell Carcinoma Reveals Novel Actionable Mutations and Correlations With Pathological Parameters.

BackgroundTargeted next-generation sequencing (NGS) is increasingly applied in clinical oncology to advance personalized treatment. Despite success in many other tumour types, use of targeted NGS panels for assisting diagnosis and treatment of head and neck squamous cell carcinomas (HNSCC) is still limited.AimThe focus of this study was to establish a robust NGS panel targeting most frequent cancer mutations in long-term preserved formalin-fixed paraffin-embedded (FFPE) tissue samples of HNSCC from routine diagnostics.Materials and MethodsTumour DNA obtained from archival FFPE tissue blocks of HNSCC patients treated at Haukeland University Hospital between 2003-2016 (n=111) was subjected to mutational analysis using a custom made AmpliSeq Library PLUS panel targeting 31 genes (Illumina). Associations between mutational burden and clinical and pathological parameters were investigated. Mutation and corresponding clinicopathological data from HNSCC were extracted for selected genes from the Cancer Genome Atlas (TCGA) and used for Chi-square and Kaplan-Meier analysis.ResultsThe threshold for sufficient number of reads was attained in 104 (93.7%) cases. Although the specific number of PCR amplified reads detected decreased, the number of NGS-annotated mutations did not significantly change with increased tissue preservation time. In HPV-negative carcinomas, mutations were detected mainly in TP53 (73.3%), FAT1 (26.7%) and FLG (16.7%) whereas in HPV-positive, the common mutations were in FLG (24.3%) FAT1 (17%) and FGFR3 (14.6%) genes. Other less common pathogenic mutations, including well reported SNPs were reproducibly identified. Presence of at least one cancer-specific mutations was found to be positively associated with an extensive desmoplastic stroma (p=0.019), and an aggressive type of invasive front (p=0.035), and negatively associated with the degree of differentiation (p=0.041). Analysis of TCGA data corroborated the association between cancer-specific mutations and tumour differentiation and survival analysis showed that tumours with at least one mutation had shorter disease-free and overall survival (p=0.005).ConclusionsA custom made targeted NGS panel could reliably detect several specific mutations in archival samples of HNSCCs preserved up to 17 years. Using this method novel associations between mutational burden and clinical and pathological parameters were detected and actionable mutations in HPV-positive HNSCC were discovered.

Linc8087 predicts favorable prognosis and inhibits cell migration and invasion in NSCLC

BackgroundNon-small cell lung cancer (NSCLC) is the most common cancer and has poor prognosis. Long non-coding RNA(LncRNA) plays important roles in the regulation of cell migration in various types of cancer. In this study, we aimed to demonstrate the function of linc8087 in regulating cell migration and invasion in NSCLC cells. MethodsA lncRNA microarray was used to identify differentially expressed lncRNAs between NSCLC tissues and normal tissues. RT-qPCR was used to confirm the expression of linc8087 in tumor tissues. The association between linc8087 expression and clinicopathological characteristics was analyzed. RNA fluorescence in situ hybridization (FISH) was performed to observe the subcellular localization of linc8087. We investigated the effects of linc8087 expression on cell migration and invasion by wound healing assay, Transwell and invasion assays. The Human Tumor Metastasis RT2 Profiler PCR Array was used to detect and analyze the mRNA levels of 84 genes involved in metastasis. ResultsWe found that linc8087 expression was obviously decreased in both NSCLC tissues and cell lines compared with paired normal tissues and a normal bronchial epithelium cell line. Low expression of linc8087 was significantly associated with poor survival. In addition, linc8087 was an independent risk factor for survival. Overexpressed linc8087 inhibited cell migration and invasion in A549 and PC9 cell lines. Knockdown of linc8087 promoted cell migration and invasion. The result of RT2 Profiler PCR Array showed that overexpressed linc8087 upregulated the expression of the COL4A2, CST7 and FAT1 genes and led to the downregulation of SERPINE1. ConclusionsThese results indicate that linc8087 plays a key role in the progression of NSCLC, and it may serve as a meaningful prognostic biomarker as well as a latent therapeutic target in NSCLC patients.

Abstract 321: Identification of proteins associated with FAT1 mutations which potentially contribute to oncogenesis and progression of head and neck cancers

Abstract Objectives: The Cancer Genome Atlas (TCGA) reported a FAT1 mutation rate of 23% in head and neck squamous cell carcinoma (HNSCC), which was higher in human papilloma virus (HPV)-negative cancer. Wild-type FAT1 exhibits a tumor suppressor activity, and the majority of mutated FAT1 genes are inactivated by missense or truncation, leading to altered or terminated gene products. However, the roles of FAT1 mutants in HNSCC oncogenesis and progression remain poorly understood. Methods: We investigated the landscape of altered protein and gene expression associated with FAT1 mutation, which predicted clinical outcomes of 329 HNSCC patients. The relationships of FAT1 mutation with clinical information from HNSCC datasets of TCGA were tested using Chi-Square or Fisher's exact test. The association of FAT1 mutation with more than 200 proteins or phosphorylated sites detected by Reverse Phase Protein Arrays (RPPA) was revealed using Student's T-test. Overall survival (OS) and progression-free survival (PFS) were analyzed with Kaplan Meier method, Log-rank tests, and COX model. Hierarchical clustering classified HNSCC subgroups, and the underlying signaling pathways and regulatory networks were revealed by Ingenuity Pathway Analysis (IPA). Results: Clinically, the prevalence of FAT1 mutation was significantly associated with HPV(-), female, or older patients, and enriched in the oral cavity and larynx/hypopharynx primary tumor sites. FAT1 mutation was significantly associated with lower FAT1 gene expression and increased protein expression of HER3_pY1289, IRS1, CAVEOLIN1 and VEGFR2, while twelve proteins were upregulated in patients with wild type FAT1, including immune check point molecule PD1. Poorer OS or PFS was observed in patients with FAT1 mutation and/or altered HER3_pY1289, IRS1, CYCLINE2, RET_pY905, P16INK4A, CMYC, MTORpS2448, or CAVEOLIN. IPA revealed dominant ERBB/neuregulin pathways, and protein signature panels divulged the heterogeneity of patient subgroups and predicted PFS. Alteration of FAT1 appears to involve mostly HPV-unrelated HNSCC and may contribute to the resistance to EGFR-targeted therapy and immunotherapy. Summary: We revealed dozens of actionable proteins associated with FAT1 mutation, which provide important insight into possible prospective clinical investigation with therapeutic relevance. (Supported by grants from a pilot Winship Invests Grant of Emory University Winship Cancer Institute to GZC and NFS; and NIDCD/NIH intramural projects Z01-DC-00016, 73, 74 to ZC, and partially by the University of Illinois Cancer Center Biostatistics Shared Resource Core). Citation Format: Zhuo Georgia Chen, Zhengjia Chen, Chao Zhang, Jianhong Chen, Dongsheng Wang, Nabil F. Saba, Zhong Chen. Identification of proteins associated with FAT1 mutations which potentially contribute to oncogenesis and progression of head and neck cancers [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 321.

CRISPR/Cas9-Mediated Specific Integration of Fat-1 and IGF-1 at the pRosa26 Locus.

Many researchers have focused on knock-in pigs for site-specific integration, but little attention has been given to genetically modified pigs with the targeted integration of multiple recombinant genes. To establish a multigene targeted knock-in editing system, we used the internal ribosome entry site (IRES) and self-cleaving 2A peptide technology to construct a plasmid coexpressing the fatty acid desaturase (Fat-1) and porcine insulin-like growth factor-1 (IGF-1) genes at equal levels. In this study, pigs were genetically modified with multiple genes that were precisely inserted into the pRosa26 locus by using the clustered regularly spaced short palindrome repeat sequence (CRISPR)/CRISPR-related 9 (Cas9) system and somatic cell nuclear transfer technology (SCNT) in combination. Single copies of the Fat-1 and IGF-1 genes were expressed satisfactorily in various tissues of F0-generation pigs. Importantly, gas chromatography analysis revealed a significantly increased n-3 polyunsaturated fatty acid (PUFA) level in these genetically modified pigs, which led to a significant decrease of the n-6 PUFA/n-3 PUFA ratio from 6.982 to 3.122 (*** p < 0.001). In conclusion, the establishment of an editing system for targeted double-gene knock-in in this study provides a reference for the precise integration of multiple foreign genes and lays a foundation for the development of new transgenic pig breeds with multiple excellent phenotypes.

Abstract 3175: FAT1 and the immunosuppressive milieu in glioblastoma tumors

Abstract INTRODUCTION: Glioblastoma tumors harbor an immunosuppressive microenvironment which is sustained by key players such as TGF-β1/2 and IL-10 cytokines; and mediators of the PD-1/PD-L1/2 pathway. Previously, our lab has shown the role of a new gene, FAT1, in promoting gliomagenesis by supporting the production of pro-inflammatory cytokines (IL-1β, IL-6, etc.) in glioma cells. Here, we show a potent role of FAT1 gene in regulation of anti-inflammatory genes in human glioma tumors, cell lines and primary cultures. Our findings indicate an additional way in which FAT1 might contribute to gliomagenesis. METHODS: q-PCR was used to analyse transcript levels of FAT1 and genes of the TGF-β1/2, IL-10 and PD-L1/2 pathways in fresh-frozen glioblastoma samples relative to normal brain RNA. Gene expression correlation was analysed using Spearman's test and semi-supervised clustering. TCGA expression data was extracted to analyse the correlation of FAT1 with anti-inflammatory genes. Primary tumor cultures and cell lines were transfected with FAT1-specific siRNA to check the effect on expression of anti-inflammatory cytokines, as well as effect on migratory properties of monocytes towards FAT1-knockdown tumor cells (supernatants). RESULTS: Significant positive correlation was found between upregulation of FAT1 and that of anti-inflammatory genes in fresh-frozen glioblastoma samples as well as TCGA datasets. Upon siRNA-mediated knockdown of FAT1 in in-vitro glioma cell cultures, we observed decreased anti-inflammatory genes' expression. We also found increased transmigration of monocytes towards the supernatants from siFAT1-transfected cells. CONCLUSION: FAT1 favors the immunosuppressive milieu in glioblastoma by supporting expression of anti-inflammatory genes. This is an emerging role of the new oncogene FAT1 and crucial to our understanding of the contributions of FAT1 to hallmarks of cancer. FUNDING: SERB Start-Up Grant (DST, India); DHR Young Scientist Scheme (ICMR, India) Citation Format: Khushboo Irshad, Chitrangda Srivastava, Nargis Malik, Yakhlesh Gupta, Vaishali Suri, Swati Mahajan, Deepak Gupta, Ashish Suri, Parthaprasad Chattopadhyay, Subrata Sinha, Kunzang Chosdol. FAT1 and the immunosuppressive milieu in glioblastoma tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3175.

Whole Exome Sequencing Is the Minimal Technological Approach in Probands Born to Consanguineous Couples.

We report on two siblings suffering from different pathogenic conditions, born to consanguineous parents. A multigene panel for brain malformations and microcephaly identified the homozygous splicing variant NM_005886.3:c.1416+1del in the KATNB1 gene in the older sister. On the other hand, exome sequencing revealed the homozygous frameshift variant NM_005245.4:c.9729del in the FAT1 gene in the younger sister, who had a more complex phenotype: in addition to bilateral anophthalmia and heart defects, she showed a right split foot with 4 toes, 5 metacarpals, second toe duplication and preaxial polydactyly on the right hand. These features have been never reported before in patients with pathogenic FAT1 variants and support the role of this gene in the development of limb buds. Notably, each parent was heterozygous for both of these variants, which were ultra-rare and rare, respectively. This study raises awareness about the value of using whole exome/genome sequencing rather than targeted gene panels when testing affected offspring born to consanguineous couples. In this way, exomic data from the parents are also made available for carrier screening, to identify heterozygous pathogenetic and likely pathogenetic variants in genes responsible for other recessive conditions, which may pose a risk for subsequent pregnancies.

The Proteomic Landscape of Growth Factor Signaling Networks Associated with FAT1 Mutations in Head and Neck Cancers.

FAT1 is frequently mutated in head and neck squamous cell carcinoma (HNSCC), but the biological and clinical effects of FAT1 mutations in HNSCC remain to be fully elucidated. We investigated the landscape of altered protein and gene expression associated with FAT1 mutations and clinical outcomes of patients with HNSCC. FAT1 mutation was stratified with clinical information from The Cancer Genome Atlas HNSCC databases with more than 200 proteins or phosphorylated sites. FAT1 mutation was significantly more prevalent among HPV(-), female, and older patients and was enriched in oral, larynx, and hypopharynx primary tumors. FAT1 mutation was also significantly associated with lower FAT1 gene expression and increased protein expression of HER3_pY1289, IRS1, and CAVEOLIN1. From an independent International Cancer Genome Consortium dataset, FAT1 mutation in oral cancer co-occurred with top mutated genes TP53 and CASP8. Poorer overall survival or progression-free survival was observed in patients with FAT1 mutation or altered HER3_pY1289, IRS1, or CAVEOLIN1. Pathway analysis revealed dominant ERBB/neuregulin pathways linked to FAT1 mutations in HNSCC, and protein signature panels uncovered the heterogeneity of patient subgroups. Decreased pEGFR, pHER2, and pERK and upregulated pHER3 and HER3 proteins were observed in two FAT1 knockout HNSCC cell lines, supporting that FAT1 alterations lead to altered EGFR/ERBB signaling. In squamous cancers of the lung and cervix, a strong association of FAT1 and EGFR gene expressions was identified. Collectively, these results suggest that alteration of FAT1 appears to involve mostly HPV(-) HNSCC and may contribute to resistance to EGFR-targeted therapy. SIGNIFICANCE: Integrative bioinformatics and statistical analyses reveal a panel of genes and proteins associated with FAT1 mutation in HNSCC, providing important insights into prospective clinical investigations with targeted therapies.

Prognostic value of tumor mutational burden in patients with oral cavity squamous cell carcinoma treated with upfront surgery

BackgroundOral cavity is the most prevalent site of head and neck squamous cell carcinomas (HNSCCs). Most often diagnosed at a locally advanced stage, treatment is multimodal with surgery as the cornerstone. The aim of this study was to explore the molecular landscape of a homogenous cohort of oral cavity squamous cell carcinomas (OCSCCs), and to assess the prognostic value of tumor mutational burden (TMB), along with classical molecular and clinical parameters.Patients and methodsOne hundred and fifty-one consecutive patients with OCSCC treated with upfront surgery at the Institut Curie were analyzed. Sequencing of tumor DNA from frozen specimens was carried out using an in-house targeted next-generation sequencing panel (571 genes). The impact of molecular alterations and TMB on disease-free survival (DFS) and overall survival (OS) was evaluated in univariate and multivariate analyses.ResultsPathological tumor stage, extranodal spread, vascular emboli, and perineural invasion were associated with both DFS and OS. TP53 was the most mutated gene (71%). Other frequent molecular alterations included the TERT promoter (50%), CDKN2A (25%), FAT1 (17%), PIK3CA (14%), and NOTCH1 (15%) genes. Transforming growth factor-β pathway alterations (4%) were associated with poor OS (P = 0.01) and DFS (P = 0.02) in univariate and multivariate analyses. High TMB was associated with prolonged OS (P = 0.01 and P = 0.02, in the highest 10% and 20% TMB values, respectively), but not with DFS. Correlation of TMB with OS remained significant in multivariate analysis (P = 0.01 and P = 0.005 in the highest 10% and 20% TMB values, respectively). Pathological tumor stage combined with high TMB was associated with good prognosis.ConclusionOur results suggest that a high TMB is associated with a favorable prognosis in patients with OCSCC treated with upfront surgery.

Next-generation sequencing in children of West India with suspected inherited tubulopathies

Background: The renal tubules maintain homeostasis through an array of proteins coded by multiple genes, which directly or indirectly transport water and solutes in the tubules. Malfunction of these transport proteins leads to inherited tubulopathies. Objective: The objective of the study was to analyze the clinical utility of next-generation sequencing (NGS) in the diagnosis and management of children with tubulopathies. Materials and Methods: This retrospective study was conducted in a tertiary care nephrology center in West India between September 2016 and September 2020. All children ?18 years with suspected inherited tubulopathy, where an NGS was sent, were included. Clinical exome sequencing (CES) covering 6670 genes was done in 14 children. CES included 73 tubulopathy genes. The test result was interpreted as per American College of Medical Genetics classification: No pathogenic variant, variant of unknown significance (VUS), and likely pathogenic and pathogenic variant. Results: The median age (IQR) of the cohort at the onset of disease was 18 months (4–65). History of consanguinity was present in 2 children (14.2%). Five children (35.7%) had Fanconi syndrome, two had distal renal tubular acidosis (d-RTA), two had Bartter/Gitelman syndrome, two had rickets, two had nephrocalcinosis, and one had low-molecular-weight proteinuria. Nine children (64%) had pathogenic variants detected in eight genes: ATP6V0A4, CTNS1, FAH1, PHEX, SLC12A1, SLC4A1, SLCA2, and CLDN 16. Five (35.7%) were novel variants. Two children had three VUS in FAT1, EYA1, and KCNJ1 gene. Three children (21.4%) had no genetic variant. Bartter syndrome type 1 and d-RTA were the most common genetic diagnoses with two patients each. Other diagnoses were tyrosinemia type 1, nephropathic cystinosis, Fanconi Bickel syndrome, X-linked hypophosphatemic rickets, and familial hypomagnesemia, hypercalciuria, and nephrocalcinosis in one each. Conclusion: The yield of NGS in children with tubulopathy was remarkably high. NGS was useful in the diagnosis and management in these children.

Endogenous conversion of n-6 to n-3 polyunsaturated fatty acids facilitates the repair of cardiotoxin-induced skeletal muscle injury in fat-1 mice.

In this study, we investigated the beneficial effects of high endogenous levels of n-3 polyunsaturated fatty acids (PUFAs) on skeletal muscle repair and regeneration using a mouse cardiotoxin (CTX, 20 μM/200 μL) -induced gastrocnemius muscle injury model. Transgenic fat-1 mice expressing the Caenorhabditis elegans fat-1 gene, encoding n-3 fatty acid desaturase, showed higher n-3 PUFA levels and lower n-6/n-3 PUFA ratios in gastrocnemius muscle tissues. Hematoxylin and eosin and Masson’s trichrome staining of gastrocnemius sections revealed increased muscle fiber size and reduced fibrosis in fat-1 mice on days 7 and 14 after CTX injections. Gastrocnemius muscle tissues from fat-1 mice showed reduced inflammatory responses and increased muscle fiber regeneration reflecting enhanced activation of satellite cells on day 3 after cardiotoxin injections. Gastrocnemius muscle tissues from cardiotoxin-treated fat-1 mice showed reduced levels of pro-apoptotic proteins (Caspase 3 and Bax) and increased levels of anti-apoptotic proteins (Bcl-2 and Survivin). Moreover, eicosapentaenoic acid (EPA) reduced the incidence of apoptosis among cardiotoxin-treated C2C12 mouse myoblasts. These findings demonstrate that higher endogenous n-3 PUFA levels in fat-1 mice enhances skeletal muscle repair and regeneration following cardiotoxin-induced injury.

Effect of FAT1 gene expression on the prognosis of medulloblastoma in children: A protocol for systematic review and meta-analysis.

Background:It was reported that cloning human adipose atypical cadherin 1 (FAT1) has an effect on the prognosis of medulloblastoma (MB), while the conclusion still needs to be further proved. Therefore, this study attempted to explore the effect of the high expression of FAT1 on the prognosis of MB children.Methods:The database was retrieved from China National Knowledge Infrastructure (CNKI), Chinese Biomedical literature Database (CBM), Chinese Scientific and Journal Database (VIP), Wan Fang database, PubMed, and EMBASE. Hazard ratios (HRs) and its 95% confidence intervals (CIs) were applied to assess the prognostic effect of FAT1 on overall survival (OS) and disease-free survival (DFS). RevMan 5.3 and STATA 16.0 software were used to perform the meta-analysis.Results:The results of the study would be published in peer-reviewed journals or at relevant meetings.Conclusion:Our findings revealed the effect of the high expression of FAT1 on the prognosis of MB children. Such studies may find a new prognostic marker for MB children and help clinicians and health professionals make clinical decisions.OSF Registration Number:DOI 10.17605/OSF.IO/5FN8M.

Identification of vital genes and pathways associated with mucosal melanoma in Chinese

Mucosal melanoma is a rare malignant melanoma with more aggressive and poorer outcomes. The incidence of mucosal melanoma varies greatly among different ethnic groups. We herein sought to characterize the vital genes and pathways of Chinese mucosal melanoma patients. By whole-exome sequencing in six patients with mucosal melanoma, we detected a total of 21,733 CNVs and 2372 SNPs. The CNV/SNP burden varies greatly between individuals, including recurrent CNV targeting PIK3 family, KRAS, APC and BRCA1. Significantly mutated genes were NUDT5, ZBTB18, NEURL4, ZNF430, RBM44, GAK, PCDHA13, STK38 and UBR5. Besides, FAT1 gene was identified frequently mutated in anorectal melanoma patients (3/3, 100%). Moreover, our result showed that HPV infection may be associated with mucosal melanoma. In conclusion, this study indicated that mucosal melanomas have a low SNPs burden and a high number of CNVs and expand the spectrum of mucosal melanoma variants, also provided an insight for the pathological mechanism of mucosal melanoma.