Impact of essential genes on the success of genome editing experiments generating 3313 new genetically engineered mouse lines

The International Mouse Phenotyping Consortium (IMPC) systematically produces and phenotypes mouse lines with presumptive null mutations to provide insight into gene function. The IMPC now uses the programmable RNA-guided nuclease Cas9 for its increased capacity and flexibility to efficiently generate null alleles in the C57BL/6N strain. In addition to being a valuable novel and accessible research resource, the production of 3313 knockout mouse lines using comparable protocols provides a rich dataset to analyze experimental and biological variables affecting in vivo gene engineering with Cas9. Mouse line production has two critical steps - generation of founders with the desired allele and germline transmission (GLT) of that allele from founders to offspring. A systematic evaluation of the variables impacting success rates identified gene essentiality as the primary factor influencing successful production of null alleles. Collectively, our findings provide best practice recommendations for using Cas9 to generate alleles in mouse essential genes, many of which are orthologs of genes linked to human disease

Human and mouse essentiality screens as a resource for disease gene discovery

The identification of causal variants in sequencing studies remains a considerable challenge that can be partially addressed by new gene-specific knowledge. Here, we integrate measures of how essential a gene is to supporting life, as inferred from viability and phenotyping screens performed on knockout mice by the International Mouse Phenotyping Consortium and essentiality screens carried out on human cell lines. We propose a cross-species gene classification across the Full Spectrum of Intolerance to Loss-of-function (FUSIL) and demonstrate that genes in five mutually exclusive FUSIL categories have differing biological properties. Most notably, Mendelian disease genes, particularly those associated with developmental disorders, are highly overrepresented among genes non-essential for cell survival but required for organism development. After screening developmental disorder cases from three independent disease sequencing consortia, we identify potentially pathogenic variants in genes not previously associated with rare diseases. We therefore propose FUSIL as an efficient approach for disease gene discovery. Discovery of causal variants for monogenic disorders has been facilitated by whole exome and genome sequencing, but does not provide a diagnosis for all patients. Here, the authors propose a Full Spectrum of Intolerance to Loss-of-Function (FUSIL) categorization that integrates gene essentiality information to aid disease gene discovery

Georgia Quality Improvement Programs Multi-Institutional Collection of Postoperative Opioid Data Using ACS-NSQIP Abstraction

Background Excessive postoperative opioid prescribing contributes to opioid misuse throughout the US. The Georgia Quality Improvement Program (GQIP) is a collaboration of ACS-NSQIP participating hospitals. GQIP aimed to develop a multi-institutional opioid data collection platform as well as understand our current opioid-sparing strategy (OSS) usage and postoperative opioid prescribing patterns. Methods This study was initiated 7/2019, when 4 custom NSQIP variables were developed to capture OSS usage and postoperative opioid oral morphine equivalents (OMEs). After pilot collection, our discharge opioid variable required optimization for adequate data capture and was expanded from a free text option to 4 drop-down selection variables. Data collection then continued from 2/2020-5/2021. Logistic regression was used to determine associations with OSS usage. Average OMEs were calculated for common general surgery procedures and compared to national guidelines. Results After variable optimization, the percentage where a total discharge prescription OME could be calculated increased from 26% to 70% ( P &lt; .001). The study included 820 patients over 10 operations. There was a significant variation in OSS usage between GQIP centers. Laparoscopic cases had higher odds of OSS use (1.92 (1.38-2.66)) while OSS use had lower odds in black patients on univariate analysis (.69 (.51-.94)). On average 7 out of the 10 cases had higher OMEs prescribed compared to national guidelines recommendations. Conclusion Developing a multi-institutional opioid data collection platform through ACS-NSQIP is feasible. Preselected drop-down boxes outperform free text variables. GQIP future quality improvement targets include variation in OSS use and opioid overprescribing. </jats:sec

Safety and Efficacy of Transcatheter Aortic Valve Replacement in the Treatment of Pure Aortic Regurgitation in Native Valves and Failing Surgical Bioprostheses: Results From an International Registry Study

The aim of this study was to evaluate the use of transcatheter heart valves (THV) for the treatment of noncalcific pure native aortic valve regurgitation (NAVR) and failing bioprosthetic surgical heart valves (SHVs) with pure severe aortic regurgitation (AR)

Extensive identification of genes involved in congenital and structural heart disorders and cardiomyopathy

Clinical presentation of congenital heart disease is heterogeneous, making identification of the disease-causing genes and their genetic pathways and mechanisms of action challenging. By using in vivo electrocardiography, transthoracic echocardiography and microcomputed tomography imaging to screen 3,894 single-gene-null mouse lines for structural and functional cardiac abnormalities, here we identify 705 lines with cardiac arrhythmia, myocardial hypertrophy and/or ventricular dilation. Among these 705 genes, 486 have not been previously associated with cardiac dysfunction in humans, and some of them represent variants of unknown relevance (VUR). Mice with mutations in Casz1, Dnajc18, Pde4dip, Rnf38 or Tmem161b genes show developmental cardiac structural abnormalities, with their human orthologs being categorized as VUR. Using UK Biobank data, we validate the importance of the DNAJC18 gene for cardiac homeostasis by showing that its loss of function is associated with altered left ventricular systolic function. Our results identify hundreds of previously unappreciated genes with potential function in congenital heart disease and suggest causal function of five VUR in congenital heart disease

Impact of essential genes on the success of genome editing experiments generating 3313 new genetically engineered mouse lines.

Acknowledgements: We thank all technical personnel at the IMPC production centres for their contributions. H.E., E.A., M.G., L.L., C.M., and L.M.J.N. were supported by Ontario Genomics and Genome Canada OGI-051, OGI-090, OGI-137 and the Canada Foundation for Innovation. M-C.B. and Y.H. were supported by the Université de Strasbourg, the CNRS, the INSERM and the ‘Investissements d’avenir’ programs (ANR-10-IDEX-0002-02, ANR-10-LABX-0030-INRT and ANR-10-INBS-07 PHENOMIN). A.C., G.C., M.M., L.T., and S.W. were supported by the Medical Research Council MC_UP_1502/3 International Mouse Phenotyping Consortium—building a functional catalogue of a mammalian genome. B.D., G.D., E.R., H.W.-J., D.A., A.B., R.R.-S., and W.S. were supported by the Wellcome Trust. P.M. and H.P. were supported by European Molecular Biology Laboratory core funding. P.K. and R.S. used services of the Czech Centre for Phenogenomics supported by the Czech Academy of Sciences RVO 68378050 and project LM2018126 Czech Centre for Phenogenomics provided by Ministry of Education, Youth and Sports of the Czech Republic, LM2015040 Czech Centre for Phenogenomics by MEYS, CZ.1.05/2.1.00/19.0395 Higher quality and capacity for transgenic models by MEYS and ERDF, CZ.1.05/1.1.00/02.0109 Biotechnology and Biomedicine Centre of the Academy of Sciences and Charles University in Vestec (BIOCEV) by MEYS and ERDF, CZ.02.1.01/0.0/0.0/16_013/0001789 Upgrade of the Czech Centre for Phenogenomics by MEYS and ESIF. Research reported in this publication was supported by the NIH Common Fund, the Office of The Director and the National Human Genomic Research Institute of the National Institutes of Health (U42OD011174 and UMIHG006348 supported A.C., D.L., G.C., F.D., I.L., M.M., J.S., L.T., S.W., M.D., and J.D.H.; U42OD011175 and UM1OD023221 supported M.G., L.L., C.M., L.M.J.N., B.W., J.A.W., M.R., and K.L.; U42OD011185 and UM1OD023222 supported L.G., K.P., R.B., J.K.W., and S.A.M.; UM1HG006370 supported A.-M.M. and H.P.). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.Funder: Wellcome Trust; doi: http://dx.doi.org/10.13039/100010269The International Mouse Phenotyping Consortium (IMPC) systematically produces and phenotypes mouse lines with presumptive null mutations to provide insight into gene function. The IMPC now uses the programmable RNA-guided nuclease Cas9 for its increased capacity and flexibility to efficiently generate null alleles in the C57BL/6N strain. In addition to being a valuable novel and accessible research resource, the production of 3313 knockout mouse lines using comparable protocols provides a rich dataset to analyze experimental and biological variables affecting in vivo gene engineering with Cas9. Mouse line production has two critical steps - generation of founders with the desired allele and germline transmission (GLT) of that allele from founders to offspring. A systematic evaluation of the variables impacting success rates identified gene essentiality as the primary factor influencing successful production of null alleles. Collectively, our findings provide best practice recommendations for using Cas9 to generate alleles in mouse essential genes, many of which are orthologs of genes linked to human disease