Proteomic and Clinical Impact of Human Knockouts in British South Asians

Human loss-of-function (LoF) variants affecting both copies of a gene ('human knockouts') provide a unique opportunity to directly study function and clinical impact of genes but are very rare in most populations sequenced to date. Here we study 1,569 British Bangladeshi and Pakistani adults who were recalled for plasma sampling for proteomic profiling using three distinct technologies (covering >12,000 proteins) from 55k whole exome sequenced Genes & Health adults - a cohort enriched for rare, biallelic (homozygous) variants due to high autozygosity. We identified 199 individuals with rare homozygous predicted LoF genotypes (pLoF) for which the respective cis-protein was measured by at least one technology, and observed extreme (> 3SDs) cis-protein underexpression in 41 individuals (median z-score = -9.72 (range: -19.61 to -4.78) and overexpression in 2 individuals (median z-score = 8.1 (range 4.80 - 11.40)), representing 19% of these variants. For missense homozygotes, we observed 158 individuals with significantly under-expressed cis-protein (median z-score = -6.95 (range: -28.16 to -3.95)) and 62 individuals over-expressed, median z-score = 5.65 (range: 4.57 to 25.08)). The majority (62%) of LoF knockout genes with an identified cis-protein effect had evidence from 2 or more platforms, highlighting the high confidence nature of these discoveries. Systematic clinical assessment of human knockouts with strong evidence of an impact on cis-protein abundance through multi-source electronic health record linkage enabled identification of 1) knockout carriers with rare disease features based on phenotypic similarity, 2) novel rare disease-causing variants, 3) evidence for reclassification of genes and variants of uncertain significance from ClinVar and rare disease panels, and 4) novel gene-phenotype associations in humans. Based on high confidence examples, we developed a machine learning model that predicted 1 in 4 pLOF and 9 in 10 missense variants are likely benign. In summary, our study provides strong human derived insights into the fundamental biology and clinical relevance of many genes and shows the value of proteogenomic studies of human knockout carriers. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement Genes & Health is/has recently been core-funded by Wellcome (WT102627, WT210561), the Medical Research Council (UK) (M009017, MR/X009777/1, MR/X009920/1), Higher Education Funding Council for England Catalyst, Barts Charity (845/1796), Health Data Research UK (for London substantive site), and research delivery support from the NHS National Institute for Health Research Clinical Research Network (North Thames). We acknowledge the support of the National Institute for Health and Care Research Barts Biomedical Research Centre (NIHR203330); a delivery partnership of Barts Health NHS Trust, Queen Mary University of London, St George's University Hospitals NHS Foundation Trust and St George's University of London Genes & Health is/has recently been funded by Alnylam Pharmaceuticals, Genomics PLC; and a Life Sciences Industry Consortium of AstraZeneca PLC, Bristol-Myers Squibb Company, GlaxoSmithKline Research and Development Limited, Maze Therapeutics Inc, Merck Sharp & Dohme LLC, Novo Nordisk A/S, Pfizer Inc, Takeda Development Centre Americas Inc. We thank Social Action for Health, Centre of The Cell, members of our Community Advisory Group, and staff who have recruited and collected data from volunteers. We thank the NIHR National Biosample Centre (UK Biocentre), the Social Genetic & Developmental Psychiatry Centre (King's College London), Wellcome Sanger Institute, and Broad Institute for sample processing, genotyping, sequencing and variant annotation. This work uses data provided by patients and collected by the NHS as part of their care and support. This research utilised Queen Mary University of London's Apocrita HPC facility, supported by QMUL Research-IT. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: G&H was approved by the London Southeast NRES Committee of the Health Research Authority (reference 14/LO/1240) on 16 Sept 2014. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. 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