Association of Toll-like receptor 7 variants with life-threatening COVID-19 disease in males

Background COVID-19 clinical presentation ranges from asymptomatic to fatal outcome. This variability is due in part to host genome specific mutations. Recently, two families in which COVID-19 segregates like an X-linked recessive monogenic disorder environmentally conditioned by SARS-CoV-2 have been reported leading to identification of loss-of-function variants in TLR7 . Objective We sought to determine whether the two families represent the tip of the iceberg of a subset of COVID-19 male patients. Methods We compared male subjects with extreme phenotype selected from the Italian GEN-COVID cohort of 1178 SARS-CoV-2-infected subjects (<60y, 79 severe cases versus 77 control cases). We applied the LASSO Logistic Regression analysis, considering only rare variants on the young male subset, picking up TLR7 as the most important susceptibility gene. Results Rare TLR7 missense variants were predicted to impact on protein function in severely affected males and in none of the asymptomatic subjects. We then investigated a similar white European cohort in Spain, confirming the impact of TRL7 variants. A gene expression profile analysis in peripheral blood mononuclear cells after stimulation with TLR7 agonist demonstrated a reduction of mRNA level of TLR7, IRF7, ISG15, IFN-□ and IFN-γ in COVID-19 patients compared with unaffected controls demonstrating an impairment in type I and II INF responses. Conclusion Young males with TLR7 loss-of-function mutations and severe COVID-19 in the two reported families represent only a fraction of a broader and complex host genome situation. Specifically, missense mutations in the X-linked recessive TLR7 disorder may significantly contribute to disease susceptibility in up to 4% of severe COVID-19. Clinical Implication In this new yet complex scenario, our observations provide the basis for a personalized interferon-based therapy in patients with rare TLR7 variants. CAPSULE SUMMARY Our results in large cohorts from Italy and Spain showed that X-linked recessive TLR7 disorder may represent the cause of disease susceptibility to COVID-19 in up to 4% of severely affected young male cases. ### Competing Interest Statement The authors have declared no competing interest. ### Clinical Trial NCT04549831 ### Funding Statement We thank private donors for the support provided to A.R. (Department of Medical Biotechnologies, University of Siena) for the COVID19 host genetics research project (D.L n.18 of March 17, 2020). We also thank the COVID19 Host Genetics Initiative (https://www.covid19hg.org/). MIUR project Dipartimenti di Eccellenza 2018/2020 to the Department of Medical Biotechnologies University of Siena, Italy. Research on the Spanish cohort received funding from the European Unions Horizon 2020 research and innovation programme under grant agreement No 824110 EASI-Genomics, to AP. ### 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: The study (GEN-COVID) was consistent with Institutional guidelines and approved by the University Hospital (Azienda Ospedaliero-Universitaria Senese) Ethical Review Board, Siena, Italy (Prot n. 16929, dated March 16, 2020). A Spanish cohort, composed of 122 COVID-19 patients, was used to expand the cohort to another representative European population highly impacted by COVID-19. The Spanish Covid HGE cohort is under IRB approval PR127/20 from Bellvitge University Hospital, Barcelona, Spain. All necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived. 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 and uploaded the relevant EQUATOR Network research reporting checklist(s) and other pertinent material as supplementary files, if applicable. Yes The data and samples referenced here are housed in the GEN-COVID Patient Registry and the GEN-COVID Biobank and are available for consultation. For further information, you may contact the corresponding author, Prof. Alessandra Renieri (e-mail: alessandra.renieri{at}unisi.it). * Ala : Alanine amino acid Asp : Asparagine amino acid CADD : Combined Annotation Dependent Depletion COVID-19 : COronaVIrus Disease 19 His : Histidine amino acid HPRT1 : hypoxanthine phosphoribosyltransferase 1 INFs : Interferons IFN-α : Interferon Alpha IFN-β1 : Interferon Beta1 IFN-γ : Interferon Gamma IL-1 : Interleukin 1 IL-6 : Interleukin 6 Ile : Isoleucine amino acid IRF7 : Interferon Regulatory Factor 7 ISG15 : Interferon-Stimulated Gene 15 LASSO : Least Absolute Shrinkage and Selection Operator LBP : LipoPolySaccharide (LPS) : Binding Protein NIG : Network of Italian Genome PBMCs : Peripheral Blood Mononuclear Cells Pro : Proline amino acid RNA : RiboNucleic Acid Ser : Serine amino acid SARS-CoV-2 : Severe Acute Respiratory Syndrome caused by CoronaVirus Thr : Threonine amino acid TLRs : Toll-like Receptors TLR4 : Toll-like Receptor 4 TLR7 : Toll-like Receptor 7 TNF-α : Tumor Necrosis Factor α Tyr : Tyrosine amino acid Val : Valine amino acid WES : Whole Exome Sequencing