Mechanisms for the Development of Lung Fibrosis in Sting-Associated Vasculopathy with Onset in Infancy (SAVI)

Adriana Almeida de Jesus¹, Louise Malle¹, Dan Yang², Bernadette Marrero¹, Yin Liu³, Gina A. Montealegre Sanchez¹, Dawn C. Chapelle⁴, Hanna Kim⁴, Michelle O'Brien⁴, Gregor Dueckers⁵, Suzanne Ramsey⁶, Joseph R. Fontana⁷, Steven M. Holland⁸, Yan Huang¹, Suvimol Hill⁹, Laisa Santiago¹⁰, Benito Gonzalez¹¹, Paul Brogan¹², Juergen Brunner¹³, Ebun Omoyinmi¹⁴, Athimalaipet V Ramanan¹⁵, Amy Paller¹⁶, Olcay Y. Jones¹⁷, Seza Ozen¹⁸, Stephen Brooks⁴, Zuoming Deng⁴, Manfred Boehm¹⁹, Raphaela Goldbach-Mansky²⁰ and Helmut Wittkowski²¹, ¹National Institute of Allergy and Infectious Diseases (NIAID), NIH, Bethesda, MD, ²National Heart, Lung, and Blood Institute (NHLBI), NIH, Bethesda, MD, ³Scientific Review Branch, NIAMS, NIH, Bethesda, MD, ⁴NIAMS/NIH, Bethesda, MD, ⁵Helios Kliniken - Kinderklinik, HELIOS Klinikum Krefeld, Krefeld, Germany, ⁶Pediatric Rheumatology, IWK Health Centre, Halifax, NS, Canada, ⁷Cardiovascular and Pulmonary Branch, NHLBI, NIH, Bethesda, MD, ⁸Laboratory of Clinical Infectious Disease, NIAID, NIH, Bethesda, MD, ⁹Radiology Department, Clinical Center, NIH, Bethesda, MD, ¹⁰Johns Hopkins All Children's Hospital Rheumatology, Saint Petersburg, FL, ¹¹Luis Calvo Mackenna Hospital, Santiago, Chile, ¹²UCL Institute of Child Health and Great Ormond Street Hospital NHS Foundation Trust, London, United Kingdom, ¹³Department of Pediatrics, Medical University Innsbruck, Innsbruck, Austria, ¹⁴University College London Institute of Child Health, London, United Kingdom, ¹⁵University Hospitals Bristol NHS Foundation Trust, Bristol, United Kingdom, ¹⁶Departments of Dermatology and Pediatrics, Feinberg School of Medicine, Northwestern University, Chicago, IL, USA;, Chicago, IL, ¹⁷Pediatrics, Walter Reed National Military Medical Center, Bethesda, MD, ¹⁸Department of Pediatrics, Division of Rheumatology, Hacettepe University Faculty of Medicine, ANKARA, Turkey, ¹⁹Laboratory of Cardiovascular Regenerative Medicine, National Heart, Lung, and Blood Institute (NHLBI), NIH, Bethesda, MD, ²⁰Translational Autoinflammatory Disease Studies, National Institute of Allergy and Infectious Diseases (NIAID), NIH, Bethesda, MD, ²¹Department of Pediatric Rheumatology and Immunology, University Hospital of Muenster, Münster, Germany

Meeting: 2016 ACR/ARHP Annual Meeting

Date of first publication: September 28, 2016

Keywords: Autoinflammatory Disease, endothelial cells and interferons, Gene Expression, Lung Disease

Session Information

Date: Tuesday, November 15, 2016

Title: Pediatric Rheumatology – Pathogenesis and Genetics - Poster

Session Type: ACR Poster Session C

Session Time: 9:00AM-11:00AM

Background/Purpose: STING-Associated Vasculopathy with Onset in Infancy (SAVI) is a monogenic autoinflammatory interferonopathy caused by gain-of-function mutations in TMEM173/STING, a nucleic acid sensor adaptor linked to IFNβ transcription. Patients (Pts.) develop acral vasculitis and variable severity of interstitial lung disease (ILD)(Liu Y, NEJM, 2014). We hypothesized that a common STING variant modulates lung disease severity, and investigated a mechanistic role of STING in ILD.

Methods: We assessed chest computed tomography (CT) and pulmonary function tests (PFTs) in 12 SAVI pts. (N154S, V155M, V147L, V147M); lung biopsies were available for 7 pts. DNA samples were typed for the STING SNP R232H, rs1131769 (Yi G, PLoSOne 2013). Transfection studies in HEK293T cells with wildtype or “SAVI mutations” constructs on the R232 and H232 haplotypes, and stimulations of pt. fibroblast and endothelial cell lines (HUVECs) were performed. IFNB1luciferase reporter activity was assessed upon stimulation with the endogenous: 2’3’cGAMP, or the microbial: 3’3’cGAMP, c-di-AMP and c-di-GMP, STING activators. Gene expression (q-RT-PCR, RNA-seq), cytokine production, and endothelial-mesenchymal transition (endo-MT, by cell morphology and gene expression studies), were assessed.

Results: The presence of R232 in cis with the disease-causing STING mutations is associated with severe lung disease. Of 9 pts. with severe ILD, 4 pts. succumbed to pulmonary complications; all but one were homozygous for R232/R232. The pt. with no ILD was homozygous for the H232 allele (H232/H232); 2 pts. with mild ILD were heterozygous (R232/H232) for the SNP. Transfection of HEK293T cells with mutant STING on the R232 and H232 haplotypes showed increased IFNB1 expression upon stimulation with microbial stimulants but not endogenous cGAMP; which was only significant when the SAVI mutations were on the R232 but not on the H232 haplotype. Similarly, IFNB1transcription in response to exogenous stimuli in fibroblasts from 4 pts. were higher in cells with R232/R232 than in cells with H232/H232 haplotype. RNA-seq of stimulated pt. and control fibroblasts showed no expression of myofibroblast and extracellular matrix (ECM) markers ruling out that STING pathway activation would induce myofibroblast-differentiation as a mechanism for pulmonary fibrosis. Trichrome stains of SAVI lung tissues (n=2) revealed localized fibrosis around the pulmonary vessels, suggesting that STING pathway activation may induce endo-MT. Endogenous cGAMP stimulation on endothelial cells induced morphologic transition to fibroblasts with increased expression of the mesenchymal marker αSMA and the pro-fibrotic markers SLUG and TWIST. EndoMT was TGFβ independent and not induced by microbial STING stimuli. However, IFNβ significantly increased the endo-MT process over 2’3’cGAMP stimulation alone.

Conclusion: The minor allele H232 of a STING SNP confers a protective role by mitigating IFNβ responses in the context of STING activating infections while the R232 haplotype accelerates endo-MT amplified by IFNβ. Our findings suggest a novel mechanism for inducing endo-MT as a cause for the development of lung fibrosis in SAVI through endogenous STING activation.

Disclosure: A. Almeida de Jesus, None; L. Malle, None; D. Yang, None; B. Marrero, None; Y. Liu, None; G. A. Montealegre Sanchez, None; D. C. Chapelle, None; H. Kim, None; M. O'Brien, None; G. Dueckers, None; S. Ramsey, None; J. R. Fontana, None; S. M. Holland, None; Y. Huang, None; S. Hill, None; L. Santiago, None; B. Gonzalez, None; P. Brogan, None; J. Brunner, None; E. Omoyinmi, None; A. V. Ramanan, None; A. Paller, None; O. Y. Jones, None; S. Ozen, None; S. Brooks, None; Z. Deng, None; M. Boehm, None; R. Goldbach-Mansky, None; H. Wittkowski, None.

To cite this abstract in AMA style:

Almeida de Jesus A, Malle L, Yang D, Marrero B, Liu Y, Montealegre Sanchez GA, Chapelle DC, Kim H, O'Brien M, Dueckers G, Ramsey S, Fontana JR, Holland SM, Huang Y, Hill S, Santiago L, Gonzalez B, Brogan P, Brunner J, Omoyinmi E, Ramanan AV, Paller A, Jones OY, Ozen S, Brooks S, Deng Z, Boehm M, Goldbach-Mansky R, Wittkowski H. Mechanisms for the Development of Lung Fibrosis in Sting-Associated Vasculopathy with Onset in Infancy (SAVI) [abstract]. Arthritis Rheumatol. 2016; 68 (suppl 10). https://acrabstracts.org/abstract/mechanisms-for-the-development-of-lung-fibrosis-in-sting-associated-vasculopathy-with-onset-in-infancy-savi/. Accessed .

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