0567: Gain-of-Function Mutations in Lyn Kinase Cause a Novel Autoinflammatory Disease with Small Vessel Vasculitis and Liver Fibrosis

Adriana Almeida de Jesus¹, Guibin Chen², Dan Yang², Tomas Brdicka³, Natasha Ruth⁴, David Bennin⁵, Dita Cebecauerova⁶, Hana Malcova⁷, Helen Freeman⁸, Neil Martin⁹, Karel Svojgr⁶, Murray Passo⁴, Farzana Bhuyan¹⁰, Sara Alehashemi¹¹, Andre Rastegar¹², Kat Uss¹², Lela Kardova¹², Iris Duric³, Ebun Omoyinmi¹³, Petra Peldova⁶, Chyi-Chia Richard Lee¹⁴, David Kleiner¹⁴, Colleen Hadigan¹⁵, Stephen Hewitt¹⁴, Stefania Pittaluga¹⁴, Carmelo Carmona-Rivera¹⁶, Katherine R. Calvo¹⁵, Nirali Shah¹⁴, Miroslava Balascakova⁶, Danielle Fink¹⁷, Radana Kotalova⁶, Zuzana Parackova⁶, Lucie Peterkova⁶, Daniela Kuzilkova⁶, Vit Campr⁶, Lucie Sramkova⁶, Stephen Brooks¹⁶, Eric Meffre¹⁸, Rebecca Harper², Hyesun Kuehn¹⁵, Mariana Kaplan¹⁶, Paul Brogan¹⁹, Sergio Rosenzweig¹⁵, Zuoming Deng¹⁶, Anna Huttenlocher⁵, Susan Moir¹², Douglas Kuhns¹⁷, Manfred Boehm², Karolina Skvarova Kramarzova²⁰ and Raphaela Goldbach-Mansky²¹, ¹NIAID, NIH, Silver Spring, MD, ²National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, MD, ³Laboratory of Leukocyte Signaling, Institute of Molecular Genetics of the Academy of Sciences of Czech Republic (ASCR), Prague, Czech Republic, ⁴Medical University of South Carolina, Charleston, SC, ⁵Departments of Pediatrics and Medical Microbiology and Immunology, University of Wisconsin-Madison, Madison, WI, ⁶Second Faculty of Medicine, Charles University/University Hospital Motol, Prague, Czech Republic, ⁷Second Faculty of Medicine, Charles University/University Hospital Motol, Praha, Czech Republic, ⁸Raigmore Hospital, Inverness, Scotland, United Kingdom, ⁹Royal Hospital for Children, Glasgow, Scotland, United Kingdom, ¹⁰National Institutes of Health, Bethesda, MD, ¹¹NIH/NIAID/TADS, Clarksville, MD, ¹²National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, ¹³Great Ormond Street Hospital for Children NHS Foundation Trust, London, United Kingdom, ¹⁴National Cancer Institute, National Institutes of Health, Bethesda, MD, ¹⁵Clinical Center, National Institutes of Health, Bethesda, MD, ¹⁶National Institute of Arthritis and Musculoskeletal and Skin Diseases, National Institutes of Health, Bethesda, MD, ¹⁷National Institute of Allergy and Infectious Diseases, National Institutes of Health, Frederick, MD, ¹⁸Yale University, New Haven, CT, ¹⁹UCL Institute of Child Health and Great Ormond Street Hospital NHS Foundation Trust, London, United Kingdom, ²⁰Second Faculty of Medicine, Charles University/University Hospital Motol, Prague, ²¹NIH/NIAID, Potomac, MD

Background/Purpose: Lyn kinase is a member of the Src family of tyrosine kinases that regulates innate and adaptive immune responses. Lyn C-terminal tail tyrosine residue, p.Y508, inactivates wildtype Lyn kinase upon phosphorylation. We describe 3 unrelated patients with de novo LYN mutations affecting p.Y508, whose disease sheds light on the pathogenesis of neutrophilic small vessel vasculitis.

Methods: All patients (Pts) were enrolled into an IRB-approved natural history protocol (NCT02974595). The effects of the LYN mutations were investigated in Pts' and healthy control (HC) blood cells and serum, in transfection and immunological studies and in induced pluripotent stem cells (iPS cells)-derived endothelial cells (iECs).

Results: The three Pts presented with perinatal purpuric rash and systemic inflammation, hepatosplenomegaly, periorbital erythema, abdominal and testicular pain. Pts 1 and 3 had early onset liver fibrosis. Transiently elevated autoantibodies (ANA, anti-Sm, anti-SSA) were present in Pt 1. All patients had a skin biopsy showing neutrophilic small vessel vasculitis. The 3 Pts were found to have de novo mutations in LYN: p.Y508* in Pt 1, p.Y508F in Pt 2 and p.Q507* in Pt 3 and responded partially (Pts 1 and 3) or fully (Pt 2) to the TNF inhibitor etanercept. Combination therapy with the Src kinase inhibitor, dasatinib, led to resolution of liver fibrosis in Pt 1. Transfection of wildtype and mutant Lyn into HEK293FT cells showed constitutive phosphorylation of the activating tyrosine, p.Y397, and absent phosphorylation of the inhibitory tyrosine, p.Y508. Skin biopsies showed a dense neutrophilic infiltrate with small vessel wall destruction and neutrophil extracellular traps (NETs) formation; ICAM-1 was upregulated and focally colocalized with the EC marker CD31. All Pts showed elevated pro-inflammatory cytokines, and/or markers of neutrophil (L-selectin) and EC activation (ICAM-1, E-selectin) in serum. Monocyte-derived macrophages released increased levels of TNF-a, IL-6, CXCL10 and sICAM-1 upon LPS stimulation. Neutrophils from Pt 1 showed signs of constitutive activation, including high surface expression of b2-integrins. Expression of ICAM-1 in areas of adherent neutrophils were increased in Pt 1 iEC/HC neutrophil co-cultures compared to isogenic and HC iEC. Neutrophil adhesion and ICAM-1 clusters were blocked with dasatinib and less with the TNF inhibitor. Transendothelial migration of HC neutrophils through Pt 1 iECs was increased compared to isogenic or HC iECs and reduced by treatment with dasatinib or TNF inhibitor.

Conclusion: We describe a novel autoinflammatory disease which we called Lyn associated vasculopathy and liver fibrosis (LAVLI). De novo activating LYN mutations link Lyn kinase to regulating dermal vascular permeability, restricting inflammatory signals in immune cells, and promoting hepatic fibrosis. Functional data support a stepwise approach to treatment of reserving dasatinib for patients with liver fibrosis and insufficient responses to TNF inhibition. This study identifies Lyn kinase as potential treatment target in neutrophilic small vessel vasculitis and early liver fibrosis.


This study was supported in part by the IRP of NIAID, NIH.

Disclosures: A. Almeida de Jesus, None; G. Chen, None; D. Yang, None; T. Brdicka, None; N. Ruth, None; D. Bennin, None; D. Cebecauerova, None; H. Malcova, None; H. Freeman, None; N. Martin, None; K. Svojgr, None; M. Passo, None; F. Bhuyan, None; S. Alehashemi, None; A. Rastegar, None; K. Uss, None; L. Kardova, None; I. Duric, None; E. Omoyinmi, None; P. Peldova, None; C. Lee, None; D. Kleiner, None; C. Hadigan, None; S. Hewitt, None; S. Pittaluga, None; C. Carmona-Rivera, None; K. Calvo, None; N. Shah, None; M. Balascakova, None; D. Fink, None; R. Kotalova, None; Z. Parackova, None; L. Peterkova, None; D. Kuzilkova, None; V. Campr, None; L. Sramkova, None; S. Brooks, None; E. Meffre, None; R. Harper, None; H. Kuehn, None; M. Kaplan, None; P. Brogan, None; S. Rosenzweig, None; Z. Deng, None; A. Huttenlocher, None; S. Moir, None; D. Kuhns, None; M. Boehm, None; K. Skvarova Kramarzova, None; R. Goldbach-Mansky, None.