New York School of Medicine Ann Arbor, MI, United States
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Jasmine Shwetar1, Katie Preisinger1, Devyn Zaminski2, Philip Carlucci1, Kristina Deonaraine1, Qian Xiao3, Joseph Mears4, Siddarth Gurajala3, Izmirly peter5, Judith James6, Joel Guthridge6, Andrea Fava7, Brad Rovin8, Wade DeJager6, Ming Wu9, Deepak Rao10, Chaim Putterman11, Betty Diamond12, Derek Fine13, Jose Monroy-Trujillo13, Kristin Haag14, H Michael Belmont2, William Apruzzese10, Anne Davidson12, Fernanda Payan-Schober15, Richard Furie16, Paul Hoover10, Celine Berthier17, Maria Dall'Era18, Kerry Cho19, Diane L. Kamen20, Kenneth Kalunian21, Jennifer Anolik22, Soumya Raychaudhuri10, Nir Hacohen23, Michelle Petri24, Robert Clancy25, David Wofsy18, Arnon Arazi26, Kelly Ruggles9, Jill Buyon25 and The Accelerating Medicines Partnership SLE/RA27, 1New York University School of Medicine, New York, NY, 2NYU School of Medicine, New York, NY, 3Harvard Medical School, Boston, MA, 4Michigan University, Ann Arbor, MI, 5NYU, New York, NY, 6Oklahoma Medical Research Foundation, Oklahoma City, OK, 7Johns Hopkins University, Baltimore, MD, 8Ohio State University, Columbus, OH, 9NYU Langone, New York, NY, 10Brigham and Women's Hospital, Boston, MA, 11Albert Einstein College of Medicine, Bronx, NY, 12Feinstein Institutes for Medical Research, Manhasset, NY, 13Johns Hopkins School of Medicine, Baltimore, MD, 14Thomas Jefferson University, Philadelphia, PA, 15Texas Tech University Health Sciences Center, El Paso, TX, 16Northwell Health, Manhasset, NY, 17University of Michigan, Ann Arbor, MI, 18University of California San Francisco, San Francisco, CA, 19UCSF Health, San Francisco, CA, 20Medical University of South Carolina, Charleston, SC, 21University of California San Diego, La Jolla, CA, 22University of Rochester Medical Center, Rochester, NY, 23Broad Institute of MIT and Harvard, Cambridge, MA, 24Department of Medicine, Division of Rheumatology, Johns Hopkins University School of Medicine, Timonium, MD, 25NYU Grossman School of Medicine, New York, NY, 26Broad Institute of MIT and Harvard, Melrose, MA, 27University of Colorado Anschutz Medical Campus, Aurora, CO
Background/Purpose: Lupus Nephritis (LN) significantly reduces the survival and life expectancy of patients with SLE. Given this, considerable effort has gone into characterizing the histologic classes that confer the highest standardized mortality ratios. Although Class II LN is considered a milder form of disease which often requires less aggressive treatment, a growing body of clinical evidence suggests that these patients can progress to more advanced LN classes. Accordingly, this study leveraged the multi-center LN Accelerating Medicine Partnership (AMP) Network to focus on the transcriptome of Class II kidney biopsies with comparison to other histologic classes to provide insights into the molecular underpinnings of early disease and potential drivers of progression.
Methods: LN patients were enrolled in AMP at the time of a clinically indicated kidney biopsy and followed for one year. scRNA-seq was performed using the 10x genomics sequencing platform and quality control was performed to retain high-quality cells. Using the AMP consortium dataset which includes patients with membranous, mixed and proliferative LN, a reference-based mapping approach was employed to identify shared cell states and insights into Class II pathogenesis and outcomes.
Results: scRNA-Seq from 12 patients with Class II LN biopsies yielded 25,483 high-quality cells comprised of 11 parenchymal and three immune cell types (Fig 1). Reference mapping identified B, T and myeloid cells which are infrequently found in Class II LN by conventional microscopy (Fig 2A). In the myeloid compartment, cell type proportion analysis revealed a distinct signature with fewer monocytes and a greater number of differentiated and phagocytic macrophages in Class II compared to control and other LN classes (Fig 2B). This distinct myeloid signature was also identified in proliferative LN samples with high activity and low chronicity (activity index >= 4, chronicity index = 0). In both Class II and proliferative LN with high activity, monocytes and phagocytic macrophage subtypes were contracted and expanded relative to biopsies from healthy donors, respectively (Fig. 2C). The stromal compartment of Class II unexpectedly had a higher proportion of fibroblasts and myofibroblasts compared to all other classes (3A-B). These myofibroblasts were characterized by high expression of canonical markers, such as ACTA2 as well as markers of immune activation such as CTGF, CCL19, and CCL21 (3C). The proportion of myofibroblasts was higher in Class II patients whose UPCR fell below 0.5 and creatinine remained normal or did not exceed 125% of baseline compared to Class II patients considered non-responders at 52 weeks (3D, p = 0.048).
Conclusion: These data support that histology provides an incomplete picture of the molecular and cellular landscape in Class II LN. Increased phagocytic macrophage activity and immune-induced myofibroblast differentiation were associated with higher disease activity, highlighting these cellular composition changes as potential drivers of non-responsiveness and progression.
Figure 1: UMAP representation of all cells in AMP dataset from control and membranous, proliferative, and mixed LN biopsies (left); UMAP representation generated by Symphony reference mapping of all cells from Class II LN biopsies (right).
Figure 2: (A) UMAP representation of myeloid cells in AMP (left); UMAP representation generated by Symphony reference mapping of Class II myeloid cells (right). (B) Per disease condition proportion of cells in each myeloid subcluster (control = light gray; proliferative, membranous or mixed LN = dark gray; Class II LN = red) (C) Per sample proportion of cells in various monocyte (top) and macrophage (bottom) subclusters. All comparisons of the median proportion of cells in Class II LN versus control were significant (p<0.05) using Mann-U Whitney test. When comparing Class II LN with high activity low chronicity proliferative LN, only the porportion of differentiating CD16+ Monocytes and Phagocytic macrophages were found to be significant (p = 0.033, 0.003 respectively). Control = red; Class II LN = green; High activity low chronicity proliferative LN = blue.
Figure 3: (A) UMAP representation of stromal cells in AMP dataset (left); UMAP representation generated by Symphony reference mapping of Class II stromal cells (right). (B) Proportion of fibroblast and myofibroblast cells in each disease class. P-values calculated using Mann-U Whitney test. (C) Heatmap of markers in Class II fibroblasts (red) and myofibroblasts (blue). (D) Proportion of myofibroblasts in Class II LN patients that were complete responders to treatment (CR) and non-responders to treatment (NR). P-value comparing median myofibroblast proportion using Mann-U Whitney test.
J. Shwetar: None; K. Preisinger: None; D. Zaminski: None; P. Carlucci: None; K. Deonaraine: None; Q. Xiao: None; J. Mears: None; S. Gurajala: None; I. peter: None; J. James: Bristol-Myers Squibb(BMS), 5, GlaxoSmithKlein(GSK), 2, Novartis, 2, Progentec Biosciences, 5; J. Guthridge: None; A. Fava: Annexon Biosciences, 2, Sanofi, 1; B. Rovin: AstraZeneca, 2, 5, Aurinia, 2, 5, Biogen, 2, F. Hoffmann-La Roche Ltd, 2, Genentech, 2, GlaxoSmithKlein(GSK), 2, Novartis, 2; W. DeJager: None; M. Wu: None; D. Rao: AstraZeneca, 2, Bristol-Myers Squibb, 2, 5, GlaxoSmithKlein(GSK), 2, Hifibio, 2, Janssen, 5, Merck, 5, Scipher Medicine, 2; C. Putterman: Equillium, 2, KidneyCure, 1, Progentec, 2; B. Diamond: Alpine, 12, DSMB, DBV, 2, 2, IMT, 2, Kyverna, 2, Nighthawk, 2, ONO, 2; D. Fine: None; J. Monroy-Trujillo: None; K. Haag: None; H. Belmont: Alexion, 6, Aurinia, 6; W. Apruzzese: None; A. Davidson: None; F. Payan-Schober: None; R. Furie: Biogen, 2, 5; P. Hoover: None; C. Berthier: None; M. Dall'Era: Annexon Biosciences, 2, 5, AstraZeneca, 2, Aurinia, 2, Biogen, 2, GlaxoSmithKlein, 2, 5, Pfizer, 2; K. Cho: None; D. Kamen: None; K. Kalunian: AbbVie/Abbott, 2, Amgen, 5, AstraZeneca, 2, Aurinia, 2, Bristol-Myers Squibb(BMS), 2, Eli Lilly, 2, EquilliumBio, 2, Genentech, 2, Gilead, 2, Janssen, 2, KezarBio, 1, Merck/MSD, 2, Novartis, 2, Pfizer, 2, Remegene, 2, Roche, 2, UCB, 5; J. Anolik: None; S. Raychaudhuri: AbbVie, 6, Janssen, 1, Mestag, Inc, 2, 8, Pfizer, 1, Sanofi, 1, Sonoma, 1, 8; N. Hacohen: None; M. Petri: Alexion, 1, Amgen, 1, AnaptysBio, 1, Annexon Bio, 1, Argenx, 1, Arhros-Focus Med/Ed, 6, AstraZeneca, 1, 5, Aurinia, 1, 5, 6, Axdev, 1, Biogen, 1, Boxer Capital, 2, Cabaletto Bio, 2, Caribou Biosciences, 2, CVS Health, 1, Eli Lilly, 1, 5, Emergent Biosolutions, 1, Exagen, 5, Exo Therapeutics, 2, Gilead Biosciences, 2, GlaxoSmithKlein(GSK), 1, 5, 6, Horizon Therapeutics, 2, Idorsia Pharmaceuticals, 2, IQVIA, 1, Janssen, 1, 5, Kira Pharmaceuticals, 2, MedShr, 6, Merck/EMD Serono, 1, Momenta Pharmaceuticals, 2, Nexstone Immunology, 2, Nimbus Lakshmi, 2, Proviant, 2, Sanofi, 2, Sinomab Biosciences, 2, Thermofisher, 5, UCB, 2; R. Clancy: None; D. Wofsy: Amgen, 7, Novartis, 7; A. Arazi: None; K. Ruggles: None; J. Buyon: Bristol-Myers Squibb(BMS), 2, GlaxoSmithKlein(GSK), 2, Related Sciences, 1; T. SLE/RA: None.
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