Atypical hemolytic uremic syndrome (aHUS) is frequently associated in humans with loss-of-function mutations in complement-regulating proteins or gain-of-function mutations in complement-activating proteins. Thus, aHUS provides an archetypal complement-mediated disease with which to model new therapeutic strategies and treatments. Herein, we show that, when transferred to mice, an aHUS-associated gain-of-function change (D1115N) to the complement-activation protein C3 results in aHUS. Homozygous C3 p.D1115N (C3KI) mice developed spontaneous chronic thrombotic microangiopathy together with hematuria, thrombocytopenia, elevated creatinine, and evidence of hemolysis. Mice with active disease had reduced plasma C3 with C3 fragment and C9 deposition within the kidney. Therapeutic blockade or genetic deletion of C5, a protein downstream of C3 in the complement cascade, protected homozygous C3KI mice from thrombotic microangiopathy and aHUS. Thus, our data provide in vivo modeling evidence that gain-of-function changes in complement C3 drive aHUS. They also show that long-term C5 deficiency is not accompanied by development of other renal complications (such as C3 glomerulopathy) despite sustained dysregulation of C3. Our results suggest that this preclinical model will allow testing of novel complement inhibitors with the aim of developing precisely targeted therapeutics that could have application in many complement-mediated diseases.
Kate Smith-Jackson, Yi Yang, Harriet Denton, Isabel Y. Pappworth, Katie Cooke, Paul N. Barlow, John P. Atkinson, M. Kathryn Liszewski, Matthew C. Pickering, David Kavanagh, H. Terence Cook, Kevin J. Marchbank
Across clinical trials, T cell expansion and persistence following adoptive cell transfer (ACT) have correlated with superior patient outcomes. Herein, we undertook a pan-cancer analysis to identify actionable ligand/receptor pairs capable of compromising T cell durability following ACT. We discovered that FASLG, the gene encoding the apoptosis-inducing ligand FasL, is overexpressed within the majority of human tumor microenvironments (TMEs). Further, we uncovered that Fas, the receptor for FasL, is highly expressed on patient-derived T cells used for clinical ACT. We hypothesized that a cognate Fas-FasL interaction within the TME might limit both T cell persistence and anti-tumor efficacy. We discovered that genetic engineering of Fas variants impaired in the ability to bind FADD functioned as dominant negative receptors (DNRs), preventing FasL-induced apoptosis in Fas-competent T cells. T cells co-engineered with a Fas DNR and either a T cell receptor or chimeric antigen receptor exhibited enhanced persistence following ACT, resulting in superior anti-tumor efficacy against established solid and hematologic cancers. Despite increased longevity, Fas DNR-engineered T cells did not undergo aberrant expansion or mediate autoimmunity. Thus, T cell-intrinsic disruption of Fas signaling through genetic engineering represents a potentially universal strategy to enhance ACT efficacy across a broad range of human malignancies.
Tori N. Yamamoto, Ping-Hsien Lee, Suman K. Vodnala, Devikala Gurusamy, Rigel J. Kishton, Zhiya Yu, Arash Eidizadeh, Robert Eil, Jessica Fioravanti, Luca Gattinoni, James N. Kochenderfer, Terry J. Fry, Bulent Arman Aksoy, Jeffrey Hammerbacher, Anthony C. Cruz, Richard M. Siegel, Nicholas P. Restifo, Christopher A. Klebanoff
In the stomach, chronic inflammation causes metaplasia and creates a favorable environment for the evolution of gastric cancer. Glucocorticoids are steroid hormones that repress proinflammatory stimuli but their role in the stomach is unknown. In this study, we show that endogenous glucocorticoids are required to maintain gastric homeostasis. Removal of circulating glucocorticoids in mice by adrenalectomy resulted in the rapid onset of spontaneous gastric inflammation, oxyntic atrophy, and spasmolytic polypeptide-expressing metaplasia (SPEM), a precursor of gastric cancer. SPEM and oxyntic atrophy occurred independently of lymphocytes. However, depletion of monocytes and macrophages by clodronate treatment or inhibition of gastric monocyte infiltration using the Cx3cr1 knockout mouse model prevented SPEM development. Our results highlight the requirement for endogenous glucocorticoid signaling within the stomach to prevent spontaneous gastric inflammation and metaplasia and suggest that glucocorticoid deficiency may lead to gastric cancer development.
Jonathan T. Busada, Sivapriya Ramamoorthy, Derek W. Cain, Xiaojiang Xu, Donald N. Cook, John A. Cidlowski
Background/Purpose: Plasmacytoid dendritic cells (pDC) produce large amounts of type I IFN (IFN-I), cytokines convincingly linked to systemic lupus erythematosus (SLE) pathogenesis. BIIB059 is a humanized mAb that binds BDCA2, a pDC-specific receptor that inhibits the production of IFN-I and other inflammatory mediators when ligated. A first-in-human study was conducted to assess safety, tolerability, pharmacokinetic (PK) and pharmacodynamic (PD) effects of single BIIB059 doses in healthy volunteers (HV) and patients with SLE with active cutaneous disease as well as proof of biological activity and preliminary clinical response in the SLE cohort. Methods: A randomized, double-blind, placebo-controlled, clinical trial was conducted in HV (n=54) and patients with SLE (n=12). All subjects were monitored for adverse events. Serum BIIB059 concentrations, BDCA2 levels on pDCs, and IFN-responsive biomarkers in whole blood and skin biopsies were measured. Skin disease activity was determined using the Cutaneous Lupus Erythematosus Disease Area and Severity Index Activity (CLASI-A).Results: Single doses of BIIB059 were associated with a favorable safety and PK profile. BIIB059 administration led to BDCA2 internalization on pDCs, which correlated with circulating BIIB059 levels. BIIB059 administration in patients with SLE decreased expression of IFN response genes in blood, normalized MxA expression and reduced immune infiltrates in skin lesions, and decreased CLASI-A score. Conclusion: Single doses of BIIB059 were associated with favorable safety and PK/PD profiles, and robust target engagement and biological activity, supporting further development of BIIB059 in SLE. The data suggest that targeting pDCs may be beneficial for patients with SLE, especially those with cutaneous manifestations.
Richard Furie, Victoria P. Werth, Joseph F. Merola, Lauren Stevenson, Taylor L. Reynolds, Himanshu Naik, Wenting Wang, Romy Christmann, Agnes Gardet, Alex Pellerin, Stefan Hamann, Pavan Auluck, Catherine Barbey, Parul Gulati, Dania Rabah, Nathalie Franchimont
Allergen immunotherapy for patients with allergies begins with weekly escalating doses of allergen under medial supervision to monitor and treat IgE-mast cell mediated anaphylaxis. There is currently no treatment to safely desensitize mast cells to enable robust allergen immunotherapy with therapeutic levels of allergen. Here we demonstrated that liposomal nanoparticles bearing an allergen and a high-affinity glycan ligand of the inhibitory receptor CD33 profoundly suppressed IgE-mediated activation of mast cells, prevented anaphylaxis in transgenic mice with mast cells expressing human CD33, and desensitized mice from subsequent allergen challenge for several days. We showed that high levels of CD33 were consistently expressed on human skin mast cells, and that the antigenic-liposomes with CD33 ligand prevented IgE-mediated bronchoconstriction in slices of human lung. The results demonstrated the potential of exploiting CD33 to desensitize mast cells to provide a therapeutic window for administering allergen immunotherapy without triggering anaphylaxis.
Shiteng Duan, Cynthia J. Koziol-White, William F. Jester Jr., Corwin M. Nycholat, Matthew S. Macauley, Reynold A. Panettieri Jr., James C. Paulson
Immune checkpoint inhibitors and adoptive transfer of gene-engineered T cells have emerged as novel therapeutic modalities for hard-to-treat solid tumors; however, many patients are refractory to these immunotherapies, and the mechanisms underlying tumor immune resistance have not been fully elucidated. By comparing the tumor microenvironment of checkpoint inhibition-sensitive and -resistant murine solid tumors, we observed that the resistant tumors had low immunogenicity and lacked infiltration of CD8+ T cells at the tumor site. We identified antigen presentation by CD11b+F4/80+ tumor-associated macrophages (TAMs) as a key factor correlated with immune resistance. In the resistant tumors, TAMs remained inactive and did not exert antigen-presenting activity. Targeted delivery of a long peptide antigen to TAMs by using a nano-sized hydrogel (nanogel) in the presence of a Toll-like receptor agonist activated TAMs, induced their antigen-presenting activity, and thereby transformed the resistant tumors into tumors sensitive to adaptive immune responses such as adoptive transfer of tumor-specific T cell receptor-engineered T cells. These results indicate that the status and function of TAMs have a significant impact on tumor immune sensitivity and also that manipulation of TAM functions would be an effective approach for improving the efficacy of immunotherapies.
Daisuke Muraoka, Naohiro Seo, Tae Hayashi, Yoshiro Tahara, Keisuke Fujii, Isao Tawara, Yoshihiro Miyahara, Kana Okamori, Hideo Yagita, Seiya Imoto, Rui Yamaguchi, Mitsuhiro Komura, Satoru Miyano, Masahiro Goto, Shin-ichi Sawada, Akira Asai, Hiroaki Ikeda, Kazunari Akiyoshi, Naozumi Harada, Hiroshi Shiku
Genetic variants at the PTPN2 locus, which encodes the tyrosine phosphatase PTPN2, cause reduced gene expression and are linked to rheumatoid arthritis (RA) and other autoimmune diseases. PTPN2 inhibits signaling through the T cell and cytokine receptors and loss of PTPN2 promotes T cell expansion and CD4 and CD8-driven autoimmunity. However, it remains unknown whether loss of PTPN2 in FoxP3+ regulatory T cells (Treg) plays a role in autoimmunity. Here we aimed to model human autoimmune-predisposing PTPN2 variants, which results in a partial loss of PTPN2 expression, in mouse models of RA. We identified that reduced expression of Ptpn2 enhanced the severity of autoimmune arthritis in the T cell dependent SKG mouse model and demonstrated that this phenotype was mediated through a Treg-intrinsic mechanism. Mechanistically, we found that through dephosphorylation of STAT3, Ptpn2 inhibits IL-6-driven pathogenic loss of FoxP3 after Tregs have acquired RORγt expression, at a stage when chromatin accessibility for STAT3-targeted IL-17 associated transcription factors is maximized. We conclude that PTPN2 promotes FoxP3 stability in mouse RORγt+ Tregs and that loss of function of PTPN2 in Tregs contributes to the association between PTPN2 and autoimmunity.
Mattias N.D. Svensson, Karen M. Doody, Benjamin J. Schmiedel, Sourya Bhattacharyya, Bharat Panwar, Florian Wiede, Shen Yang, Eugenio Santelli, Dennis J. Wu, Cristiano Sacchetti, Ravindra Gujar, Grégory Seumois, William B. Kiosses, Isabelle Aubry, Gisen Kim, Piotr Mydel, Shimon Sakaguchi, Mitchell Kronenberg, Tony Tiganis, Michel L. Tremblay, Ferhat Ay, Pandurangan Vijayanand, Nunzio Bottini
Antibody-mediated rejection (AMR) is a principal cause of acute and chronic failure of lung allografts. However, mechanisms mediating this oftentimes fatal complication are poorly understood. Here, we show that Foxp3+ T cells formed aggregates in rejection-free human lung grafts and accumulated within induced bronchus-associated lymphoid tissue (BALT) of tolerant mouse lungs. Using a retransplantation model, we show that selective depletion of graft-resident Foxp3+ T lymphocytes resulted in the generation of donor-specific antibodies (DSA) and AMR, which was associated with complement deposition and destruction of airway epithelium. AMR was dependent on graft infiltration by B and T cells. Depletion of graft-resident Foxp3+ T lymphocytes resulted in prolonged interactions between B and CD4+ T cells within transplanted lungs, which was dependent on CXCR5-CXCL13. Blockade of CXCL13 as well as inhibition of the CD40 ligand and the ICOS ligand suppressed DSA production and prevented AMR. Thus, we have shown that regulatory Foxp3+ T cells residing within BALT of tolerant pulmonary allografts function to suppress B cell activation, a finding that challenges the prevailing view that regulation of humoral responses occurs peripherally. As pulmonary AMR is largely refractory to current immunosuppression, our findings provide a platform for developing therapies that target local immune responses.
Wenjun Li, Jason M. Gauthier, Ryuji Higashikubo, Hsi-Min Hsiao, Satona Tanaka, Linh Vuong, Jon H. Ritter, Alice Y. Tong, Brian W. Wong, Ramsey R. Hachem, Varun Puri, Ankit Bharat, Alexander S. Krupnick, Chyi S. Hsieh, William M. Baldwin III, Francine L. Kelly, Scott M. Palmer, Andrew E. Gelman, Daniel Kreisel
Mast cells (MCs) are immune sentinels but whether they also function as antigen-presenting cells (APCs) remains elusive. Using mouse models of MC-deficiency, we report MC-dependent recruitment and activation of multiple T cell subsets to the skin and draining lymph nodes (LNs) during dengue virus (DENV) infection. Newly-recruited and locally-proliferating γδT cells were the first responding T cell subset to MC-driven inflammation and their production of IFN-γ was MC-dependent. MC-γδ T cell conjugates were observed consistently in infected peripheral tissues, suggesting a new role for MCs as non-conventional APCs for γδT cells. MC-dependent γδT cell activation and proliferation during DENV infection required TCR signaling and the non-conventional antigen presentation molecule EPCR on MCs. γδT cells, not previously implicated in DENV host defense, killed infected target dendritic cells and contributed to clearance of DENV in vivo. We believe immune synapse formation between MCs and γδT cells is a novel mechanism to induce specific and protective immunity at sites of viral infection.
Chinmay Kumar Mantri, Ashley L. St. John
Adoptive transfer of T cell receptor–engineered (TCR-engineered) T cells is a promising approach in cancer therapy but needs improvement for more effective treatment of solid tumors. While most clinical approaches have focused on CD8+ T cells, the importance of CD4+ T cells in mediating tumor regression has become apparent. Regarding shared (self) tumor antigens, it is unclear whether the human CD4+ T cell repertoire has been shaped by tolerance mechanisms and lacks highly functional TCRs suitable for therapy. Here, TCRs against the tumor-associated antigen NY-ESO-1 were isolated either from human CD4+ T cells or from mice that express a diverse human TCR repertoire with HLA-DRA/DRB1*0401 restriction and are NY-ESO-1 negative. NY-ESO-1–reactive TCRs from the mice showed superior recognition of tumor cells and higher functional activity compared with TCRs from humans. We identified a candidate TCR, TCR-3598_2, which was expressed in CD4+ T cells and caused tumor regression in combination with NY-ESO-1–redirected CD8+ T cells in a mouse model of adoptive T cell therapy. These data suggest that MHC II–restricted TCRs against NY-ESO-1 from a nontolerant nonhuman host are of optimal affinity and that the combined use of MHC I– and II–restricted TCRs against NY-ESO-1 can make adoptive T cell therapy more effective.
Lucia Poncette, Xiaojing Chen, Felix K.M. Lorenz, Thomas Blankenstein