Tumor Regression and Delayed Onset Toxicity Following B7-H4 CAR T Cell Therapy.

B7-H4 protein is frequently overexpressed in ovarian cancer. Here, we engineered T cells with novel B7-H4-specific chimeric antigen receptors (CARs) that recognized both human and murine B7-H4 to test the hypothesis that B7-H4 CAR T cell therapy can be applied safely in preclinical models. B7-H4 CAR T cells specifically secreted IFN-γ and lysed B7-H4(+) targets. In vivo, B7-H4 CAR T cells displayed antitumor reactivity against B7-H4(+) human ovarian tumor xenografts. Unexpectedly, B7-H4 CAR T cell treatment reproducibly showed delayed, lethal toxicity 6-8 weeks after therapy. Comprehensive assessment of murine B7-H4 protein distribution uncovered expression in ductal and mucosal epithelial cells in normal tissues. Postmortem analysis revealed the presence of widespread histologic lesions that correlated with B7-H4(+) expression, and were inconsistent with graft versus host disease. Lastly, expression patterns of B7-H4 protein in normal human tissue were comparable to distribution in mice, advancing our understanding of B7-H4. We conclude that B7-H4 CAR therapy mediates control of cancer outgrowth. However, long-term engraftment of B7-H4 CAR T cells mediates lethal, off-tumor toxicity that is likely due to wide expression of B7-H4 in healthy mouse organs. This model system provides a unique opportunity for preclinical evaluation of safety approaches that limit CAR-mediated toxicity after tumor destruction in vivo.

[1]  J. Cheville,et al.  Genomic Organization and Expression , 2003 .

[2]  林雨樵.  Targeting Fibroblast Activation Protein in Tumor Stroma with Chimeric Antigen Receptor T Cells Can Inhibit Tumor Growth and Augment Host Immunity without Severe Toxicity , 2015 .

[3]  Z. Eshhar,et al.  Suppression of murine colitis and its associated cancer by carcinoembryonic antigen-specific regulatory T cells. , 2014, Molecular therapy : the journal of the American Society of Gene Therapy.

[4]  S. Gottschalk,et al.  Design and development of therapies using chimeric antigen receptor‐expressing T cells , 2014, Immunological reviews.

[5]  J. Allison,et al.  Ablation of B7-H3 but Not B7-H4 Results in Highly Increased Tumor Burden in a Murine Model of Spontaneous Prostate Cancer , 2015, Cancer Immunology Research.

[6]  B. Monk,et al.  Latest research and treatment of advanced-stage epithelial ovarian cancer , 2013, Nature Reviews Clinical Oncology.

[7]  D. Powell,et al.  CD27 costimulation augments the survival and antitumor activity of redirected human T cells in vivo. , 2012, Blood.

[8]  S. Ménard,et al.  Characterization of human ovarian carcinoma‐associated antigens defined by novel monoclonal antibodies with tumor‐restricted specificity , 1987, International journal of cancer.

[9]  Z. Eshhar,et al.  Expression of immunoglobulin-T-cell receptor chimeric molecules as functional receptors with antibody-type specificity. , 1989, Proceedings of the National Academy of Sciences of the United States of America.

[10]  G. Zhu,et al.  B7-H4-deficient mice display augmented neutrophil-mediated innate immunity. , 2009, Blood.

[11]  P. Loke,et al.  B7x: A widely expressed B7 family member that inhibits T cell activation , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[12]  Matthew J. Frigault,et al.  Identification of Chimeric Antigen Receptors That Mediate Constitutive or Inducible Proliferation of T Cells , 2015, Cancer Immunology Research.

[13]  Jun Sik Lee,et al.  Host B7x Promotes Pulmonary Metastasis of Breast Cancer , 2013, The Journal of Immunology.

[14]  Bernd Hauck,et al.  Chimeric antigen receptor-modified T cells for acute lymphoid leukemia. , 2013, The New England journal of medicine.

[15]  Raphael Sandaltzopoulos,et al.  Chimeric Antigen Receptor T Cells with Dissociated Signaling Domains Exhibit Focused Antitumor Activity with Reduced Potential for Toxicity In Vivo , 2013, Cancer Immunology Research.

[16]  A. Bagg,et al.  Chimeric antigen receptor-modified T cells in chronic lymphoid leukemia. , 2011, The New England journal of medicine.

[17]  M. Lefranc,et al.  The B7 family of immunoregulatory receptors: a comparative and evolutionary perspective. , 2009, Molecular immunology.

[18]  J. Dipersio,et al.  Factors affecting human T cell engraftment, trafficking, and associated xenogeneic graft-vs-host disease in NOD/SCID beta2mnull mice. , 2007, Experimental hematology.

[19]  C. Contag,et al.  In vivo analyses of early events in acute graft-versus-host disease reveal sequential infiltration of T-cell subsets. , 2005, Blood.

[20]  G. Warnock,et al.  B7-H4 Expression in Normal and Diseased Human Islet &bgr; Cells , 2014, Pancreas.

[21]  M. Azuma,et al.  Intrinsic and extrinsic control of expression of the immunoregulatory molecule PD-L1 in epithelial cells and squamous cell carcinoma. , 2015, Oral oncology.

[22]  J. Serody,et al.  In vivo imaging of graft-versus-host-disease in mice. , 2004, Blood.

[23]  P. Loke,et al.  B7x in the Periphery Abrogates Pancreas-Specific Damage Mediated by Self-reactive CD8 T Cells , 2012, The Journal of Immunology.

[24]  Etienne Weiss,et al.  Therapeutic antibodies: successes, limitations and hopes for the future , 2009, British journal of pharmacology.

[25]  D. Powell,et al.  Novel recombinant human b7-h4 antibodies overcome tumoral immune escape to potentiate T-cell antitumor responses. , 2013, Cancer research.

[26]  D. Dangaj,et al.  Isolation and Validation of Anti-B7-H4 scFvs from an Ovarian Cancer scFv Yeast-Display Library. , 2015, Methods in molecular biology.

[27]  L. Pirofski,et al.  Tissue-Expressed B7x Affects the Immune Response to and Outcome of Lethal Pulmonary Infection , 2012, The Journal of Immunology.

[28]  Min Wang,et al.  B7-H4 expression and its role in interleukin-2/interferon treatment of clear cell renal cell carcinoma , 2014, Oncology letters.

[29]  D. Greiner,et al.  Human peripheral blood leucocyte non‐obese diabetic‐severe combined immunodeficiency interleukin‐2 receptor gamma chain gene mouse model of xenogeneic graft‐versus‐host‐like disease and the role of host major histocompatibility complex , 2009, Clinical and experimental immunology.

[30]  G. Zhu,et al.  Potential Role of Decoy B7-H4 in the Pathogenesis of Rheumatoid Arthritis: A Mouse Model Informed by Clinical Data , 2009, PLoS medicine.

[31]  J. Cheville,et al.  Genomic Organization and Expression Analysis of B7-H4, an Immune Inhibitory Molecule of the B7 Family 1 , 2003, The Journal of Immunology.

[32]  Joanne Leung.  Tumor Transplantation Model Myeloid-Derived Suppressor Cells in a 4T1 Responses through Inhibition of Host B7-H4 Regulates Antitumor T Cell , 2013 .

[33]  M. Lucia,et al.  B7-H4 overexpression in ovarian tumors. , 2006, Gynecologic oncology.

[34]  A. Rynda-Apple,et al.  NKG2D CAR T‐cell therapy inhibits the growth of NKG2D ligand heterogeneous tumors , 2013, Immunology and cell biology.

[35]  J. Cheville,et al.  B7-H4 expression in renal cell carcinoma and tumor vasculature: Associations with cancer progression and survival , 2006, Proceedings of the National Academy of Sciences.

[36]  D. Powell,et al.  Strain-dependent Lethal Toxicity in NKG2D Ligand-targeted CAR T-cell Therapy. , 2015, Molecular therapy : the journal of the American Society of Gene Therapy.

[37]  B. Levine,et al.  Multiple injections of electroporated autologous T cells expressing a chimeric antigen receptor mediate regression of human disseminated tumor. , 2010, Cancer research.

[38]  B. Kong,et al.  B7-H4 Expression Promotes Tumorigenesis in Ovarian Cancer , 2009, International Journal of Gynecologic Cancer.

[39]  K. Shroyer,et al.  B7-H4 (DD-O110) is overexpressed in high risk uterine endometrioid adenocarcinomas and inversely correlated with tumor T-cell infiltration. , 2007, Gynecologic oncology.

[40]  Yuzhang Wu,et al.  The characteristic expression of B7‐H3 and B7‐H4 in liver biopsies from patients with HBV‐related acute‐on‐chronic liver failure , 2012, Pathology international.

[41]  S. Almo,et al.  Structure and cancer immunotherapy of the B7 family member B7x. , 2014, Cell reports.

[42]  C. Dong,et al.  B7S1, a novel B7 family member that negatively regulates T cell activation. , 2003, Immunity.

[43]  D. Powell,et al.  B7-H4 as a potential target for immunotherapy for gynecologic cancers: a closer look. , 2014, Gynecologic oncology.

[44]  G. Zhu,et al.  B7-H4, a molecule of the B7 family, negatively regulates T cell immunity. , 2003, Immunity.

[45]  S. Gottschalk,et al.  CAR T Cells for Solid Tumors: Armed and Ready to Go? , 2014, Cancer journal.

[46]  P. Loke,et al.  Tissue-specific expression of B7x protects from CD4 T cell–mediated autoimmunity , 2011, The Journal of experimental medicine.

[47]  J. DeVoss,et al.  An anti-B7-H4 antibody-drug conjugate for the treatment of breast cancer. , 2015, Molecular pharmaceutics.

[48]  M. Jordana,et al.  Generation and Characterization of B7-H4/B7S1/B7x-Deficient Mice , 2006, Molecular and Cellular Biology.

[49]  Pamela A Shaw,et al.  Chimeric antigen receptor T cells for sustained remissions in leukemia. , 2014, The New England journal of medicine.

[50]  G. Coukos,et al.  Redirected antitumor activity of primary human lymphocytes transduced with a fully human anti-mesothelin chimeric receptor. , 2012, Molecular therapy : the journal of the American Society of Gene Therapy.

[51]  Songbing He,et al.  The coexpression and clinical significance of costimulatory molecules B7-H1, B7-H3, and B7-H4 in human pancreatic cancer , 2014, OncoTargets and therapy.

[52]  D. Campana,et al.  Chimeric receptors containing CD137 signal transduction domains mediate enhanced survival of T cells and increased antileukemic efficacy in vivo. , 2009, Molecular therapy : the journal of the American Society of Gene Therapy.

[53]  A. Hagenbeek,et al.  A new xenograft model for graft-versus-host disease by intravenous transfer of human peripheral blood mononuclear cells in RAG2-/- gammac-/- double-mutant mice. , 2003, Blood.

[54]  S. Rosenberg,et al.  Immune targeting of fibroblast activation protein triggers recognition of multipotent bone marrow stromal cells and cachexia , 2013, The Journal of experimental medicine.

[55]  G. Denisova,et al.  T Cells Engineered With Chimeric Antigen Receptors Targeting NKG2D Ligands Display Lethal Toxicity in Mice. , 2015, Molecular therapy : the journal of the American Society of Gene Therapy.

[56]  S. Rosenberg,et al.  Case report of a serious adverse event following the administration of T cells transduced with a chimeric antigen receptor recognizing ERBB2. , 2010, Molecular therapy : the journal of the American Society of Gene Therapy.

[57]  M. Ghosh,et al.  The immunomodulatory protein B7-H4 is overexpressed in breast and ovarian cancers and promotes epithelial cell transformation. , 2005, Experimental cell research.

[58]  Z. Ruan,et al.  Expression of the novel co-stimulatory molecule B7-H4 by renal tubular epithelial cells. , 2006, Kidney international.

[59]  B7-H4 Expression by Nonhematopoietic Cells in the Tumor Microenvironment Promotes Antitumor Immunity , 2014, Cancer Immunology Research.

[60]  K. Shroyer,et al.  Intracellular B7-H4 Suppresses Bile Duct Epithelial Cell Apoptosis in Human Primary Biliary Cirrhosis , 2011, Inflammation.

[61]  Gang Li,et al.  The costimulatory molecule B7-H4 promote tumor progression and cell proliferation through translocating into nucleus , 2013, Oncogene.

[62]  B. Seliger,et al.  B7-H4 Expression in Human Melanoma: Its Association with Patients' Survival and Antitumor Immune Response , 2011, Clinical Cancer Research.