The Optimal Antigen Response of Chimeric Antigen Receptors Harboring the CD3ζ Transmembrane Domain Is Dependent upon Incorporation of the Receptor into the Endogenous TCR/CD3 Complex

Chimeric Ag receptors (CARs) expressed in T cells permit the redirected lysis of tumor cells in an MHC-unrestricted manner. In the Jurkat T cell model system, expression of a carcinoembryonic Ag-specific CD3ζ CAR (MFEζ) resulted in an increased sensitivity of the transduced Jurkat cell to generate cytokines when stimulated through the endogenous TCR complex. This effect was driven through two key characteristics of the MFEζ CAR: 1) receptor dimerization and 2) the interaction of the CAR with the endogenous TCR complex. Mutations of the CAR transmembrane domain that abrogated these interactions resulted in a reduced functional capacity of the MFEζ CAR to respond to carcinoembryonic Ag protein Ag. Taken together, these results indicate that CARs containing the CD3ζ transmembrane domain can form a complex with the endogenous TCR that may be beneficial for optimal T cell activation. This observation has potential implications for the future design of CARs for cancer therapy.

[1]  D. Gilham,et al.  Development of a flow cytometric co‐immunoprecipitation technique for the study of multiple protein—protein interactions and its application to T‐cell receptor analysis , 2010, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[2]  Carlos Prieto,et al.  Human Gene Coexpression Landscape: Confident Network Derived from Tissue Transcriptomic Profiles , 2008, PloS one.

[3]  W. Schamel,et al.  The extracellular part of zeta is buried in the T cell antigen receptor complex. , 2008, Immunology letters.

[4]  B. Alarcón,et al.  T Cell Receptor Engagement Triggers Its CD3ε and CD3ζ Subunits to Adopt a Compact, Locked Conformation , 2008, PloS one.

[5]  E. Rieber,et al.  Targeting of tumor cells expressing the prostate stem cell antigen (PSCA) using genetically engineered T‐cells , 2007, The Prostate.

[6]  M. Slovak,et al.  Adoptive transfer of chimeric antigen receptor re-directed cytolytic T lymphocyte clones in patients with neuroblastoma. , 2007, Molecular therapy : the journal of the American Society of Gene Therapy.

[7]  Mark M Davis,et al.  Disruption of extracellular interactions impairs T cell receptor-CD3 complex stability and signaling. , 2007, Immunity.

[8]  I. Arkin,et al.  Structural Disorder of the CD3ζ Transmembrane Domain Studied with 2D IR Spectroscopy and Molecular Dynamics Simulations , 2006 .

[9]  James J. Chou,et al.  The Structure of the ζζ Transmembrane Dimer Reveals Features Essential for Its Assembly with the T Cell Receptor , 2006, Cell.

[10]  Gang Wang,et al.  A Phase I Study on Adoptive Immunotherapy Using Gene-Modified T Cells for Ovarian Cancer , 2006, Clinical Cancer Research.

[11]  S. Rosenberg,et al.  Cancer Regression in Patients After Transfer of Genetically Engineered Lymphocytes , 2006, Science.

[12]  Prabuddha Mukherjee,et al.  Picosecond dynamics of a membrane protein revealed by 2D IR. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[13]  Mark M. Davis,et al.  Deconstructing the form and function of the TCR/CD3 complex. , 2006, Immunity.

[14]  B. Baird,et al.  Transmembrane Sequences Are Determinants of Immunoreceptor Signaling 1 , 2005, The Journal of Immunology.

[15]  György Vereb,et al.  Selecting the right fluorophores and flow cytometer for fluorescence resonance energy transfer measurements , 2005, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[16]  P. Stern,et al.  The Role of Extracellular Spacer Regions in the Optimal Design of Chimeric Immune Receptors: Evaluation of Four Different scFvs and Antigens , 2005, Journal of immunotherapy.

[17]  K. Wucherpfennig,et al.  Stoichiometry of the T‐cell receptor–CD3 complex and key intermediates assembled in the endoplasmic reticulum , 2004, The EMBO journal.

[18]  S. Yamasaki,et al.  The quantity and duration of FcRgamma signals determine mast cell degranulation and survival. , 2004, Blood.

[19]  A. Lawson,et al.  Activation of Resting Human Primary T Cells with Chimeric Receptors: Costimulation from CD28, Inducible Costimulator, CD134, and CD137 in Series with Signals from the TCRζ Chain , 2004, The Journal of Immunology.

[20]  J. Irlam,et al.  T lymphocytes isolated from patients with advanced colorectal cancer are suitable for gene immunotherapy approaches , 2003, British Journal of Cancer.

[21]  Martin Wiedmann,et al.  The Organizing Principle in the Formation of the T Cell Receptor-CD3 Complex , 2002, Cell.

[22]  T. Schumacher,et al.  T-cell-receptor gene therapy , 2002, Nature Reviews Immunology.

[23]  R. Guest,et al.  Primary Polyclonal Human T Lymphocytes Targeted to Carcino-Embryonic Antigens and Neural Cell Adhesion Molecule Tumor Antigens by CD3ζ-Based Chimeric Immune Receptors , 2002, Journal of immunotherapy.

[24]  D. Engelman,et al.  Polar residues drive association of polyleucine transmembrane helices , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[25]  W. DeGrado,et al.  Polar side chains drive the association of model transmembrane peptides. , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[26]  Mark J. Smyth,et al.  Redirecting Mouse CTL Against Colon Carcinoma: Superior Signaling Efficacy of Single-Chain Variable Domain Chimeras Containing TCR-ζ vs FcεRI-γ1 , 2001, The Journal of Immunology.

[27]  L. Bolliger,et al.  Identification and functional characterization of the zeta-chain dimerization motif for TCR surface expression. , 1999, Journal of immunology.

[28]  Z. Eshhar,et al.  Loss of original antigenic specificity in T cell hybridomas transduced with a chimeric receptor containing single-chain Fv of an anti-collagen antibody and Fc epsilonRI-signaling gamma subunit. , 1998, Journal of immunology.

[29]  A. Lawson,et al.  Chimeric receptors providing both primary and costimulatory signaling in T cells from a single gene product. , 1998, Journal of immunology.

[30]  A. Kenworthy,et al.  Distribution of a Glycosylphosphatidylinositol-anchored Protein at the Apical Surface of MDCK Cells Examined at a Resolution of <100 Å Using Imaging Fluorescence Resonance Energy Transfer , 1998, The Journal of cell biology.

[31]  P. Stern,et al.  Implications for immunosurveillance of altered HLA class I phenotypes in human tumours. , 1997, Immunology today.

[32]  F. Marincola,et al.  Loss of HLA class I antigens by melanoma cells: molecular mechanisms, functional significance and clinical relevance. , 1995, Immunology today.

[33]  M. Streuli,et al.  Specific interaction of the CD45 protein-tyrosine phosphatase with tyrosine-phosphorylated CD3 zeta chain. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[34]  J. Yewdell,et al.  Identification of human cancers deficient in antigen processing , 1993, The Journal of experimental medicine.

[35]  Z. Eshhar,et al.  Specific activation and targeting of cytotoxic lymphocytes through chimeric single chains consisting of antibody-binding domains and the gamma or zeta subunits of the immunoglobulin and T-cell receptors. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[36]  J. Bonifacino,et al.  Transmembrane helical interactions: zeta chain dimerization and functional association with the T cell antigen receptor. , 1992, The EMBO journal.

[37]  B. Alarcón,et al.  Assembly and function of the T cell antigen receptor. Requirement of either the lysine or arginine residues in the transmembrane region of the alpha chain. , 1990, The Journal of biological chemistry.

[38]  S. O’Brien,et al.  Molecular cloning and chromosomal localization of the human T-cell receptor zeta chain: distinction from the molecular CD3 complex. , 1988, Proceedings of the National Academy of Sciences of the United States of America.

[39]  B. Berkhout,et al.  Assembly of the human T cell receptor-CD3 complex takes place in the endoplasmic reticulum and involves intermediary complexes between the CD3-gamma.delta.epsilon core and single T cell receptor alpha or beta chains. , 1988, The Journal of biological chemistry.

[40]  J. Lippincott-Schwartz,et al.  Failure to synthesize the T Cell CD3-ζ chain: Structure and function of a partial T cell receptor complex , 1988, Cell.

[41]  Z. Eshhar.  The T-body approach: redirecting T cells with antibody specificity. , 2008, Handbook of experimental pharmacology.

[42]  I. Arkin,et al.  Structural disorder of the CD3zeta transmembrane domain studied with 2D IR spectroscopy and molecular dynamics simulations. , 2006, The journal of physical chemistry. B.

[43]  J. Chou,et al.  The structure of the zetazeta transmembrane dimer reveals features essential for its assembly with the T cell receptor. , 2006, Cell.

[44]  J. Trapani,et al.  Redirecting mouse CTL against colon carcinoma: superior signaling efficacy of single-chain variable domain chimeras containing TCR-zeta vs Fc epsilon RI-gamma. , 2001, Journal of immunology.

[45]  T. Dull,et al.  kat: A high-efficiency retroviral transduction system for primary human T lymphocytes , 1994 .