In vitro and in vivo model of a novel immunotherapy approach for chronic lymphocytic leukemia by anti-CD23 chimeric antigen receptor Running title: CD23 CAR-targeting for CLL immunotherapy Keywords: CLL, Chimeric T-cell receptor, CD23, Immunotherapy, Cell Therapy

Authors' Affiliations: -Centro di Ricerca Matilde Tettamanti, Department of Pediatrics, University of Milano-Bicocca, San Gerardo Hospital, Monza, Milan, Italy; -Center for Cell and Gene Therapy, Baylor College of Medicine, Houston, Texas, USA -Department of Adult Hematology, San Gerardo Hospital, Monza, Italy -Pathology Unit and Lymphoma Unit, Department of Oncology, Istituto Scientifico San Raffaele, Milano, Italy Laboratory of B Cell Neoplasia and Lymphoma Unit, Università Vita-Salute San Raffaele and Istituto Scientifico San Raffaele, Milan, Italy * V.Marin and V.Hoyos equally contributed to this work **E.Biagi and G.Dotti are co-last authors and co-corresponding authors

[1]  C. Aspord,et al.  Exploration of the Lysis Mechanisms of Leukaemic Blasts by Chimaeric T-Cells , 2010, Journal of biomedicine & biotechnology.

[2]  G. Simonetti,et al.  A novel Rag2-/-gammac-/--xenograft model of human CLL. , 2010, Blood.

[3]  Arturo Molina,et al.  Phase 1/2 study of lumiliximab combined with fludarabine, cyclophosphamide, and rituximab in patients with relapsed or refractory chronic lymphocytic leukemia. , 2010, Blood.

[4]  T. Lin.  New Agents in Chronic Lymphocytic Leukemia , 2010, Current hematologic malignancy reports.

[5]  M. Brenner,et al.  Fifteen years of gene therapy based on chimeric antigen receptors: "are we nearly there yet?". , 2009, Human gene therapy.

[6]  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.

[7]  Michel Sadelain,et al.  The promise and potential pitfalls of chimeric antigen receptors. , 2009, Current Opinion in Immunology.

[8]  Jinjuan Wang,et al.  Adoptive immunotherapy for indolent non-Hodgkin lymphoma and mantle cell lymphoma using genetically modified autologous CD20-specific T cells. , 2008, Blood.

[9]  H. Heslop,et al.  Cytotoxic T lymphocytes directed to the preferentially expressed antigen of melanoma (PRAME) target chronic myeloid leukemia. , 2008, Blood.

[10]  H. Heslop,et al.  Epstein Barr virus specific cytotoxic T lymphocytes expressing the anti-CD30zeta artificial chimeric T-cell receptor for immunotherapy of Hodgkin disease. , 2007, Blood.

[11]  J. Byrd,et al.  Phase 1 Study of Lumiliximab with Detailed Pharmacokinetic and Pharmacodynamic Measurements in Patients with Relapsed or Refractory Chronic Lymphocytic Leukemia , 2007, Clinical Cancer Research.

[12]  A. Biondi,et al.  Chimeric T-cell receptors: new challenges for targeted immunotherapy in hematologic malignancies. , 2007, Haematologica.

[13]  H. Heslop,et al.  T lymphocytes redirected against the kappa light chain of human immunoglobulin efficiently kill mature B lymphocyte-derived malignant cells. , 2006, Blood.

[14]  David D. Smith,et al.  CD28 costimulation provided through a CD19-specific chimeric antigen receptor enhances in vivo persistence and antitumor efficacy of adoptively transferred T cells. , 2006, Cancer research.

[15]  M. Brenner,et al.  Addition of the CD28 signaling domain to chimeric T-cell receptors enhances chimeric T-cell resistance to T regulatory cells , 2006, Leukemia.

[16]  A. Biondi,et al.  Characterization of in vitro migratory properties of anti-CD19 chimeric receptor-redirected CIK cells for their potential use in B-ALL immunotherapy. , 2006, Experimental hematology.

[17]  F. Ravandi,et al.  Immune defects in patients with chronic lymphocytic leukemia , 2006, Cancer Immunology, Immunotherapy.

[18]  M. Andreeff,et al.  Responses to Human CD40 Ligand/Human Interleukin-2 Autologous Cell Vaccine in Patients with B-Cell Chronic Lymphocytic Leukemia , 2005, Clinical Cancer Research.

[19]  P. Bierman,et al.  Unrelated donor marrow transplantation for B-cell chronic lymphocytic leukemia after using myeloablative conditioning: results from the Center for International Blood and Marrow Transplant research. , 2005, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[20]  Roman Rydzanicz,et al.  Assembly PCR oligo maker: a tool for designing oligodeoxynucleotides for constructing long DNA molecules for RNA production , 2005, Nucleic Acids Res..

[21]  M. Andreeff,et al.  Early results of a chemoimmunotherapy regimen of fludarabine, cyclophosphamide, and rituximab as initial therapy for chronic lymphocytic leukemia. , 2005, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[22]  C. Mayr,et al.  CD23 is recognized as tumor-associated antigen (TAA) in B-CLL by CD8+ autologous T lymphocytes. , 2005, Experimental hematology.

[23]  David D. Smith,et al.  Enhanced antilymphoma efficacy of CD19-redirected influenza MP1-specific CTLs by cotransfer of T cells modified to present influenza MP1. , 2005, Blood.

[24]  M. Papamichail,et al.  Targeting of tumor cells by lymphocytes engineered to express chimeric receptor genes , 2004, Cancer Immunology, Immunotherapy.

[25]  M. Goller,et al.  Regulation of CD23 isoforms on B-chronic lymphocytic leukemia. , 2002, Leukemia research.

[26]  V. Diehl,et al.  A Phase I study with an anti-CD30 ricin A-chain immunotoxin (Ki-4.dgA) in patients with refractory CD30+ Hodgkin's and non-Hodgkin's lymphoma. , 2002, Clinical cancer research : an official journal of the American Association for Cancer Research.

[27]  M. Dettke,et al.  Notch2 is involved in the overexpression of CD23 in B-cell chronic lymphocytic leukemia. , 2002, Blood.

[28]  Richard Greil,et al.  The Role of Soluble CD23 in Distinguishing Stable and Progressive Forms of B-chronic Lymphocytic Leukemia , 2002, Leukemia & lymphoma.

[29]  C. Bollard,et al.  Targeting of GD2‐positive tumor cells by human T lymphocytes engineered to express chimeric T‐cell receptor genes , 2001, International journal of cancer.

[30]  A. Wotherspoon,et al.  High expression of CD23 in the proliferation centers of chronic lymphocytic leukemia in lymph nodes and spleen. , 1999, Human pathology.

[31]  P. Circosta,et al.  MEC1 and MEC2: two new cell lines derived from B-chronic lymphocytic leukaemia in prolymphocytoid transformation. , 1999, Leukemia research.

[32]  C. D. Wu,et al.  High incidence of relapse after autologous stem-cell transplantation for B-cell chronic lymphocytic leukemia or small lymphocytic lymphoma. , 1998, Annals of oncology : official journal of the European Society for Medical Oncology.

[33]  D. Earnshaw,et al.  CD23 (FcepsilonRII) release from cell membranes is mediated by a membrane-bound metalloprotease. , 1998, The Biochemical journal.

[34]  V. Diehl,et al.  A chimeric receptor that selectively targets membrane-bound carcinoembryonic antigen (mCEA) in the presence of soluble CEA , 1998, Gene Therapy.

[35]  H. Heslop,et al.  Production of genetically modified Epstein-Barr virus-specific cytotoxic T cells for adoptive transfer to patients at high risk of EBV-associated lymphoproliferative disease. , 1995, Journal of hematotherapy.

[36]  S. Fournier,et al.  The two CD23 isoforms display differential regulation in chronic lymphocytic leukaemia , 1995, British journal of haematology.

[37]  C. Gasche,et al.  Soluble CD23 reliably reflects disease activity in B-cell chronic lymphocytic leukemia. , 1994, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[38]  H. Eibel,et al.  Mice deficient in CD23 reveal its modulatory role in IgE production but no role in T and B cell development. , 1994, Journal of immunology.

[39]  S. Fournier,et al.  CD23 antigen regulation and signaling in chronic lymphocytic leukemia. , 1992, The Journal of clinical investigation.

[40]  J. Banchereau,et al.  Cross-linking of CD23 antigen by its natural ligand (IgE) or by anti-CD23 antibody prevents B lymphocyte proliferation and differentiation. , 1991, Journal of immunology.

[41]  R. Hardy,et al.  Fc epsilon receptor, a specific differentiation marker transiently expressed on mature B cells before isotype switching , 1986, The Journal of experimental medicine.

[42]  E. Jaffe,et al.  Detection of differentially expressed genes in lymphomas using cDNA arrays: identification of clusterin as a new diagnostic marker for anaplastic large-cell lymphomas. , 2000, Blood.

[43]  J. Roliński,et al.  Characterization of regulatory T cells in patients with B-cell chronic lymphocytic leukemia. , 2008, Oncology reports.

[44]  H. Döhner,et al.  Efficient nucleofection of primary human B cells and B-CLL cells induces apoptosis, which depends on the microenvironment and on the structure of transfected nucleic acids , 2007, Leukemia.

[45]  S. Molica,et al.  Cellular expression and serum circulating levels of CD23 in B-cell chronic lymphocytic leukemia. Implications for prognosis. , 1996, Haematologica.

[46]  S. Fournier,et al.  The in vivo expression of type B CD23 mRNA in B-chronic lymphocytic leukemic cells is associated with an abnormally low CD23 upregulation by IL-4: comparison with their normal cellular counterparts. , 1991, Leukemia research.