title: Construction and evaluation of a novel humanized HER2-specific chimeric receptor.
abstract: BACKGROUND
  The human epidermal growth factor receptor 2 (HER2) represents one of the most studied tumor-associated antigens (TAAs) for cancer immunotherapy. The monoclonal antibody (mAb) trastuzumab has improved the outcomes of patients with HER2+ breast cancer. However, a large number of HER2+ tumors are not responsive to, or become resistant to, trastuzumab-based therapy, and thus more effective therapies targeting HER2 are needed.
  
  
  METHODS
  HER2-specific T cells were generated by the transfer of genes that encode chimeric antigen receptor (CAR). Using a multistep overlap extension PCR method, we constructed a novel, humanized HER2 CAR-containing, chA21 single-chain variable fragment (scFv) region of antigen-specific mAb and T-cell intracellular signaling chains made up of CD28 and CD3ζ. An interferon γ and interleukin 2 enzyme-linked immunosorbent assay and a chromium-51 release assay were used to evaluate the antitumor immune response of CAR T cells in coculture with tumor cells. Furthermore, SKBR3 tumor-bearing nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice were treated with HER2 CAR T cells to evaluate antitumor activity. Human CD3+ T cell accumulation in tumor xenograft was detected by immunohistochemistry.
  
  
  RESULTS
  chA21-28z CAR was successfully constructed, and both CD4+ and CD8+ T cells were transduced. The expanded HER2 CAR T cells expressed a central memory phenotype and specifically reacted against HER2+ tumor cell lines. Furthermore, the SKBR3 tumor xenograft model revealed that HER2 CAR T cells significantly inhibited tumor growth in vivo. Immunohistochemical analysis showed robust accumulation of human CD3+ T cells in regressing SKBR3 lesions.
  
  
  CONCLUSIONS
  The results of this study show that novel chA21 scFv-based, HER2-specific CAR T cells not only recognized and killed HER2+ breast and ovarian cancer cells ex vivo but also induced regression of experimental breast cancer in vivo. Our data support further exploration of the HER2 CAR T-cell therapy for HER2-expressing cancers.
authors.forename: Meili
authors.initials: M
authors.lastname: Sun
authors.forename: Huan
authors.initials: H
authors.lastname: Shi
authors.forename: Chuanyong
authors.initials: C
authors.lastname: Liu
authors.forename: Jie
authors.initials: J
authors.lastname: Liu
authors.forename: Xianqiang
authors.initials: X
authors.lastname: Liu
authors.forename: Yuping
authors.initials: Y
authors.lastname: Sun
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology; D061067:Antibodies, Monoclonal, Humanized / Q000494:pharmacology; D000970:Antineoplastic Agents; D001943:Breast Neoplasms / Q000188:drug therapy* / Q000276:immunology; D018106:CD28 Antigens / Q000235:genetics / Q000276:immunology; D017252:CD3 Complex / Q000235:genetics / Q000276:immunology; D045744:Cell Line, Tumor; D002859:Chromium Isotopes / Q000737:chemistry; D019008:Drug Resistance, Neoplasm; D005260:Female; D018014:Gene Transfer Techniques; D006801:Humans; D007371:Interferon-gamma / Q000276:immunology; D007376:Interleukin-2 / Q000276:immunology; D061986:MCF-7 Cells; D051379:Mice; D016688:Mice, Inbred NOD; D016513:Mice, SCID; D009368:Neoplasm Transplantation; D010051:Ovarian Neoplasms / Q000188:drug therapy* / Q000276:immunology; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000276:immunology*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology*; D013601:T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D014183:Transplantation, Heterologous; D000068878:Trastuzumab
chemical_list: D000911:Antibodies, Monoclonal; D061067:Antibodies, Monoclonal, Humanized; D000970:Antineoplastic Agents; D018106:CD28 Antigens; D017252:CD3 Complex; D002859:Chromium Isotopes; D007371:Interferon-gamma; D007376:Interleukin-2; D018719:Erb-b2 Receptor Tyrosine Kinases; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; D000068878:Trastuzumab; C121704:CD3 antigen, zeta chain; C508594:IL2 protein, human; C508053:ERBB2 protein, human