title: Receptor affinity and extracellular domain modifications affect tumor recognition by ROR1-specific chimeric antigen receptor T cells.
abstract: OBJECTIVE
  The adoptive transfer of T cells modified to express a chimeric antigen receptor (CAR) comprised of an extracellular single-chain antibody (scFV) fragment specific for a tumor cell surface molecule, and linked to an intracellular signaling module, has activity in advanced malignancies. The receptor tyrosine kinase-like orphan receptor 1 (ROR1) is a tumor-associated molecule expressed in prevalent B-lymphoid and epithelial cancers and is absent on normal mature B cells and vital tissues, making it a candidate for CAR T-cell therapy.
  
  
  METHODS
  We constructed ROR1-CARs from scFVs with different affinities and containing extracellular IgG4-Fc spacer domains of different lengths, and evaluated the ability of T cells expressing each CAR to recognize ROR1(+) hematopoietic and epithelial tumors in vitro, and to eliminate human mantle cell lymphoma (MCL) engrafted into immunodeficient mice.
  
  
  RESULTS
  ROR1-CARs containing a short "Hinge-only" extracellular spacer conferred superior lysis of ROR1(+) tumor cells and induction of T-cell effector functions compared with CARs with long "Hinge-CH2-CH3" spacers. CARs derived from a higher affinity scFV conferred maximum T-cell effector function against primary CLL and ROR1(+) epithelial cancer lines in vitro without inducing activation-induced T-cell death. T cells modified with an optimal ROR1-CAR were equivalently effective as CD19-CAR-modified T cells in mediating regression of JeKo-1 MCL in immunodeficient mice.
  
  
  CONCLUSIONS
  Our results show that customizing spacer design and increasing affinity of ROR1-CARs enhances T-cell effector function and recognition of ROR1(+) tumors. T cells modified with an optimized ROR1-CAR have significant antitumor efficacy in a preclinical model in vivo, suggesting they may be useful to treat ROR1(+) tumors in clinical applications.
authors.affiliation: Program in Immunology, Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA. mhudecek@fhcrc.org
authors.forename: Michael
authors.initials: M
authors.lastname: Hudecek
authors.forename: Maria-Teresa
authors.initials: MT
authors.lastname: Lupo-Stanghellini
authors.forename: Paula L
authors.initials: PL
authors.lastname: Kosasih
authors.forename: Daniel
authors.initials: D
authors.lastname: Sommermeyer
authors.forename: Michael C
authors.initials: MC
authors.lastname: Jensen
authors.forename: Christoph
authors.initials: C
authors.lastname: Rader
authors.forename: Stanley R
authors.initials: SR
authors.lastname: Riddell
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D006801:Humans; D051379:Mice; D009369:Neoplasms / Q000276:immunology* / Q000473:pathology / Q000628:therapy; D017434:Protein Structure, Tertiary; D057050:Receptor Tyrosine Kinase-like Orphan Receptors / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D015398:Signal Transduction; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology*; D013601:T-Lymphocytes / Q000276:immunology / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays
chemical_list: D011948:Receptors, Antigen, T-Cell; D057127:Single-Chain Antibodies; C078716:ROR1 protein, human; D057050:Receptor Tyrosine Kinase-like Orphan Receptors