title: Heterodimeric Bispecific Single Chain Variable Fragments (scFv) Killer Engagers (BiKEs) Enhance NK-cell Activity Against CD133+ Colorectal Cancer Cells.
abstract: Natural killer (NK) cells are potent cytotoxic lymphocytes that play a critical role in tumor immunosurveillance and control. Cancer stem cells (CSC) initiate and sustain tumor cell growth, mediate drug refractory cancer relapse, and express the well-known surface marker CD133.
  
  
  
  DNA fragments from two fully humanized single chain fragment variable (scFv) antibodies recognizing CD16 on NK-cells and CD133 on CSC were genetically spliced forming a novel drug, 16 × 133 BiKE that simultaneously recognizes these antigens to facilitate an immunologic synapse. The anti-CD133 was created using a fusion protein prepared by fusing DNA fragments encoding the two extracellular domains of CD133. Immunization of mice with the resulting fusion protein generated a unique antibody that recognized the molecular framework and was species cross-reactive.
  
  
  
  In vitro chromium-51 ((51)Cr) release cytotoxicity assays at both high and low effector:target ratios demonstrated the ability of the heterodimeric biological drug to greatly enhance NK-cell killing of human Caco-2 colorectal carcinoma cells known to overexpress CD133. The tumor associated antigen specificity of the drug for CD133 even enhanced NK-cell cytotoxicity against the NK-resistant human Burkitt's lymphoma Daudi cell line, which has less than 5 % CD133 surface expression. Flow cytometry analysis revealed increases in NK-cell degranulation and Interferon-γ production upon co-culture with Caco-2 targets in the presence of the drug.
  
  
  
  These studies demonstrate that the innate immune system can be effectively recruited to kill CSC using bispecific antibodies targeting CD133 and that this anti-CD133 scFv may be useful in this bispecific platform or perhaps in the design of more complex trispecific molecules for carcinoma therapy.
authors.affiliation: University of Minnesota Masonic Cancer Center, Section of Molecular Cancer Therapeutics, Therapeutic Radiology-Radiation Oncology, University of Minnesota, Minneapolis, MN, USA.|Department for Hematology and Oncology, Medicine Department 2, University Hospital of Tuebingen, Tuebingen, 72076, Germany.
authors.forename: J U
authors.initials: JU
authors.lastname: Schmohl
authors.affiliation: University of Minnesota Masonic Cancer Center, Adult Division of Hematology, Oncology and Transplantation Oncology, University of Minnesota, Minneapolis, MN, USA.|Medical Science Liaison, Immuno-Oncology, Novartis Oncology/Sandoz, Minneapolis, MN, USA.
authors.forename: M K
authors.initials: MK
authors.lastname: Gleason
authors.affiliation: University of Minnesota Masonic Cancer Center, Section of Molecular Cancer Therapeutics, Therapeutic Radiology-Radiation Oncology, University of Minnesota, Minneapolis, MN, USA.
authors.forename: P R
authors.initials: PR
authors.lastname: Dougherty
authors.affiliation: University of Minnesota Masonic Cancer Center, Adult Division of Hematology, Oncology and Transplantation Oncology, University of Minnesota, Minneapolis, MN, USA.
authors.forename: J S
authors.initials: JS
authors.lastname: Miller
authors.affiliation: University of Minnesota Masonic Cancer Center, Section of Molecular Cancer Therapeutics, Therapeutic Radiology-Radiation Oncology, University of Minnesota, Minneapolis, MN, USA. valle001@umn.edu.
authors.forename: D A
authors.initials: DA
authors.lastname: Vallera
mesh_terms: D000071916:AC133 Antigen / Q000276:immunology*; D000818:Animals; D000951:Antigens, Neoplasm / Q000276:immunology*; D015179:Colorectal Neoplasms / Q000276:immunology*; D006801:Humans; D007694:Killer Cells, Natural / Q000276:immunology*; D051379:Mice; D057127:Single-Chain Antibodies / Q000276:immunology*
chemical_list: D000071916:AC133 Antigen; D000951:Antigens, Neoplasm; C000606024:Prom1 protein, mouse; D057127:Single-Chain Antibodies