Oncolytic Adenovirus Armed with BiTE, Cytokine, and Checkpoint Inhibitor Enables CAR T Cells to Control the Growth of Heterogeneous Tumors

Mol Ther. 2020 May 6;28(5):1251-1262. doi: 10.1016/j.ymthe.2020.02.016. Epub 2020 Feb 24.


No single cancer immunotherapy will likely defeat all evasion mechanisms of solid tumors, including plasticity of tumor antigen expression and active immune suppression by the tumor environment. In this study, we increase the breadth, potency, and duration of anti-tumor activity of chimeric antigen receptor (CAR) T cells using an oncolytic virus (OV) that produces cytokine, checkpoint blockade, and a bispecific tumor-targeted T cell engager (BiTE) molecule. First, we constructed a BiTE molecule specific for CD44 variant 6 (CD44v6), since CD44v6 is widely expressed on tumor but not normal tissue, and a CD44v6 antibody has been safely administered to cancer patients. We then incorporated this BiTE sequence into an oncolytic-helper binary adenovirus (CAdDuo) encoding an immunostimulatory cytokine (interleukin [IL]-12) and an immune checkpoint blocker (PD-L1Ab) to form CAdTrio. CD44v6 BiTE from CAdTrio enabled HER2-specific CAR T cells to kill multiple CD44v6+ cancer cell lines and to produce more rapid and sustained disease control of orthotopic HER2+ and HER2-/- CD44v6+ tumors than any component alone. Thus, the combination of CAdTrio with HER2.CAR T cells ensures dual targeting of two tumor antigens by engagement of distinct classes of receptor (CAR and native T cell receptor [TCR]), and significantly improves tumor control and survival.

Keywords: oncolytic viro-immunotherapy, CAR T cell, BiTE molecule, cytokine, checkpoint inhibitor, CD44 variant 6.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenoviridae / metabolism*
  • Animals
  • Antibodies, Bispecific / therapeutic use*
  • Female
  • Humans
  • Hyaluronan Receptors / immunology
  • Hyaluronan Receptors / metabolism
  • Immune Checkpoint Inhibitors / metabolism
  • Immune Checkpoint Inhibitors / therapeutic use*
  • Immunotherapy, Adoptive / methods*
  • Interleukin-12 / metabolism
  • Interleukin-12 / therapeutic use*
  • Male
  • Mice, Inbred NOD
  • Mice, SCID
  • Neoplasms / metabolism
  • Neoplasms / pathology
  • Neoplasms / therapy*
  • Oncolytic Virotherapy / methods*
  • Oncolytic Viruses / metabolism*
  • PC-3 Cells
  • Receptor, ErbB-2 / immunology
  • Receptor, ErbB-2 / metabolism
  • Receptors, Chimeric Antigen / immunology
  • Receptors, Chimeric Antigen / therapeutic use*
  • Treatment Outcome
  • Xenograft Model Antitumor Assays


  • Antibodies, Bispecific
  • CD44v6 antigen
  • Hyaluronan Receptors
  • Immune Checkpoint Inhibitors
  • Receptors, Chimeric Antigen
  • Interleukin-12
  • ERBB2 protein, human
  • Receptor, ErbB-2