title: Selective expansion of chimeric antigen receptor-targeted T-cells with potent effector function using interleukin-4.
abstract: Polyclonal T-cells can be directed against cancer using transmembrane fusion molecules known as chimeric antigen receptors (CARs). Although preclinical studies have provided encouragement, pioneering clinical trials using CAR-based immunotherapy have been disappointing. Key obstacles are the need for robust expansion ex vivo followed by sustained survival of infused T-cells in patients. To address this, we have developed a system to achieve selective proliferation of CAR(+) T-cells using IL-4, a cytokine with several pathophysiologic and therapeutic links to cancer. A chimeric cytokine receptor (4alphabeta) was engineered by fusion of the IL-4 receptor alpha (IL-4Ralpha) ectodomain to the beta(c) subunit, used by IL-2 and IL-15. Addition of IL-4 to T-cells that express 4alphabeta resulted in STAT3/STAT5/ERK phosphorylation and exponential proliferation, mimicking the actions of IL-2. Using receptor-selective IL-4 muteins, partnering of 4alphabeta with gamma(c) was implicated in signal delivery. Next, human T-cells were engineered to co-express 4alphabeta with a CAR specific for tumor-associated MUC1. These T-cells exhibited an unprecedented capacity to elicit repeated destruction of MUC1-expressing tumor cultures and expanded through several logs in vitro. Despite prolonged culture in IL-4, T-cells retained specificity for target antigen, type 1 polarity, and cytokine dependence. Similar findings were observed using CARs directed against two additional tumor-associated targets, demonstrating generality of application. Furthermore, this system allows rapid ex vivo expansion and enrichment of engineered T-cells from small blood volumes, under GMP-compliant conditions. Together, these findings provide proof of principle for the development of IL-4-enhanced T-cell immunotherapy of cancer.
authors.affiliation: Division of Cancer Studies, Research Oncology Section, Guy's Hospital Campus, King's College London School of Medicine, London, United Kingdom.
authors.forename: Scott
authors.initials: S
authors.lastname: Wilkie
authors.forename: Sophie E
authors.initials: SE
authors.lastname: Burbridge
authors.forename: Laura
authors.initials: L
authors.lastname: Chiapero-Stanke
authors.forename: Ana C P
authors.initials: AC
authors.lastname: Pereira
authors.forename: Siobhán
authors.initials: S
authors.lastname: Cleary
authors.forename: Sjoukje J C
authors.initials: SJ
authors.lastname: van der Stegen
authors.forename: James F
authors.initials: JF
authors.lastname: Spicer
authors.forename: David M
authors.initials: DM
authors.lastname: Davies
authors.forename: John
authors.initials: J
authors.lastname: Maher
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D005434:Flow Cytometry; D006801:Humans; D019409:Interleukin-15 / Q000494:pharmacology; D007376:Interleukin-2 / Q000494:pharmacology; D015847:Interleukin-4 / Q000494:pharmacology*; D008213:Lymphocyte Activation / Q000187:drug effects / Q000235:genetics; D051379:Mice; D010766:Phosphorylation / Q000187:drug effects; D011948:Receptors, Antigen, T-Cell / Q000235:genetics / Q000378:metabolism*; D019948:Receptors, Interleukin-4 / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D050796:STAT3 Transcription Factor / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects* / Q000378:metabolism*
chemical_list: D019409:Interleukin-15; D007376:Interleukin-2; D011948:Receptors, Antigen, T-Cell; D019948:Receptors, Interleukin-4; D011993:Recombinant Fusion Proteins; D050796:STAT3 Transcription Factor; D015847:Interleukin-4