Combined targeting of soluble latent TGF-ß and a solid tumor-associated antigen with adapter CAR T cells

Abstract

Solid tumors consist of malignant and nonmalignant cells that together create the local tumor microenvironment (TME). Additionally, the TME is characterized by the expression of numerous soluble factors such as TGF-β. TGF-β plays an important role in the TME by suppressing T cell effector function and promoting tumor invasiveness. Up to now CAR T cells exclusively target tumor-associated antigens (TAA) located on the cell membrane. Thus, strategies to exploit soluble antigens as CAR targets within the TME are needed. This study demonstrates a novel approach using Adapter CAR (AdCAR) T cells for the detection of soluble latent TGF-β within the TME of a pancreatic tumor model. We show that AdCARs in combination with the respective adapter can be used to sense soluble tumor-derived latent TGF-β, both in vitro and in vivo . Sensing of the soluble antigen induced cellular activation and effector cytokine production in AdCAR T cells. Moreover, we evaluated AdCAR T cells for the combined targeting of soluble latent TGF-β and tumor cell killing by targeting CD66c as TAA in vivo . In sum, our study broadens the spectrum of targetable moieties for AdCAR T cells by soluble latent TGF-β.

Coengineering specificity, safety, and function into T cells for cancer immunotherapy

“…Additional logic gate approaches include so‐called universal CARs in which the T cells themselves do not engage TA but instead rely on an adaptor molecule (i.e., OR‐gates) 258 . Examples of universal CARs include UniCARs, 259,260 RevCARs, 261 convertible CARs, 262 PNE (peptide neoepitope specific) CARs, 263 SpyCatcher‐based CARs, 264 and AdCARs, 265 amongst others. More recent examples include the SUPRA (split, universal, and programmable) CAR comprising zip‐CAR T cells (there is a leucine zipper in the extracellular domain) and a zipFv (a tumor‐targeting scFv adaptor with a zip that binds the zip‐CAR T cells) 266 (Figure 5F), and the Co‐LOCKR platform (the colocalization‐dependent protein switches) requiring binding to a defined combination of TA to enable activation (i.e., an AND‐gate) 267 .…”

“…SUPRA CARs, for example, comprise a zipper in their ectodomain and require administration of tumor-targeting scFv fused to a zipper (adaptor protein) 266. (G) For the inhibitory (I)CAR 268 a scFv targeting an antigen found on healthy tissue cells is fused to an inhibitory endodomain such as from PD-1 to block T-cell signaling should the 2G CAR bind to its target but offtumor.PNE (peptide neoepitope specific) CARs,263 SpyCatcher-based CARs,264 and AdCARs,265 amongst others. More recent examples include the SUPRA (split, universal, and programmable) CAR comprising zip-CAR T cells (there is a leucine zipper in the extracellular domain) and a zipFv (a tumor-targeting scFv adaptor with a zip that binds the zip-CAR T cells) 266 (Figure5F), and the Co-LOCKR platform (the colocalization-dependent protein switches) requiring binding to a defined combination of TA to enable activation (i.e.,…”

Mechanisms and strategies for safe chimeric antigen receptor T‐cell activity control

“…Tag‐specific or anti‐tag CAR T‐cells recognize a tag which is chemically, enzymatically, or genetically coupled to a tumor‐targeting moiety 124‐126 such as the synthetic dye fluorescein isothiocyanate (FITC), 77‐81 biotin 82‐86 or peptide tags 80,87‐93 . The introduction of these tags by genetic fusion requires additional quality control steps of the adaptor molecule and increases the risk of immunogenicity.…”

“…FITC‐specific CAR T‐cells are currently evaluated in a Phase I trial in osteogenic sarcoma (NCT05312411). The so‐called AdCAR T‐cell system is redirected to surface antigens via biotin‐labeled adapter molecules and was tested in aggressive lymphoma models, 84 in AML 85 and for the detection of soluble latent TGF‐β within the TME of a pancreatic tumor model 86 . Another well‐established system is the UniCAR strategy, which is based on an anti‐epitope scFv used in the modular BiTE format UniMab and UniCAR T‐cells recognizing the peptide epitope 127 .…”

“…75,76 However, these CAR T-cells cannot discriminate between administered monoclonal and endogenous antibodies, raising concerns of autoimmune side effects. 75,97 Tag-specific or anti-tag CAR T-cells recognize a tag which is chemically, enzymatically, or genetically coupled to a tumor-targeting moiety [124][125][126] such as the synthetic dye fluorescein isothiocyanate (FITC), [77][78][79][80][81] biotin [82][83][84][85][86] or peptide tags. 80,[87][88][89][90][91][92][93] The introduction of these tags by genetic fusion requires additional quality control steps of the adaptor molecule and increases the risk of immunogenicity.…”