CD28 cosignalling does not affect the activation threshold in a chimeric antigen receptor-redirected T-cell attack

Abstract

Adoptive immunotherapy of cancer using chimeric antigen receptor (CAR)-engineered T cells with redirected specificity showed efficacy in recent trials. In preclinical models, 'second-generation' CARs with CD28 costimulatory domain in addition to CD3z performed superior in redirecting T-cell effector functions and survival. Whereas CD28 costimulation sustains physiological T-cell receptor (TCR)-CD3 activation of naïve T cells, the impact of CD28 cosignalling on the threshold of CAR-mediated activation of pre-stimulated T cells without B7-CD28 recruitment remained unclear. Using CARs of different binding affinities, but same epitope specificity, we demonstrate that CD28 cosignalling neither lowered the antigen threshold nor the binding affinity for redirected T-cell activation. 'Affinity ceiling' above which increase in affinity does not increase T-cell activation was not altered. Accordingly, redirected tumor cell killing depended on the binding affinity but was likewise effective for CD3z and CD28-CD3z CARs. In contrast to CD3z, CD28-CD3z CAR-driven activation was not increased further by CD28-B7 engagement. However, CD28 cosignalling, which is required for interleukin-2 induction could not be replaced by high-affinity CD3z CAR binding or high-density antigen engagement. We conclude that CD28 CAR cosignalling does not alter the activation threshold but redirects T-cell effector functions.

Efficacy of CAR T-cell Therapy in Large Tumors Relies upon Stromal Targeting by IFNγ

“…1C). CAR-Ts can secrete IL2 when stimulated through their endogenous CD28 receptor (22) or the chimeric CD28-z receptor (19). Accordingly, we found that CAR-Ts engineered with 9-28-z but not z-CARs secreted IL2 (Fig.…”

“…Here, we first established that eradication of large established tumors can be achieved by HER2-specific CAR-Ts if provided with costimulatory CD28 signaling (28-z-CAR; ref. 19). This rejection was associated with sustained accumulation, proliferation, and differentiation of CAR-Ts to effector memory (TEM) cell type at the tumor site.…”

Coexpressed Catalase Protects Chimeric Antigen Receptor–Redirected T Cells as well as Bystander Cells from Oxidative Stress–Induced Loss of Antitumor Activity

“…The CARs used in this study are specific for Her2 and CEA and have been shown to eliminate tumor cells with the respective targets in a specific fashion (28,36). The Her2 and CEA are highly expressed in a variety of breast cancer and colorectal cancer (CRC) lesions, respectively (37,38), both of which have increased ROS production and local oxidative stress (15,39).…”

“…CAR-CAT were based on the CEA-specific CAR BW431/26scFv-IgG1-CD28-CD3z and the Her2-specific CAR C6-B1.D2-IgG1-CD28-CD3z (27,28) by inserting the full-length human catalase cDNA downstream of the internal ribosome entry site (Fig. 1A).…”

Treatment of solid tumors with chimeric antigen receptor-engineered T cells: current status and future prospects

“…Next, second-generation CARs contain costimulatory signaling domains derived from T cell costimulatory, with receptors such as CD28 being the most common choice. CD28-mediated CAR signaling is preferred in inducing IL-2 secretion and promoting T cell amplification (Chmielewski et al, 2011). Besides CD28, other costimulatory molecules, such as tumor necrosis factor receptor superfamily member 9 (TNFRSF9, 4-1BB), tumor necrosis factor receptor superfamily, member 4 (TNFRSF9, OX40), inducible T-cell CO Stimulator (ICOS), and CD27, may be included in CAR construction to take advantage of their associated functions in regulating T cell proliferation, survival, and antitumor response.…”