Three-dimensional colon cancer organoids model the response to CEA-CD3 T-cell engagers

Abstract

Rationale: The CEA-CD3 T cell bispecific antibody cibisatamab (CEA-TCB) is currently undergoing clinical trials. Here we study its performance against three-dimensional tumor organoids in cocultures with T cells as compared to a higher affinity CEACAM5-CD3 (CEACAM5-TCB) bispecific antibody using time-lapse confocal microscopy. Methods: Pre-labelled spheroids derived from colon cancer cell lines and primary organoids derived from four colorectal cancer surgical specimens, which expressed different graded levels of CEA, were exposed in cocultures to T lymphocytes. Cocultures were treated with CEA-CD3 T-cell engagers and were followed by live confocal microscopy. Caspase 3 activation detected in real-time was used as an indicator of tumor cell death. Co-cultures were also set up with autologous tumor-associated fibroblasts to test the co-stimulatory effect of a fibroblast activated protein (FAP)- targeted 4-1BBL bispecific antibody fusion protein currently undergoing clinical trials. Results: Tumor-cell killing of 3D colon carcinoma cultures was dependent on the levels of surface CEA expression, in such a way that the lower affinity agent (CEA-TCB) did not mediate killing by human preactivated T cells below a certain CEA expression threshold, while the high affinity construct (CEACAM5-TCB) remained active on the low CEA expressing organoids. Modelling heterogeneity in the levels of CEA expression by coculturing CEA high and low organoids showed measurable but weak bystander killing. Cocultures of tumor organoids, autologous fibroblasts and T cells allowed to observe a costimulatory effect of anti-FAP-4-1BBL both to release IFNγ and to attain more efficacious tumor cell killing. Conclusion: Three-dimensional tumor cocultures with T cells using live confocal microscopy provide suitable models to test the requirements for colon-cancer redirected killing as elicited by CEA-targeted T-cell engagers undergoing clinical trials and treatment allow combinations to be tested in a relevant preclinical system.

A first-in-human study of the fibroblast activation protein–targeted, 4-1BB agonist RO7122290 in patients with advanced solid tumors

“…Clinical activity was observed in combination with atezolizumab but was mainly limited to cancer patients without prior CPI treatment. A phase 1b study of RO7122290 in combination with cibisatamab [carcinoembryonic antigen × CD3 bispecific mAb ( 51 )] is ongoing, evaluating the concept of synergistic T cell engagement (signal 1) and costimulation (signal 2) as a potential therapeutic modality for advanced colorectal cancer (NCT04826003).…”

LY6G6D is a selectively expressed colorectal cancer antigen that can be used for targeting a therapeutic T-cell response by a T-cell engager

“…Although this is the main mechanism mediating tumor control by TcE, some preclinical studies have reported the evidence of a TcE-bystander killing of target negative cells ( 20 ). Cytokine release, mainly IFNγ and TNFα ( 19 , 27 , 28 ), but also the Fas/FasL pathway, have been described to be the main mechanisms involved in bystander killing ( 18 , 20 ). In 2D cell cocultures, LY6G6D/CD3 TcE induced activation of T cells and killing of LY6G6D-negative cells even at a low percentage of 20% LY6G6D-positive cells.…”

“…The mechanism overlying the bystander effect in TcE and CAR T cell therapeutics has been studied recently. Fas/FasL, TNFa and IFNg signaling have been described as possible mechanisms, albeit none (18)(19)(20) sufficient to explain the bystander effect in all circumstances or models [18][19][20]. To understand the mechanism responsible for the bystander killing in our system, we blocked pharmacologically and individually the three pathways in our tumor cell 3D cocultures containing LS1034 and NCM460D-GFP cells.…”

Flourishing tumor organoids: History, emerging technology, and application

“…208,209 Currently, organoid models are used in tumor immunology studies including exploring potential immune mechanisms, 210,211 immunotherapy modeling, 161,212,213 and personalized immunotherapy testing. [214][215][216] T cells are key players in immunotherapy, and the coculture of autologous tumor organoids and peripheral blood lymphocytes could be used from mismatch repair-deficient colorectal cancer and peripheral blood from patients with nonsmall cell lung cancer to enrich tumor-reactive T cells and assess the efficiency of T-cellmediated killing of matched tumor organoids. 217 This co-culture model can also be used to assess the sensitivity of patients to immunotherapy and explain the possible mechanisms of drug resistance (Figure 7b).…”

“…Therefore, we can choose to actively target specific antigens on the tumor or augment the host immune system, including monoclonal antibodies, adoptive transfer of TILs, allogeneic cell‐based vaccines, chimeric antigen receptors T (CAR‐T) cells, and immune checkpoint inhibition when the strength of the immune response induced by tumor cell neoantigens is insufficient 208,209 . Currently, organoid models are used in tumor immunology studies including exploring potential immune mechanisms, 210,211 immunotherapy modeling, 161,212,213 and personalized immunotherapy testing 214–216 . T cells are key players in immunotherapy, and the co‐culture of autologous tumor organoids and peripheral blood lymphocytes could be used from mismatch repair‐deficient colorectal cancer and peripheral blood from patients with nonsmall cell lung cancer to enrich tumor‐reactive T cells and assess the efficiency of T‐cell‐mediated killing of matched tumor organoids 217 .…”