Pancreatic Ductal Adenocarcinoma (PDAC) is a highly aggressive form of cancer, responsible for nearly half a million deaths annually worldwide. Despite advancements in treatments, PDAC remains difficult to treat with a poor survival rate. Chimeric Antigen Receptor (CAR) T-cell therapy has shown success and applicability in treating B-cell malignancies and blood tumors, which raised hopes for CAR T-cells to be used in solid tumors. However, its application in solid tumors is challenging due to antigen escape, heterogeneity observed and off-target effect.
Therefore, in this study we demonstrate the use of Boolean logic to target multiple antigens for enhanced tumor discrimination and limited off-target effects. Mesothelin (MSLN) and Epidermal Growth Factor Receptor (EGFR) were the antigens chosen using the Antigen Explorer database. Gene delivery will be achieved using the piggyBac transposon system, an emerging non-viral method, using the pNB328-meso3 CAR plasmid and the pIRII-CAR.EGFR(EGFv1) CAR plasmid to transfect and engineer the T-cells. Furthermore, CAR T-cells were activated using co-stimulation signals that got split into two separately expressed CARs directed against the two selected antigens. In the split-dual CAR system, the first CAR (EGFR) provides the primary activation signal (CD3ζ) and the second CAR (MSLN) delivers the costimulatory CD28 signal. Full T-cell activation only occurs when both antigens are expressed, ultimately reducing the risk of off-tumor toxicity and increasing the specificity for solid tumors. This AND-Gated CAR T-cell approach may present a promising strategy to overcome PDAC heterogeneity and improve the precision of immunotherapy for PDAC.