Chimeric antigen receptor (CAR)-engineered cell therapies have shown remarkable promise, yet development is hindered by the lengthy process of identifying functional antigen-specific single-chain variable fragments (scFvs). To overcome this bottleneck, we developed a high-throughput, mRNA-based CAR-NK platform that enables rapid screening and optimization of scFvs targeting mesothelin (MSLN), a tumor-associated antigen commonly overexpressed in solid cancers. A key challenge in MSLN-directed therapy is the presence of soluble MSLN (solMSLN), resulting from proteolytic shedding of the distal MSLN domain, which can impair therapeutic efficacy. Using a yeast surface-display scFv library, we identified shed-resistant scFvs that selectively bind membrane-proximal MSLN epitopes. These scFvs were transiently expressed via electroporation of linear mRNA into primary NK cells for live-cell cytotoxicity assays. Final candidates were incorporated into circular mRNA (circRNA)-encoded CAR constructs and co-engineered with immune-enhancing modules to improve persistence and anti-tumor function. The optimized CAR-MS10-NK cells exhibited enhanced in vitro cytotoxicity and significantly reduced tumor burden in MSLN-positive, solMSLN-rich pancreatic tumor models, outperforming lentiviral CAR-NK counterparts. This modular, non-viral platform accelerates the discovery of functional CARs against challenging antigens like MSLN and supports scalable, cost-effective immunotherapy development.