Patient-derived organoids (PDOs) serve as invaluable 3D tumor models, retaining the histological complexity and genetic heterogeneity found in primary tumors. However, the limitation of small sample volumes and the lack of tailored platforms have hindered the research using PDOs. Within the tumor microenvironment, cancer-associated fibroblasts play a pivotal role in influencing drug sensitivity. In this study, we introduce an agarose microwell platform designed for PDO-based tumor and tumor microenvironment models, enabling rapid drug screening and resistance studies with small sample volumes. These microwells, constructed using 3D printing molds, feature a U-shaped bottom and 200 μm diameter. We successfully generated co-culture spheroids of non-small cell lung carcinoma (NSCLC) cells, including NCI-H358 or A549, and NSCLC PDOs F231 or F671 with fibroblast cell line, WI-38. Our results demonstrate the production of uniformly-sized spheroids (coefficient of variation <30%), high viability (>80% after 1 week), and fibroblast-induced drug resistance. The PDOs maintained their viability (>81% after 2 weeks) and continued to proliferate. Notably, when exposed to adagrasib, a KRASG12C inhibitor, we observed reduced cytotoxicity in KRASG12C-mutant spheroids when co-cultured with fibroblasts or their supernatant. The fibroblast supernatant sustained proliferative signals in tumor models. Taking into account the physical features, viability, and drug resistance acquired through supernatants from the fibroblasts, our platform emerges as a suitable platform for in vitro tumor modeling and the evaluation of drug efficacy using patient-derived tissues.