MicroRNA-873 Suppresses Viability and Invasion of Colorectal Cancer Through KRAS/MAPK Signaling and Sensitizes Tumor Spheroids to 5-Fluorouracil in a 3D Microwell Model

Colorectal cancer (CRC) ranks third in incidence among all malignancies and is highly lethal in advanced stages. Combination chemotherapy regimens based on 5-fluorouracil (5-FU) remain the mainstay of colorectal cancer treatment alongside surgical resection. Even though new treatment modalities are emerging, many are either ineffective against KRAS-mutant tumors or prone to therapy resistance. Therefore, there is a critical need for new targeted therapies that may overcome the KRAS-driven chemoresistance and enhance the effect of chemotherapy. MicroRNAs can modulate several oncogenic pathways at once and can strengthen chemotherapy. In this study, we identified miR-873 as a potential chemosensitizer that modulates KRAS/MAPK signaling in CRC. We found that KRAS is overexpressed in metastatic versus primary tissues and in a large CRC patient cohort (n = 1,061), high KRAS expression was associated with worse overall survival (HR = 1.27; 95% CI, 1.04–1.56; log-rank p = 0.018). In vitro inhibition of KRAS by siRNA reduced clonogenic growth (HCT116, p = 0.0023; RKO, p = 0.0018) and invasion (p ≤ 0.0001). In silico prediction (TargetScan/miRWalk) analyses showed a conserved binding site between miR-873 and KRAS 3′UTR. Consistent with this prediction, miR-873 mimic transfection reduced KRAS protein expression and phenocopied KRAS knockdown by suppressing colony formation (p ≤ 0.0021) and invasion (p ≤ 0.0001) in KRAS-mutant HCT116 and KRAS-wild-type RKO cells. Dose-matrix screening and SynergyFinder+ analysis revealed synergistic inhibition of spheroid viability with miR-873 + 5-FU, including a low-dose pair (25 nM miR-873 + 12.5 μM 5-FU) showing positive synergy across ZIP/HSA/Bliss/Loewe models. In a poly(ethylene glycol)diacrylate(PEGDA) microwell 3D platform that generates uniform, size-controlled CRC spheroids, this combination produced the strongest suppression of spheroid expansion (day-5/day-3 area: HCT116, 0.61 ± 0.18 vs control, 2.08 ± 0.49; RKO, 0.66 ± 0.04 vs control, 2.08 ± 0.31) and reduced the live-cell fraction to ∼41% in both lines. Moreover, western blot analysis showed decreased KRAS and MAPK pathway activity (reduced p-ERK and context-dependent p-MEK), reduced Cyclin D1, and increased apoptotic readouts (cleaved PARP and a Bax/Bcl-2 shift). Together, these results position miR-873 treatment as a potential targeting approach to suppress KRAS/MAPK signaling and sensitize CRC to 5-FU and validate our PEGDA microwell 3D platform as a practical, translational testbed for miRNA–chemotherapy combinations.