The mucosal barrier remains a major barrier in the pulmonary drug delivery system, as mucociliary clearance in the airway accelerates the removal of inhaled nanoparticles (NPs). Herein, we designed and developed the inhalable Pluronic F127-modified silk fibroin NPs loading with quercetin (marked as QR-SF (PF127) NPs), aiming to solve the airway mucus barrier and improve the cancer therapeutic effect of QR. The PF127 coating on the SF NPs could attenuate the interaction between NPs and mucin proteins, thus facilitating the diffusion of SF(PF127) NPs in the mucus layer. The QR-SF (PF127) NPs had particle sizes of approximately 200 nm with negatively charged surfaces and showed constant drug release properties. Fluorescence recovery after photobleaching (FRAP) assay and transepithelial transport test showed that QR-SF (PF127) NPs exhibited superior mucus-penetrating ability in artificial mucus and monolayer Calu-3 cell model. Notably, a large amount of QR-SF (PF127) NPs distributed uniformly in the mice airway section, indicating the good retention of NPs in the respiratory tract. The mice melanoma lung metastasis model was established, and the therapeutic effect of QR-SF (PF127) NPs was significantly improved . PF127-modified SF NPs may be a promising strategy to attenuate the interaction with mucin proteins and enhance mucus penetration efficiency in the pulmonary drug delivery system.