Biomineralized PCAL Microspheres Trigger Synergistic Calcicoptosis-Ferroptosis for Enhanced Non-Small Cell Lung Cancer Therapy.

Non-small cell lung cancer (NSCLC) is characterized by significant tumor heterogeneity and the development of drug resistance, which greatly limit the efficacy of conventional targeted therapies. Emerging evidence has increasingly highlighted the therapeutic potential of inducing regulated cell death modalities such as calcicoptosis and ferroptosis in overcoming these challenges. In this study, we developed an inhalable, pH-responsive, multifunctional drug delivery system (PCAL microspheres) to address the limitations of traditional single-target therapies. The PCAL microspheres consist of a core composed of poly(D, L-lactide-co-glycolide) (PLGA: P) loaded with erlotinib (ERL) and artesunate (ART), encapsulated within a calcium phosphate (CaP: CA)-based mineralized shell mediated by bovine serum albumin. The surface of the microspheres is further functionalized with iron-saturated lactoferrin (Holo-Lf: L), enabling active targeting of lung cancer cells. Upon delivery, ERL released from PCAL inhibits tumor cell proliferation by suppressing epidermal growth factor receptor (EGFR) activation. ART induces intracellular Ca²⁺ accumulation by inhibiting sarcoplasmic/endoplasmic reticulum Ca²⁺-ATPase (SERCA) and promoting CaP degradation, leading to endoplasmic reticulum stress and mitochondrial dysfunction. Concurrently, ART and Holo-Lf jointly induce ferroptosis, resulting in plasma membrane pore formation and further amplification of Ca²⁺ influx. This 'calcicoptosis-ferroptosis' dual pathway generated a synergistic antitumor effect. In a mouse model of lung cancer, inhalation of PCAL significantly inhibited tumor growth. Moreover, the treatment exhibited favorable safety profiles without detectable systemic toxicity. These findings demonstrate that nebulized PCAL microspheres provide a promising and innovative strategy for the effective treatment of NSCLC. STATEMENT OF SIGNIFICANCE: Non-small cell lung cancer (NSCLC) remains one of the most challenging cancers to treat, largely due to tumor heterogeneity and rapid development of drug resistance. In this work, we developed inhalable PCAL microspheres, a targeted drug delivery system that acts directly in the lungs. Unlike conventional single-pathway therapies, PCAL combines erlotinib and artesunate within a PLGA core, coated with calcium phosphate and modified with iron-saturated lactoferrin for tumor targeting. This design activates a dual mechanism-calcicoptosis (calcium overload-induced cell death) and ferroptosis (iron-dependent lipid peroxidation)-leading to a powerful synergistic antitumor effect. In vivo studies demonstrated significant tumor regression without systemic toxicity, suggesting that PCAL as a promising and impactful strategy for NSCLC treatment.