Dual-targeted pH-responsive nanoliposome (M5/NMN/DEAP/iRGD-Lip): Immunostimulatory-photodynamic synergy for breast cancer therapy.
Photodynamic therapy (PDT) holds promise for combination antitumor therapies by triggering immunogenic cell death (ICD). ICD is defined as the process by which tumor cells, upon death induced by external stimuli, convert from a non‑immunogenic to an immunogenic state, thereby mediating an anti‑tumor immune response in the host. But the poor aqueous solubility and inadequate tumor targeting of photosensitizers hinder their clinical translation. This study focuses on a novel BODIPY photosensitizer (M5) and aims to improve its antitumor efficacy via efficient tumor-targeted delivery and controllable release. Herein, we successfully synthesized iRGD-functionalized DSPE-PEG2000-iRGD and pH-sensitive HA-g-DEAP polymers, and further fabricated multifunctional M5/NMN/DEAP/iRGD-Lip liposomes via the thin-film dispersion method, which possess pH responsiveness and enhanced tumor-targeting ability. β-Nicotinamide Mononucleotide (NMN), a NAD + precursor, exerts a potent stimulatory effect on T-cell activation; 3-(Diethylamino)propylamine (DEAP) and hyaluronic acid (HA) can form pH-responsive HA-g-DEAP; the iRGD peptide (CRGDK/RGPDC), upon hydrolysis at its C‑terminus, exposes a motif that binds to neuropilin‑1 (NRP1), thereby conferring tumor‑targeting and tissue‑penetrating properties, endowing the liposomes with tumor-targeting and tissue-penetrating capabilities. M5 exhibits a high molar absorption coefficient of 5.33 × 104 M⁻¹ cm⁻¹ and a singlet oxygen quantum yield of 0.3854. In vitro cellular assays showed IC50 values of 104.1 nM and 72.68 nM in breast cancer MDA-MB-231 and 4T1 cells, respectively. Treatment with M5/NMN/DEAP/iRGD-Lip induced apoptosis rates of 56.84% and 55.6% in MDA-MB-231 and 4T1 cells, respectively. T-cell co-culture assays showed that M5/NMN/DEAP/iRGD-Lip increased the proportions of CD4+ and CD8+ T cells while reducing the proportion of regulatory T cells (Tregs) among CD4+ T cells. Collectively, the multifunctional M5/NMN/DEAP/iRGD-Lip liposomes integrate targeted delivery, pH-controlled release, and synergistic PDT-immunotherapy, effectively addressing key limitations of conventional photosensitizers. This work provides a promising nanoplatform for the development of novel combination therapies against breast cancer, laying a foundation for future preclinical and clinical translations.