Taming the Tumor Stroma: A Two-Stage Targeted Nanocapsule for Potent Deep Chemo-Immunotherapy in Triple-Negative Breast Cancer

<b>Background</b>: The tumor microenvironment (TME) poses significant challenges to effective therapy, with cancer-associated fibroblasts (CAFs) playing a key role in tumor progression and drug resistance in triple-negative breast cancer (TNBC). Herein, a TME responsive nanocapsule, NPC-ABS/FDS, was developed utilizing baicalein, a CAFs modulator, and the cytotoxic drug doxorubicin to selectively target CAFs and tumor cells, respectively, in a stepwise manner. <b>Methods</b>: NPC-ABS/FDS was designed with CD13-mediated primary targeting for tumor accumulation and secondary targeting via σ-receptor binding (ABS nanoparticles) for CAFs and folate modification (FDS nanoparticles) for cancer cells. Physicochemical properties were assessed using TEM, particle size, and ζ-potential analyses. Fluorescence imaging evaluated tumor retention, while cellular uptake and TME modulation were analyzed in vitro and in vivo. <b>Results</b>: The successful preparation of NPC-ABS/FDS was demonstrated by its uniform morphology, stable characteristics, charge reversal, and increased particle size. Fluorescence imaging confirmed prolonged peritumoral retention. Cellular uptake increased 2.5-fold for baicalein in CAFs and 4.3-fold for doxorubicin in cancer cells. NPC-ABS/FDS downregulated α-SMA and FAP, reducing CAFs activation, improving intratumoral drug penetration, and enhancing CD8⁺ and CD4⁺ T cell infiltration while decreasing regulatory T cells. <b>Conclusions</b>: NPC-ABS/FDS effectively modulates multiple TME components, including CAFs and immune cells, and improves drug delivery in TNBC. These findings may support the development of improved therapeutic approaches for TNBC.