Tumor Microenvironment-modulated Metal Organic Framework For Breast Cancer Therapy

Breast cancer is the most prevalent malignant tumor among women and has remained an important topic of oncological research.Currently,chemotherapy and immunotherapy are the main pharmacological modalities for inhibiting breast cancer progression in clinical practice.However,due to the complexity and heterogeneity of the tumor microenvironment（TME）,a single drug delivery regimen is usually constrained by the TME,resulting in inefficient delivery of chemotherapeutic and immunotherapeutic drugs,poor therapeutic efficacy,and an inadequate immune response.The therapeutic strategy of utilizing biometabolic molecules in the TME to achieve oxidation of reactive oxide species（ROS）through cascade catalysis is non-invasive,less prone to side effects and drug resistance,and holds greater potential in the treatment of tumors.Therefore,a rational approach involves combining enzymes,chemotherapeutics,and functional molecules to up-regulate ROS levels in the TME through cascade catalysis,thereby aiding in improving drug resistance of tumor cells and enhancing the efficiency of tumor therapy.In this study,we firstly designed cascade-responsive nanoparticles,GLDFe/Z-FA,to up-regulate the ROS level in tumors by regulating the tumor metabolic mode and reversing multidrug resistance.The Fe²⁺-doped metal-organic framework（MOF）Fe/ZIF-8 was constructed and loaded with Glucose oxidase（Gox）,L-arginine（L-arg）and doxorubicin hydrochloride（Dox）.Additionally,the nanocarriers were functionalized with folic acid（FA）to enable GLDFe/Z-FA to actively target the highly expressed folate receptor on the surface of breast cancer cells.The prepared GLDFe/Z-FA particles,with a particle size of about 170 nm and a potential of-20.7 m V,rapidly released Dox in media with p H 6.2 and 5.0,with cumulative release rates of 59.6%and99.1%,respectively,for 30 h.FA increased the dispersion and stability of the nanoparticles,while also endowing them with an active targeting ability.In cellular experiments,GLDFe/Z-FA nanoparticles showed a higher uptake rate and a stronger inhibitory effect on tumor cells.Gox-loaded nanoparticles catalyzed endogenous glucose catabolism and in situ hydrogen peroxide（H₂O₂）generation,leading to a decrease in intracellular glucose levels and adenosine triphosphate（ATP）content,thereby reducing glucose utilization and inducing tumor starvation.Fe²⁺catalyzed the generation of·OH from H₂O₂,inducing ROS overload while implementing chemokinetic therapy.Excessive intracellular ROS oxidized L-arg to release nitric oxide（NO）,which inhibited the expression of P-glycoprotein（P-gp）,a Dox efflux protein,on the cell membrane,thus accumulating intracellular Dox content and enhancing cytotoxicity.Moreover,GLDFe/Z-FA catalyzes the oxidation of glutathione（GSH）,further disrupting intracellular redox homeostasis and induced cell death.In the in vivo drug evaluation study in Balb/c mice,Fe/Z-FA was actively enriched at the tumor site and prolonged the circulation time of the drug in vivo.GLDFe/Z-FA showed the highest tumor inhibition efficiency under the synergistic triple strategy（starvation,chemodynamic,and gas therapy）in antitumor experiments and followed the iron death pathway in vitro and in vivo,effectively reversing tumor multidrug resistance with good biocompatibility.Metastasis and recurrence of breast cancer are the primary factors contributing to poor prognosis and mortality of breast cancer patients.The TME-responsive nanoparticles developed above achieve a synergistic effect by combining starvation therapy,chemodynamic therapy,and gas therapy to reverse the multidrug-resistant environment of tumors,thereby exhibiting a potent anti-tumor effect.While Dox has been demonstrated to induce immunogenic cell death（ICD）,the immunogenic efficacy induced by chemotherapeutic drugs is limited and is not able to to sustain a long-term anti-tumor immune response.Photothermal therapy（PTT）promotes tumor ablation by transforming"cold tumor"into"hot tumor",and increases drug penetration and immune cell infiltration.Dying tumor cells release damage-related molecular patterns（DAMPs）under the action of light and heat,enhancing the irritability of natural tumor antigens.The pro-inflammatory cytokines produced further stimulate the immune system.In addition,tumor cells killed by PTT release tumor-associated antigens as substrates for in situ autologous vaccines,enhancing the ICD-induced anti-tumor immune response and establishing long-term anti-tumor immune memory,thereby completely eradicating residual tumor cells throughout the body.Therefore,in the second part of the study,nano-adjuvants with PTT and cascade catalytic activities were designed to induce intracellular redox imbalance,remodel the tumor microenvironment,sensitizeαPD-L1 therapeutic effects,and enhance the comprehensive immune treatment of breast cancer.The multifunctional nano-adjuvant GITFe/Z-HA developed in this chapter is based on Fe³⁺-coordinated ZIF-8 modified with target hyaluronic acid（HA）,loaded with Gox,the phototherapeutic agent new indocyanine green（IR820）,and the oxygen depletion activating precursor tirazamine（TPZ）.The multifunctional nano-adjuvant GITFe/Z-HA has a particle size of about189.7 nm and a potential of-23.6 m V,possessing a crystal structure,acid sensitivity,and good stability.Moreover,it maintains a hemolysis rate of less than 5%at high concentrations,meeting the safety standard for intravenous drug delivery.Validated by in vitro and cellular experiments,the nanoadjuvant can achieve a cascade reaction mediated by Gox,Fe³⁺and IR820 in vitro and in tumor cells,depleting intracellular glucose and oxygen,and promoting the elevation of ROS levels.Meanwhile,tumor hypoxia was exacerbated under photodynamic therapy（PDT）,providing a favorable environment for TPZ to exert its therapeutic effects.The combination therapy of PTT,PDT and chemotherapy demonstrated a high tumor inhibition rate and good synergistic effects.Additionally,the ROS/GSH imbalance induced by this therapeutic approach combined with the PTT strategy induced ICD,triggering changes in DAMPs（CRT,HMGB1,and ATP）,and promoting T-lymphocyte infiltration.The results of in vivo distribution experiments confirmed that Fe/Z-HA could prolong the in vivo circulation time and allow better drug accumulation at the tumor site compared to free IR820.In the in vivo pharmacodynamics experiments,after laser irradiation in the GITFe/Z-HA group of unilateral subcutaneous tumor model in mice,the highest percentages of CD8⁺T lymphocytes and CD4⁺T lymphocytes were observed,and the percentage of regulatory T cells（Tregs）decreased significantly,which successfully stimulated the anti-tumor immune response.In a bilateral subcutaneous tumor model,GITFe/Z-HA in combination withαPD-L1 significantly activated T cell killing capacity under laser irradiation,ablated proximal and suppressed distal tumors.Increased infiltration of splenic CD4⁺Tmem cells and CD8⁺Tmem cells confirmed more effective activation of long-term immune memory under the combination,offering the possibility of tumor vaccine construction.In the in vivo lung metastasis model,CD8⁺T lymphocyte infiltration sensitized theαPD-L1 response,activating the systemic immune response,eliminating the metastatic cancer cells in the lungs,and improving the survival rate of mice.In summary,the development of TME-responsive antitumor nanoplatforms based on free radical oxidation in this study overcomes the limitations of single treatment modalities by environmentally driven disruption of redox homeostasis in tumor cells.It addresses the challenges of drug resistance and metastasis in breast cancer,thereby improving the prognosis of tumor patients.This strategy of microenvironmental remodeling through combination therapy is not only confined to breast cancer treatment,but also provides novel insights and approaches for the management of other types of cancers.