Exploration Of Novel Pharmaceuticals For Cancer Chemotherapy And Vaccination

Cancer is one of the most lethal diseases which keep threatening the health and life of people, and the development of efficient anticancer therapy is of great significance. Chemotherapy and immunotherapy hold the most promising potential due to their wide applicability. Focusing on the primary restrictions in these two areas, novel insoluble chemo-drug nanocrystallite and biomimetic vaccine formulations were proposed and developed by combining the latest theoretical and technical breakthrough.This thesis mainly included the following issues:1. PTX nanocrystallites of uniform and controllable size were prepared by developing a novel droplet-confined/cryodesiccation-driven crystallization approach, and the size effect on anticancer performance were evaluated in vitro. By confining paclitaxel (PTX) molecules inside separated emulsion droplets (Oil-in-Water emulsion), drug molecules of defined amount were engaged during the crystallization process, thus leading to the successful preparation of PTX nanocrystallites with controllable sizes. The performances of PTX nanocrystallites in two different average sizes (～10 nm and ～70 nm) on tumor cellular uptake, cytotoxicity, and tissue penetration were systematically evaluated in comparison with Taxol(?). The results showed that PTX nanocrystallites outperformed Taxol(?) on all the above aspects. Further study carried out in 3D multicellular tumor spheroids (MCTSs) revealed that the ultra-nano size contributed greatly to the tissue penetrating ability in the assistance of iRGD, as nanocrystallites of ～10 nm size (iNDs) exhibited the most efficient infiltration and cell killing ability in MCTSs.2. The effects of PTX nanocrystallites on anticancer therapy and safety were further explored in murine model, and the superiority of ultra-small sized PTX nanocrystallites was demonstrated. Systematical in vivo experiments revealed that the use of PTX nanocrystallites could significantly reduce the drug distribution in liver, and benefit the drug enrichment inside tumor. Taking advantages of iRGD promotion and ultra-nano size, iNDs could achieve successful tumor tissue penetration and arrive at cancer stem cell, thus leading to much more effective tumor suppression, lifetime extension, and metastasis prevention over Taxol(?). Besides, the use of PTX nanocrystallites was demonstrated to greatly reduce adverse effects including hypersensitivity reactions, hematological toxicity, and neurotoxicity.3. Biomimetic vaccine was constructed from Bacillus Lactobacillus casei via optimized hydrothermal treatment, and its performance and mechanism on immune stimulation were evaluated in vitro. Through optimized pretreatment and hydrothermal treatment, biomimetic vaccine backbones (DB) were transformed from Bacillus Lactobacillus casei. These DB thus inherited both surface ligands and pathogen-like physical properties from their archetypes. Subsequently, model antigen OVA and adjuvant CpG could be efficiently loaded in virtue of the hollow and porous structure of DB, leading to the successful construction of biomimetic vaccine which closely resembles the natural bacteria. In vitro research showed that DB could be rapidly recognized, captured, and internalized by antigen presenting cells (APCs) via the introduction of mannose receptor, and mainly resided in early endosomes after cellular uptake. Owing to highly efficient APC uptake and special intracellular trafficking, the biomimetic vaccine could promote enhanced APC activation and antigen delivery/presentation, leading to the potent proliferation of CD8+ T cells and their differentiation into antigen-specific cytotoxic T lymphocyte (CTL).4. By employing murine tumor model, the biological safety and efficacy in anticancer therapy/prophylaxis of the biomimetic vaccine were systematically investigated. The results demonstrated that the injection of biomimetic vaccine led to no acute inflammatory reaction and injury to visceral organs, confirming the excellent safety of the current formulation. After administration, the biomimetic vaccine could recruit patrolling APCs to the injection site to load up antigen, and promote their subsequent migration to the draining lymph nodes, where effector cells were then efficiently primed. The anticancer investigation showed that the use of biomimetic vaccine greatly enhanced the CTL infiltration in tumor microenvironment, and achieved more potent anticancer effects than traditional attenuated live vaccine in aspects of tumor suppression, metastasis prevention, lifetime extension, and health condition improvement. In combination with early treatment, the biomimetic vaccine could even achieve complete tumor elimination. Besides, the biomimetic vaccine could also induce a strong and long-lasting immune memory, thus achieving excellent performance in tumor prophylaxis.In summary, this thesis solved the primary issues which currently hamper the development of chemotherapy and immunotherapy by exploratory research, and very promising therapeutic strategies were proposed. These will break new ground for the anticancer therapy development in the future.