Dendritic Cell-Derived Exosomes Driven Drug Co-Delivery Biomimetic Nanosystem for Effective Combination of Malignant Melanoma Immunotherapy and Gene Therapy.

Purpose: Malignant melanoma (MM), the most lethal skin cancer, is highly invasive and metastatic. These qualities are related to not only genetic mutations in MM itself but also the interaction of MM cells with the immune system and microenvironment. This study aimed to construct a combined immunotherapy and gene therapy drug delivery system for the effective treatment of MM. Methods: Mature dendritic cell (mDC) exosomes (mDexos) with immune induction functions were used as carriers. BRAF siRNA (siBRAF) with the ability to silence mutated BRAF in MM was encapsulated in mDexos by electroporation to construct a biomimetic nanosystem for the codelivery of immunotherapy and gene therapy drugs (siBRAF-mDexos) to the MM microenvironment. Then, we investigated the nanosystem's serum stability and biocompatibility, uptake efficiency in mouse melanoma cells (B16-F10 cells), cytotoxicity against B16-F10 cells and inhibitory effect on BRAF expression. Furthermore, we evaluated its antimelanoma activity and safety in vivo. Results: SiBRAF-mDexos were nanosized. Compared to siBRAF, siBRAF-mDexos displayed significantly increased serum stability, biocompatibility, uptake efficiency in B16-F10 cells, and cytotoxicity to B16-F10 melanoma cells; they also had a significantly greater inhibitory effect on BRAF expression and induced T-lymphocyte proliferation. Moreover, compared with siBRAF, siBRAF-mDexos showed significantly enhanced anti-MM activity and a high level of safety in vivo. Conclusion: The study suggests that the siBRAF-mDexo biomimetic drug codelivery system can be used to effectively treat MM, which provides a new strategy for combined gene therapy and immunotherapy for MM.

Effects of the surface charge of polyamidoamine dendrimers on cellular exocytosis and the exocytosis mechanism in multidrug-resistant breast cancer cells.

BACKGROUND: Polyamidoamine (PAMAM) dendrimer applications have extended from tumor cells to multidrug-resistant tumor cells. However, their transportation in multidrug-resistant tumor cells remains unclear. Herein, we investigated the exocytosis rule and mechanism of PAMAM dendrimers in multidrug-resistant tumor cells. RESULTS: Using a multidrug-resistant human breast cancer cell model (MCF-7/ADR), we performed systematic analyses of the cellular exocytosis dynamics, pathways and mechanisms of three PAMAM dendrimers with different surface charges: positively charged PAMAM-NH2, neutral PAMAM-OH and negatively charged PAMAM-COOH. The experimental data indicated that in MCF-7/ADR cells, the exocytosis rate was the highest for PAMAM-NH2 and the lowest for PAMAM-OH. Three intracellular transportation processes and P-glycoprotein (P-gp) participated in PAMAM-NH2 exocytosis in MCF-7/ADR cells. Two intracellular transportation processes, P-gp and multidrug resistance (MDR)-associated protein participated in PAMAM-COOH exocytosis. P-gp and MDR-associated protein participated in PAMAM-OH exocytosis. Intracellular transportation processes, rather than P-gp and MDR-associated protein, played major roles in PAMAM dendrimer exocytosis. PAMAM-NH2 could enter MCF-7/ADR cells by forming nanoscale membrane holes, but this portion of PAMAM-NH2 was eliminated by P-gp. Compared with PAMAM-OH and PAMAM-COOH, positively charged PAMAM-NH2 was preferentially attracted to the mitochondria and cell nuclei. Major vault protein (MVP) promoted exocytosis of PAMAM-NH2 from the nucleus but had no effect on the exocytosis of PAMAM-OH or PAMAM-COOH. CONCLUSIONS: Positive charges on the surface of PAMAM dendrimer promote its exocytosis in MCF-7/ADR cells. Three intracellular transportation processes, attraction to the mitochondria and cell nucleus, as well as nuclear efflux generated by MVP led to the highest exocytosis observed for PAMAM-NH2. Our findings provide theoretical guidance to design a surface-charged tumor-targeting drug delivery system with highly efficient transfection in multidrug-resistant tumor cells. Especially, to provide more DNA to the nucleus and enhance DNA transfection efficiency in multidrug-resistant tumor cells using PAMAM-NH2, siRNA-MVP or an inhibitor should be codelivered to decrease MVP-mediated nuclear efflux.