Therapeutic Strategy Of Nano Optimized Metabolic Inhibitor Combined With Innate Immune Activator For Hepatocellular Carcinoma

Hepatocellular carcinoma(HCC)is a serious threat to human health.The traditional treatments of liver cancer mainly includes surgical resection,radiotherapy,chemotherapy and immunotherapy.But they all have their own disadvantages.The treatment method of blocking glucose metabolism or energy metabolism is a new tumor therapy.This therapy inhibits the relevant metabolic pathways of tumor cells through relevant small molecule inhibitors,so as to achieve the purpose of inhibiting tumor growth.Among these metabolic blockers,3-bromopyruvate(3-BPA)and 6,8-bis(benzylthio)octanoic acid(CPI-613)have been widely studied.In recent years,with the development of nanotechnology,nano materials are more and more widely used in biomedicine.As drug carriers,nano materials have the advantages of protecting drugs,reducing drug systemic toxicity,improving drug uptake,sustained-release and targeted drug delivery.In addition,the innate immune signaling pathway mediated immunotherapy mediated by cyclic guanosine monophosphate synthase/interferon gene stimulating factor(c GAS/STING)has attracted more and more attention.In this study,(3-BPA+CPI-613)@PLGA nanoparticles(BCP NPs)were synthesized by encapsulating 3-BPA and CPI-613 into PLGA nano carrier at the same time.The composite nanoparticles have synergistic anti HCC ability both in vivo and in vitro.At the same time,the combination of the composite nanoparticles and STING agonist ABZI enhanced the effect of tumor inhibition.The specific contents are divided into the following three parts:Part I:Synthesis and characterization of BCP NPs.We prepared double drug loaded PLGA composite nanoparticles(BCP NPs)by emulsion solvent evaporation method.Then the particle size and potential parameters were measured by DLS method.The morphology and crystal form of the drug with the best ratio were studied by TEM and XRD.The in vitro stability and drug release ability of BCP nanoparticles were studied.Part II:Antitumor effect of BCP NPs in vitro.We evaluated the antitumor effect of BCP NPs in vitro by MTT assay,apoptosis assay and mitochondrial membrane potential assay.In addition,we explored the anti-tumor mechanism of composite nanoparticles by measuring metabolism related indicators.The results showed that BCP composite nanoparticles could effectively inhibit tumor cells.Its antitumor mechanism is that BCP composite nanoparticles greatly enhance metabolic inhibition by enhancing the inhibition of ATP and lactic acid and reducing glucose consumption.BCP NPs can also induce the dimerization of STING in tumor cells and activate the STING signal pathway.Part III:Antitumor effect of BCP NPs in vivo.We evaluated the antitumor properties of BCP nanoparticles on Hepa1-6 subcutaneous tumor bearing mouse model.The results showed that BCP nanoparticles could significantly inhibit tumor growth;During the treatment,the weight of mice did not decrease significantly,indicating that the systemic toxicity of the drug was low;We further studied the antitumor effect of BCP NPs combined with STING agonist(ABZI).In vivo experiments showed that the combination of BCP NPs and ABZI significantly enhanced the ability to inhibit tumor.Flow cytometry showed that the combined use of BCP nanoparticles and abzi enhanced the activation of CD8~+T cells in mice.In conclusion,we have developed a new type of bifunctional nanoparticles.The cocktail strategy combined with immune activators can effectively inhibit tumor growth.Specifically,we synthesized BCP nanoparticles,which can effectively reduce the metabolism of tumor cells and inhibit the proliferation of tumor cells.At the same time,we found that Hepa1-6 cells treated with BCP NPs had obvious TBK1 phosphorylation and STING dimerization.We speculate that the underlying mechanism is that BCP nanoparticles cause mitochondrial disorder and release mitochondrial DNA(mt DNA),thus activating the intracellular c GAS/STING signaling pathway.Finally,we combined BCP nanoparticles with STING agonists to further activate the immune system of tumor microenvironment and achieve an efficient cascade of enhanced combined therapy.The combination of immunosuppression and metabolic inhibition in this study has high theoretical reference value and clinical transformation prospect.