Study On The Pulmonary Process And Anti-lung Cancer Efficacy Of Pulmonary Delivery Of Peptide Self-assembled Nanoparticle

In order to improve therapeutic efficacy for the treatment of lung cancer,inhaled nanomedicines have been increasingly drawing attention in the last two decades.Extensive work has demonstrated that there are several advantages associated with inhalation delivery of nanoparticles,including improved lung bioavailability,reduced potential of metastasis and minimized systemic adverse effects when compared to systemic administration.The tumor accumulation of drug and/or nanoparticles has to overcome lung clearances and to enhance the transport to tumors.However,nanoparticles with single size could not simultaneously achieve prolonged lung retention and high tumor accumulation and penetration.To deal with this dilemma,the present thesis developed a sizetransformable nano drug delivery system with mucus penetration ability to overcome various biological barriers of pulmonary delivery and thereby enhancing antitumor efficacy.In addition,very little is currently known about how inhaled nanomedicine for lung cancer treatment overcomes biological barriers hampering the tumor availability of drug and nanoparticles.To fill this gap,we set out to quantify the pulmonary and tumor fate of nanoparticles with a view to elucidating their biological features relevant to lung retention,transport to tumor,and tumor accumulation and penetration.Firstly,the thesis synthesized the linear-dendritic block copolymer modified with nearinfrared fluorescent dye indocyanine green derivative(ICGD),called K4 monomer,and its self-assembly could form K4 NPs to load antineoplastic agent 7-Ethyl-10hydroxycamptothecin(SN38)with particle size of 28 nm.K4 NPs could be coated with hyaluronic acid(HA)modified with dopamine and folic acid via phenylboric acid on its surface to form HA-K4 NPs with particle size of about 120 nm.Under conditions mimicking tumor microenvironment in vitro,HA-K4 NPs could be transformed to smallsized K4 NPs in a responsive manner.In addition,the nebulization performance of K4 NPs and HA-K4 NPs showed good inhalability.Then the thesis evaluated the mucus and tumor penetration abilities of K4 and HA-K4 NPs in vitro and in vivo and their cytotoxicity against Lewis lung carcinoma cells(LLCs)in vitro.Compared with small-size K4 NPs,HA-K4 NPs showed stronger mucus penetration and lung retention.In vitro and in vivo tumor penetration results showed that K4 NPs showed better tumor penetration at the initial stage,while HA-K4 NPs penetrate the tumor tissues in a delayed manner.In addition,both nanoparticles have strong sonodynamic transformation ability and could produce reactive oxygen species(ROS)under ultrasonic conditions.The cytotoxicity of K4 and HA-K4 NPs was further enhanced after ultrasound treatment,demonstrating a synergistic or additive effect of chemo-sonodynamic therapy on the cytotoxicity in vitro.Next,the thesis investigated the pulmonary and tumor pharmacokinetics of K4 and HAK4 NPs after intratracheal instillation to mice bearing orthotopic Lewis lung carcinoma tumors.It was found that the AUC values of SN38 and fluorescence intensity in lung tissues from instilled HA-K4 NPs were 43.7-99.7 folds of that by injected HA-K4 NPs,indicating that pulmonary delivery could significantly increase lung tissue exposure.Relative to small-size nanoparticles,the large-size counterparts exhibit superior ability of mucus penetration and lung retention,leading to an increase in lung bioavailability of SN38 by 58.5%.In addition,HA-K4 NPs could enhance the tumor accumulation via direct access or transcytosis of nanoparticles and diffusion of the released drug,and the large nanoparticles penetrate within tumor tissues after size-transformation,leading to increases in tumor bioavailability of sonosensitizer by 5.99-fold and SN38 by 1.99-fold compared with the small-size nanoparticles.HA-K4 NPs exhibits good safety profile,and the sustained and high levels of sonosensitizer and SN38 conferred in the tumor after pulmonary delivery triggered chemo-sonodynamic combination therapy,extending the survival time of mice bearing orthotopic Lewis lung carcinoma tumors from 15 days to 22 days,which achieved significant anti-tumor efficacy.Finally,to explore the effect of macrophage phagocytosis on pulmonary delivery of nanoparticles,the thesis synthesized three polypeptide derivatives modified with different targeting groups,which self-assembled to form three nanocarriers,namely phenylboric acid modified ICP nanoparticles for tumor cell targeting,phenylboric acid and folic acid dual-modified FCP nanoparticles for enhanced tumor cell targeting and phenylboric acid and mannose dual-modified MCP nanoparticles for macrophage targeting.The particle size distribution,zeta potential,morphology and critical micellar concentration of the three nanoparticles were similar,and only the target groups affected the fate of the nanoparticles after pulmonary administration.Upon establishing a sodium alginate/PVA hydrogel 3D LLC and macrophage co-culture system,the competitive uptake of nanoparticles by cells was evaluated in vitro and the results showed that FCP NPs could be selectively uptake by tumor cells and MCP NPs could be selectively uptake by macrophages.Subsequently,in vivo efficacy evaluation in mice bearing orthotopic Lewis lung carcinoma tumors showed that there was no significant difference in tumor weight between MCP NPs and the control group,while ICP NPs inhibited tumor growth to a certain extent with an inhibition rate of 29.46%.The tumor inhibition rate of FCP NPs was 48.51%,markedly higher than the other two nanoparticles.The antitumor effect of MCP NPs was significantly weaker than that of FCP NPs and ICP NPs,indicating that targeting macrophages decreased the anti-tumor efficacy of pulmonary delivery of nanoparticles.In summary,this thesis has investigated the effect of particle size and ligand modification of inhaled nanoparticles on the pulmonary and tumor fates.The present results demonstrated that macrophage phagocytosis may restrict the drug availability to tumor cells,reducing the anti-tumor efficacy of inhaled nanomedicine.In addition,an inhaled size-transformable nanosystem could not only restrict mucociliary clearance by immediate penetration through the mucus,but also retard absorption clearances by delaying the drug release and translocation of nanoparticles into the bloodstream and extrapulmonary organs,and improved the tumor accumulation and penetration through size transformation.Pulmonary delivery of such a size-transformable nanocarrier represented a promising means to co-deliver chemotherapeutics and sonosensitizers for noninvasive lung cancer management.