Near-infrared Light-responsive Drug-loaded Hydrogel For Photothermal-chemo Synergistic Lung Cancer Therapy

Objective: We constructed injectable sodium alginate(ALG)hydrogel,which co-encapsulated the hydrophilic chemotherapy drug Dorubicin hydrochloride(DOX·HCl)and gold nanorods(Au/Ag NRs)with apparent absorption in the near-infrared region II(IR-II)to achieve the strategy of one injection and multiple light treatment and achieved the goal of photothermal-chemotherapy synergistic effect under 1064-nm laser irradiation in the treatment of non-small cell lung cancer.Methods:1.Preparation and characterization of Au / Ag NRs.Au/Ag NRs with strong absorbance in the NIR-II region were synthesized via seed-mediated growth procedures.DDTC was covalently coated on the first metal core(gold)surface to precisely control the second metal(Ag)growth to prepare a corn-like bimetallic core-shell structure with strong absorption in NIR-II.Next,Au/Ag NRs were characterized.The morphology,length,diameter,and dispersion of Au/Ag NRs were characterized by transmission electron microscopy(TEM).Then the absorption peaks of Au/Ag NRs were determined by UV-vis-NIR.The photothermal conversion ability and photothermal stability of Au/Ag NRs were calculated by 1064-nm near infrared-laser irradiation.2.Preparation and characterization of NR/DOX/ALG hydrogels.The gelation of sodium alginate with various concentrations in PBS buffer containing 1.8 n M Ca Cl2 was evaluated.And then,the NR/DOX/ALG hydrogels were characterized.The morphology of the hydrogel was observed by scanning electron microscopy(SEM).The rheological and Young's modulus were used to test the mechanical properties of the hydrogels.Next,the swelling ratio and in vitro degradation properties of the hydrogels were detected.The thermal curve evaluated the photothermal conversion ability and photothermal stability of NR/DOX/ALG hydrogels under 1064-nm near-infrared laser irradiation.Finally,the release of DOX·HCl from the NR/DOX/ALG hydrogels was measured by spectrofluorometer.3.Cell experiment Confocal microscopy(CLSM)and flow cytometry were used to test the uptake of DOX·HCl in hydrogel by A549 cells.Cell Counting Kit-8(CCK-8)was used to evaluate in vitro biosafety of hydrogels and the pernicious effect of photothermal properties on A549 cells.The cell killing assay was used to assess the synergistic effect of NR/DOX/ALG hydrogels on A549 cells.4.Animal experiments We observed the gelation of NR/DOX/ALG hydrogels in mice and evaluated the biosafety of hydrogels in vivo by weight monitoring,HE staining,and peripheral blood indexes.The degradation of hydrogels in vivo was detected at the corresponding time points by weighting the quality of hydrogels.Next,we constructed the mouse subcutaneous tumor model of the A549 tumor.The tumor growth curve,tumor quality,and TUNEL staining were used to evaluate the treatment effect.The mouse's systemic toxicity treated by NR/DOX/ALG hydrogels was tested by monitoring body weight and peripheral blood biochemical indexes of the mouse.Results:1.TEM images showed that Au/Ag NRs were bimetallic core-shell structures with corn-like shapes.The length and diameter of Au/Ag NRs were 146.84 ± 15.59 nm,53.06 ± 5.92 nm,respectively.The zeta potential was 35.1 MV.The longitudinal local surface plasmon resonance(LSPR)of Au/Ag NRs shifted from 820 nm to 1064 nm.The photothermal conversion efficiency of Au/Ag NRs solution irradiated by 1064 nm near-infrared laser is 33.7%.The temperature increased in a concentration-dependent and NIR-II light intensity-dependent manner and showed good photothermal stability.2.Notably,the hydrogel formed quickly when the ALG concentration was more significant than or equal to 5.0 mg/m L.From the SEM image,the resultant NR/DOX/ALG hydrogels exhibited a porous network microstructure.Compared with ALG and NR/ALG hydrogels,NR/DOX/ALG hydrogel has the highest hardness and a minor elasticity.The weight of NR/DOX/ALG increased three times after incubated with water for 24 h.But in vitro,it showed a slow degradation trend.Under the irradiation of 1064-nm near-infrared light,The NR/DOX/ALG hydrogels showed Au concentration-dependent and laser power-dependent photothermal effect and had good photothermal stability.The trapped DOX·HCL was released from the NR/DOX/ALG hydrogels gradually and sustainable.1064-nm laser irradiation efficiently triggered DOX·HCl release,exhibiting a greatly improved release rate.3.The total uptake of DOX·HCl in NR/DOX/ALG hydrogels by A549 increased with prolonged incubation time.The photothermal effect could promote the uptake of DOX·HCl by A549 after receiving 1064-nm near-infrared light.The prepared hydrogel had good biological safety,and the killing effect of NR/ALG hydrogel on A549 cells increased with the rising of Au content.Moreover,the NR/DOX/ALG hydrogels plus irradiation exhibited the best anticancer activity due to the combination of PTT and released DOX·HCl-mediated chemotherapy.4.The hydrogel forms 30 min after subcutaneous injection of the precursor solution.Besides,the formed hydrogel degraded as the implantation time increased.A mild inflammatory reaction was observed around the hydrogel,but it did not lead to systemic toxicity and hemolytic reaction.Collectively,these results confirmed the excellent biosafety of the injectable ALG hydrogels.In vivo tumor inhibition experiments confirmed that NR/DOX/ALG hydrogels achieved the long-term retention of therapeutic agents in tumors and one injection and repeated treatment strategy.Excitingly,the photothermal-chem synergistic therapy eliminated the tumor target.Conclusion:1.In this experiment,a corn cob-like bimetallic Au/Ag core-shell nanostructure with strong absorption in the second region of the near-infrared was prepared by the seeding method.The synthesized Au/Ag NRs have good photothermal conversion efficiency and photothermal stability.2.The NR/DOX/ALG hydrogels were successfully prepared.The prepared hydrogel had a porous structure,good mechanical properties,good swelling,and in vitro degradation properties.The Au/Ag NRs encapsulated in the hydrogel still had good photothermal performance and photothermal stability.3.The photothermal effect under 1064-nm laser irradiation could effectively promote the release of the small molecule drug DOX? HCl,which provided an excellent formulation for realizing the “on-demand administration” strategy.4.The uptake of doxorubicin from NR/DOX/ALG hydrogels by A549 cells increased in a time-dependent manner.The photothermal effect promoted the release of doxorubicin,which increased the uptake of doxorubicin by A549 cells.5.Sodium alginate hydrogel had good biological safety in vitro.The photothermal effect could kill tumor cells on a content dependent of Au/Ag NRs in the NR/ALG hydrogels.6.Compared with single photothermal therapy or single chemotherapy,NR/DOX/ALG hydrogels had photothermal-chemotherapy synergistically treatment under NIR-II laser irradiation,to achieve "1+1>2" effect 7.The composite nano-hydrogel precursor solution had gelled 30 minutes after being injected into mice.The hydrogel could be gradually degraded in the body.The formed hydrogels could cause mild local inflammation with no hemolytic reaction.During the whole experiment,the body weight of the mice did not change significantly,and there were no abnormalities in the biochemical indicators of the peripheral blood.In summary,the prepared hydrogels had good biological safety in vivo.8.The results of animal experiments confirmed that NR/DOX/ALG hydrogels achieved the strategy of one injection and multiple light treatment and achieved the goal of photothermal-chemotherapy synergistic effect under 1064-nm laser irradiation.