Cathepsin B Response Aggregation Nanotheranostic Agents For Dual-mode Imaging-guided Triple-therapy For Cancer

Photothermal therapy(PTT)is a promising strategy for the local thermal ablation of tumors.Lasers are used to irradiate tumor sites,exploiting their high energy to induce heating and thus cause cell damage.For example,Nd:YAG lasers have been used in clinic for endoscopic irradiation of obstructing endobronchial cancers.After PTT,the damaged cells released damage-associated molecular patterns such as calreticulin(CRT),high mobility group protein B1(HMGB1)and adenosine triphosphate(ATP),which act as immunogenic signals to promote dendritic cells to engulf tumor cells and activate the host’s immune system to fight tumors.To further improve the effect of PTT,contrast-enhanced PTT using near-infrared(NIR)photosensitizers were employed,which showed advantages of lower power,better selectivity and superior therapeutic effects.Despite there promises,accurate tumor positioning for lasers irradiation is a prerequisite for effective PTT,which remians a challenge in positioning tumor location during the treatment process.Otherwise,the residual or tumor metastasis tumor cells outside the laser irradiation range lead to certain limitations of single PTT in tumor treatment.To ensure the precise localization of laser irradiation,nanotheranostics provide an effective approach.Nanotheranostics have been developed to achieve diversified image and therapeutics strategies.Nanotheranostics agents integrate imaging agents and therapeutic drugs into one space,and targeted accumulation in tumor tissue,which is expected to enhance imaging capabilities and achieve optimal therapeutic effects.Combining PTT with nanotheranostics is a promising strategy for precise localization of laser irradiation.Precise directional guidance and real-time monitoring could provide key information for local thermal ablation of tumors.Thus,the development of PTT based nanotheranostics is of great significance for PTT accurate tumor positioning and showed an important impact on the clinical application.In clinical,magnetic resonance imaging(MRI),computed tomography(CT),positron emission tomography(PET),near-infrared fluorescence imaging(NIRFI)and other imaging modality have been used for localization and auxiliary diagnosis of various solid malignant tumors.Currently,multimodality imaging has been broadly concerned in research works and clinical applications to improve the accuracy of medical diagnosis to obtain precise and reliable information at the disease sites.The MRI/NIRFI dual-mode system realized the complementarity of diagnostic parameters,which significantly improves the quantification accuracy and spatial resolution of tumor positioning.Here,we propose a two-wave variable nanotheranostics strategy to enhance MRI/NIRFI dual-mode image-guided photo-induced triple-therapy(Figure0-1).Nanotheranostics agents were composed of thermosensitive liposomes(TSLs)co-loaded with SPIONs,IR780 and chemotherapeutic drug Abemaciclib(SIA-TSLs).The peptide ACKFRGD and click group CABT were co-modified on the surface of SIA-TSLs to form SIA-αTSLs.The SIA-αTSLs could achieve the following functions:i)precisely localized light irradiation:the RGD moiety in ACKFRGD peptide could target to the tumor site.The KFRGD moiety in ACFKRGD was cleaved off to expose AC with the high expression of cathepsin B in tumor tissue.1,2-thiolamino groups on AC occurred a click cycloaddition with the contiguous cyano groups on CABT,leading to the aggregation of SIA-αTSLs.SPIONs and IR780 act as imaging agents for MRI and NIRFI,respectively.After the aggregation of SIA-αTSLs in tumors,MRI/NIRFI imaging capability was significantly improved,achieving precisely localized light irradiation.ii)light-controlled drug release:after the accurately localized laser irradiation,IR780/SPIONs convert light energy into thermal energy to cause localized hyperthermia in the tumor.The local tumor temperature was higher than the phase transition temperature of thermosensitive liposomes,resulting in enhanced lipid membranes fluidity and permeability,inducing morphological changes in SIA-αTSLs.Those promoted the release of Abemaciclib in tumor tissues.This MRI/NIRFI image-guided photo-induced triple-therapy strategy exhibited great potential in achieving precise tumor localization therapy.Eradicating tumors completely by PTT alone can not be sufficient,and multimodal therapy combined with PTT is promising for cancer treatment.Several proposed combination partners for PTT,such as chemotherapy or immunotherapy,have been already well-established.PTT ablated specific tumors and induces ICD by localized photothermal.To enhance the effect of PTT,chemotherapy can not only enhanced the local tumor treatment effect,but also eliminate the the residual tumor cells.Meanwhile,immunotherapy could enhance PTT-induced ICD effect and further exerted systemic antitumor immunity.Therefore,we propose a dual-mode imaging-guided photo-induced photo/chemo/immuno-therapy to enhance the efficiency of tumor treatment.ⅰ)After the dual-mode imaging guided near-infrared laser irradiation,the local heating caused by SPIONs/IR780 promoted the ICD,accompanied by the release of tumor-associated antigens and damage-associated molecular patterns(DAMPs).ⅱ)The local heating promoted the release of Abemaciclib,which induce G1 cycle arrest of tumor cells through inhibiting the phosphorylation of retinoblastoma(Rb).Abemaciclib could also effectively reduce the proportion of Tregs and the secretion of IL-10 and TGF-β in tumor tissue,alleviating the immunosuppressive tumor microenvironment.ⅲ)Dendritic cells(DCs)are the most effective antigen-presenting cells,which could uptake,process and present antigens to trigger effective antitumor T cell response.To enhance the PTT-induced immune response,Granulocyte-macrophage colony-stimulating factor(GM-CSF)was injected intratumorally to recruit DCs and initiate antitumor immunity.This dual-mode imaging-guided photo-induced triple-therapy strategy was expected to achieve suprior efficient tumor combination therapy.Main contents and results are listed as follows in this study:1.Determination method for Abemaciclib and IR780DocetaxelThe content determination methodology of Abemaciclib and IR780 was established,and its methodology was investigated.2.Preparation and characterization of SIA-αTSLsThermosensitive liposomes co-loaded with Abemaciclib,IR780 and SPIONs were prepared by thin film dispersion method,and the carrier was functionally modified.Single factor experiments were carried out to investigate the optimal preparation process,and the particle size,potential,drug loading and enzyme cross-linking ability were also investigated.The function of the prepared carrier was verified,and the in vitro MRI/NIRFI imaging ability,heating ability and Abemaciclib release ability were investigated.3.The imaging ability of SIA-αTSLsThe accumulation ability of SIA-αTSLs in the tumor site and the MRI/NIRFI imaging ability were investigated,and the SIA-αTSLs photothermal conversion ability in mice was also investigated.4.Study on antitumor efficacy and safety for SIA-αTSLsThe cytotoxicity test,apoptosis test,cell cycle arrest test and in vitro immune effect evaluation were carried out at the cell level in vitro.The results showed that photothermal therapy had significant cytotoxicity,and CT26 cells all exhibited dose-dependent cytotoxicity to Abemaciclib.At the same time,in promoting apoptosis and inhibiting cell cycle,photothermotherapy showed the best effect of promoting apoptosis,and the apoptosis rate of SIA-αTSLs+L group increased to 40.50 ± 5.67%.The group containing Abemaciclib effectively inhibited the proliferation of tumor cells.In immunological evaluation,SIA-αTSLs could reduce the proportion of Treg in tumor tissue,and promote the infiltration of CD4+ and CD8+T cells,increase the levels of immune activating factors IFN-γ and IL-2,and reduce the immunosuppressive factors IL-4,IL-10 and IL-2.The level of TGF-β triggers an effective anti-tumor immune response.Meanwhile,in CT-26 tumor-bearing mice,SIA-αTSLs combined with GM-CSF exhibited excellent antitumor efficiency and induced long-term immune memory.The main organs of the mice with good preliminary safety of the preparation did not have obvious lesions.Collectively,based on MRI/NIRFI imaging-guided photo-induce triple therapy strategy,thermosensitive theranostics liposomes co-loaded with IR780,SPIONs and Abemaciclib(SIA-αTSLs)was constructed for precise localization of tumors.SIA-αTSLs showed a good dual-mode localization-guided tumor treatment effect in CT26 tumor-bearing mice,which provided a good reference for the realization of precise localization and treatment of tumors,and brought new prospects for the development of diagnostic preparations.