Tumor-targeted self-assembled micelles reducing PD-L1 expression combined with ICIs to enhance chemo-immunotherapy of TNBC

Immune checkpoint inhibitors(ICIs)therapy targeting programmed cell death ligand 1(PD-L1)and programmed death protein 1(PD-1)had exhibited significant clinical benefits for cancer treatment such as triple negative breast cancer(TNBC).However,the relatively low anti-tumor immune response rate and ICIs drug resistance highlight the necessity of developing ICIs combination therapy strategies to improve the anti-tumor effect of immunotherapy.Herein,the immunomodulator epigallocatechin gallate palmitate(PEGCG)and the immunoadjuvant metformin(MET)self-assembled into tumor-targeted micelles via hydrogen bond and electrostatic interaction,which encapsulated the therapeutic agents doxorubicin(DOX)-loaded PEGCG-MET micelles(PMD)and combined with ICIs(anti-PD-1 antibody)as therapeutic strategy to reduce the endogenous expression of PD-L1 and improve the tumor immunosuppressive microenvironment.The results presented that PMD integrated chemotherapy and immunotherapy to enhance antitumor efficacy in vitro and in vivo,compared with DOX or anti-PD-1 antibody for the therapy of TNBC.PMD micelles might be a potential candidate,which could remedy the shortcomings of antibody-based ICIs and provide synergistic effect to enhance the antitumor effects of ICIs in tumor therapy.

Advances in aggregatable nanoparticles for tumor-targeted drug delivery

Recent days,aggregatable nanoparticles,which can specifically respond to certain stimulus,have shown great potential in tumor-targeted drug delivery with prolonged retention and deeper penetration.In this review,we summarize recent advances in design of aggregatable nanoparticles by different stimuli.Internal(pH and enzyme)and external(light,temperature and ROS)stimuli are introduced for a comprehensive description.Moreover,the aggregated nanoparticles usually exhibit photothermal,photoacoustic,PET and enhanced MRI contrast,which is also described.In the end,we discuss about the potential applications and challenges for the future clinical translation.

Macrophage-mediated tumor-targeted delivery of engineered Salmonella typhimurium VNP20009 in anti-PD1 therapy against melanoma

Bacterial antitumor therapy has great application potential given its unique characteristics,including genetic manipulation, tumor targeting specificity and immune system modulation. However,the nonnegligible side effects and limited efficacy of clinical treatment limit their biomedical applications. Engineered bacteria for therapeutic applications ideally need to avoid their accumulation in normal organs and possess potent antitumor activity. Here, we show that macrophage-mediated tumor-targeted delivery of Salmonella typhimurium VNP20009 can effectively reduce the toxicity caused by administrating VNP20009 alone in a melanoma mouse model. This benefits from tumor-induced chemotaxis for macrophages combined with their slow release of loaded strains. Inspired by changes in the tumor microenvironment, including a decrease in intratumoral dysfunctional CD8+T cells and an increase in PDL1 on the tumor cell surface, macrophages were loaded with the engineered strain VNP-PD1nb, which can express and secrete anti-PD1 nanoantibodies after they are released from macrophages. This novel triple-combined immunotherapy significantly inhibited melanoma tumors by reactivating the tumor microenvironment by increasing immune cell infiltration, inhibiting tumor cell proliferation, remodeling TAMs to an M1-like phenotype and prominently activating CD8+T cells. These data suggest that novel combination immunotherapy is expected to be a breakthrough relative to single immunotherapy.

Lipoprotein-biomimetic nanostructure enables tumor-targeted penetration delivery for enhanced photo-gene therapy towards glioma

Glioma is one of the most malignant primary tumors affecting the brain.The efficacy of therapeutics for glioma is seriously compromised by the restriction of blood-brain barrier(BBB),interstitial tumor pressure of resistance to chemotherapy/radiation,and the inevitable damage to normal brain tissues.Inspired by the natural structure and properties of high-density lipoprotein(HDL),a tumor-penetrating lipoprotein was prepared by the fusion tLyP-1 to apolipoprotein A-I-mimicking peptides(D4F),together with indocyanine green(ICG)incorporation and lipophilic small interfering RNA targeted HIF-1α(siHIF)surface anchor for site-specific photo-gene therapy.tLyP-1 peptide is fused to HDL-surface to facilitate BBB permeability,tumor-homing capacity and-site accumulation of photosensitizer and siRNA.Upon NIR light irradiation,ICG not only served as real-time targeted imaging agent,but also provided toxic reactive oxygen species and local hyperthermia for glioma phototherapy.The HIF-1αsiRNA in this nanoplatform downregulated the hypoxia-induced HIF-1αlevel in tumor microenvironment and enhanced the photodynamic therapy against glioma.These studies demonstrated that the nanoparticles could not only efficiently across BBB and carry the payloads to orthotopic glioma,but also modulate tumor microenvironment,thereby inhibiting tumor growth with biosafety.Overall,this study develops a new multifunctional drug delivery system for glioma theranostic,providing deeper insights into orthotopic brain tumor imaging and treatment.

Molecular regulation of vasculogenic mimicry in tumors and potential tumor-target therapy

"Vasculogenic mimicry(VM)",is a term that describes the unique ability of highly aggressive tumor cells to express a multipotent,stem cell-like phenotype,and form a pattern of vasculogenic-like networks in threedimensional culture.As an angiogenesis-independent pathway,VM and/or periodic acid-schiff-positive patterns are associated with poor prognosis in tumor patients.Moreover,VM is resistant to angiogenesis inhibitors.Here,we will review the advances in research on biochemical and molecular signaling pathways of VM in tumors and on potential anti-VM therapy strategy.

Redox dual-stimuli responsive drug delivery systems for improving tumor-targeting ability and reducing adverse side effects

Cancer is a big challenge that has plagued the human beings for ages and one of the most effective treatments is chemotherapy. However, the low tumor-targeting ability limits the wide clinical application of chemotherapy. The microenvironment plays a critical role in many aspects of tumor genesis. It generates the tumor vasculature and it is highly implicated in the progression to metastasis. To maintain a suitable environment for tumor progression, there are special microenvironment in tumor cell, such as low pH, high level of glutathione(GSH) and reactive oxygen species(ROS), and more special enzymes, which is different to normal cell. Microenvironment-targeted therapy strategy could create new opportunities for therapeutic targeting. Compared to other targeting strategies, microenvironment-targeted therapy strategy will control the drug release into tumor cells more accurately. Redox responsive drug delivery systems(DDSs) are developed based on the high level of GSH in tumor cells. However, there are also GSH in normal cell though its level is lower. In order to control the release of drugs more accurately and reduce side effects, other drug release stimuli have been introduced to redox responsive DDSs. Under the synergistic reaction of two stimuli, redox dual-stimuli responsive DDSs will control the release of drugs more accurately and quickly and even increase the accumulation. This review summarizes strategies of redox dual-stimuli responsive DDSs such as pH, light, enzyme, ROS, and magnetic guide to delivery chemotherapeutic agents more accurately, aiming at providing new ideas for further promoting the drug release,enhancing tumor-targeting and improving anticancer effects. To better illustrate the redox dual-stimuli responsive DDS, preparations of carriers are also briefly described in the review.

Amino acid transporters: Emerging roles in drug delivery for tumor-targeting therapy

Amino acid transporters,which play a vital role in transporting amino acids for the biosynthesis of mammalian cells,are highly expressed in types of tumors.Increasing studies have shown the feasibility of amino acid transporters as a component of tumortargeting therapy.In this review,we focus on tumor-related amino acid transporters and their potential use in tumor-targeting therapy.Firstly,the expression characteristics of amino acid transporters in cancer and their relationship with tumor growth are reviewed.Secondly,the recognition requirements are discussed,focusing on the“acidbase”properties,conformational isomerism and structural analogues.Finally,recent developments in amino acid transporter-targeting drug delivery strategies are highlighted,including prodrugs and nanocarriers,with special attention to the latest findings of molecular mechanisms and targeting efficiency of transporter-mediated endocytosis.We aim to offer related clues that might lead to valuable tumor-targeting strategies by the utilization of amino acid transporters.

Folic Acid-Polyethyleneimine-Ethosome as a Potential Tumor Targeting Carrier: Preparation, Characterization and in vitro Evaluation

A novel composite carrier of folic acid(FA)-polyethyleneimine(PEI)-ethosome(Eth)(FA-PEI-Eth)was developed for the treatment of cancers through loading and targeting delivery of multidrug(including gene and other drugs)into cancer cells.Physical and chemical property tests were done to prove the grafting of the composite.Gel retardation test was done to determine the optimal ratio of DNA@PEI complex,and cytocompatibility tests and tumor cell uptake tests were done to evaluate the efficiency of the composite.The results demonstrated that the FA-PEI-Eth could effectively deliver a gene and other drugs into tumor cells simultaneously,and suggested that this composite would be a promising carrier in tumor-targeted therapy applications.

A multi-functional nanoplatform for efficacy tumor theranostic applications

Nanomaterials with multiple functions have become more and more popular in the domain of cancer research. MoS2 has a great potential in photothermal therapy, X-ray/CT imaging and drug delivery. In this study, a water soluble MoS2 nanosystem(MoS2-PEG) was synthesized and explored in drug delivery, photothermal therapy(PTT) and X-ray imaging.Doxorubicin(DOX) was loaded onto MoS2-PEG with a high drug loading efficiency(~69%)and obtained a multifunctional drug delivery system(MoS2-PEG/DOX). As the drug delivery, MoS2-PEG/DOX could efficiently cross the cell membranes, and escape from the endosome via NIR light irradiation, lead to more apoptosis in MCF-7 cells, and afford higher antitumor efficacy without obvious toxic effects to normal organs owing to its prolonged blood circulation and 11.6-fold higher DTX uptake of tumor than DOX. Besides, MoS2-PEG/DOX not only served as a drug delivery system, but also as a powerful PTT agent for thermal ablation of tumor and a strong X-ray contrast agent for tumor diagnosis. In the in vitro and in vivo studies, MoS2-PEG/DOX exhibited excellent tumor-targeting efficacy, outstanding synergistic anti-cancer effect of photothermal and chemotherapy and X-ray imaging property,demonstrating that MoS2-PEG/DOX had a great potential for simultaneous diagnosis and photothermal-chemotherapy in cancer treatment.

Photo-induced crosslinked and anti-PD-L1 peptide incorporated liposomes to promote PD-L1 multivalent binding for effective immune checkpoint blockade therapy

Immune checkpoint blockade(ICB)therapy targeting PD-L1 via monoclonal antibody(m Ab)has shown extensive clinical benefits in the diverse types of advanced malignancies.However,most patients are completely refractory to ICB therapy owing to the PD-L1 recycling mechanism.Herein,we propose photo-induced crosslinked and anti-PD-L1 peptide incorporated liposomes(immune checkpoint blockade liposomes;ICB-LPs)to promote PD-L1 multivalent binding for inducing lysosomal degradation of PD-L1 in tumor cells.The ICB-LPs are prepared by formulation of DC_(8,9)PC with photo-polymerized diacetylenic moiety,1,2-dipalmitoylphosphatidylcholine(DPPC)and anti-PD-L1peptide(D-form NYSKPTDRQYHF)-conjugated DSPE-PEG_(2k)(anti-PD-L1-DSPE-PEG_(2k))in a molar ratio of 45:45:10,followed by cross-linking of liposomal bilayer upon UV irradiation.The 10 mol% antiPD-L1-DSPE-PEG_(2k)incorporated ICB-LPs have a nano-sized lipid bilayer structure with an average diameter of 137.7±1.04 nm,showing a high stability in serum condition.Importantly,the ICB-LPs efficiently promote the multivalent binding with PD-L1 on the tumor cell membrane,which are endocytosed with aim to deliver PD-L1 to the lysosomes,wherein the durable PD-L1 degradation is observed for72 h,in contrast to anti PD-L1 m Abs showing the rapid PD-L1 recycling within 9 h.The in vitro coculture experiments with CD8^(+)T cells show that ICB-LPs effectively enhance the T cell-mediated antitumor immune responses against tumor cells by blocking the PD-L1/PD-1 axis.When ICB-LPs are intravenously injected into colon tumor-bearing mice,they efficiently accumulate within the targeted tumor tissues via both passive and active tumor targeting,inducing a potent T cell-mediated antitumor immune response by effective and durable PD-L1 degradation.Collectively,this study demonstrates the superior antitumor efficacy of crosslinked and anti-PD-L1 peptide incorporated liposome formulation that promotes PD-L1 multivalent binding for trafficking of PD-L1 toward the lysosomes instead of the recycling endosomes.

Emerging transporter-targeted nanoparticulate drug delivery systems

Transporter-targeted nanoparticulate drug delivery systems(nano-DDS) have emerged as promising nanoplatforms for efficient drug delivery. Recently, great progress in transporter-targeted strategies has been made, especially with the rapid developments in nanotherapeutics. In this review, we outline the recent advances in transporter-targeted nano-DDS. First, the emerging transporter-targeted nano-DDS developed to facilitate oral drug delivery are reviewed. These include improvements in the oral absorption of protein and peptide drugs, facilitating the intravenous-to-oral switch in cancer chemotherapy. Secondly, the recent advances in transporter-assisted brain-targeting nano-DDS are discussed,focusing on the specific transporter-based targeting strategies. Recent developments in transportermediated tumor-targeting drug delivery are also discussed. Finally, the possible transport mechanisms involved in transporter-mediated endocytosis are highlighted, with special attention to the latest findings of the interactions between membrane transporters and nano-DDS.

Tumor microenvironment-responsive docetaxel-loaded micelle combats metastatic breast cancer

Efficient tumor-targeting drug delivery systems are urgently needed for treating metastatic breast cancer.In this work, a docetaxel(DTX)-loaded micelle(pDM) as the tumor-microenvironment-responsive delivery platform is developed. The micelle is composed of a pH-sensitive amphiphilic copolymer,poly((1,4-butanediol)-diacrylate-b-N,N-diisopropylethylenediamine)-polyethyleneimine(BD-PEI), and a matrix metalloproteinase(MMP)-responsive polymer, poly((1,4-butanediol)-diacrylate-b-N,N-diisopropy lethylenediamine)-peptide-polyethylene glycol(PEG)(BD-peptide-PEG). The PEG block of BD-peptidePEG will be split by MMPs at the tumor microenvironment, which leads to the change of the surface charge and particle size of the micelle to more positive and smaller one. Owing to this transformation and enhanced permeability and retention(EPR) effect, pDM delivers more DTX into tumor tissues and is internalized more efficiently by tumor cells than the non-MMP-sensitive micelles in the 4 T1 tumorbearing mice model. In addition, DTX is released in acidic endo/lysosomes due to the dissociation of the micelle, triggered by the protonation of the hydrophobic block of BD-PEI. As a result, the DTX-loaded micelle inhibits primary tumor growth and pulmonary metastasis effectively. Thus, this pH/MMP-dual-sensitive drug delivery system, which simultaneously attains three keypoints: prolonged circulation time, directional and efficient uptake into tumor cells, and speedy intracellular drug release, is a promising strategy for metastatic breast cancer therapy.

GLUT1-mediated effective anti-miRNA21 pompon for cancer therapy

Oncogenic microRNAs are essential components in regulating the gene expression of cancer cells. Especially miR21, which is a major player involved of tumor initiation, progression, invasion and metastasis in several cancers. The delivery of anti-miR21 sequences has significant potential for cancer treatment. Nevertheless, since anti-miR21 sequences are extremely unstable and they need to obtain certain concentration to function, it is intensely difficult to build an effective delivery system for them.The purpose of this work is to construct a self-assembled glutathione(GSH)-responsive system with tumor accumulation capacity for effective anti-miR21 delivery and cancer therapy. A novel drug delivery nanosphere carrying millions of anti-miR21 sequences was developed through the rolling circle transcription(RCT) method. GSH-responsive cationic polymer polyethyleneimine(pOEI) was synthesized to protect the nanosphere from degradation by Dicer or other RNase in normal cells and optimize the pompon-like nanoparticle to suitable size. Dehydroascorbic acid(DHA), a targeting molecule, which is a substrate of glucose transporter 1(GLUT 1) and highly expressed on malignant tumor cells, was connected to pOEI through PEG, and then the polymer was used for contracting a RNA nanospheres into nanopompons. The anti-miR21 nanopompons showed its potential for effective cancer therapy.

NIR light-induced tumor phototherapy using ICG delivery system based on platelet-membrane-camouflaged hollow bismuth selenide nanoparticles

Near-infrared(NIR)light-triggered photothermal therapy(PTT)is a promising treatment strategy for treating cancer.The combination of nanotechnology and NIR has been widely applied.However,the therapeutic efficacy of the drug-delivery system depends on their ability to avoid phagocytosis of endothelial system,cross the biological barriers,prolong circulation life,localize and rapidly release the therapeutic at target sites.In this work,we designed a platelet membrane(PM)-camouflaged hollow mesoporous bismuth selenide nanoparticles(BS NPs)loading with indocyanine green(ICG)(PM@BS-ICG NPs)to achieve the above advantages.PM-coating has active tumor-targe ting ability which could preve nt drug leakage and provide drug long circulation,causing drug delivery systems to accumulate in tumor sites effectively.Moreover,as a type of the photothermal sensitizers,BS NPs are used as the inner cores to improve ICG stability and are served as scaffolds to enhance the hardness of this drug delivery system.For one hand,the thermal vibration of BS NPs under NIR laser irradiation causes tumor inhibition through hyperthermia.For another hand,this hyperthermia process could damage PM and let ICG rapid release from PM@BS-ICG NPs.The in vitro and in vivo results showed that this biomimetic nano-drug delivery system exhibits obvious antitumor activity which has good application prospect.

Overcoming the obstacles of current photodynamic therapy in tumors using nanoparticles

Photodynamic therapy(PDT)has been applied in clinical treatment of tumors for a long time.However,insufficient supply of pivotal factors including photosensitizer(PS),light,and oxygen in tumor tissue dramatically reduces the therapeutic efficacy of PDT.Nanoparticles have received an influx of attention as drug carriers,and recent studies have demonstrated their promising potential to overcome the obstacles of PDT in tumor tissue.Physicochemical optimization for passive targeting,ligand modification for active targeting,and stimuli-responsive release achieved efficient delivery of PS to tumor tissue.Various trials using upconversion NPs,two-photon lasers,X-rays,and bioluminescence have provided clues for efficient methods of light delivery to deep tissue.Attempts have been made to overcome unfavorable tumor microenvironments via artificial oxygen generation,Fenton reaction,and combination with other chemical drugs.In this review,we introduce these creative approaches to addressing the hurdles facing PDT in tumors.In particular,the studies that have been validated in animal experiments are preferred in this review over proof-of-concept studies that were only performed in cells.

Novel fibronectin-targeted nanodisk drug delivery system displayed superior efficacy against prostate cancer compared with nanospheres

Currently,prostate cancer is the most frequently diagnosed cancer in males and chemotherapy is often essential for treating advaneed prostate cancer.However,common chemotherapies for prostate cancer suffer from serious adverse effects due to poor drug targeting ability and tissue penetration,even with the help of conventional drug delivery systems.Here,encouraged by recent studies showing possible drug retention and tissue pen etration advan tages of unconventional non-spherical nan oparticles over conventional spherical nan oparticles,we design and construct a novel non-spherical nano disk drug delivery system for treating prostate cancer.norder to enhance tumor-targeting capability,these nanodisks are further modified with targeting peptide,Cys-Arg-Glu丄ys?Ala peptide with N-methylated Glu(CR(NMe)EKA),which recognizes extracellular matrix fibronectin and its complexes specifically expressed on the walls of tumor vessels and in tumor stroma.Compared with conventional nano spheres,the nano disks achieve much higher drug accumulati on at prostate tumor sites.When loaded with paclitaxel,the CR(NMe)EKA-modified nan odisks display superior an titumor efficacy to free paclitaxel,unmodified nan odisks and nano spheres.In summary,our study provides an attractive therapeutic strategy for targeted therapy against prostate cancer with simple preparation,high efficiency and low toxicity,and supplements a theoretical support for treatments realized by different shaped nano platforms.Our study also offers valuable data for understa nding biological effects of non-spherical nano disks and highlights the great pote ntial of unconventional nan oparticles in biomedical applicati ons.

Dextran Gadolinium Complex Containing Folate Groups as a Potential Magnetic Resonance Imaging Contrast Agent

Folate-containing dextran ligand （FA-Dextran-DTPA） was synthesized by the incorporation of diethylenetriamine- pentaacetic acid （DTPA） and folate （FA） as a tumor-targeting group into dextran as a polymer carrier. This ligand was further reacted with gadolinium chloride to make a dextran gadolinium complex FA-Dextran-DTPA-Gd. The ligand and its gadolinium complex were characterized by 1H-NMR, FTIR, UV-Vis, average particle sizes and zeta potential, as well. In vitro properties including relaxivity, cytotoxicity assay, cellular uptake assay, and magnetic resonance imaging （MRI） were also evaluated. Compared with Gd-DTPA, FA-Dextran-DTPA-Gd possessed obviously higher relaxation effectiveness and lower cytotoxicity to HeLa cells. FA-Dextran-DTPA-Gd had a high affinity to the H460 and MDA-MB-231 tumor cells and can be taken up selectively by these tumor cells. Moreover, FA-Dextran-DTPA-Gd showed enhanced signal intensities （SI） of MRI and enhanced the contrast of MR images of tumor cells. These results indicated that FA-Dextran-DTPA-Gd showed the potential as a tumor-targeting contrast agent in MRI.

Co-delivery of TRAIL and paclitaxel by fibronectin-targeting liposomal nanodisk for effective lung melanoma metastasis treatment

Melanoma is a highly aggressive cancer which often forms metastatic tumors in the lung,leading to sharply reduced patients'survival rate.Effectively treating these tumors thus could improve late stage melanoma with lung metastasis.In this study,we fabricated a Cys-Arg-Glu-Lys-Ala with N-methylated Glu(CR(NMe)EKA)decorated disk shaped nano vehicle to co-deliver tumor necrosis factor-related apoptosis-inducing ligand(TRAIL)and paclitaxel(PTX)to lung melanoma tumor sites(TRAIL-[ND-PTX]^(CR(NMe)EKA)).These nanodisks displayed better tumor-targeting and penetration capability than spherical nanoparticles,while the fibronectin-targeting CR(NMe)EKA motif also increased the tumor accumulation of loaded drugs.The combined usage of TRAIL and PTX both killed tumor cells and reduced local nutrition supply,leading to stronger overall anti-tumor effect.This TRAIL-[ND-PTX]^(CR(NMe)EKA)system performed remarkably better than free paclitaxel and also significantly elongated survival rate of melanoma lung metastasis bearing mice,without displaying significant toxicity.Hence,this designing strategy and the fabricated nanoplatform possess potential for further development.

Self-delivery of a peptide-based prodrug for tumor- targeting therapy

A novel self-delivered prodrug system was fabricated for tumor-targeting therapy. In this nanosystem, the Arg-Gly-Asp-Ser （RGDS） tetrapeptide was used to improve the therapeutic index to integrin-overexpressing tumor cells. The antitumorous drug camptothecin was further appended to the ε-amino group of lysine by 20-O-succinyl linkage and controllably released via hydrolytic cleavage. Prodrug molecules self-assembled into fibrillar nano-architectures and achieved the capability of self-delivery after being injected subcutaneously into mice. Introduction of hydrophobic myristic add favored the self-assembly and enhanced the cellular internalization of the prodrugs. In vitro and in vivo studies demonstrated that the self-assembled nanofibers could effectively target integrin- overexpressing tumorous cells and inhibit tumor growth via RGD-mediated specific targeting. Therefore, the traditional idea that fibrillar structures hold low therapeutic efficacy due to poor cell uptake can be challenged.

The Preparation of Cationic Folic Acid and Its Application in Drug Delivery System

The cationic folic acid（CFA） was prepared by introducing triethylenetetramine into folic acid with EDCI/NHS and characterized by IR, NMR and mass spectra. It was found that approximately one of two carboxyls in the folic acid molecule was substituted to form CFA. The conversion of γ-carboxyl is found to be 59% higher than 30% of γ-carboxyl. The CFA and doxorubicin hydrochloride can be loaded on the ionic shell of PTX-encapsulated micelle to form CFA loaded binary drug carrier via static interaction in aqueous solutions. The successful loading was demonstrated by zeta potential measurement and the drug load amount（DLA） of CFA was measured by HPLC. In vitro cytotoxicity results revealed the CFA drug carrier showed higher cytotoxicity to cancer cell MDA-MB-321 than the binary drug carrier without CFA and the positive control, while it showed lower cytotoxicity to normal cell HUVEC than the positive control, and similar cytotoxicity with the binary drug carrier without CFA. These results as well as confocal laser scanning microscopy observation indicate the synthesized CFA drug carrier possesses active tumor-targeting property.