Boosting synergism of chemo- and immuno-therapies via switching paclitaxel-induced apoptosis to mevalonate metabolism-triggered ferroptosis by bisphosphonate coordination lipid nanogranules.

Conventional chemotherapy based on cytotoxic drugs is facing tough challenges recently following the advances of monoclonal antibodies and molecularly targeted drugs. It is critical to inspire new potential to remodel the value of this classical therapeutic strategy. Here, we fabricate bisphosphonate coordination lipid nanogranules (BC-LNPs) and load paclitaxel (PTX) to boost the chemo- and immuno-therapeutic synergism of cytotoxic drugs. Alendronate in BC-LNPs@PTX, a bisphosphonate to block mevalonate metabolism, works as both the structure and drug constituent in nanogranules, where alendronate coordinated with calcium ions to form the particle core. The synergy of alendronate enhances the efficacy of paclitaxel, suppresses tumor metastasis, and alters the cytotoxic mechanism. Differing from the paclitaxel-induced apoptosis, the involvement of alendronate inhibits the mevalonate metabolism, changes the mitochondrial morphology, disturbs the redox homeostasis, and causes the accumulation of mitochondrial ROS and lethal lipid peroxides (LPO). These factors finally trigger the ferroptosis of tumor cells, an immunogenic cell death mode, which remodels the suppressive tumor immune microenvironment and synergizes with immunotherapy. Therefore, by switching paclitaxel-induced apoptosis to mevalonate metabolism-triggered ferroptosis, BC-LNPs@PTX provides new insight into the development of cytotoxic drugs and highlights the potential of metabolism regulation in cancer therapy.

Remodeling of intestinal epithelium derived extracellular vesicles by nanoparticles and its bioeffect on tumor cell migration.

Extracellular vesicles (EVs) are an effective tool to elucidate the bioeffect of nanomedicines. To clarify the interaction between oral nanomedicines and intestinal epithelial cells, and their bioeffects on downstream cells, polystyrene nanoparticles (PS-NPs) with different sizes were used as the model nanomedicines for EVs induction. Caco-2 monolayers were selected as the model of the intestinal epithelium and DLD-1 cells as the colorectal cancer model proximal to the gastrointestinal tract. It is found that compared with small-sized (25, 50, 100 nm) PS-NPs, the large-sized (200 and 500 nm) exhibited higher co-localization with multivesicular bodies and lysosomes, and more significant reduction of lysosomal acidification in Caco-2 cells. Proteomic and western-blotting analysis showed that the EVs remodeled by large-sized PS-NPs exhibited a higher extent of protein expression changes. The in vitro and in vivo signaling pathway detection in DLD-1 cells and DLD-1 cell xenograft nude mice showed that the remodeled EVs by large-sized PS-NPs inhibited the activation of multiple signaling pathways including Notch3, EGF/EGFR, and PI3K/Akt pathways, which resulted in the inhibition of tumor cell migration. These results primarily clarify the regulation mechanisms of nanomedicines-EVs-receptor cells chain. It provides a new perspective for the rational design and bioeffect evaluation of oral drug nanomaterials and sets up the fundamental knowledge for novel tumor therapeutics in the future.

A Trinity Nano-Vaccine System with Spatiotemporal Immune Effect for the Adjuvant Cancer Therapy after Radiofrequency Ablation.

Cancer vaccine gains great attention with the advances in tumor immunology and nanotechnology, but its long-term efficacy is restricted by the unsustainable immune activity after vaccination. Here, we demonstrate the vaccine efficacy is negatively correlated with the tumor burden. To maximum the vaccine-induced immunity and prolong the time-effectiveness, we design a priming-boosting vaccination strategy by combining with radiofrequency ablation (RFA), and construct a bisphosphonate nanovaccine (BNV) system. BNV system consists of nanoparticulated bisphosphonates with dual electric potentials (BNV(+&-)), where bisphosphonates act as the immune adjuvant by blocking mevalonate metabolism. BNV(+&-) exhibits the spatial and temporal heterogeneity in lymphatic delivery and immune activity. As the independent components of BNV(+&-), BNV(-) is drained to the lymph nodes, and BNV(+) is retained at the injection site. The alternately induced immune responses extend the time-effectiveness of antitumor immunity and suppress the recurrence and metastasis of colorectal cancer liver metastases after RFA. As a result, this trinity system integrated with RFA therapy, bisphosphonate adjuvant, and spatiotemporal immune effect provides an orientation for the sustainable regulation and precise delivery of cancer vaccines.

Oral administration of nanoformulated indoximod ameliorates ulcerative colitis by promoting mitochondrial function and mucosal healing.

Ulcerative colitis (UC) is a chronic relapsing inflammatory bowel disease with serious mucosal inflammation mainly in the colon and rectum. Currently, there is no effective therapeutics for UC. Indoximod (IND) is a water-insoluble inhibitor for indolamine 2, 3-dioxygenase (IDO) and has been mainly reported in cancer therapy. Here, we prepared orally administrated IND nanoparticles (IND-NPs) for UC treatment and investigated their functions and mechanisms in cellular and animal inflammatory models. Confocal imaging demonstrated that IND-NPs maintained the expression level of ZO-1, Occludin and E-cadherin, thereby stabilizing of intercellular junction in Caco-2 cells. It was found that IND-NPs could lower the ROS level and increase mitochondrial membrane potential as well as ATP level, indicating that IND-NPs could restore DSS-induced mitochondrial dysfunction. In the mice model with DSS-induced colitis, IND-NPs were found to alleviate UC-associated symptoms, inhibit inflammatory response, and improve the integrity of epithelial barrier. The untargeted metabolomics analysis validated that IND-NPs also contributed to regulate the metabolite levels to normal. As an agonist of aryl hydrocarbon receptor (AhR), IND-NPs might repair mucosa via the AhR pathway. These findings demonstrated that IND-NPs prominently ameliorated DSS-induced colonic injury and inflammation and preserved intestinal barrier integrity, showing a promising potential in UC treatment.

An ameliorated anti-hTNF-α therapy for arthritis via carrier-free macromolecular nanoparticles consisted of infliximab.

Infliximab (INF) is intravenously used for the clinical treatment of rheumatoid arthritis. However, it can cause serious side effects, which are mainly associated with systemic exposure and high doses. Here, we developed a modified hydrophobic ion-pairing complexes (INF HIPC) through the sequential introduction of bovine lactoferrin (BLF) and hyaluronic acid (HA) with opposite charges into the INF solution. INF and BLF were found to be not only integrally responsible for the structural integrity of HIPC but also were determined to have respective biological activities by binding human tumor necrosis factor-alpha (hTNF-α) or promoting the proliferation of osteoblasts. The INF HIPC had good stability, high drug-loading efficiency, and long-term retention effects. Whether via knee joint injection or intravenous injection, INF HIPC resulted in lower hTNF-α levels and less cartilage destruction than INFs in the transgenic mouse model. At the same time, INF HIPC could reduce toxicity based on body weight changes in transgenic mice. Our findings provide a simple and promising avenue to develop advanced delivery systems for other antibodies and macromolecules.

An enzyme-responsive and transformable PD-L1 blocking peptide-photosensitizer conjugate enables efficient photothermal immunotherapy for breast cancer.

Mild photothermal therapy combined with immune checkpoint blockade has received increasing attention for the treatment of advanced or metastatic cancers due to its good therapeutic efficacy. However, it remains a challenge to facilely integrate the two therapies and make it potential for clinical translation. This work designed a peptide-photosensitizer conjugate (PPC), which consisted of a PD-L1 antagonist peptide (CVRARTR), an MMP-2 specific cleavable sequence, a self-assembling motif, and the photosensitizer Purpurin 18. The single-component PPC can self-assemble into nanospheres which is suitable for intravenous injection. The PPC nanosphere is cleaved by MMP-2 when it accumulates in tumor sites, thereby initiating the cancer-specific release of the antagonist peptide. Simultaneously, the nanospheres gradually transform into co-assembled nanofibers, which promotes the retention of the remaining parts within the tumor. In vivo studies demonstrated that PPC nanospheres under laser irradiation promote the infiltration of cytotoxic T lymphocytes and maturation of DCs, which sensitize 4T1 tumor cells to immune checkpoint blockade therapy. Therefore, PPC nanospheres inhibit tumor growth efficiently both in situ and distally and blocked the formation of lung metastases. The present study provides a simple and efficient integrated strategy for breast cancer photoimmunotherapy.

Optimize the combination regimen of Trastuzumab and Nab-paclitaxel in HER2-positive tumors via modulating Caveolin-1 expression by lovastatin.

The combination regimen of trastuzumab (Tras) plus Nab-paclitaxel (Nab) is recommended to treat HER2-positive (HER2+) cancers. However, they exert effects in different mechanisms: Tras need to stay on cell membranes, while Nab need to be endocytosed, therefore the concurrent combination regimen may not be the best one in HER2+ tumors treatment. Caveolin-1 (Cav-1) is a key player in mediating their endocytosis and is associated with their efficacy, but few researches noticed the opposite effect of Cav-1 expression on the combination efficacy. Herein, we systematically studied the Cav-1 expression level on the combination efficacy and proposed an optimized and clinically feasible combination regimen for HER2+ Cav-1High tumor treatment. In the regimen, lovastatin (Lova) was introduced to modulate the Cav-1 expression and the results indicated that Lova could downregulate Cav-1 expression, increase Tras retention on cell membrane and enhance the in vitro cytotoxicity of Tras in HER2+ Cav-1High cells but not in HER2+ Cav-1Low cells. Therefore, by exchanging the dosing sequence of Nab and Tras, and by adding Lova at appropriate time points, the precise three-drug-sequential regimen (PTDS, Nab(D1)-Lova(D2)-Lova & Tras(D2+12 h)) was established. Compared with the concurrent regimen, the PTDS regimen exhibited a higher in vitro cytotoxicity and a stronger tumor growth inhibition in HER2+ Cav-1High tumors, which might be a promising combination regimen for these patients in clinics.

Nanoparticulates reduce tumor cell migration through affinity interactions with extracellular migrasomes and retraction fibers.

Nano-tumor interactions are fundamental for cancer nanotherapy, and the cross-talk of nanomedicines with the extracellular matrix (ECM) is increasingly considered essential. Here, we specifically investigate the nano-ECM interactivity using drug-free nanoparticulates (NPs) and highly metastatic cancer cells as models. We discover with surprise that NPs closely bind to specific types of ECM components, namely, retraction fibers (RFs) and migrasomes, which are located at the rear of tumor cells during their migration. This interaction is observed to alter cell morphology, limit cell motion range and change cell adhesion. Importantly, NPs are demonstrated to inhibit tumor cell removal in vitro, and their anti-metastasis potential is preliminarily confirmed in vivo. Mechanically, the NPs are found to coat and form a rigid shell on the surface of migrasomes and retraction fibers via interaction with lipid raft/caveolae substructures. In this way, NPs block the recognition, endocytosis and elimination of migrasomes by their surrounding tumor cells. Thereby, NPs interfere with the cell-ECM interaction and reduce the promotion effect of migrasomes on cell movement. Additionally, NPs trigger alteration of the expression of proteins related to cell-cell adhesion and cytoskeleton organization, which also restricts cell migration. In summary, all the findings here provide a potential target for anti-tumor metastasis nanomedicines.

One-pot preparation of nanodispersion with readily available components for localized tumor photothermal and photodynamic therapy.

Photothermal (PTT) and photodynamic (PDT) combined therapy has been hindered to clinical translation, due to the lack of available biomaterials, difficult designs of functions, and complex chemical synthetic or preparation procedures. To actualize a high-efficiency combination therapy for cancer via a feasible approach, three readily available materials are simply associated together in one-pot, namely the single-walled carbon nanohorns (SWCNH), zinc phthalocyanine (ZnPc), and surfactant TPGS. The established nanodispersion is recorded as PCT. The association of SWCNH/ZnPc/TPGS was confirmed by energy dispersive spectrum, Raman spectrum and thermogravimetric analysis. Under lighting, PCT induced a temperature rising up to about 60 °C due to the presence of SWCNH, production a 7-folds of singlet oxygen level elevation because of ZnPc, which destroyed almost all 4T1 tumor cells in vitro. The photothermal effect of PCT depended on both laser intensity and nanodispersion concentration in a linear and nonlinear manner, respectively. After a single peritumoral injection in mice and laser treatment, PCT exhibited the highest tumor temperature rise (to 65 °C) among all test groups, completely destroyed primary tumor without obvious toxicity, and inhibited distant site tumor. Generally, this study demonstrated the high potential of PCT nanodispersion in tumor combined therapy.

A common strategy to improve transmembrane transport in polarized epithelial cells based on sorting signals: Guiding nanocarriers to TGN rather than to the basolateral plasma membrane directly.

The intestinal barrier has always been the rate-limiting step in the oral administration process. To overcome the intestinal barrier, researchers have widely adopted nanocarriers, especially active-targeting nanocarriers strategies. However, most of these strategies focus on the ligand decoration of nanocarriers targeting specific receptors, so their applications are confined to specific receptors or specific cell types. In this study, we tried to investigate more common strategies in the field of transmembrane transport enhancement. Trans-Golgi network (TGN) is the sorting center of biosynthetic route which could achieve polarized localization of proteins in polarized epithelial cells, and the basolateral plasma membrane is where all transcytotic cargos have to pass through. Thus, it is expected that guiding nanocarriers to TGN or basolateral plasma membrane may improve the transcytosis. Hence, we choose sorting signal peptide to modify micelles to guide micelles to TGN (named as BAC decorated micelles, BAC-M) or to basolateral plasma membrane (named as STX decorated micelles, STX-M). By incorporating coumarin-6 (C6) or Cy5-PEG-PCL in the micelles to indicate the behavior of micelles, the effects of these two strategies on the transcytosis were investigated. To our surprise, BAC-M and STX-M behaved quite differently when crossing biological barriers. BAC-M showed significant superiority in colocalization with TGN, transmembrane transport and even in vivo absorption, while STX-M had no significant difference from blank micelles. Further investigation revealed that the strategy of directly guiding nanocarriers to the basolateral plasma membrane (STX-M) only caused the stack of vesicles near the basolateral plasma membrane. So, we concluded that guiding nanocarriers to TGN which related to secretion may contribute to the transmembrane transport. This common strategy based on the physiological function of TGN in polarized epithelial cells will have broad application prospects in overcoming biological barrier.

The role of caveolin-1 in the biofate and efficacy of anti-tumor drugs and their nano-drug delivery systems.

As one of the most important components of caveolae, caveolin-1 is involved in caveolae-mediated endocytosis and transcytosis pathways, and also plays a role in regulating the cell membrane cholesterol homeostasis and mediating signal transduction. In recent years, the relationship between the expression level of caveolin-1 in the tumor microenvironment and the prognostic effect of tumor treatment and drug treatment resistance has also been widely explored. In addition, the interplay between caveolin-1 and nano-drugs is bidirectional. Caveolin-1 could determine the intracellular biofate of specific nano-drugs, preventing from lysosomal degradation, and facilitate them penetrate into deeper site of tumors by transcytosis; while some nanocarriers could also affect caveolin-1 levels in tumor cells, thereby changing certain biophysical function of cells. This article reviews the role of caveolin-1 in tumor prognosis, chemotherapeutic drug resistance, antibody drug sensitivity, and nano-drug delivery, providing a reference for the further application of caveolin-1 in nano-drug delivery systems.

A biomimetic antitumor nanovaccine based on biocompatible calcium pyrophosphate and tumor cell membrane antigens.

Currently, the cancer immunotherapy has made great progress while antitumor vaccine attracts substantial attention. Still, the selection of adjuvants as well as antigens are always the most crucial issues for better vaccination. In this study, we proposed a biomimetic antitumor nanovaccine based on biocompatible nanocarriers and tumor cell membrane antigens. Briefly, endogenous calcium pyrophosphate nanogranules with possible immune potentiating effect are designed and engineered, both as delivery vehicles and adjuvants. Then, these nanocarriers are coated with lipids and B16-OVA tumor cell membranes, so the biomembrane proteins can serve as tumor-specific antigens. It was found that calcium pyrophosphate nanogranules themselves were compatible and possessed adjuvant effect, while membrane proteins including tumor associated antigen were transferred onto the nanocarriers. It was demonstrated that such a biomimetic nanovaccine could be well endocytosed by dendritic cells, promote their maturation and antigen-presentation, facilitate lymph retention, and trigger obvious immune response. It was confirmed that the biomimetic vaccine could induce strong T-cell response, exhibit excellent tumor therapy and prophylactic effects, and simultaneously possess nice biocompatibility. In general, the present investigation might provide insights for the further design and application of antitumor vaccines.

Dual-targeting nanovesicles enhance specificity to dynamic tumor cells in vitro and in vivo via manipulation of αvß3-ligand binding.

The dynamic or flowing tumor cells just as leukemia cells and circulating tumor cells face a microenvironment difference from the solid tumors, and the related targeting nanomedicines are rarely reported. The existence of fluidic shear stress in blood circulation seems not favorable for the binding of ligand modified nanodrugs with their target receptor. Namely, the binding feature is very essential in this case. Herein, we utilized HSPC, PEG-DSPE, cholesterol and two αvß3 ligands (RGDm7 and DT4) with different binding rates to build dual-targeting nanovesicles, in an effort to achieve a "fast-binding/slow-unbinding" function. It was demonstrated that the dual-targeting nanovesicles actualized efficient cellular uptake and antitumor effect in vitro both for static and dynamic tumor cells. Besides, the potency of the dual-targeting vesicles for flowing tumor cells was better than that for static tumor cells. Then, a tumor metastasis mice model and a leukemia mice model were established to detect the killing ability of the drug-loaded dual-targeting vesicles to dynamic tumor cells in vivo. The therapy efficacy of the dual-targeting system was higher than other controls including single-targeting ones. Generally, it seems possible to strengthen drug-targeting to dynamic tumor cells via the control of ligand-receptor interaction.

A combined "eat me/don't eat me" strategy based on extracellular vesicles for anticancer nanomedicine.

A long-term and huge challenge in nanomedicine is the substantial uptake and rapid clearance mediated by the mononuclear phagocyte system (MPS), which enormously hinders the development of nanodrugs. Inspired by the natural merits of extracellular vesicles, we therefore developed a combined "eat me/don't eat me" strategy in an effort to achieve MPS escape and efficient drug delivery. Methodologically, cationized mannan-modified extracellular vesicles derived from DC2.4 cells were administered to saturate the MPS (eat me strategy). Then, nanocarriers fused to CD47-enriched exosomes originated from human serum were administered to evade phagocytosis by MPS (don't eat me strategy). The nanocarriers were also loaded with antitumor drugs and functionalized with a novel homing peptide to promote the tumour tissue accumulation and cancer cell uptake (eat me strategy). The concept was proven in vitro as evidenced by the reduced endocytosis of macrophages and enhanced uptake by tumour cells, whereas prolonged circulation time and increased tumour accumulation were demonstrated in vivo. Specially, the strategy induced a 123.53% increase in tumour distribution compared to conventional nanocarrier. The study both shed light on the challenge overcoming of phagocytic evasion and provided a strategy for significantly improving therapeutic outcomes, potentially permitting active drug delivery via targeted nanomedicines.

A magnetism/laser-auxiliary cascaded drug delivery to pulmonary carcinoma.

Although high-efficiency targeted delivery is investigated for years, the efficiency of tumor targeting seems still a hard core to smash. To overcome this problem, we design a three-step delivery strategy based on streptavidin-biotin interaction with the help of c(RGDfK), magnetic fields and lasers. The ultrasmall superparamagnetic iron oxide nanoparticles (USIONPs) modified with c(RGDfK) and biotin are delivered at step 1, followed by streptavidin and the doxorubicin (Dox) loaded nanosystems conjugated with biotin at steps 2 and 3, respectively. The delivery systems were proved to be efficient on A549 cells. The co-localization of signal for each step revealed the targeting mechanism. The external magnetic field could further amplify the endocytosis of USPIONs based on c(RGDfK), and magnify the uptake distinctions among different test groups. Based on photoacoustic imaging, laser-heating treatment could enhance the permeability of tumor venous blood vessels and change the insufficient blood flow in cancer. Then, it was noticed in vivo that only three-step delivery with laser-heating and magnetic fields realized the highest tumor distribution of nanosystem. Finally, the magnetism/laser-auxiliary cascaded delivery exhibited the best antitumor efficacy. Generally, this study demonstrated the necessity of combining physical, biological and chemical means of targeting.

Regulating Interactions Between Targeted Nanocarriers and Mononuclear Phagocyte System via an Esomeprazole-Based Preconditioning Strategy.

PURPOSE: The mononuclear phagocyte system (MPS) presents a formidable obstacle that hampers the delivery of various nanopreparations to tumors. Therefore, there is an urgent need to improve the off-MPS targeting ability of nanomedicines. In the present study, we present a novel preconditioning strategy to substantially increase the circulation times and tumor targeting of nanoparticles by regulating nanocarrier-MPS interactions. METHODS: In vitro, the effect of different vacuolar H+-ATPase inhibitors on macrophage uptake of targeted or nontargeted lipid vesicles was evaluated. Specifically, the clinically approved proton-pump inhibitor esomeprazole (ESO) was selected as a preconditioning agent. Then, we further investigated the blocking effect of ESO on the macrophage endocytosis of nanocarriers. In vivo, ESO was first intravenously administered into A549-tumor-bearing nude mice, and 24 h later, the c(RGDm7)-modified vesicles co-loaded with doxorubicin and gefitinib were intravenously injected. RESULTS: In vitro, ESO was found to reduce the interactions between macrophages and c(RGDm7)-modified vesicles by interfering with the latter's lysosomal trafficking. Studies conducted in vivo confirmed that ESO pretreatment greatly decreased the liver and spleen distribution of the targeted vesicles, enhanced their tumor accumulation, and improved the therapeutic outcome of the drug-loaded nanomedicines. CONCLUSION: Our findings indicate that ESO can regulate the nanoparticle-MPS interaction, which provides a feasible option for enhancing the off-MPS targeting of nanomedicines.

Tumour sensitization via the extended intratumoural release of a STING agonist and camptothecin from a self-assembled hydrogel.

Tumours with an immunosuppressive microenvironment respond poorly to therapy. Activation of the stimulator of interferon genes (STING) pathway can enhance intratumoural immune activation, but STING agonists are associated with high toxicity and degrade prematurely, which limits their effectiveness. Here, we show that the extended intratumoural release of the STING agonist cyclic di-AMP transforms the tumour microenvironment from immunosuppressive to immunostimulatory, increasing the efficacy of antitumour therapies. The STING agonist was electrostatically complexed with nanotubes comprising a peptide-drug conjugate (a peptide that binds to the protein neuropilin-1, which is highly expressed in tumours, and the chemotherapeutic agent camptothecin) that self-assemble in situ into a supramolecular hydrogel. In multiple mouse models of murine tumours, a single low dose of the STING agonist led to tumour regression and increased animal survival, and to long-term immunological memory and systemic immune surveillance, which protected the mice against tumour recurrence and the formation of metastases. Locally delivered STING agonists could help to reduce tumour immunosuppression and enhance the efficacy of a wide range of cancer therapies.

Supramolecular prodrug hydrogelator as an immune booster for checkpoint blocker-based immunotherapy.

Immune checkpoint blockers (ICBs) have shown great promise at harnessing immune system to combat cancer. However, only a fraction of patients can directly benefit from the anti-programmed cell death protein 1 (aPD1) therapy, and the treatment often leads to immune-related adverse effects. In this context, we developed a prodrug hydrogelator for local delivery of ICBs to boost the host's immune system against tumor. We found that this carrier-free therapeutic system can serve as a reservoir for extended tumoral release of camptothecin and aPD1 antibody, resulting in an immune-stimulating tumor microenvironment for boosted PD-1 blockade immune response. Our in vivo results revealed that this combination chemoimmunotherapy elicits robust and durable systemic anticancer immunity, inducing tumor regression and inhibiting tumor recurrence and metastasis. This work sheds important light into the use of small-molecule prodrugs as both chemotherapeutic and carrier to awaken and enhance antitumor immune system for improved ICBs therapy.

Anisotropic active ligandations in siRNA-Loaded hybrid nanodiscs lead to distinct carcinostatic outcomes by regulating nano-bio interactions.

Active targeting modification is one of the foremost nanomedicine strategies for the efficacy improvement. Compared to the homogeneous ligandation on spherical nanocarriers, non-spherical nanomedicines usually make the ligand modification more complicated. The modified ligands always exhibit anisotropy and heterogeneity. However, there is very little systematic study on these diversified anisotropic modifications. The efficacy difference and underlying mechanism were still unclear. Here, we separately fabricated hybrid nanodiscs (NDs) conjugated with cRGD on the edge and plane surfaces to engineer two anisotropic targeting nanocarriers (E-cRGD-NDs and P-cRGD-NDs, respectively) for gene delivery. The ligand anisotropy endowed NDs with diversified cellular interactions, and caused different efficacies between E-cRGD-NDs and P-cRGD-NDs. Of note, E-cRGD-NDs showed significant superiority in siRNA loading, cellular uptake, silence efficiency, protein expression and even in vivo efficacy. The mechanism investigation revealed the functional anisotropy specifically for E-cRGD-NDs. The edge modification of cRGD efficiently separated the targeting and siRNA loading domains, maximizing their respective functions. These findings reflected the unique effect of ligand anisotropy, also provided a new strategy for the targeting screening of extensive nanomedicines.