Gas station in blood vessels: An endothelium mimicking, self-sustainable nitric oxide fueling stent coating for prevention of thrombosis and restenosis.

Stenting is the primary treatment for vascular obstruction-related cardiovascular diseases, but it inevitably causes endothelial injury which may lead to severe thrombosis and restenosis. Maintaining nitric oxide (NO, a vasoactive mediator) production and grafting endothelial glycocalyx such as heparin (Hep) onto the surface of cardiovascular stents could effectively reconstruct the damaged endothelium. However, insufficient endogenous NO donors may impede NO catalytic generation and fail to sustain cardiovascular homeostasis. Here, a dopamine-copper (DA-Cu) network-based coating armed with NO precursor L-arginine (Arg) and Hep (DA-Cu-Arg-Hep) is prepared using an organic solvent-free dipping technique to form a nanometer-thin coating onto the cardiovascular stents. The DA-Cu network adheres tightly to the surface of stents and confers excellent NO catalytic activity in the presence of endogenous NO donors. The immobilized Arg functions as a NO fuel to generate NO via endothelial nitric oxide synthase (eNOS), while Hep works as eNOS booster to increase the level of eNOS to decompose Arg into NO, ensuring a sufficient supply of NO even when endogenous donors are insufficient. The synergistic interaction between Cu and Arg is analogous to a gas station to fuel NO production to compensate for the insufficient endogenous NO donor in vivo. Consequently, it promotes the reconstruction of natural endothelium, inhibits smooth muscle cell (SMC) migration, and suppresses cascading platelet adhesion, preventing stent thrombosis and restenosis. We anticipate that our DA-Cu-Arg-Hep coating will improve the quality of life of cardiovascular patients through improved surgical follow-up, increased safety, and decreased medication, as well as revitalize the stenting industry through durable designs.

Biomimicking design of artificial periosteum for promoting bone healing.

Background: Periosteum is a vascularized tissue membrane covering the bone surface and plays a decisive role in bone reconstruction process after fracture. Various artificial periosteum has been developed to assist the allografts or bionic bone scaffolds in accelerating bone healing. Recently, the biomimicking design of artificial periosteum has attracted increasing attention due to the recapitulation of the natural extracellular microenvironment of the periosteum and has presented unique capacity to modulate the cell fates and ultimately enhance the bone formation and improve neovascularization. Methods: A systematic literature search is performed and relevant findings in biomimicking design of artificial periosteum have been reviewed and cited. Results: We give a systematical overview of current development of biomimicking design of artificial periosteum. We first summarize the universal strategies for designing biomimicking artificial periosteum including biochemical biomimicry and biophysical biomimicry aspects. We then discuss three types of novel versatile biomimicking artificial periosteum including physical-chemical combined artificial periosteum, heterogeneous structured biomimicking periosteum, and healing phase-targeting biomimicking periosteum. Finally, we comment on the potential implications and prospects in the future design of biomimicking artificial periosteum. Conclusion: This review summarizes the preparation strategies of biomimicking artificial periosteum in recent years with a discussion of material selection, animal model adoption, biophysical and biochemical cues to regulate the cell fates as well as three types of latest developed versatile biomimicking artificial periosteum. In future, integration of innervation, osteochondral regeneration, and osteoimmunomodulation, should be taken into consideration when fabricating multifunctional artificial periosteum.The Translational Potential of this Article: This study provides a holistic view on the design strategy and the therapeutic potential of biomimicking artificial periosteum to promote bone healing. It is hoped to open a new avenue of artificial periosteum design with biomimicking considerations and reposition of the current strategy for accelerated bone healing.

A universal biocompatible coating for enhanced lubrication and bacterial inhibition.

Antibacterial coatings that inhibit bacterial adhesion are essential for many implanted medical devices. A variety of antibacterial strategies, such as repelling or killing bacteria, have been developed, but not yet been completely successful. Here, we develop a universal biocompatible coating for enhanced lubrication and bacterial inhibition. The coating is designed based on mussel-inspired surface-attachable dopamine bases and consists of lubricating zwitterionic polymers poly(2-methacryloxyethyl phosphorylcholine) (MPC) and a bacterial membrane destroying anti-bacteria molecule poly(3-hydroxybutyric acid) (PHB). The coating boasts strong adhesion to surfaces of various materials (such as polydimethylsiloxane (PDMS)/ceramic/316L stainless steel (316L SS); it is biocompatible, and cell/platelet/bacteria repelling, significantly inhibiting bacterial growth. We envision that our strategy represents a universal strategy for surface functionalization of a variety of biomedical devices and implants.

3D Bioprinting Photo-Crosslinkable Hydrogels for Bone and Cartilage Repair.

Three-dimensional (3D) bioprinting has become a promising strategy for bone manufacturing, with excellent control over geometry and microarchitectures of the scaffolds. The bioprinting ink for bone and cartilage engineering has thus become the key to developing 3D constructs for bone and cartilage defect repair. Maintaining the balance of cellular viability, drugs or cytokines' function, and mechanical integrity is critical for constructing 3D bone and/or cartilage scaffolds. Photo-crosslinkable hydrogel is one of the most promising materials in tissue engineering; it can respond to light and induce structural or morphological transition. The biocompatibility, easy fabrication, as well as controllable mechanical and degradation properties of photo-crosslinkable hydrogel can meet various requirements of the bone and cartilage scaffolds, which enable it to serve as an effective bio-ink for 3D bioprinting. Here, in this review, we first introduce commonly used photo-crosslinkable hydrogel materials and additives (such as nanomaterials, functional cells, and drugs/cytokine), and then discuss the applications of the 3D bioprinted photo-crosslinkable hydrogel scaffolds for bone and cartilage engineering. Finally, we conclude the review with future perspectives about the development of 3D bioprinting photo-crosslinkable hydrogels in bone and cartilage engineering.

pH-Triggered Copper-Free Click Reaction-Mediated Micelle Aggregation for Enhanced Tumor Retention and Elevated Immuno-Chemotherapy against Melanoma.

Natural killer (NK) cell-based immunotherapy presents a promising antitumor strategy and holds potential for combination with chemotherapy. However, the suppressed NK cell activity and poor tumor retention of therapeutics hinder the efficacy. To activate NK cell-based immuno-chemotherapy and enhance the tumor retention, we proposed a pH-responsive self-aggregated nanoparticle for the codelivery of chemotherapeutic doxorubicin (DOX) and the transforming growth factor-ß (TGF-ß)/Smad3 signaling pathway inhibitor SIS3. Polycaprolactone-poly(ethylene glycol) (PCL-PEG2000) micelles modified with dibenzylcyclooctyne (DBCO) or azido (N3) and coated with acid-cleavable PEG5000 were established. This nanoplatform, namely, M-DN@DOX/SIS3, could remain well dispersed in the neutral systemic circulation, while quickly respond to the acidic tumor microenvironment and intracellular lysosomes, triggering copper-free click reaction-mediated aggregation, leading to the increased tumor accumulation and reduced cellular efflux. In addition, the combination of DOX with SIS3 facilitated by the aggregation strategy resulted in potent inhibition of melanoma tumor growth and significantly increased NK cells, NK cell cytokines, and antitumor T cells in the tumor. Taken together, our study offered a new concept of applying copper-free click chemistry to achieve nanoparticle aggregation and enhance tumor retention, as well as a promising new combined tumor treatment approach of chemotherapy and immunotherapy.

Antibacterial nanosystems for cancer therapy.

Bacteria and cancer cells share a unique symbiotic relationship in the process of cancer development and treatment. It has been shown that certain bacteria can mediate cancer and thrive inside cancerous tissues. Moreover, during cancer treatment, microbial infections have been shown to impair the therapeutic efficacy and lead to serious complications. In the past decades, the application of antibiotics has achieved great success in fighting numerous bacteria but the administration route, low localization effects and related drug resistance limit the further utilization of antibiotics. Recently, advances in nanotechnology have made a significant impact in the medical field, which enhance the drug solubility and can target lesion sites, and some nanomaterials can even be applied as the therapeutic agent itself. In this review, we introduce anti-bacterial nanosystems for cancer therapy in the aspects of spontaneous and triggered anti-bacterial action, and our notions, as well as proposed research directions for the further development of this field, are discussed.

Nitric Oxide-Producing Cardiovascular Stent Coatings for Prevention of Thrombosis and Restenosis.

Cardiovascular stenting is an effective method for treating cardiovascular diseases (CVDs), yet thrombosis and restenosis are the two major clinical complications that often lead to device failure. Nitric oxide (NO) has been proposed as a promising small molecule in improving the clinical performance of cardiovascular stents thanks to its anti-thrombosis and anti-restenosis ability, but its short half-life limits the full use of NO. To produce NO at lesion site with sufficient amount, NO-producing coatings (including NO-releasing and NO-generating coatings) are fashioned. Its releasing strategy is achieved by introducing exogenous NO storage materials like NO donors, while the generating strategy utilizes the in vivo substances such as S-nitrosothiols (RSNOs) to generate NO flux. NO-producing stents are particularly promising in future clinical use due to their ability to store NO resources or to generate large NO flux in a controlled and efficient manner. In this review, we first introduce NO-releasing and -generating coatings for prevention of thrombosis and restenosis. We then discuss the advantages and drawbacks on releasing and generating aspects, where possible further developments are suggested.

Autophagy inhibition changes the disposition of non-viral gene carriers during blood-brain barrier penetration and enhances TRAIL-induced apoptosis in brain metastatic tumor.

Non-viral gene delivery systems have proven to be a promising approach in the treatment of brain metastatic cancers but facing delivery difficulties. Due to the existence of blood-brain barrier, non-viral gene carriers must pass through brain capillary endothelial cells to accumulate at the brain tumor sites. However, during this process, most of them trap into brain capillary endothelial cells and fail to penetrate to the brain tumor sites. Autophagy is involved in dynamic disposition of both intracellular and extracellular components, which theoretically affects intracellular fate of non-viral gene carriers during BBB penetration. In the present study, R6dGR peptide-modified PEGylated polyethyleneimine that carry therapeutic gene encoding human tumor necrosis factor-related apoptosis-inducing ligand (PPR/pTRAIL) are established as model non-viral gene delivery system and applied in breast cancer brain metastasis therapy. Autophagy-mediated lysosome degradation pathway is found to be involved in the degradation of PPR/pTRAIL in brain capillary endothelial cells and prevents them from BBB penetration. Pre-inhibiting BBB autophagy level by wortmannin loaded liposomes (Wtmn-Lip) can increase brain accumulation of non-viral gene carrier PPR without damaging BBB tight junctions. Besides, Wtmn-Lip synergistically induces apoptosis with TRAIL via different signaling pathways. Herein, pre-treatment of Wtmn-Lip might solve delivery difficulties of non-viral gene carriers in the treatment of brain metastatic cancers.

Multifunctional polymeric micelle-based chemo-immunotherapy with immune checkpoint blockade for efficient treatment of orthotopic and metastatic breast cancer.

Immunotherapy has become a highly promising paradigm for cancer treatment. Herein, a chemo-immunotherapy was developed by encapsulating chemotherapeutic drug doxorubicin (DOX) and Toll-like receptor 7 agonist imiquimod (IMQ) in low molecular weight heparin (LMWH)-d-α-tocopheryl succinate (TOS) micelles (LT). In this process, LMWH and TOS were conjugated by ester bond and they were not only served as the hydrophilic and hydrophobic segments of the carrier, but also exhibited strong anti-metastasis effect. The direct killing of tumor cells mediated by DOX-loaded micelles (LT-DOX) generated tumor-associated antigens, initiating tumor-specific immune responses in combination with IMQ-loaded micelles (LT-IMQ). Furthermore, the blockade of immune checkpoint with programmed cell death ligand 1 (PD-L1) antibody further elevated the immune responses by up-regulating the maturation of DCs as well as the ratios of CD8+ CTLs/Treg and CD4+ Teff/Treg. Therefore, such a multifunctional strategy exhibited great potential for inhibiting the growth of orthotopic and metastatic breast cancer.

Size-adjustable micelles co-loaded with a chemotherapeutic agent and an autophagy inhibitor for enhancing cancer treatment via increased tumor retention.

Autophagy plays a key role in the stress response of tumor cells, which contributes to cancer cell survival and resistance to chemotherapy by degrading cytoplasmic proteins to provide energy and clear the hazardous substances. Therefore, combined treatment of chemotherapeutics and autophagy inhibitors is thought to obtain a desirable antitumor effect. Nanoparticles (NPs) show potential in tumor-targeting drug delivery because of the enhanced permeability and retention (EPR) effect. However, NPs with fixed particle size cannot achieve optimal delivery effect. Herein, a strategy based on Cu (I)-catalyzed click chemistry-triggered aggregation of azide/alkyne-modified micelles was developed for the co-delivery of the chemotherapeutic drug doxorubicin (Dox) and the autophagy inhibitor wortmannin (Wtmn). In vitro experiments showed that the size of micelles increased in a time-dependent manner, which enhanced micelle accumulation in both B16F10 and 4 T1 cells. The fluorescence resonance energy transfer (FRET) experiment and biodistribution study further demonstrated that the aggregation of micelles through click cycloaddition significantly improved the accumulation of drug-loading micelles at the tumor region. Furthermore, the decreased amount of autophagosomes observed by transmission electron microscopy (TEM), the declined expression of LC3-II, and the increased level of p62 by western blotting and immunohistochemistry (IHC) confirmed the obvious inhibition of autophagy induced by Dox/Wtmn co-loaded size-adjustable micelles, which had a synergistic effect in cancer suppression. In addition, the co-loaded size-adjustable micelles showed outstanding cytotoxicity and antitumor effect. Therefore, this strategy effectively suppressed melanoma and breast cancer in mice. STATEMENT OF SIGNIFICANCE: The therapeutic effects of chemotherapy can be limited by autophagy; hence, combined use of autophagy inhibitors with chemotherapeutics achieves desirable anticancer efficacy. In the present study, we designed size-adjustable micelles by modifying the click reaction substrate azide group and the alkyne group on the surface of micelles, and subsequently, the autophagy inhibitor wortmannin and the chemotherapeutic drug doxorubicin were co-loaded. The micelles could aggregate by click reaction at the tumor site when the catalysts were intratumorally injected. The results showed that the size-adjustable micelles achieved efficient drug delivery, penetration, and retention in tumors; through the combined effect of wortmannin-mediated autophagy inhibition and doxorubicin-mediated cytotoxicity, this strategy exerted significant anticancer effect in melanoma and breast cancer treatment.

Low Molecular Weight Heparin-Coated and Dendrimer-Based Core-Shell Nanoplatform with Enhanced Immune Activation and Multiple Anti-Metastatic Effects for Melanoma Treatment.

High-efficiency treatment for tumor is not easy to achieve owing to the existence of metastasis, which remains the arch-criminal of most tumor deaths. Conventional chemotherapy exhibits insufficient inhibitory efficiency on tumor metastasis and more powerful strategies to conquer metastatic tumors are urgently needed. In this study, a rational chemoimmunotherapy strategy was adopted to treat highly aggressive melanoma based on a newly developed multifunctional nanoplatform. Firstly, immunoadjuvant cytosine-phosphate-guanine oligonucleotides (CpG ODNs) were used to boost the doxorubicin (DOX)-elicited immune responses, which synergistically suppressed tumor growth and metastasis. And the anti-metastatic low molecular weight heparin (LMWH) was also integrated, thus multiple anti-metastatic effects to against tumor metastasis were achieved. Methods: G4 PAMAM was serving as the main support to conjugate DOX by pH-sensitive hydrazone bond (PPD) and the synthesized conjugates were confirmed by 1H-NMR spectra, IR spectra and HRMS. Immunoadjuvant CpG ODNs were loaded by electrostatic adsorption to formulate PPD/CpG. After the coating of anti-metastatic LMWH, the designed LMWH/PPD/CpG was fabricated and characterized. The platelets-related and platelets-unrelated anti-metastatic mechanisms were investigated on B16F10 the immune activation effects, anti-tumor and anti-metastatic efficacy of LMWH/PPD/CpG were evaluated on a B16F10 melanoma xenograft model. Results: DOX elicited tumor-specific immune responses by ICD, and the immunological effects could be further promoted by CpG ODNs, exhibiting enhanced maturation of dendritic cells (DCs) and increased level of cytolytic T lymphocytes (CTLs) in vivo. Owing to the coating of LMWH, the platelets-induced epithelial-mesenchymal-like transition of tumor cells was hindered and the actin cytoskeletal arrangement of tumor cells was affected, thus the migration ability of tumor cells was further inhibited. This multifunctional nanoplatform showed enhanced treatment efficiency on melanoma primary tumor and pulmonary metastasis. Conclusion: The immune activation and multiple anti-metastatic effects of LMWH/PPD/CpG establish a novel therapeutic strategy for melanoma. This anti-metastatic nanoplatform could be broadly applied for the co-delivery of other nucleic acids and chemotherapeutic drugs to treat highly aggressive tumors.

PD-L1 knockdown via hybrid micelle promotes paclitaxel induced Cancer-Immunity Cycle for melanoma treatment.

The Cancer-Immunity Cycle is a series of anticancer immune responses initiated and allowed to proceed and expand iteratively. Paclitaxel (PTX) is a classic chemotherapeutic agent, which could induce immunogenic cell death (ICD) to trigger the Cancer-Immunity Cycle. However, the Cycle is severely impaired by tumor cell immunosuppression of host T cell antitumor activity through the programmed cell death receptor 1 (PD-1) and programmed cell death ligand 1 (PD-L1) (PD-1/PD-L1) immune checkpoint pathway. Here, we demonstrated that PTX mediated the Cancer-Immunity Cycle could be enhanced by PD-L1 knockdown (KD) and followed mTOR pathway inhibition in tumor cells. PD-L1 siRNA (siP) and the hydrophobic chemotherapy drug PTX were co-delivered with a rationally designed hybrid micelle (HM). We showed clear evidence that the HM-siP/PTX is capable of delivering siP and PTX simultaneously to the B16F10 cells both in vitro and in vivo. We demonstrated that HM-PTX/siP reduced the expression of PD-L1 and p-S6K (a marker of mTOR pathway activation) both in vitro and in melanoma-bearing mice and attenuated synergistically tumor growth by chemical toxicity, promoting cytotoxic T-cell immunity and suppressing the mTOR pathway.

Dual receptor recognizing liposomes containing paclitaxel and hydroxychloroquine for primary and metastatic melanoma treatment via autophagy-dependent and independent pathways.

Autophagy acts as a cytoprotective mechanism for malignant tumors, thus maintaining the survival and promoting proliferation and metastasis of malignant tumors. Recent studies have showed that autophagy inhibitors can enhance the chemotherapeutic efficacy of anti-tumor growth. However, the antimetastasis effects and the possible mechanisms of chemotherapeutics combined with autophagy inhibitors have not been thoroughly explored. Here, we prepared R8-dGR peptide modified paclitaxel (PTX) and hydroxychloroquine (HCQ) co-loaded liposomes (PTX/HCQ-R8-dGR-Lip) for enhanced delivery by recognizing integrin αvß3 receptors and neuropilin-1 receptors on B16F10 melanoma cells. Our results showed that R8-dGR modified liposomes (R8-dGR-Lip) enhanced tumor-targeting delivery in vitro and in vivo. Besides, PTX/HCQ-R8-dGR-Lip exhibited the optimum inhibitory effects on migration, invasion and anoikis resistance of B16F10 cells in vitro, and showed enhanced efficiency on inhibiting primary tumor growth and reducing lung metastasis in vivo. Meanwhile, the antimetastasis mechanism studies confirmed that the combination of the chemotherapeutic PTX and the autophagy inhibitor HCQ further suppressed the degradation of paxillin, the expression of MMP9 and MMP2. Moreover, HCQ disturbed the CXCR4/CXCL12 axis which could induce invasion and metastasis of malignant melanoma in an autophagy-independent way.

Effective treatment of the primary tumor and lymph node metastasis by polymeric micelles with variable particle sizes.

Nanoparticles (NPs) offer new solutions for the diagnosis and treatment of tumors. However, the anti-tumor effect has not been greatly improved. Tumors are easily spread through the lymphatic system while the traditional NPs (~100 nm) can hardly reach lymph nodes for the treatment of metastasis. In addition, the NPs with fixed particle size cannot achieve efficient "penetration" and long-term "retention" simultaneously. Herein, we established "transformable" micelles modified with azide/alkyne groups for click chemical reaction. Not surprisingly, the small micelles (~25 nm) could effectively target lymph nodes, limiting the growth of the metastases associated with their size-regulated abilities to extravasate from the vasculature. Tumor lymph node metastasis dropped by 66.7%. After reaching primary tumors, cycloaddition reaction occurred between groups on micelles, resulting in the formation of aggregates. The strategy resulted in improved retention of the micelles in 4 T1 cells both in vitro and in vivo owing to the decreasing of nanoparticle exocytosis and minimizing the backflow to the bloodstream. Enhanced cytotoxicity on 4 T1 cells and improved antitumor efficacy were also observed. S-PTX (+) exhibited 76.23% tumor suppression, and tumor mass at the end of the treatment also showed the best tumor inhibitory effect. In conclusion, this drug delivery system provides a strategy for effective treatment of the primary tumor and lymphatic metastasis.

Enhanced Tumor Retention Effect by Click Chemistry for Improved Cancer Immunochemotherapy.

Because of the limited drug concentration in tumor tissues and inappropriate treatment strategies, tumor recurrence and metastasis are critical challenges for effectively treating malignancies. A key challenge for effective delivery of nanoparticles is to reduce uptake by reticuloendothelial system and to enhance the permeability and retention effect. Herein, we demonstrated Cu(I)-catalyzed click chemistry triggered the aggregation of azide/alkyne-modified micelles, enhancing micelles accumulation in tumor tissues. In addition, combined doxorubicin with the adjuvant monophosphoryl lipid A, an agonist of toll-like receptor4, generated immunogenic cell death, which further promoted maturity of dendritic cells, antigen presentation and induced strong effector T cells in vivo. Following combined with anti-PD-L1 therapy, substantial antitumor and metastasis inhibitory effects were achieved because of the reduced PD-L1 expression and regulatory T cells. In addition, effective long-term immunity from memory T cell responses protected mice from tumor recurrence.

Ligand-Mediated and Enzyme-Directed Precise Targeting and Retention for the Enhanced Treatment of Glioblastoma.

Glioblastoma (GBM), one of the most lethal cancers, remains as a hard task to handle. The major hurdle of nanostructured therapeutic agents comes from the limited retention at the GBM site and poor selectivity. In this study, we reported dual-functional gold nanoparticles (AuNPs) to figure out the biological barrier and improve their accumulation in GBM. The nanoparticles, AuNP-A&C-R, were composed of two functional particles: one was Ala-Ala-Asn-Cys-Asp (AK) and R8-RGD-comodified AuNPs (AuNP-AK-R) and the other was 2-cyano-6-amino-benzothiazole and R8-RGD-comodified AuNPs (AuNP-CABT-R). AuNP-A&C-R could aggregate in the presence of legumain, resulting in a size increase from 41.4 ± 0.6 to 172.9 ± 10.2 nm after 8 h incubation. After entering the circulatory system, AuNP-A&C-R actively targeted the integrin αvß3 receptor on blood-brain barrier (BBB), mediated transcytosis of particles across BBB, and then targeted the receptor on the GBM cells. Once AuNP-A&C-R entered into GBM, they formed further aggregates with increased size extracellularly or intracellularly because of the overexpressed legumain, which in turn blocked their backflow to the bloodstream or limited their exocytosis by cells. In vivo optical imaging demonstrated that AuNP-A&C-R were efficiently delivered to the GBM site and retained with high selectivity. We further confirmed that AuNP-A&C-R acquired a higher accumulation at the GBM site than AuNP-A&C and AuNP-R because of the synergistic effect. More importantly, the doxorubicin (DOX)-loaded AuNP-A&C-R showed an improved chemotherapeutic effect to C6 GBM-bearing mice, which significantly prolonged the median survival time by 1.22-fold and 1.27-fold compared with the DOX-loaded AuNP-A&C and the DOX-loaded AuNP-R, respectively. These results suggested that the dual-functional nanoplatform is promising for the GBM treatment.