Curcumin loaded sub-30 nm targeting therapeutic lipid nanoparticles for synergistically blocking nasopharyngeal cancer growth and metastasis.

Systemic chemotherapy is still the primary treatment for advanced-stage nasopharyngeal carcinoma (NPC), but only limited therapeutic success has been achieved in the past decade because of drug resistance and systemic toxicity. Curcumin (Cur) is an effective alternative to chemotherapeutics because it showed remarkable therapeutic potential in the treatment of NPC. However, lack of tissue specificity and poor penetration in solid tumors are the major obstacles to effective therapy. Therefore, in this work, a self-assembled sub-30 nm therapeutic lipid nanoparticle loaded with Cur, named as Cur@α-NTP-LN, was constructed, specifically targeting scavenger receptor class B member 1 (SR-B1) and enhancing its therapeutic effects on NPC in vivo. Our results showed that Cur@α-NTP-LNs were effective and superior to free Cur on NPC cell-specific targeting, suppressing cell proliferation and inducing cell apoptosis. In vivo and ex vivo optical imaging revealed that Cur@α-NTP-LNs exerted high targeting efficiency, specifically accumulating in NPC xenograft tumors and delivering Cur into the tumor center after systemic administration. Furthermore, Cur@α-NTP-LNs exhibited a remarkable inhibitory effect on the growth of NPC subcutaneous tumors, with over 71 and 47% inhibition compared to Cur- and α-NTP-LNs-treated groups, respectively. In addition, Cur@α-NTP-LNs almost blocked NPC metastasis in a lung metastasis model of NPC and significantly improved the survival rate. Thus, the sub-30 nm Cur@α-NTP-LNs enhanced the solubility of Cur and demonstrated the ability of targeted Cur delivery into the center of the solid NPC tumor, performing synergistic inhibitory effects on the growth of NPC tumor and its metastasis with high efficiency.

Investigation on Adaptability and Applicability of Polymer-Mediated Cell Surface Engineering by Ligation with Transglutaminase.

Polymer-mediated cell surface engineering can be a powerful tool to modify the cell's biological behavior, but a simple ligation strategy must be identified. This manuscript assessed the use of transglutamination as a versatile and adaptable approach for cell surface engineering in various cellular models relevant to biomedical applications. This enzymatic approach was evaluated for its feasibility and potential for conjugating polymers to diverse cell surfaces and its biological effects. Transglutaminase-mediated ligation was successfully performed at temperatures ranging from 4 to 37 °C in as quickly as 30 min, while maintaining biocompatibility and preserving cell viability. This approach was successfully applied to nine different cell surfaces (including adherent cells and suspension cells) by optimizing the enzyme source (guinea pig liver vs microbial), buffer compositions, and incubation conditions. Finally, polymer-mediated cell surface engineering using transglutaminase exhibited immunocamouflage abilities for endothelial cells, T cells, and red blood cells by preventing the recognition of cell surface proteins by antibodies. Employing transglutaminase in polymer-mediated cell surface engineering is a promising approach to maximize its application in cell therapy and other biomedical applications.

Long-Term Prevention of Biofilm Formation by Polycatechol-Based Supramolecular Assemblies with Low Molecular Weight Polymers on Surfaces.

Achievement of a stable surface coating with long-term resistance to biofilm formation remains a challenge. Catechol-based polymerization chemistry and surface deposition are used as tools for surface modification of diverse materials. However, the control of surface deposition of the coating, surface coverage, coating properties, and long-term protection against biofilm formation remain to be solved. We report a new approach based on supramolecular assembly to generate long-acting antibiofilm coating. Here, we utilized catechol chemistry in combination with low molecular weight amphiphilic polymers for the generation of such coatings. Screening studies with diverse low molecular weight (LMW) polymers and different catechols are utilized to identify lead compositions, which resulted in a thick coating with high surface coverage, smoothness, and antibiofilm activity. We have identified that small supramolecular assemblies (∼10 nm) formed from a combination of polydopamine and LMW poly(N-vinyl caprolactam) (PVCL) resulted in relatively thick coating (∼300 nm) with excellent surface coverage in comparison to other polymers and catechol combinations. The coating properties, such as thickness (10-300 nm) and surface hydrophilicity (with water contact angle: 20-60°), are readily controlled. The optimal coating composition showed excellent antibiofilm properties with long-term (>28 days) antibiofilm activity against both Gram-negative (Escherichia coli) and Gram-positive (Staphylococcus aureus) strains. We further utilized the combination of optimal binary coating with silver to generate a coating with sustained release of silver ions, resulting in killing both adhered and planktonic bacteria and preventing long-term surface bacterial colonization. The new coating method utilizing LMW polymers opens a new avenue for the development of a novel class of thick, long-acting antibiofilm coatings.

Sulfated Hyperbranched and Linear Polyglycerols Modulate HMGB1 and Morphological Plasticity in Neural Cells.

The objective of this study was to establish if polyglycerols with sulfate or sialic acid functional groups interact with high mobility group box 1 (HMGB1), and if so, which polyglycerol could prevent loss of morphological plasticity in excitatory neurons in the hippocampus. Considering that HMGB1 binds to heparan sulfate and that heparan sulfate has structural similarities with dendritic polyglycerol sulfates (dPGS), we performed the experiments to show if polyglycerols can mimic heparin functions by addressing the following questions: (1) do dendritic and linear polyglycerols interact with the alarmin molecule HMGB1? (2) Does dPGS interaction with HMGB1 influence the redox status of HMGB1? (3) Can dPGS prevent the loss of dendritic spines in organotypic cultures challenged with lipopolysaccharide (LPS)? LPS plays a critical role in infections with Gram-negative bacteria and is commonly used to test candidate therapeutic agents for inflammation and endotoxemia. Pathologically high LPS concentrations and other stressful stimuli cause HMGB1 release and post-translational modifications. We hypothesized that (i) electrostatic interactions of hyperbranched and linear polysulfated polyglycerols with HMGB1 will likely involve sites similar to those of heparan sulfate. (ii) dPGS can normalize HMGB1 compartmentalization in microglia exposed to LPS and prevent dendritic spine loss in the excitatory hippocampal neurons. We performed immunocytochemistry and biochemical analyses combined with confocal microscopy to determine cellular and extracellular locations of HMGB1 and morphological plasticity. Our results suggest that dPGS interacts with HMGB1 similarly to heparan sulfate. Hyperbranched dPGS and linear sulfated polymers prevent dendritic spine loss in hippocampal excitatory neurons. MS/MS analyses reveal that dPGS-HMGB1 interactions result in fully oxidized HMGB1 at critical cysteine residues (Cys23, Cys45, and Cys106). Triply oxidized HMGB1 leads to the loss of its pro-inflammatory action and could participate in dPGS-mediated spine loss prevention. LPG-Sia exposure to HMGB1 results in the oxidation of Cys23 and Cys106 but does not normalize spine density.

Rapid Assembly of Infection-Resistant Coatings: Screening and Identification of Antimicrobial Peptides Works in Cooperation with an Antifouling Background.

Bacterial adhesion and the succeeding biofilm formation onto surfaces are responsible for implant- and device-associated infections. Bifunctional coatings integrating both nonfouling components and antimicrobial peptides (AMPs) are a promising approach to develop potent antibiofilm coatings. However, the current approaches and chemistry for such coatings are time-consuming and dependent on substrates and involve a multistep process. Also, the information is limited on the influence of the coating structure or its components on the antibiofilm activity of such AMP-based coatings. Here, we report a new strategy to rapidly assemble a stable, potent, and substrate-independent AMP-based antibiofilm coating in a nonfouling background. The coating structure allowed for the screening of AMPs in a relevant nonfouling background to identify optimal peptide combinations that work in cooperation to generate potent antibiofilm activity. The structure of the coating was changed by altering the organization of the hydrophilic polymer chains within the coatings. The coatings were thoroughly characterized using various surface analytical techniques and correlated with the efficiency to prevent biofilm formation against diverse bacteria. The coating method that allowed the conjugation of AMPs without altering the steric protection ability of hydrophilic polymer structure results in a bifunctional surface coating with excellent antibiofilm activity. In contrast, the conjugation of AMPs directly to the hydrophilic polymer chains resulted in a surface with poor antibiofilm activity and increased adhesion of bacteria. Using this coating approach, we further established a new screening method and identified a set of potent surface-tethered AMPs with high activity. The success of this new peptide screening and coating method is demonstrated using a clinically relevant mouse infection model to prevent catheter-associated urinary tract infection (CAUTI).

Enzymatically releasable polyethylene glycol - host defense peptide conjugates with improved activity and biocompatibility.

Host defense peptides (HDPs) have been the subject of great interest for the treatment of multidrug-resistant bacterial infections due to their multimodal activity and low induction of resistance. However, aggregation, toxicity, and short biological half-life have limited their applicability for clinical treatment. Many methods have been explored to alleviate these issues, such as polymer (e.g., polyethylene glycol (PEG)) conjugation, but these are often accompanied by reductions in the activity of the HDP. Here, we detail the design of a novel PEG-HDP conjugate incorporating an enzymatic cleavage sequence targeting matrix metalloproteinases (MMPs) that accumulate at sites of inflammation and infection. Addition of the cleavage sequence onto either the N- or the C-terminal region of the parent peptide (peptide 73, a derivative of the HDP aurein 2.2) was explored to determine the location for optimal antimicrobial activity following MMP cleavage; furthermore, the susceptibility of the peptide to MMP cleavage after conjugation to 2 kDa or 5 kDa PEG was examined. The top candidate, L73, utilized an N-terminal cleavage site that was subsequently conjugated to a 2 kDa PEG polymer. Both L73 and the conjugate exhibited no antimicrobial activity in vitro until cleaved by purified MMP, which liberated a peptide fragment with 16- or 63-fold improved activity, respectively, corresponding to a minimum inhibitory concentration (MIC) of 8 µg/mL, comparable to that of peptide 73 (4 µg/mL). Furthermore, PEG conjugation improved the blood compatibility and reduced the aggregation tendency of the HDP in vitro, indicating enhanced biocompatibility. When administered as a single subcutaneous dose (~3.6 mg, or a peptide concentration of 142 mg/kg) in a mouse abscess model of high-density methicillin-resistant Staphylococcus aureus (MRSA) infection, the conjugate displayed strong activity, reducing abscess size and bacterial load by 73.3% and 58-fold, respectively. This activity was completely lost when the cleavage site was rendered resistant to MMPs by the substitution of two d-amino acids, supporting the hypothesis that antimicrobial activity was dependent on cleavage by MMPs, which were shown here to increasingly accumulate at the abscess site up to 18 h post infection. Finally, the conjugate displayed biocompatibility in vivo, with no identifiable toxicity or aggregation.

Prevention of vascular-allograft rejection by protecting the endothelial glycocalyx with immunosuppressive polymers.

Systemic immunosuppression for the mitigation of immune rejection after organ transplantation causes adverse side effects and constrains the long-term benefits of the transplanted graft. Here we show that protecting the endothelial glycocalyx in vascular allografts via the enzymatic ligation of immunosuppressive glycopolymers under cold-storage conditions attenuates the acute and chronic rejection of the grafts after transplantation in the absence of systemic immunosuppression. In syngeneic and allogeneic mice that received kidney transplants, the steric and immunosuppressive properties of the ligated polymers largely protected the transplanted grafts from ischaemic reperfusion injury, and from immune-cell adhesion and thereby immunocytotoxicity. Polymer-mediated shielding of the endothelial glycocalyx following organ procurement should be compatible with clinical procedures for transplant preservation and perfusion, and may reduce the damage and rejection of transplanted organs after surgery.