Hyaluronic Acid Conjugates Research Articles

Preparation and in vitro evaluation of radiolabeled HA-PLGA nanoparticles as novel MTX delivery system for local treatment of rheumatoid arthritis

Radiosynovectomy is a technique used to decrease inflammation of the synovial tissue by intraarticular injection of a β-emitting radionuclide, such as 177Lu, which is suitable for radiotherapy due to its decay characteristics. Drug-encapsulating nanoparticles based on poly lactic‑co‑glycolic acid (PLGA) polymer are a suitable option to treat several arthritic diseases, used as anti-inflammatory drugs transporters of such as methotrexate (MTX), which has been widely used in the arthritis treatment (RA), and hyaluronic acid (HA), which specifically binds the CD44 and hyaluronan receptors overexpressed on the inflamed synovial tissue cells. The 1,4,7,10‑Tetraazacyclododecane‑1,4,7,10‑tetraacetic acid (DOTA) was used as complexing agent of Lutetium-177 for radiotherapy porpoises. The aim of this research was to synthesize 177Lu-DOTA-HA-PLGA(MTX) as a novel, smart drug delivery system with target-specific recognition, potentially useful in radiosynovectomy for local treatment of rheumatoid arthritis. The polymeric nanoparticle system was prepared and chemically characterized. The MTX encapsulation and radiolabelling were performed with suitable characteristics for its in vitro evaluation. The HA-PLGA(MTX) nanoparticle mean diameter was 167.6 nm ± 57.4 with a monomodal and narrow distribution. Spectroscopic techniques demonstrated the effective conjugation of HA and chelating agent DOTA to the polymeric nanosystem. The MTX encapsulation was 95.2% and the loading efficiency was 6%. The radiochemical purity was 96 ± 2%, determined by ITLC. Conclusion: 177Lu-DOTA-HA-PLGA(MTX) was prepared as a biocompatible polymeric PLGA nanoparticle conjugated to HA for specific targeting. The therapeutic nanosystem is based on bi-modal mechanisms using MTX as a disease-modifying antirheumatic drug (DMARD) and 177Lu as a radiotherapeutic component. The 177Lu-DOTA-HA-PLGA(MTX) nanoparticles showed properties suitable for radiosynovectomy and further specific targeted anti-rheumatic therapy.

Enhanced Transfection of Human Mesenchymal Stem Cells Using a Hyaluronic Acid/Calcium Phosphate Hybrid Gene Delivery System

Human mesenchymal stem cells (hMSCs) show enormous potential in regenerative medicine and tissue engineering. However, current use of hMSCs in clinics is still limited because there is no appropriate way to control their behavior in vivo, such as differentiation to a desired cell type. Genetic modification may provide an opportunity to control the cells in an active manner. One of the major hurdles for genetic manipulation of hMSCs is the lack of an efficient and safe gene delivery system. Herein, biocompatible calcium phosphate (CaP)-based nanoparticles stabilized with a catechol-derivatized hyaluronic acid (dopa-HA) conjugate were used as a carrier for gene transfection to hMSCs for improved differentiation. Owing to the specific interactions between HA and CD44 of bone marrow-derived hMSCs, dopa-HA/CaP showed significantly higher transfection in hMSCs than branched polyethylenimine (bPEI, MW 25 kDa) with no cytotoxicity. The co-delivery of a plasmid DNA encoding bone morphogenetic protein 2 (BMP-2 pDNA) and micro RNA 148b (miRNA-148b) by dopa-HA/CaP achieved significantly improved osteogenic differentiation of hMSCs.

Hyaluronic acid - dihydroartemisinin conjugate: Synthesis, characterization and in vitro evaluation in lung cancer cells

In recent years, a great deal of attention has been given towards re-purposing and re-innovating the potential drugs. In this regard, dihydroartemisinin (DHA) has been reported to demonstrate anti-proliferative effects on various cancerous cells viz. breast, liver and lung. However, it is associated with some limitations, such as low bioavailability which is hampered by its poor aqueous solubility and its rapid metabolism in systemic circulation. Therefore, in order to overcome these limitations, we synthesized a novel hyaluronic acid-dihydroartemisinin conjugate in which the hydroxyl group of DHA was covalently linked to carboxylic group of hyaluronic acid (HA). The conjugate was successfully characterized using 1H NMR, Fourier transform infrared spectroscopy (FT-IR) and gel permeation chromatography (GPC). The synthesized conjugate self-assembled into nanoparticles in aqueous solution. The developed nanoparticles were characterized for their average size, zeta potential, Transmission Electron Microscopy (TEM), X-ray Powder Diffraction (XRD) and loading efficiency. The nanoparticles were cytotoxic to lung cancer (A549) cell line which was determined using CCK-8 cell viability assay. This was further supported by Annexin-V-FITC-Propidium iodide apoptosis assay, reactive oxygen species (ROS) production and mitochondrial membrane potential (MMP) loss. Conclusively, present findings demonstrate hyaluronic acid conjugates can be used to improve the therapeutic outcomes of anticancer drugs.

Albumin Based Nanoparticles for Detection of Pancreatic Cancer Cells.

Molecular imaging of cancer cells using effective drug targeting systems are most interested research area in recent years. Albumin protein is a soluble and most abundant protein in circulatory system. It has a ligand-binding function and acts as a transport protein. Researchers are interested in developing albumin based nanostructured specific anti-tumor drugs in cancer therapy. Pancreatic cancer treatment or drug design for targeted pancreatic cancer cell has great importance due to it has a high mortality rate comparing other cancer types. In this article, our goal is to develop new targeting nanoparticles based on the conjugation of albumin and Hyaluronic Acid (HA) for pancreatic cancer cells. In this article, we proposed a new technique for conjugation of albumin (BSA) and HA in nano formation. Firstly, cationic BSA is synthesized. Then, BSA-HA conjugation is obtained by interacted cationic BSA with 1000 ppm HA. Secondly, nano BSA-HA particles and nano BSA particles were synthesized according to AmiNoAcid Decorated and Light Underpinning Conjugation Approach (ANADOLUCA) method which provides a special cross-linking strategy for biomolecules using ruthenium-based amino acid monomer haptens. After characterization studies, in vitro cytotoxic activity of synthesized nano BSA-HA particles were determined for PANC-1 ATCC® CRL146 cells. According to the data, nano BSA and nano BSA-HA particles synthesized uniquely using special ruthenium-based amino acid decorated cross-linking agent, (MATyr)2-Ru-(MATyr)2.based on ANDOLUCA method. Characterization results showed that there was not any change in protein folding structures during nano formation process. In addition, nano protein particles gained fluorescence feature. When interacting synthesized nano BSA and nano BSA-HA particles with pancreatic cells, it was found that BSA nanoparticles were usually around cells and membranes, but BSA-HA nanoparticles were identified around the cells, in the cytoplasm inside the cell, and next to the cell nucleus. So, nano BSA-HA particles could be used as cancer cell imaging agent for PANC-1 ATCC® CRL146 cells. The satisfactory conclusion of this study is that synthesized nano BSA-HA particles are fundamental materials for targeting pancreatic cancer cells due to HA receptors located on pancreatic cancer cells and imaging agents due to fluorescence feature of the BSA-HA nanoparticles.

Targeted Photodynamic Therapy Activities of Surface-Enhanced Raman Scattering-Active Theranostic System Based on Folate/Hyaluronic Acid-Functionalized Gold Nanochains.

Light-based diagnostics and therapies have become indispensable tools in the field of cancer medicine. Nanotheranostics, namely the integration of diagnostic elements with therapeutic agents into a single nanosized-regimen, have been the recent focus of intense exploration with the aim to develop more specific, individualized therapies and to combine diagnostics and therapeutic capabilities into a single agent. In this study, gold nanochains (AuNCs) with worm-like nanostructures were developed as a light-triggered theranostic system for effective multiplex detection and photodynamic therapy (PDT). We fabricated the AuNC system through assembling of citrate-stabilized gold nanoparticles using hyaluronic acid-hydrocaffeic acid (HA-HCA) conjugates as templates. Raman reporters, photosensitizers (PSs), and active tumor targeting ligands were also conjugated onto the surfaces of the AuNCs. The length of the AuNCs could be controlled by the HCA composition ratio in the HA-HCA conjugates. The AuNCs with worm-like nanostructures exhibited significant surface-enhanced Raman scattering (SERS) properties, and their optical properties depended on their length. The AuNCs conjugated with a Raman reporter, PS, and active targeting ligand (HA-HCA2.0-Au-NPT-FA-Ce6) exhibited excellent selectivity and showed marked phototoxicity even at low concentration of PS compared with free PS after laser irradiation. This tumor-targeting HA-HCA2.0-Au-NPT-FA-Ce6 AuNC system could be a promising approach for applications in cancer theranostics, integrating SERS detection, fluorescence imaging, and PDT.

Hyaluronic acid modified doxorubicin loaded Fe3O4 nanoparticles effectively inhibit breast cancer metastasis.

Tumor-associated macrophages (TAMs) play a critical role in tumor survival and metastasis. Iron(ii,iii) oxide (Fe3O4) nanoparticles have been shown to induce M1 macrophage polarization to initiate antitumor immunity and inhibit tumor metastasis. Hyaluronic acid (HA) modified doxorubicin (DOX) loaded Fe3O4 nanoparticles (Fe3O4-DOX-HA) have been constructed to mediate specific delivery of Fe3O4 nanoparticles to CD44-positive 4T1 tumor cells and tumor associated macrophages. Covalent conjugation of HA with DOX rendered nanoparticles with pH sensitivity, and further contributed to the prolonged circulation and enhanced tumor-specific accumulation in vivo. Furthermore, combining the M1 polarization effect of the Fe3O4 nanoparticles and enhanced cytotoxicity, Fe3O4-DOX-HA demonstrated enhanced antitumor and anti-metastasis effects both in vitro and in vivo.

Fluorescent Nanoclusters for Imaging of Cells/Stem Cells.

Imaging of cancer cells and cancer stem cells (CSCs) plays an important role in studying cell biology and tracking cancer development and metastasis. There is a wide interest in targeting cancer cells using fluorescent nanoclusters (NCs) capped in protein due to excellent cell viability and photostability, one-step synthesis route, large Stokes shift, good aqueous stability, and easy functionalization capability with long lifetime. Since CD44 is a CSC marker as well as a transmembrane receptor for hyaluronic acid (HA) and many other extracellular matrix (ECM) components, in this protocol, a biocompatible platform was synthesized by conjugation of HA onto luminescent platinum nanoclusters (Pt NCs) in human hemoglobin (Hb) (Hb/Pt NCs). This bioplatform could be used for specific imaging via an efficient targeting of CD44-overexpressing cancer cells and cancer stem cells.

Synergistic antitumor efficacy of hybrid micelles with mitochondrial targeting and stimuli-responsive drug release behavior.

The term synergism means that the overall therapeutic benefits should be greater than the sum of the effects of individual agents and that the optimal therapeutic efficacy can be achieved at reduced doses. Micellar systems usually fail to deliver multiple drugs to target sites at synergistic doses and thus are not able to maximize the antitumor efficacy. In the current study, we demonstrate a strategy to coordinate the release of camptothecin (CPT) and α-tocopheryl succinate (TOS) from hybrid micelles for nucleus and mitochondrion interferences. TOS is decorated with cationic triphenylphosphonium (TPP) to promote the targeting capability of TOS-TPP to mitochondria. The combination of CPT and TOS-TPP shows strong synergistism with a combination index of 0.186. Hyaluronic acid (HA) is conjugated with CPT or TOS-TPP via disulfide linkages for tumor cell targeting and intracellular reduction-triggered release. Both conjugates either separately self-assemble into MC and MT micelles, or are blended at different ratios to form MC-T hybrid micelles. In response to elevated intracellular glutathione levels, the coordinated release of CPT and TOS-TPP from MC-T results in a combination index of 0.26 and the dose-reduction indexes of CPT and TOS are 7.7 and 3.4, respectively. Compared with MC and MT, MC-T micelles with 5 fold lower doses exhibit higher intracellular reactive oxygen species (ROS) levels, comparable tumor growth inhibition and animal survival, indicating no hematologic and intestinal toxicities. Moreover, the HA conjugates of MC-T are linked to polylactide via acid-labile linkages and electrospun into short fibers (MC-T@SF) as an injectable depot to release MC-T in response to the acidic tumor microenvironment. At a predetermined synergistic ratio, MC-T@SF with 5 fold lower doses achieves antitumor profiles comparable to those of individual micelle-loaded short fibers. Therefore, the hybrid micelles and micelle-releasing short fibers represent a feasible strategy to synergistically enhance the therapeutic efficacy and enable significant reduction in effective doses of chemotherapeutic agents.

Bioreducible Zinc(II)-Dipicolylamine Functionalized Hyaluronic Acid Mediates Safe siRNA Delivery and Effective Glioblastoma RNAi Therapy.

RNA interference (RNAi) is an emerging therapeutic modality for tumors. However, lack of a safe and efficient small interfering RNA (siRNA) delivery system limits its clinical application. Here, we report a bioreducible and less-cationic siRNA delivery carrier by conjugating Zn(II)-dipicolylamine complexes (Zn-DPA) onto hyaluronic acid (HA) via a redox-sensitive disulfide (-SS-) linker. Such polymer conjugates can formulate stable siRNA nanomedicines via coordination between zinc ions of DPA and the anionic phosphate of siRNA. After the conjugates are taken up by cells, intracellular reduction stimulus subsequently triggers the release of siRNAs and elucidates the desired RNAi effect. Our studies showed the formulated siRNA nanomedicines can be efficiently delivered into tumor cells/tissues and mediates less cytotoxicities both in vitro and in vivo. More importantly, when applied in a xenograft glioblastoma tumor model, this siRNA nanomedicine demonstrated significantly enhanced antitumor ability comparing to naked siRNA. This work demonstrates that such bioreducible Zn-DPA-functionalized HA conjugates without using cationic material as a siRNA carrier represents a promising direction for RNAi-based cancer therapy.

Specific cancer stem cell-therapy by albumin nanoparticles functionalized with CD44-mediated targeting

BackgroundCancer stem cells (CSCs) are highly proliferative and tumorigenic, which contributes to chemotherapy resistance and tumor occurrence. CSCs specific therapy may achieve excellent therapeutic effects, especially to the drug-resistant tumors.ResultsIn this study, we developed a kind of targeting nanoparticle system based on cationic albumin functionalized with hyaluronic acid (HA) to target the CD44 overexpressed CSCs. All-trans-retinoic acid (ATRA) was encapsulated in the nanoparticles with ultrahigh encapsulation efficiency (EE%) of 93% and loading content of 8.37%. TEM analysis showed the nanoparticles were spherical, uniform-sized and surrounded by a coating layer consists of HA. Four weeks of continuously measurements of size, PDI and EE% revealed the high stability of nanoparticles. Thanks to HA conjugation on the surface, the resultant nanoparticles (HA-eNPs) demonstrated high affinity and specific binding to CD44-enriched B16F10 cells. In vivo imaging revealed that HA-eNPs can targeted accumulate in tumor-bearing lung of mouse. The cytotoxicity tests illustrated that ATRA-laden HA-eNPs possessed better killing ability to B16F10 cells than free drug or normal nanoparticles in the same dose, indicating its good targeting property. Moreover, HA-eNPs/ATRA treatment decreased side population of B16F10 cells significantly in vitro. Finally, tumor growth was significantly inhibited by HA-eNPs/ATRA in lung metastasis tumor mice.ConclusionsThese results demonstrate that the HA functionalized albumin nanoparticles is an efficient system for targeted delivery of antitumor drugs to eliminate the CSCs.

Oral siRNA delivery using dual transporting systems to efficiently treat colorectal liver metastasis

Oral siRNA delivery is an ideal way to translate siRNA therapeutic effects in the clinic due to its ability to be administered in convenient and multiple dosages. However, an effective oral delivery system requires overcoming both a hostile gastrointestinal (GI) environment and non-specific targeting. Here, an HTsRP-NC system is a new oral siRNA delivery system consisting of a siRNA/protamine (sRP) nano-complex protected by a multi-functional hyaluronic acid-taurocholic acid (HA-TCA) conjugate. The HTsRP-NC promotes cell penetration and enhances endosomal escape in cancer cells. Moreover, protection of the sRP by HA-TCA from the hostile GI environment helps the AKT siRNA complex to reach the liver through the utilization of a TCA-mediated enterohepatic bile acid recycling system. AKT siRNA was released by 90% in presence of hyaluronidase in the tumor cells which indicate the potential use of HTsRP-NCs for siRNA delivery to treat tumor. After HA receptor (CD44)-mediated cellular uptake of the HTsRP-NC by the liver cancer cells, functional expression of AKT siRNA leads to the suppression of metastatic liver cancer growth in a colorectal liver metastasis (CLM) murine model. Tumor nodules were reduced by more than 1 mm size compared to control group and tumor cells were suppressed by 50% after HTsRP-NCs treatment with AKT siRNA. Overall, oral administration of the HTsRP-NC supports its potential in therapeutic applications for the effective treatment of CLM.

Hyaluronan magnetic nanoparticle for mitoxantrone delivery toward CD44-positive cancer cells.

Hyaluronic acid (HA) is increasingly investigated for biomedical applications such as regenerative medicine, aesthetic medicine, and drug delivery. Accordingly, conjugation of HA to PEGylated MNPs could increase the active targeting ability of nano-drug carriers toward CD44 receptors and be useful in a clinical setting such as drug delivery. So, we chemically conjugated mitoxantrone (MTX) to HA-PEGylated MNPs to use concurrent advantages such as prolong the circulation time, decrease the side effects and delivery toward special tumor cells. Size of the Fe3O4-DPA-PEG-HA-MTX NPs was determined ∼200 nm utilizing FESEM and DLS. Stability analysis confirmed that prepared MNPs were stable in physiological conditions even after 8 days and only 47.3% of MTX was liberated from nanocarriers, in the event that, acidic condition and also presence of protease enzyme accelerated the amount of MTX release to 100% after 8 days of incubation. The in vitro cytotoxicity analysis by MTT assay revealed that viable cell numbers were reduced by 32% when MTX-HA-MNPs were applied against MDA-MB-231 cell lines, while they showed significant decreased cellular cytotoxic effects on cell viability in the MCF-7 cell lines which express lower levels of CD44 receptor at the cell surface. Also, results of flow cytometry analysis following 24 h exposure confirmed that MTX-HA-MNPs have significant induction of apoptosis in MDA-MB-231 cell lines (70.3%) which contains high levels of CD44 expression, whereas there was little effect on the induction of apoptosis in MCF-7 cell lines (5%). The obtained binding models through molecular docking simulation related to each single moieties of prepared MNPs clearly confirmed that MTX-HA-MNPs can easily be bonded to the CD44 receptor with more affinity value in comparison to HA ligand, and so conjugation of HA to MNPs can be a good way for MTX delivery toward special tumor cells or tissues.

Hyaluronic Acid Conjugates of TLR7/8 Agonists for Targeted Delivery to Secondary Lymphoid Tissue.

Immunogens carried in lymphatic fluid drain via afferent vessels into regional lymph nodes and facilitate the efficient induction of appropriate immune responses. The lymphatic system possesses receptors recognizing hyaluronic acid (HA). Covalent conjugates of small-molecule TLR7/8 agonists with HA are entirely devoid of immunostimulatory activity in vitro. In murine models of immunization, however, such conjugates traffic to lymph nodes, where they are "unmasked", releasing the small molecule TLR7/8 agonist from the carrier polysaccharide. The resulting focal immunostimulation is manifested in potent adjuvantic effects with negligible systemic exposure. The efficient delivery of immunogens has been a major challenge in the development of subunit vaccines, and enhancing targeted delivery of immunogens to secondary lymphoid organs might be a promising approach for improving vaccine efficacy, as well as safety.

Abstract 5811: Evaluation of antitumor efficacy of CA102N, H-Nim and hyaluronic acid (HA) conjugate, alone or in combination with 5-fluorouracil (5-FU) in in vivo human colorectal cancer models

Abstract CA102N is a conjugate of HA and H-Nim (N-(4-Amino-2-phenoxyphenyl, methanesulfonamide), a derivative of cyclooxygenase-2 inhibitor Nimesulide, and a CD44 receptor targeted system currently in preclinical development for metastatic colorectal cancer. The in vivo antitumor activities of CA102N was evaluated singly or combined with 5-FU in CR5038 (Patient Derived Xenograft (PDX) and HT29 Xenograft models. Inhibitory activity was assessed according to tumor-volume changes. CA102N (i.v. 200mg/kg, BIW) monotherapy did not show significant tumor growth inhibition at either the end of the treatment (21 days) or prolonged observational (45 days) period, in CR5038 PDX-SCID metastatic model in comparison to vehicle control or 5-FU (i.v. 50mg/kg, QW). Notably, the combination therapy displayed a marked reduction in tumor growth (>70%) compared to 5-FU alone (p=0.0003) or vehicle control (p<0.0001) in addition to prolonged median survival (p<0.05). Alternatively, HT29 xenografts nude mice were found to have significantly reduced tumor burden treated with either single agents CA102N (400 mg/kg, QW) or 5-FU (50 mg/kg, QW), (p<0.012) compared to control vehicle. The drug combination further displayed a synergistic antitumoral effect and greater median survival (p <0.05) in HT29 model. CA102N was well tolerated alone or in the combination therapy. Based on the results of this study, combined treatment of CA102N and 5-FU resulted in a complete response at the regimen given. The combination of CA102N and 5-FU represents a promising potential therapeutic strategy and a new approach to reverse drug resistant in the treatment of colorectal cancer. Studies presently are ongoing to further evaluate its efficacy in CT26 murine colorectal models. Citation Format: Eskouhie Tchaparian, Louis Lin. Evaluation of antitumor efficacy of CA102N, H-Nim and hyaluronic acid (HA) conjugate, alone or in combination with 5-fluorouracil (5-FU) in in vivo human colorectal cancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5811.

Magnetic resonance imaging contrast enhancement in vitro and in vivo by octanuclear iron-oxo cluster-based agents

A water-soluble octanuclear cluster, [Fe8], was studied with regard to its properties as a potential contrast enhancing agent in magnetic resonance imaging (MRI) in magnetic fields of 1.3, 7.2 and 11.9 T and was shown to have transverse relaxivities r2 = 4.01, 10.09 and 15.83 mM s−1, respectively. A related hydrophobic [Fe8] cluster conjugated with 5 kDa hyaluronic acid (HA) was characterized by 57Fe-Mössbauer and MALDI-TOF mass spectroscopy, and was evaluated in aqueous solutions in vitro with regard to its contrast enhancing properties [r2 = 3.65 mM s−1 (1.3 T), 26.20 mM s−1 (7.2 T) and 52.18 mM s−1 (11.9 T)], its in vitro cellular cytotoxicity towards A-549 cells and COS-7 cells and its in vivo enhancement of T2-weighted images (4.7 T) of a human breast cancer xenografted on a nude mouse. The physiologically compatible [Fe8]-HA conjugate was i.v. injected to the tumor-bearing mouse, resulting in observable, heterogeneous signal change within the tumor, evident 15 min after injection and persisting for approximately 30 min. Both molecular [Fe8] and its HA-conjugate show a strong magnetic field dependence on r2, rendering them promising platforms for the further development of T2 MRI contrast agents in high and ultrahigh magnetic fields.

Blood-clotting mimetic behavior of biocompatible microgels

Recent advances in hydrogel chemistry have led to the development of various soft materials capable of self-healing. Nevertheless, their self-healing capabilities are primitive compared to the responsive and adaptive strategies of blood clotting and wound healing in the human body. We developed a novel microgel system that mimics the process of blood clotting. Electrospun polylactic acid (PLA) fibers were entrapped inside the microgels of a hyaluronic acid conjugate with hesperidin side groups crosslinked by Fe ions; the resulting hydrogels showed fast self-healing and magnetic responsive properties. By applying an external magnetic field, which mimicked blood flow, the microgels successfully aggregated at target sites, like platelets. The aggregates were stable, as demonstrated by resistance to sonication for 30min, and their moduli spanned tens to hundreds of kPa, demonstrating the mechanical integrity of the artificial clots. Like fibrin, the PLA fibers successfully strengthened the aggregates due to formation of uniform fiber-reinforced hydrogels; the artificial clots containing fibers had a 300% improved modulus and 50% increased hardness relative to the hydrogels without fibers. This unique intelligent system can be utilized in future self-healing systems, delivery systems, and devices with microfluidic channels.

PH-Sensitive Coiled-Coil Peptide-Cross-Linked Hyaluronic Acid Nanogels: Synthesis and Targeted Intracellular Protein Delivery to CD44 Positive Cancer Cells.

The clinical translation of protein drugs that act intracellularly is limited by the absence of safe and efficient intracellular protein delivery vehicles. Here, pH-sensitive coiled-coil peptide-cross-linked hyaluronic acid nanogels (HA-cNGs) were designed and investigated for targeted intracellular protein delivery to CD44 overexpressing MCF-7 breast cancer cells. HA-cNGs were obtained with a small size of 176 nm from an equivalent mixture of hyaluronic acid conjugates with GY(EIAALEK)3GC (E3) and GY(KIAALKE)3GC (K3) peptides, respectively, at pH 7.4 by nanoprecipitation. Circular dichroism (CD) proved the formation of coiled-coil structures between E3 and K3 peptides at pH 7.4 while fast uncoiling at pH 5.0. HA-cNGs showed facile loading of cytochrome C (CC) and greatly accelerated CC release under mild acidic conditions (18.4%, 76.8%, and 91.4% protein release in 24 h at pH 7.4, 6.0, and 5.0, respectively). Confocal microscopy and flow cytometry displayed efficient internalization of CC-loaded HA-cNGs and effective endosomal escape of CC in MCF-7 cancer cells. Remarkably, HA-cNGs loaded with saporin, a ribosome inactivating protein, exhibited significantly enhanced apoptotic activity to MCF-7 cells with a low IC50 of 12.2 nM. These coiled-coil peptide-cross-linked hyaluronic acid nanogels have appeared as a simple and multifunctional platform for efficient intracellular protein delivery.

Targeted delivery of SNX-2112 by polysaccharide-modified graphene oxide nanocomposites for treatment of lung cancer

Graphene oxide (GO) is a promising material for biomedical applications, particularly in drug delivery, due to its exceptional chemical and physical properties. In this work, an innovative GO-based carrier was developed by modifying GO with chitosan (CHI) to improve the biocompatibility, and followed by the conjugation of hyaluronic acid (HA), the target ligand for CD44, to realize the specific recognition of tumor cells and improve the efficiency of anti-tumor drug delivery. The resulting product GO-CHI-HA was loaded with an anti-cancer drug SNX-2112, which is the Hsp90 inhibitor. The total release amount and release rate of SNX-2112 were significantly higher in acidic condition than in physiological condition. GO-CHI-HA with a low concentration had little impact on the lysis of red blood cells (RBCs) and blood coagulation and showed low toxicity in A549 cells and NHBE cells. The GO-CHI-HA/SNX-2112 proved to be effective in inhibiting and killing A549 cells while having lower cytotoxicity against normal human bronchial epithelial cells (NHBE cells). Furthermore, in vivo toxicity of the materials towards vital organs in SD rats were also studied through histological examinations and blood property analyses, the results of which showed that although inflammatory response was developed in the short-term, GO-CHI-HA/SNX-2112 caused no severe long-term injury. Therefore, this drug delivery system showed great potential as an effective and safe drug delivery system with little adverse side effects for cancer therapy.

Self-assembled thermoresponsive nanostructures of hyaluronic acid conjugates for osteoarthritis therapy.

Under pathological conditions, joints and skin are often affected by an imbalance in the breakdown and production of hyaluronic acid (HA). The unique biochemical and biomechanical properties provided by HA must be restored for the long-term lubrication and cushioning effects. To overcome the inconvenience of repeated injections and the rapid degradation of exogenous HA treatments, HA is conjugated to a thermosensitive polymer, enabling the spontaneous formation of nanoparticles (HA Nano) at body temperature. Three HA Nano preparations are tested for their injectability, sensitivity to enzymatic degradation and cytocompatibility. One of them is delivered via subcutaneous and intra-articular injections to healthy mice and tested in a murine osteoarthritis (OA) model. It is found to be biocompatible, to offer a prolonged residence time at the injection site, have the ability to protect cartilage, to reduce pro-inflammatory cytokines and to preserve epiphysis thickness. In this study, HA Nano spontaneously forms nanoparticles at body temperature in vivo and is a promising candidate for the next generation of the sustainable/long-lasting treatment of OA and potentially also dermatological conditions.

Core-shell patterning of synthetic hydrogels via interfacial bioorthogonal chemistry for spatial control of stem cell behavior.

A new technique is described for the patterning of cell-guidance cues in synthetic extracellular matrices (ECM) for tissue engineering applications. Using s-tetrazine modified hyaluronic acid (HA), bis-trans-cyclooctene (TCO) crosslinkers and monofunctional TCO conjugates, interfacial bioorthogonal crosslinking was used to covalently functionalize hydrogels as they were synthesized at the liquid-gel interface. Through temporally controlled introduction of TCO conjugates during the crosslinking process, the enzymatic degradability, cell adhesivity, and mechanical properties of the synthetic microenvironment can be tuned with spatial precision. Using human mesenchymal stem cells (hMSCs) and hydrogels with a core-shell structure, we demonstrated the ability of the synthetic ECM with spatially defined guidance cues to modulate cell morphology in a biomimetic fashion. This new method for the spatially resolved introduction of cell-guidance cues for the establishment of functional tissue constructs complements existing methods that require UV-light or specialized equipment.