Hyaluronic Acid Conjugates Research Articles

Development of a dexamethasone-hyaluronic acid conjugate with selective targeting effect for acute lung injury therapy

Acute lung injury (ALI), a critical complication of COVID-19, is characterized by widespread inflammation and severe pulmonary damage, necessitating intensive care for those affected. Although glucocorticoids (GCs), such as dexamethasone (Dex), have been employed clinically to lower mortality, their nonspecific systemic distribution has led to significant side effects, limiting their use in ALI treatment. In this study, we explored the conjugation of Dex to hyaluronic acid (HA) to achieve targeted delivery to inflamed lung tissues. We achieved a conjugation efficiency exceeding 98 % using a cosolvent system, with subsequent ester bond cleavage releasing the active Dex, as verified by liquid chromatography. Biodistribution and cellular uptake studies indicated the potential of the HA conjugate for cluster of differentiation 44 (CD44)-mediated targeting and accumulation. In a lipopolysaccharide-induced ALI mouse model, intravenous (IV) HA-Dex administration showed superior anti-inflammatory effects compared to free Dex administration. Flow cytometry analysis suggested that the HA conjugate preferentially accumulated in lung macrophages, suggesting the possibility of reducing clinical Dex dosages through this targeted delivery approach.

Sustainable Chemical Modification of Natural Polysaccharides: Mechanochemical, Solvent-Free Conjugation of Pectins and Hyaluronic Acid Promoted by Microwave Radiations.

The modern chemistry has the main focus of saving resources and developing synthetic strategies characterized by intrinsic efficiency, ease and safety in operation, short reaction time, reduced energy, and waste. Natural polysaccharides are largely distributed in plant/animal cells; in other words, they are often provided by renewable sources. This characteristic makes them suitable compounds to be investigated for their employment as biodegradable material. In addition, natural polysaccharides have been proven to have a wide range of applications, and this prompted researchers to investigate their chemical modifications in order to modulate their properties. Herein we discuss the development of conjugation strategies of some polysaccharides with natural substrates and the effects of the structural modification on their bioactivities. Finally, this work intends to provide suggestions and perspectives on the development of safe and sustainable synthetic processes on polysaccharides.

Hyaluronic acid conjugates of glycine peptides and L-tryptophan

This work reports about the conjugation of glycine C-terminal ethyl and methyl ester peptides and L-tryptophan methyl ester with sodium hyaluronate in aqueous solutions using the peptide coupling agent DMTMM (or short DMT, 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methyl-morpholinium chloride). Detailed infrared (IR) absorbance and 1H and 13C (2D) NMR studies (heteronuclear multi-bond correlation spectroscopy, HMBC) confirmed covalent and regioselective amide bonds with the D-glucuronate, but also proves the presence of DMT traces in all conjugates. The ethyl ester`s methyl protons on the peptides` C-terminal could be used to quantify the degree of substitution of the peptide on the hyaluronate scaffold by NMR. The ester group also proved stable during conjugation and work-up, and could in some cases be selectively cleaved in water whilst leaving the amide bond intact as shown by potentiometric charge titration, NMR and IR. The conjugates did not influence the capability of human umbilical vein endothelial cells (HUVECs) to reduce MTS (5-[3-(carboxymethoxy)phenyl]-3-(4,5-dimethyl-2-thiazolyl)-2-(4-sulfophenyl)-2H-tetrazolium inner salt) to a formazan dye, which points towards a low cytotoxicity for the obtained products. The conjugation method and products could be tested for tissue engineering gels or drug delivery purposes with alternative, biologically active peptides.

Characterization of a Delivery System Based on a Hyaluronic Acid 3D Scaffold and Gelatin Microparticles.

The objective of this study was to develop and characterize a novel hyaluronic acid (HA) 3D scaffold integrated with gelatin microparticles for sustained-delivery applications. To achieve this goal, the delivery microparticles were synthesized and thoroughly characterized, focusing on their crosslinking mechanisms (vanillin and genipin), degradation profiles, and release kinetics. Additionally, the cytotoxicity of the system was assessed, and its impact on the cell adhesion and distribution using mouse fibroblasts was examined. The combination of both biomaterials offers a novel platform for the gradual release of various factors encapsulated within the microparticles while simultaneously providing cell protection, support, and controlled factor dispersion due to the HA 3D scaffold matrix. Hence, this system offers a platform for addressing injure repair by continuously releasing specific encapsulated factors for optimal tissue regeneration. Additionally, by leveraging the properties of HA conjugates with small drug molecules, we can enhance the solubility, targeting capabilities, and cellular absorption, as well as prolong the system stability and half-life. As a result, this integrated approach presents a versatile strategy for therapeutic interventions aimed at promoting tissue repair and regeneration.

Inhibiting the cGAS-STING Pathway in Ulcerative Colitis with Programmable Micelles.

Ulcerative colitis is a chronic condition in which a dysregulated immune response contributes to the acute intestinal inflammation of the colon. Current clinical therapies often exhibit limited efficacy and undesirable side effects. Here, programmable nanomicelles were designed for colitis treatment and loaded with RU.521, an inhibitor of the cyclic GMP-AMP synthase-stimulator of interferon genes (cGAS-STING) pathway. STING-inhibiting micelles (SIMs) comprise hyaluronic acid-stearic acid conjugates and include a reactive oxygen species (ROS)-responsive thioketal linker. SIMs were designed to selectively accumulate at the site of inflammation and trigger drug release in the presence of ROS. Our in vitro studies in macrophages and in vivo studies in a murine model of colitis demonstrated that SIMs leverage HA-CD44 binding to target sites of inflammation. Oral delivery of SIMs to mice in both preventive and delayed therapeutic models ameliorated colitis's severity by reducing STING expression, suppressing the secretion of proinflammatory cytokines, enabling bodyweight recovery, protecting mice from colon shortening, and restoring colonic epithelium. In vivo end points combined with metabolomics identified key metabolites with a therapeutic role in reducing intestinal and mucosal inflammation. Our findings highlight the significance of programmable delivery platforms that downregulate inflammatory pathways at the intestinal mucosa for managing inflammatory bowel diseases.

ROS-responsive sprayable hydrogel as ROS scavenger and GATA6+ macrophages trap for the prevention of postoperative abdominal adhesions

Postoperative abdominal adhesions are a common clinical problem after surgery and can cause many serious complications. Current most commonly used antiadhesion products are less effective due to their short residence time and focus primary on barrier function. Herein, we developed a sprayable hydrogel barrier (sHA-ADH/OHA-E) with self-regulated drug release based on ROS levels at the trauma site, to serve as a smart inflammatory microenvironment modulator and GATA6+ macrophages trap for non-adherent recovery from abdominal surgery. Sulfonated hyaluronic acid (HA) conjugates modified with adipic dihydrazide (sHA-ADH), and oxidized HA conjugates grafted with epigallocatechin-3-gallate (EGCG) via ROS-cleavable boronate bonds (OHA-E) were synthesized. sHA-ADH/OHA-E hydrogel was facilely fabricated within 5 s after simply mixing sHA-ADH and OHA-E through forming dynamic covalent acylhydrazones. With good biocompatibility, appropriate mechanical strength, tunable shear-thinning, self-healing, asymmetric adhesion, and reasonable in vivo retention time, sHA-ADH/OHA-E hydrogel meets the requirements of a perfect physical barrier. Intriguingly, sulfonic acid groups endowed the hydrogel with satisfactory anti-fibroblast and macrophage attachment capability, and were demonstrated for the first time to act as polyanion traps to prevent GATA6+ macrophages aggregation. Importantly, EGCG could be intelligently released by ROS triggering to alleviate oxidative stress and promote proinflammatory M1 macrophage polarize to antiinflammatory M2 phenotype. Further, the fibrinolytic system balance was restored to reduce fibrosis. Thanks to the above advantages, the sHA-ADH/OHA-E hydrogel exhibited excellent anti-adhesion effects in a rat sidewall defect–cecum abrasion model and is expected to be a promising and clinically translatable antiadhesion barrier.

In silico validation of hyaluronic acid – drug conjugates based targeted drug delivery for the treatment of COVID-19

The impact of COVID-19 urges scientists to develop targeted drug delivery to manage Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) viral infections with a fast recovery rate. The aim of the study is to develop Hyaluronic Acid (HA) drug conjugates of viral drugs to target two important enzymes (Mpro and PLpro) of SARS-CoV-2. Three antiviral drugs, namely Dexamethasone (DEX), Favipiravir (FAV), and Remdesivir (REM), were chosen for HA conjugation due to their reactive functional groups. Free forms of drugs (DEX, FAV, REM) and HA drug conjugates (HA-DEX, HA-FAV, HA-REM, HA-RHA, HA-RHE) were validated against Mpro (PDB ID 6LU7) and PLpro (PDB 7LLZ), which play an essential role in the replication and reproduction of the SARS-CoV-2 virus. The results of the present study revealed that HA-drug conjugates possess higher binding affinity and the best docking score towards the Mpro and PLpro target proteins of SARS-CoV-2 than their free forms of drugs. ADMET screening resulted that HA-drug conjugates exhibited better pharmacokinetic profiles than their pure forms of drugs. Further, molecular dynamic simulation studies, essential dynamics and free energy landscape analyses show that HA antiviral drug conjugates possess good trajectories and energy status, with the PLpro target protein (PDB 7LLZ) of SARS-CoV-2 through long-distance (500 ns) simulation screening. The research work recorded the best drug candidate for Cell-Targeted Drug Delivery (CTDD) for SARS-CoV-2-infected cells through hyaluronic acid conjugates of antiviral drugs. Communicated by Ramaswamy H. Sarma

Immunomodulatory and anticytokine therapeutic potential of three Indian spices constituents and its hyaluronic acid conjugates for prevention and post COVID-19 complications: a computational modeling approach.

Targeted drug delivery to SARS-CoV-2 host target proteins for preventing or blocking COVID-19 infection is making serious concern during COVID-19 pandemic and its consequent waves around the globe. People seek reliable, effective folkloric preventive medication for immediate and precautionary relief from COVID-19. These folkloric medicines were effective and saved many patients during the past COVID-19 pandemic. The current research study aims to deliver antiviral Indian spices phytocompounds and their hyaluronic acid conjugates to human host target proteins (ACE-2, TNF-α, IL-6, IL-1β, PAR-1) of SARS-CoV-2 to inhibit virus propagation and also to regulate early clinical complications of COVID-19. Targeted drug delivery of hyaluronic acid conjugated traditional natural bioactive agent produces more effective and fewer side effects in delivering novel drugs to human host proteins of COVID-19. In silico molecular docking study of six phytocompounds from three Indian spices and standard drug atazanavir and its hyaluronic acid conjugates reveals that phytocompounds and its hyaluronic acid conjugates possess high affinity to binding pockets of SARS-CoV-2 human host targets with more binding affinity scores. Most notably HA cyclocurcumin exhibit a docking score -9.9 kcal/mol against ACE-2 (PDB ID 1R42) target protein similarly HA-Hydrazinocurcumin exhibit a docking score -9.8 kcal/mol against PAR-1 (PDB ID 3VW7). ADMET validation of phytocompounds and their hyaluronic acid conjugates reveals its best pharmacokinetic profile over standard antiviral drug especially HA cyclocurcumin conjugate possesses high HIA (86%) and good pharmacokinetic profiles. DFT analysis affirms the reason behind the higher binding affinity of hyaluronic acid conjugates of spices phytocompounds towards all screened target proteins especially HA-hydrazinocurcumin conjugate possess high softness (19.1570 eV) and low hardness (0.0522 eV) values. Finally, MD simulation of best-docked compounds against ACE-2 and PAR-1 target protein revealed that hyaluronic acid conjugates of Indian spices compounds exhibit stable RMSD values and more protein-ligand interactions during simulation than hyaluronic acid conjugates of drug atazanavir.Communicated by Ramaswamy H. Sarma.

A self-assembled, genetically engineered, irradiated tumor cell debris vaccine.

Vaccine-based therapeutics for cancers face several challenges including lack of immunogenicity and tumor escape pathways for single antigen targets. It has been reported that radiotherapy has an in situ vaccine effect that provides tumor antigens following irradiation, helping to activate antigen-presenting cells (APCs). Herein, a new vaccine approach is developed by combining genetically engineered irradiated tumor cell debris (RTD) and hyaluronic acid (HA), termed HA@RTD. A cancer cell line is developed that overexpresses granulocyte-macrophage colony-stimulating factor (GM-CSF). A hydrogel was developed by covalent conjugation of HA with RTD proteins that acted as a potent vaccine system, the effects which were probed with T cell receptor sequencing. The engineered vaccine activated antitumor immunity responses and prevented tumor growth in mice even with a single immunization. HA@RTD vaccine efficacy was also assessed in therapeutic settings with established tumors and in combination with immune checkpoint blockade.

Targeted Delivery of Geraniol via Hyaluronic Acid-Conjugation Enhances Its Anti-Tumor Activity Against Prostate Cancer.

Targeted delivery systems have been developed to improve cancer treatment by reducing side effects and enhancing drug efficacy. Geraniol, a natural product, has demonstrated promising anti-cancer effects in various cancer types, including prostate cancer, which is the most commonly diagnosed cancer in men. Hyaluronic acid (HA), a natural carrier targeting CD44-positive prostate cancer cells, can be utilized in a targeted delivery system. This study investigated the efficacy of a conjugate of HA and geraniol linked via a disulfide bond linker (HA-SS-Geraniol) in prostate cancer. The cytotoxicity of HA-SS-Geraniol was evaluated on human PC-3 prostate cancer cells. Flow cytometry was used to assess its effects on mitochondrial membrane potential, apoptosis, and cell cycle arrest. Additionally, proteomic analysis was conducted to explore the underlying mechanism of action induced by HA-SS-Geraniol treatment. A subcutaneous xenograft tumor model was established in nude mice to evaluate the toxicity and efficacy of HA-SS-Geraniol in vivo. The results demonstrated that HA-SS-Geraniol exhibited potent cytotoxicity against PC-3 prostate cancer cells by inducing mitochondrial membrane potential loss and apoptosis in vitro. The proteomic analysis further supported the hypothesis that HA-SS-Geraniol induces cell death through mitochondria-mediated apoptosis, as evidenced by differential protein expression. The in vivo mouse model confirmed the safety of HA-SS-Geraniol and its ability to inhibit tumor growth. HA-SS-Geraniol holds promise as a biologically safe and potentially effective therapeutic agent for prostate cancer treatment. Its targeted delivery system utilizing HA as a carrier shows potential for improving the efficacy of geraniol in cancer therapy.

Improving Cancer Targeting: A Study on the Effect of Dual-Ligand Density on Targeting of Cells Having Differential Expression of Target Biomarkers.

Silica nanoparticles with hyaluronic acid (HA) and folic acid (FA) were developed to study dual-ligand targeting of CD44 and folate receptors, respectively, in colon cancer. Characterization of particles with dynamic light scattering showed them to have hydrodynamic diameters of 147-271 nm with moderate polydispersity index (PDI) values. Surface modification of the particles was achieved by simultaneous reaction with HA and FA and results showed that ligand density on the surface increased with increasing concentrations in the reaction mixture. The nanoparticles showed minimal to no cytotoxicity with all formulations showing ≥ 90% cell viability at concentrations up to 100 µg/mL. Based on flow cytometry results, SW480 cell lines were positive for both receptors, the WI38 cell line was positive for CD44 receptor, and Caco2 was positive for the folate receptor. Cellular targeting studies demonstrated the potential of the targeted nanoparticles as promising candidates for delivery of therapeutic agents. The highest cellular targeting was achieved with particles synthesized using folate:surface amine (F:A) ratio of 9 for SW480 and Caco2 cells and at F:A = 0 for WI38 cells. The highest selectivity was achieved at F:A = 9 for both SW480:WI38 and SW480:Caco2 cells. Based on HA conjugation, the highest cellular targeting was achieved at H:A = 0.5-0.75 for SW480 cell, at H:A = 0.75 for WI38 cell and at H:A = 0.5 for Caco2 cells. The highest selectivity was achieved at H:A = 0 for both SW480:WI38 and SW480:Caco2 cells. These results demonstrated that the optimum ligand density on the nanoparticle for targeting is dependent on the levels of biomarker expression on the target cells. Ongoing studies will evaluate the therapeutic efficacy of these targeted nanoparticles using in vitro and in vivo cancer models.

A mitochondria-targeting ROS-activated nanoprodrug for self-augmented antitumor oxidation therapy.

Mitochondrion is an ideal target for amplifying ROS attack in antitumor treatment. Benefiting from distinctive properties of mitochondria, the precise delivery of ROS generator to mitochondria could maximumly utilize ROS for oxidation therapy. Herein, we prepared an innovative ROS-activatable nanoprodrug (HTCF) which dually targets tumor cells and mitochondria for antitumor therapy. Cinnamaldehyde (CA) was conjugated to ferrocene (Fc) and triphenylphosphine by thioacetal linker, to synthesize mitochondria-targeting ROS-activated prodrug (TPP-CA-Fc), which subsequently self-assembled into nanoprodrug via host-guest interactions between TPP-CA-Fc and cyclodextrin-decorated hyaluronic acid conjugate. Under mitochondrial high ROS condition, especially in tumor cells, HTCF selectively initiate in-situ Fenton reaction to catalyze H2O2 into highly cytotoxic •OH, ensuring maximum generation and utilization of •OH for precision CDT. Meanwhile, the mitochondrial high ROS trigger thioacetal bond cleavage and CA release. The released CA stimulate mitochondrial oxidative stress aggravation and H2O2 regeneration, which in turn react with Fc for more •OH generation, forming self-amplifying positive feedback cycle of CA release and ROS burst. With self-augmented Fenton reaction and mitochondria-specific destruction, HTCF ultimately induce intracellular ROS burst and severe mitochondrial dysfunction for amplified ROS-mediated antitumor therapy. Such an ingenious organelles-specialized nanomedicine exhibited prominent antitumor effect both in vitro and in vivo, revealing underlying perspectives to amplify tumor-specific oxidation therapy.

What Is Wrong with Hyaluronic Acid Chemistry? A 15N/13C Solid-State NMR Re-Evaluation of Its Dopamine Conjugates

In this work, a systematic 15N/13C solid-state NMR investigation is performed on three dopamine (DA) conjugates of hyaluronic acid, considered in both its native (HA) and NaIO4-oxidized (HAOx) forms. Two of them, here named HAEDC-DA and HAOx-DA, have been previously introduced as covalent conjugates involving DA amine nitrogen: the former by EDC-mediated amide bond formation, and the latter by reaction of the Schiff base with the aldehyde moieties presumed to exist in HAOx. The third conjugate, HA-DA, is reported here for the first time; it is obtained by simply mixing hyaluronan with DA∙HCl at pH 5. The 15N ss-NMR spectra were found to be consistent in all the systems, and the DA molecules were found to be in their charged -NH3+ form, which contradicts the HAEDC-DA/HAOx-DA covalent bonding schemes proposed in the literature. The 13C ss-NMR results add useful new insights into the structure and interaction patterns of the conjugates. All of our findings are relevant for future practical applications, for instance in developing novel HA-based hydrogels. In addition, the present study demonstrates the importance of using the most appropriate analytical tools when investigating composite systems due to the complexity of hyaluronic acid conjugates. Solid-state NMR proved essential to answering the question in the title: actually, there is nothing wrong with hyaluronic acid chemistry; the claimed covalent bonds between DA and the HA(HAOx) chain do not exist in these systems, because the conditions for their formation do not hold in practice.

Formulation and optimization of aceclofenac loaded hyaluronic-oleic acid based micellar gel for the management of osteoarthritis

Aceclofenac is a non-steroidal anti-inflammatory drug used to manage osteoarthritis. However, its limited solubility, short half-life, and gastrointestinal side effects on chronic use limit its delivery via the oral route. This study used hyaluronic acid tailored with oleic acid via esterification to formulate micelles for aceclofenac topical delivery. The prepared micelles were characterized for average particle size, size distribution, surface charge, drug loading, entrapment efficiency, and surface morphology. The drug loading, entrapment efficiency, mean particle size, and zeta potential of micelles were 8.21 ± 0.3%, 81.25 ± 2.98%, 245.93 ± 7.68 nm, and −17.6 ± 1.95 mV, respectively. The prepared micelles were found to be spherical. The micelles were further loaded in chitosan gel and evaluated for drug release, skin permeation, and deposition. The hydrogel showed shear thinning behavior, sustained drug release, enhanced skin permeation, and deposition of aceclofenac. Further, the pharmacodynamic study revealed a significant reduction in pain and inflammatory response with an improved radiological and histopathological condition in the animal models. These results show the potential of hyaluronic acid oleic acid conjugate micellar gel as a promising carrier to deliver the poorly soluble small molecules like aceclofenac topically to manage the inflammatory condition.

Abstract 2007: Evaluation of the antitumor activity of ca102l, a hyaluronic acid and lenalidomide conjugate, in multiple myeloma

Abstract Multiple Myeloma (MM) is a rare and incurable hematological malignancy. Despite emerging treatments, there is still an unmet medical need for novel therapeutics to target the inherent and acquired drug resistance associated with MM. CA102L is a conjugate of hyaluronic acid and an anticancer drug designed to deliver its payload, Lenalidomide (Len), to MM cells via the CD44 receptor. Lenalidomide is used as a front-line therapy in MM; The CD44 receptor is highly expressed in MM and is associated with advanced clinical stages and poor survival. In addition to the enhancement of the therapeutic efficacy of Len, a reduction in off-target toxicity is suggested. The present study was designed to evaluate the efficacy of CA102L in MM and identify its potential mechanism of action. The antitumor efficacy of CA102L was tested (in vitro and in vivo) in two MM cancer models, IM-9 and NCI-H929. Protein expression was used to identify the potential targets of CA102L in MM. Both in in vitro and in vivo investigations, CA102L as a single agent displayed an improvement in potency in MM models. In CB-17 SCID mice bearing either IM-9 or NCI-H929 cells, tumor growth inhibition was significantly higher (P&lt 0.05) in the CA102L (25 mg/kg) treated group compared to that of Len (25 mg/kg). Further, CD31 and hematoxylineosin (H&E) staining of tumor sections from the IM-9 xenograft mice treated with CA102L resulted in greater antiangiogenic and antiproliferative effects compared to the other groups. Prolonged survival was also observed in CA102L monotherapy in H929 models relative to Len (56% vs 11%). Interestingly, in H929 cell viability studies, CA102L and Len at 400μM showed comparable cytotoxicity with a 60% cell growth inhibition, while superior activity was noted against IM-9-treatments (CA102L 60%, Len 20%). Western blot studies confirmed CRBN as the primary target of CA102L. In summary CA102L as a single agent, exhibited greater efficacy than Len in preclinical MM models, thus representing a promising potential therapeutic strategy for MM. Studies presently are ongoing to further evaluate CA102L’s preclinical profile to advance the candidate into the clinic. Citation Format: Eskouhie Tchaparian, Star Lin. Evaluation of the antitumor activity of ca102l, a hyaluronic acid and lenalidomide conjugate, in multiple myeloma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2007.

Development of triamcinolone acetonide-hyaluronic acid conjugates with selective targeting and less osteoporosis effect for rheumatoid arthritis treatments

Rheumatoid arthritis (RA) is a common systemic autoimmune disease in developed countries. In clinical treatment, steroids have been used as bridging and adjunctive therapy after disease-modifying anti-rheumatic drug administration. However, the severe side effects caused by the nonspecific targeting of organs followed by long-term administration have limited their usage in RA. In this study, poorly water-soluble triamcinolone acetonide (TA), a highly potent corticosteroid for intra-articular injection, is conjugated on hyaluronic acid (HA) for intravenous purposes with increased specific drug accumulation in inflamed parts for RA. Our results demonstrate that the designed HA/TA coupling reaction reveals >98 % conjugation efficiency in the dimethyl sulfoxide/water system, and the resulting HA-TA conjugates show lower osteoblastic apoptosis compared with that in free TA-treated osteoblast-like NIH3T3 cells. Furthermore, in a collagen-antibody-induced arthritis animal study, HA-TA conjugates enhanced the initiative targeting ability to inflame tissue and reduce the histopathological arthritic changes (score = 0). Additionally, the level of bone formation marker P1NP in HA-TA-treated ovariectomized mice (303.6 ± 40.6 pg/mL) is significantly higher than that in the free TA-treated group (143.1 ± 3.9 pg/mL), indicating the potential for osteoporotic reduction using an efficient HA conjugation strategy for the long-term administration of steroids against RA.

Combination of hyaluronic acid conjugates with immunogenic cell death inducer and CpG for glioblastoma local chemo-immunotherapy elicits an immune response and induces long-term survival

The efficacy of standard glioblastoma (GBM) treatments has been limited due to the highly immunosuppressive tumor immune microenvironment, interpatient tumor heterogenicity and anatomical barriers, such as the blood brain barrier. In the present work, we hypothesized that a new local therapy based on the combination of doxorubicin (DOX) as an immunogenic cell death (ICD) inducer and CpG, a Toll-like receptor (TLR)-9 agonist, would act synergistically to eradicate GBM. DOX and CpG were first tested in an orthotopic GL261 GBM model showing enhanced survival. To improve the outcome with a reduced dose, we designed bioresponsive hyaluronic acid (HA)-drug conjugates for effective in situ chemoimmunotherapy. HA was derivatized with CpG. The new HA-CpG conjugate showed high efficacy in re-educating protumoral M2-like microglia into an antitumoral M1-like phenotype, inducing the expression of immune-stimulatory cytokines. DOX was also conjugated to HA. DOX conjugation increased ICD induction in GL261 cells. Finally, a combination of the conjugates was explored in an orthotopic GL261 GBM model. The local delivery of combined HA-DOX + HA-CpG into the tumor mass elicited antitumor CD8+ T cell responses in the brain tumor microenvironment and reduced the infiltration of M2-like tumor-associated macrophages and myeloid-derived suppressor cells. Importantly, the combination of HA-DOX and HA-CpG induced long-term survival in >66% of GBM-bearing animals than other treatments (no long-term survivor observed), demonstrating the benefits of conjugating synergistic drugs to HA nanocarrier. These results emphasize that HA-drug conjugates constitute an effective drug delivery platform for local chemoimmunotherapy against GBM and open new perspectives for the treatment of other brain cancers and brain metastasis.

Ultrasound-mediated delivery of flexibility-tunable polymer drug conjugates for treating glioblastoma.

Effective chemotherapy delivery for glioblastoma multiforme (GBM) is limited by drug transport across the blood-brain barrier and poor efficacy of single agents. Polymer-drug conjugates can be used to deliver drug combinations with a ratiometric dosing. However, the behaviors and effectiveness of this system have never been well investigated in GBM models. Here, we report flexible conjugates of hyaluronic acid (HA) with camptothecin (CPT) and doxorubicin (DOX) delivered into the brain using focused ultrasound (FUS). In vitro toxicity assays reveal that DOX-CPT exhibited synergistic action against GBM in a ratio-dependent manner when delivered as HA conjugates. FUS is employed to improve penetration of DOX-HA-CPT conjugates into the brain in vivo in a murine GBM model. Small-angle x-ray scattering characterizations of the conjugates show that the DOX:CPT ratio affects the polymer chain flexibility. Conjugates with the highest flexibility yield the highest efficacy in treating mouse GBM in vivo. Our results demonstrate the association of FUS-enhanced delivery of combination chemotherapy and the drug-ratio-dependent flexibility of the HA conjugates. Drug ratio in the polymer nanocomplex may thus be employed as a key factor to modulate FUS drug delivery efficiency via controlling the polymer flexibility. Our characterizations also highlight the significance of understanding the flexibility of drug carriers in ultrasound-mediated drug delivery systems.

Comparative study of dual delivery of gemcitabine and curcumin using CD44 targeting hyaluronic acid nanoparticles for cancer therapy

Dual-delivery nanoparticles of gemcitabine (GEM) and curcumin (CUR) conjugated on hyaluronic acid (HA) were successfully developed for targeting to cancer cells. The polymer-drug conjugates (PDCs) were synthesized by grafting GEM onto CUR (hydrazone)-HA (ChH) conjugates using ester (non-pH-sensitive) and hydrazone (pH-sensitive) bonds to form two different chemical structures; random graft: GeChH and block graft: (GhC)hH, respectively. The GeChH and (GhC)hH nanoparticles were prepared by solvent diffusion evaporation. The (GhC)hH nanoparticles showed superior characteristics with smaller particle size (221 nm), narrower size distribution (PDI 0.01) and lower critical aggregation concentration (CAC 0.28 mg/mL), as compared to the GeChH nanoparticles. The release profile of GEM and CUR from both nanoparticles was specific and fast in the acidic microenvironment (pH 5.0–6.5) while retarded at physiological pH (pH 7.4), indicating pH-dependence. In particular, the GEM released from (GhC)hH nanoparticles at pH 7.4 was less than that from the GeChH nanoparticles, suggesting the suitability for anticancer drug delivery. In vitro cytotoxicity study revealed that the (GhC)hH nanoparticles had higher toxicity and synergistic effect in all cell lines (PANC-1, A549, HCT116 and Caco-2 cells) when compared to the GeChH nanoparticles and the GEM/CUR solution. The in vitro cellular uptake study supported that both nanoparticles could be internalized into the cells and deposited around the nuclei. Moreover, the %uptake of the (GhC)hHFITC nanoparticles was higher than that of the GeChHFITC nanoparticles in both A549 and HCT116 cells. After HA blockade treatment, the %uptake of the nanoparticles significantly decreased, indicating that HA targeting to CD44 receptors played an important role to enhance specificity to the cancer cells. In summary, the developed (GhC)hH nanoparticles could provide dual delivery of GEM and CUR which would be beneficial for cancer therapy.

Hyaluronic acid and cholecalciferol conjugate based nanomicelles: Synthesis, characterization, and cytotoxicity against MCF-7 breast cancer cells

Recently, growing and conclusive evidences showed the particularly close ties between the vitamin D (VD) and breast cancer (BC). However, few researches were focused on the delivery system against breast cancer which based on cholecalciferol (VD3). Herein, we designed a series of self-assembled micelles. The conjugate (HA-VD) of VD3 with hyaluronic acid (HA) actively targeting breast cancer can efficiently delivery doxorubicin (DOX). The HA-VD conjugate was successfully synthesized, which was confirmed using 1H nuclear magnetic resonance (NMR) and Fourier transform infrared (FITR). Preliminary studies of its physical and chemical properties indicated that VD3 was successfully grafted with the HA (HAVD). When the molar ratio was 1:2 and the degree of substitution (DS) was 18.6%, the minimum critical micelle concentration (CMC) value was 0.0137 mg/mL. For DOX-loaded HAVD micelles (DOX/HAVD), the drug loading (DL) was 6.2% and drug encapsulation efficiency (EE) was 79.5%. The DOX/HAVD micelles remain stable in serum for 48 h. The in vitro release rate of DOX decreased after encapsulating compared with that without HAVD encapsulation. The decrease with IC50 of DOX/HAVD micelle was significantly stronger than that of free DOX (p < 0.05) and blank micelles (p < 0.01). At 50% cell inhibition rate, VD3 and DOX acted synergistically on BC cells. Hence, the HAVD micelles can significantly improve the therapeutic effect of DOX against BC cells. In a word, the DOX/HAVD micelles have great potential in treating breast cancer.