Double Emulsion Research Articles

Folate receptor-mediated delivery system based on chitosan coated polymeric nanoparticles for combination therapy of breast cancer

Combination therapy using two or more drugs with different mechanisms of action is an effective strategy for treating cancer. This is because of the synergistic effect of complementary drugs that enhances their effectiveness. However, this approach has some limitations, such as non-specific distribution of the drugs in the tumor and the occurrence of dose-dependent toxicity to healthy tissues. To overcome these issues, we have developed a folate receptor-mediated co-delivery system that improves the access of chemotherapy drugs to the tumor site. We prepared a nanoplatform by encapsulating paclitaxel (PTX) and curcumin (CUR) in poly(caprolactone)-poly(ethylene glycol)-poly(caprolactone) (PCL-PEG-PCL) co-polymer using a double emulsion method and coating nanoparticles with pH-responsive chitosan-folic acid (CS-FA) conjugate. The nanocarrier’s physicochemical properties were studied, confirming successful preparation with appropriate size and morphology. PTX and CUR could be released synchronously in a controlled and acid-facilitated manner. The dual drug-loaded nanocarrier exhibited excellent anti-tumor efficiency in MDA-MB-231 cells in vitro. The active targeting effect of FA concluded from the high inhibitory effect of dual drug-loaded nanocarrier on MDA-MB-231 cells, which have overexpressed folate receptors on their surface, compared to Human umbilical vein endothelial cells (HUVEC). Overall, the nanoengineered folate receptor-mediated co-delivery system provides great potential for safe and effective cancer therapy.

Polymersomes as the Next Attractive Generation of Drug Delivery Systems: Definition, Synthesis and Applications.

Polymersomes are artificial nanoparticles formed by the self-assembly process of amphiphilic block copolymers composed of hydrophobic and hydrophilic blocks. They can encapsulate hydrophilic molecules in the aqueous core and hydrophobic molecules within the membrane. The composition of block copolymers can be tuned, enabling control of characteristics and properties of formed polymersomes and, thus, their application in areas such as drug delivery, diagnostics, or bioimaging. The preparation methods of polymersomes can also impact their characteristics and the preservation of the encapsulated drugs. Many methods have been described, including direct hydration, thin film hydration, electroporation, the pH-switch method, solvent shift method, single and double emulsion method, flash nanoprecipitation, and microfluidic synthesis. Considering polymersome structure and composition, there are several types of polymersomes including theranostic polymersomes, polymersomes decorated with targeting ligands for selective delivery, stimuli-responsive polymersomes, or porous polymersomes with multiple promising applications. Due to the shortcomings related to the stability, efficacy, and safety of some therapeutics in the human body, polymersomes as drug delivery systems have been good candidates to improve the quality of therapies against a wide range of diseases, including cancer. Chemotherapy and immunotherapy can be improved by using polymersomes to deliver the drugs, protecting and directing them to the exact site of action. Moreover, this approach is also promising for targeted delivery of biologics since they represent a class of drugs with poor stability and high susceptibility to in vivo clearance. However, the lack of a well-defined regulatory plan for polymersome formulations has hampered their follow-up to clinical trials and subsequent market entry.

One-step fabrication of microfluidic W/O/W droplets as fat-reduced high internal phase emulsions: Microstructure, stability and 3D printing performance

High internal phase emulsions (HIPEs) have been served as effective fat replacements, but they still remain calorie-dense. Herein, we described microfluidic fabrication of W/O/W mononuclear high internal phase double emulsions (MHIPDEs) with a reduced oil fraction ranging from 38 vol% to 53 vol%. These emulsions were composed of oil drops (∼170 μm) that contained smaller water droplets (117.22–136.33 μm). The incorporation of gellan gum in the internal water droplets and palm oil in the oil phase facilitated the stabilization of MHIPDEs. All the emulsions demonstrated favorable stability during storage at both 4 °C and 25 °C, maintaining their intact structure for a duration of 90 days. This feature enabled the emulsions to effectively protect lipids from oxidation, resulting in a notable two-thirds reduction in the production of MDA during stored at 4 °C. All of these emulsions maintained a uniform appearance after undergoing freeze-thaw processing eight times, and MHIPDEs with the highest oil content (62 vol%) remained structurally intact. Additionally, the apparent viscosity and elastic modulus of MHIPDEs increased with higher oil content, suggesting the formation of a stiffer lipid crystal shell surrounding the water droplet. MHIPDEs with a moderate oil fraction (54 vol%) possessed optimal 3D printing performance due to their appropriate viscoelasticity and high yield stress (6.3%). These findings propose a fresh perspective on the design and manufacturing of fat-reduced high internal phase emulsions with desirable physicochemical properties.

Zinc-Boron-PLGA biocomposite material: preparation, structural characterization, and in vitro assessment.

Nowadays, the state-of-the-art discoveries in the field of delivery systems for therapeutic purposes have redefined the importance of biocompatible and biodegradable poly(lactic-co-glycolic acid (PLGA) nanocomposites. The study aimed to obtain a biocomposite material, with improved properties of its constituents [zinc-boron (Zn-B) complex and PLGA], by a simple, cost-effective method. The water∕oil∕water double emulsion technique allowed the adjustment of the synthesis parameters, to maximize the degree of Zn-B complex encapsulation. The morphological aspects of the samples were established by scanning electron microscopy (SEM). Particle size distribution was determined by dynamic light scattering (DLS). Morphology was typical for PLGA, spherical one. Depending on the synthesis conditions, the obtained particles have diameters between 10-450 nm. Zeta potential (ZP) showed that the particles have electronegative surface charge, offering a favorable perspective on aggregation, flocculation, and dispersion phenomena. It was observed, applying the design of experiments, that the particles size increased with increasing amounts of PLGA and polyvinyl alcohol (PVA), while ZP increased with higher PLGA and smaller PVA amounts in the formulation. The encapsulation efficiency was determined by ultra-high performance liquid chromatography∕mass spectrometry (UHPLC∕MS). The in vitro assessment was performed using Vero CCL-81 epithelial cell line and the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) test. Zn-B-PLGA biocomposite has promising characteristics and can be used for future biomedical applications.

Design automation of microfluidic single and double emulsion droplets with machine learning

Droplet microfluidics enables kHz screening of picoliter samples at a fraction of the cost of other high-throughput approaches. However, generating stable droplets with desired characteristics typically requires labor-intensive empirical optimization of device designs and flow conditions that limit adoption to specialist labs. Here, we compile a comprehensive droplet dataset and use it to train machine learning models capable of accurately predicting device geometries and flow conditions required to generate stable aqueous-in-oil and oil-in-aqueous single and double emulsions from 15 to 250 μm at rates up to 12000 Hz for different fluids commonly used in life sciences. Blind predictions by our models for as-yet-unseen fluids, geometries, and device materials yield accurate results, establishing their generalizability. Finally, we generate an easy-to-use design automation tool that yield droplets within 3 μm (<8%) of the desired diameter, facilitating tailored droplet-based platforms and accelerating their utility in life sciences.

Intramuscular injection of palmitic acid-conjugated Exendin-4 loaded multivesicular liposomes for long-acting and improving in-situ stability

ABSTRACT Background Exendin-4 (Ex4) is a promising drug for diabetes mellitus with a half-life of 2.4 h in human bodies. Besides, the Ex4 formulations currently employed in the clinic or under development have problems pertaining to stability. In this study, palmitic acid-modified Ex4 (Pal-Ex4) was prepared and purified to extend the half-life of Ex4. In addition, Pal-Ex4-MVLs were further designed and optimized as a long-acting delivery system for intramuscular injection. Methods Pal-Ex4 was encapsulated within multivesicular liposomes (MVLs) via a two-step double emulsification process. The formulated products were then assessed for their vesicle size, encapsulation efficiency, and in vitro and in vivo. Results Pal-Ex4-MVLs with a notable encapsulation efficiency of 99.18% were successfully prepared. Pal-Ex4-MVLs, administered via a single intramuscular injection in Sprague-Dawley rats, sustained stable plasma concentrations for 168 h, presenting extended half-life (77.28 ± 12.919 h) and enhanced relative bioavailability (664.18%). MVLs protected Ex4 through providing stable retention and slow release. This approach considerably improved the in-situ stability of the drug for intramuscular administration. Conclusions The combination of palmitic acid modification process with MVLs provides dual protection for Ex4 and can be a promising strategy for other hydrophilic protein/polypeptide-loaded sustained-release delivery systems with high drug bioactivity.

Nanoparticle‐based sustained drug delivery for the treatment of ocular neovascular diseases

Aims/Purpose: Nanoparticles have shown great potential as a drug delivery system for the treatment of neovascular ocular diseases characterized by the growth of abnormal blood vessels in the retina. The current standard of therapy for neovascular ocular diseases involves administering monoclonal antibody‐based anti‐VEGF drugs via intravitreal injections on a periodic basis. The short half‐lives of these drugs necessitate frequent injections, which can limit the effectiveness of the treatments. Hence, there is a need to develop new approaches for enhanced bioavailability and sustained and long‐lasting delivery of anti‐VEGF drugs. The aim of this study was to utilize poly (glycerol sebacate) (PGS) nanoparticles to achieve an extended release of anti‐VEGF (anti‐vascular endothelial growth factor) drugs, with the objective of reducing the frequency of intravitreal injections required for treatment.Methods: Double emulsion solvent evaporation method was utilized for the synthesis of PGS nanoparticles. Nanoparticle size and polydispersity index (PDI) were determined with dynamic light scattering method.Results: Nanoparticles were synthesized with a size of approximately 150 nm and PDI less than 0.3. The encapsulation efficiency of nanoparticles loaded with anti‐VEGF drugs was found to be 74%. The 30% drug release from nanoparticles in the first 2 months indicates that nanoparticles are a promising candidate for sustained drug release applications. Cell‐viability studies showed that anti‐VEGF drug‐loaded nanoparticles did not exhibit toxicity to ARPE‐19 cells, but they did exhibit anti‐angiogenesis properties by inhibiting the growth of HUVEC cells in a concentration‐dependent manner.Conclusions: The development of new‐generation materials for the storage and preservation of anti‐VEGF drugs, as well as their enhancement with prolonged release, will considerably improve the efficacy of long‐term treatment strategies for ocular neovascular diseases.

Identification of the Vulnerability of Atherosclerotic Plaques by a Photoacoustic/Ultrasonic Dual-Modal cRGD Nanomolecular Probe.

To explore the feasibility of using cRGD-GNR-PFP-NPs to assess plaque vulnerability in an atherosclerotic plaque mouse model by dual-modal photoacoustic/ultrasonic imaging. A nanomolecular probe containing gold nanorods (GNRs) and perfluoropentane (PFP) coated with the cyclic Arg-Gly-Asp (cRGD) peptide were prepared by double emulsion solvent evaporation and carbodiimide methods. The morphology, particle size, potential, cRGD conjugation and absorption features of the nanomolecular probe were characterized, along with its in vitro phase transformation and photoacoustic/ultrasonic dual-modal imaging properties. In vivo fluorescence imaging was used to determine the distribution of cRGD-GNR-PFP-NPs in vivo in apolipoprotein E-deficient (ApoE-/-) atherosclerotic plaque model mice, the optimal imaging time was determined, and photoacoustic/ultrasonic dual-modal molecular imaging of integrin αvβ3 expressed in atherosclerotic plaques was performed. Pathological assessments verified the imaging results in terms of integrin αvβ3 expression and plaque vulnerability. cRGD-GNR-PFP-NPs were spherical with an appropriate particle size (average of approximately 258.03±6.75 nm), a uniform dispersion, and a potential of approximately -9.36±0.53 mV. The probe had a characteristic absorption peak at 780~790 nm, and the surface conjugation of the cRGD peptide reached 92.79%. cRGD-GNR-PFP-NPs were very stable in the non-excited state but very easily underwent phase transformation under low-intensity focused ultrasound (LIFU) and had excellent photoacoustic/ultrasonic dual-modal imaging capability. Mice fed a high-fat diet for 20 weeks had obvious hyperlipidemia with larger, more vulnerable plaques. These plaques could be specifically targeted by cRGD-GNR-PFP-NPs as determined by in vivo fluorescence imaging, and the enrichment of nanomolecular probe increased with the increasing of plaque vulnerability; the photoacoustic/ultrasound signals of the plaques in the high-fat group were stronger. The pathological assessments were in good agreement with the cRGD-GNR-PFP-NPs plaque accumulation, integrin αvβ3 expression and plaque vulnerability results. A phase variant photoacoustic/ultrasonic dual-modal cRGD nanomolecular probe was successfully prepared and can be used to identify plaque vulnerability safely and effectively.

Microfluidic fabrication of monodisperse microcapsules with gas cores.

A facile strategy for efficient and continuous fabrication of monodisperse gas-core microcapsules with controllable sizes and excellent ultrasound-induced burst performances is developed based on droplet microfluidics and interfacial polymerization. Monodisperse gas-in-oil-in-water (G/O/W) double emulsion droplets with a gas core and monomer-contained oil layer are fabricated in the upstream of a microfluidic device as templates, and then water-soluble monomers are added into the aqueous continuous phase in the downstream to initiate rapid interfacial polymerization at the O/W interfaces to prepare monodisperse gas-in-oil-in-solid (G/O/S) microcapsules with gas cores. The sizes of both microbubbles and G/O/W droplet templates can be precisely controlled by adjusting the gas supply pressure and the fluid flow rates. Due to the very thin shells of G/O/S microcapsules fabricated via interfacial polymerization, the sizes of the resultant G/O/S microcapsules are almost the same as those of the G/O/W droplet templates, and the microcapsules exhibit excellent deformable properties and ultrasound-induced burst performances. The proposed strategy provides a facile and efficient route for controllably and continuously fabricating monodisperse microcapsules with gas cores, which are highly desired for biomedical applications.

Flow pattern maps of double emulsions transporting through bifurcation microchannels.

The transportation behaviors of compound droplets in confined channels are widespread phenomena while the physical mechanisms are far from being completely unraveled. In this work, behaviors of double emulsions flowing through bifurcation microchannels are experimentally studied with the aim of building universal flow pattern maps. Three flow patterns are categorized according to different features of daughter droplets in terms of size, uniformity, and shell thickness. A detailed analysis of the dynamics of interfacial evolutions in different patterns is carried out and the coupling interaction between interfaces is found to affect the minimum tail distance during transportation. It is feasible to obtain the threshold of the occurrence of the coupling interaction, due to the different variation tendencies in the two states, which relies on three dimensionless parameters, i.e. droplet length, length ratio, and capillary number. Furthermore, a novel physical model is proposed to build the flow pattern map, with the two transition boundaries being expressed as different relationships in terms of the three identified parameters. The physical mechanisms are summarized with the aid of force analysis. An excellent agreement is shown between the model and experimental results in different liquid systems and bifurcation structures, indicating the generality of the proposed model.

Double emulsion microencapsulation of ionic liquids for carbon capture

Double emulsion microencapsulation of ionic liquids for carbon capture

Autophagic blockage by metformin-loaded PLGA nanoparticles causes cell cycle arrest of HepG2 cells.

Aim: To fabricate and characterize metformin-loaded PLGA nanoparticles andinvestigate theirinhibitory effect on HepG2 cells. Materials & methods: The nanoparticles were prepared using a double emulsification method, then characterized and subjected to a series of in vitro assays on HepG2 cells. Results: The nanoparticles were ~277.9nm in size, and the entrapment efficiency and drug loading of metformin were 31.3 and 14.4%, respectively. In vitro studies suggested that the nanoparticles showed a higher inhibitory effect on HepG2 cells compared with metformin alone, mainly attributed to its blockage of autophagy, and ultimately result in cell cycle inhibition. Conclusion: The metformin-loaded PLGA nanoparticles could inhibit mTOR activity, increase p53 levels and decrease HIF1A levels, which ultimately caused HepG2 cell cycle arrest.

Enhancing oral bioavailability of Ca-DTPA by self double emulsifying drug delivery system (SDEDDS)

Objective BCS class III drug (highly soluble, poorly permeable) possesses low oral bioavailability. The research work highlights the utility of self-double emulsifying drug delivery system (SDEDDS) which are stable isotropic mixture of w/o primary emulsion and hydrophilic surfactants for improving oral bioavailability of Ca-DTPA (Calcium diethylenetriamine pentaacetate). Upon oral administration, SDEDDS rapidly emulsifies into w/o/w double emulsions in the aqueous gastrointestinal environment, with hydrophilic drugs entrapped inside oil reservoirs. Methods SDEDDS formulation was successfully developed using excipients, that is, medium chain triglycerides, oleic acid, phospholipids, Span 80, Tween 80 using double emulsification technique. Results The optimized formulation F4 (Aq. phase: 11.6%w,w; MCT & oleic acid: 70.9%w/w; Span 80:17.5%w/w; Lecithin:16%w/w and Tween 80 (10%w/w)) appeared bright yellow liquid which upon dilution appeared milky white within 2 min, droplet size (501.7 nm), pdi value (0.044), zeta potential (-52 mV), entrapment efficiency (79.6 ± 1.63), viscosity (72.2 ± 1.8 mpA.s), significant high cumulative in vitro drug permeation (CDP) and 2.17-fold increase in apparent permeability coefficient. Pharmacokinetic studies in rats showed 1.17-fold increases in AUC of F4 and comparatively higher plasma levels (Cmax) compared with pure drug administered orally. The Absolute (OF4, OD) and Relative bioavailability was found to be 14.52%, 12.35%, and 117.47%, respectively. Conclusion The present studies have clearly demonstrated that SDEDDS could readily form w/o/w double emulsions in vivo with enhanced in vitro and in vivo oral bioavailability. Therefore, considerable augmentation in the rate and extent of oral drug absorption ratified the better performance of the SDEDDS in enhancing the bioavailability of Ca-DTPA.

Microfluidic Giant Polymer Vesicles Equipped with Biopores for High-Throughput Screening of Bacteria.

Understanding the mechanisms of antibiotic resistance is critical for the development of new therapeutics. Traditional methods for testing bacteria are often limited in their efficiency and reusability. Single bacterial cells can be studied at high throughput using double emulsions, although the lack of control over the oil shell permeability and limited access to the droplet interior present serious drawbacks. Here, a straightforward strategy for studying bacteria-encapsulating double emulsion-templated giant unilamellar vesicles (GUVs) is introduced. This microfluidic approach serves to simultaneously load bacteria inside synthetic GUVs and to permeabilize their membrane with the pore-forming peptide melittin. This enables antibiotic delivery or the influx of fresh medium into the GUV lumen for highly parallel cultivation and antimicrobial efficacy testing. Polymer-based GUVs proved to be efficient culture and analysis microvessels, as microfluidics allow easy selection and encapsulation of bacteria and rapid modification of culture conditions for antibiotic development. Further, a method for in situ profiling of biofilms within GUVs for high-throughput screening is demonstrated. Conceivably, synthetic GUVs equipped with biopores can serve as a foundation for the high-throughput screening of bacterial colony interactions during biofilm formation and for investigating the effect of antibiotics on biofilms.

Fabrication of novel W/O/W emulsion gels using beeswax stabilized W/O: Preparation, characterization and co-delivery of phycocyanin and astaxanthin

Double emulsions (DEs), one of the most promising dispersion systems, have attracted much attention in the food industry. However, instability and the need for synthetic surfactants hinder the application of DEs. The development of clean-label DEs with good stability remains a challenge. Herein, novel DE gels (DEGs) with water-in-oil-in-water morphology were developed by combining multiple gelation and emulsification processes, which can stabilize the dispersion system without the synthetic surfactants used in conventional emulsions. Gelatin and beeswax were used to gelatinize the internal aqueous and oil phases respectively. Sodium caseinate acted as the external aqueous phase emulsifier, while a blend of gellan gum (GG) and sodium alginate (SA) was introduced to create DEGs. The networks formed by SA and GG in the water phase inhibited the movement of droplets in the emulsions and improved their stability. The DEGs can withstand up to five freeze-thaw cycles and at least 100 d of storage. In addition, the textural properties of DEGs were characterized and the results suggested that DEGs could be used as a cheese simulant. DEGs could be used as carriers for the co-delivery of phycocyanin and astaxanthin. Their encapsulation efficiency was greater than 90%. Furthermore, in vitro digestion showed that DEGs significantly improved the bioavailability of astaxanthin and phycocyanin, which increased from 18.5 ± 0.15% and 16.3 ± 0.45% to 43.2 ± 0.35% and 41.6 ± 0.5%, respectively. In conclusion, we have demonstrated a novel strategy for the preparation of clean-label DEGs, which could be used as a carrier for the co-delivery of hydrophilic and hydrophobic substances.

Investigation of vanillin-loaded O/W, W/O, and W/O/W emulsion systems as a proof of concept for sugar reduction strategy: Preparation, characterization, and evaluation

Achieving food sugar reduction based on odor-taste cross-modal interaction requires careful design of food emulsion systems, which are a promising delivery mechanism with controlled flavor release. Hence, we designed the single (O/W, W/O) and W/O/W double emulsions, which distributed the flavor component (vanillin) and sugar substitute (d-allulose) in the oil and aqueous phase, respectively. In this work. we acquired the optimal formulations for three emulsions and characterized the physical properties of each emulsion. Furthermore, we investigated the stability of three emulsions under various environmental stresses and found that high-temperature short-time (HTST) sterilization had no effect on the quality of the three emulsions, but autoclaving, heat treatment, and freeze-thawing all caused structural integrity damage to the emulsions. Additionally, the storage results indicated that 4 °C might be the optimal temperature for all emulsions. According to these results, W/O emulsions possess the best overall stability, which has great potential for applications to meet the current sugar reduction strategy.

Optimisation and characterization of double emulsion derived from rice starch, rice protein isolates and rice bran oil

This study focused on the development and evaluation of a stable emulsion system using rice starch, protein isolates, and bran oil as food ingredients. This was performed using a one-factor-at-a-time (OFAT) experimental strategy, which was subsequently refined using response surface modeling (RSM), and the emulsion was then characterized. The resulting quadratic polynomial model effectively captured all the responses, with regression coefficients exceeding 0.90. The emulsion activity index, emulsion stability index, and droplet size were 86.29 %, 93.44 %, and 100.59 nm, respectively. Turbidity was significantly influenced by particle size, with the double emulsion (sample B) exhibiting higher turbidity index (43,250.34 ± 0.046) than the reference sample (29,433.303 ± 0.018). The viscosity of the emulsion increased with the addition of the protein isolates. This inferred that a stable emulsion system derived from rice could serve a multifunctional purpose as carriers, fillers, and binders to enhance the physical, functional and sensory properties of foods.

Preparation of Bifunctional Nanoparticles and Evaluation of Therapeutic Function in Glioma Cells

Small interfering RNA (siRNA)drugs have great potential in the field of anti-tumor, but they have poor biological stability and are easily degraded by nucleases in vivo, resulting in poor bioavailability. In this study, a novel albumin Poly(lactide-co-glycolide) (PLGA) nanocarrier was prepared to efficiently deliver siRNA to glioma cells to achieve tumor-targeted therapy. Recombinant VAR2CSA protein (RVP) targeting albumin PLGA nanocarriers (RVP–APN–siRNA) loaded with EGFR-siRNA were prepared by a double emulsion method. Determine its characterization performance, loading rate, in vitro release and cell uptake, and investigate the effects of nanoparticles on the cytotoxicity, proliferation and apoptosis of brain glioma cells. The prepared nanoparticles are spherical with uniform particle size, the average particle size distribution is 120–180 nm, the average potential distribution is 20–30 mv, and the loading rate is 82.36 ± 1.84%. The release rate of siRNA within 24 h is 93%, with low cytotoxicity and targeting performance, effectively inhibiting the activity of glioma cells. The prepared albumin nanoparticles have good physical and chemical properties, and have dual functions of targeted recognition of glioma cells and tumor treatment. They are efficient and safe nano-carriers for tumor-targeted treatment, and have good potential in the application of siRNA-targeted drug delivery.

Tumor Microenvironment Modulating CaCO3 -Based Colloidosomal Microreactors Can Generally Reinforce Cancer Immunotherapy.

Tumor hypoxia and acidity, two general features of solid tumors, are known to have negative effect on cancer immunotherapy by directly causing dysfunction of effector immune cells and promoting suppressive immune cells inside tumors. Herein, a multifunctional colloidosomal microreactor is constructed by encapsulating catalase within calcium carbonate (CaCO3 ) nanoparticle-assembled colloidosomes (abbreviated as CaP CSs) via the classic double emulsion method. The yielded CCaP CSs exhibit well-retained proton-scavenging and hydrogen peroxide decomposition performances and can thus neutralize tumor acidity, attenuate tumor hypoxia, and suppress lactate production upon intratumoral administration. Consequently, CCaP CSs treatment can activate potent antitumor immunity and thus significantly enhance the therapeutic potency of coloaded anti-programmed death-1 (anti-PD-1) antibodies in both murine subcutaneous CT26 and orthotopic 4T1 tumor xenografts. In addition, such CCaP CSs treatment also markedly reinforces the therapeutic potency of epidermal growth factor receptor expressing chimeric antigen receptor T (EGFR-CAR-T) cells toward a human triple-negative breast cancer xenograft by promoting their tumor infiltration and effector cytokine secretion. Therefore, this study highlights that chemical modulation of tumor acidity and hypoxia can collectively reverse tumor immunosuppression and thus significantly potentiate both immune checkpoint blockade and CAR-T cell immunotherapies toward solid tumors.

Gelatin/polyethylene glycol/g-C3N4 hydrogel with olive oil as a sustainable and biocompatible nanovehicle for targeted delivery of 5-fluorouracil

5-fluorouracil (5-FU) is an anticancer drug with poor water solubility and stability, leading to short plasma half-time and undesired toxic effects on normal tissues. Therefore, novel drug delivery systems (DDS) are required to improve its therapeutic efficiency. The use of nanoparticles in DDS has emerged as a promising method for controlling the release of chemotherapy drugs. In this study, a novel, sustainable and pH-sensitive nanocomposite was designed for the targeted delivery of 5-FU against breast cancer cells. The nanodelivery system based on biocompatible and biodegradable graphitic carbon nitride (g-C3N4) coated with Gelatin (G)/Polyethylene glycol (PEG) biopolymers was prepared by W/O/W double emulsion technique using olive oil and span 80 surfactant. Successful loading of 5-FU into the nanovehicle was confirmed by infrared spectroscopy and X-ray diffraction analyses. Dynamic light scattering corroborated the monodispersity of the nanocarriers, with a typical size between 250 and 300 nm. Scanning electron microscopy images revealed the homogeneity of the nanocomposite and its spherical shape, which was appropriate for DDS. The average zeta potential was + 35.4 mV, demonstrating the good stability of the nanocomposite. The drug loading (47 %) and encapsulation efficiency (89 %) were higher than those reported for other 5-FU nanocarriers. A continuous and controlled 5-FU release from the nanovehicle was observed, and the release rate at pH 5.4 (simulating a cancerous environment) was higher than at pH 7.4 (physiological conditions). Kinetic studies suggested a diffusion controlled mechanism of 5-FU release from the nanocarrier in an acidic environment, while an anomalous transport prevails at neutral pH. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay revealed that G/PEG/g-C3N4 had a significant cytotoxicity on MCF-7 breast cancer cells. Flow cytometry indicated that G/PEG/g-C3N4/5-FU led to much higher apoptotic percentage than free 5-FU, while the free drug showed higher percentage of necroptotic cells. This novel bio-nanocomposite shows great potential as a pH-sensitive DDS against breast cancer cells.