Small Hydrophilic Molecules Research Articles

Selecting Counterions to Improve Ionized Hydrophilic Drug Encapsulation in Polymeric Nanoparticles.

Hydrophobic ion pairing (HIP) can successfully increase the drug loading and control the release kinetics of ionizable hydrophilic drugs, addressing challenges that prevent these molecules from reaching the clinic. Nevertheless, polymeric nanoparticle (PNP) formulation development requires trial-and-error experimentation to meet the target product profile, which is laborious and costly. Herein, we design a preformulation framework (solid-state screening, computational approach, and solubility in PNP-forming emulsion) to understand counterion-drug-polymer interactions and accelerate the PNP formulation development for HIP systems. The HIP interactions between a small hydrophilic molecule, AZD2811, and counterions with different molecular structures were investigated. Cyclic counterions formed amorphous ion pairs with AZD2811; the 0.7 pamoic acid/1.0 AZD2811 complex had the highest glass transition temperature (Tg; 162 °C) and the greatest drug loading (22%) and remained as phase-separated amorphous nanosized domains inside the polymer matrix. Palmitic acid (linear counterion) showed negligible interactions with AZD2811 (crystalline-free drug/counterion forms), leading to a significantly lower drug loading despite having similar log P and pKa with pamoic acid. Computational calculations illustrated that cyclic counterions interact more strongly with AZD2811 than linear counterions through dispersive interactions (offset π-π interactions). Solubility data indicated that the pamoic acid/AZD2811 complex has a lower organic phase solubility than AZD2811-free base; hence, it may be expected to precipitate more rapidly in the nanodroplets, thus increasing drug loading. Our work provides a generalizable preformulation framework, complementing traditional performance-indicating parameters, to identify optimal counterions rapidly and accelerate the development of hydrophilic drug PNP formulations while achieving high drug loading without laborious trial-and-error experimentation.

A Novel Curcumin Arginine Salt: A Solution for Poor Solubility and Potential Anticancer Activities.

Curcumin is a natural polyphenolic compound with well-known anticancer properties. Poor solubility and permeability hamper its use as an anticancer pharmaceutical product. In this study, L-arginine, a basic amino acid and a small hydrophilic molecule, was utilized to form a salt with the weak acid curcumin to enhance its solubility and potentiate the anticancer activities of curcumin. Two methods were adopted for the preparation of curcumin: L-arginine salt, namely, physical mixing and coprecipitation. The ion pair or salt was characterized for docking, solubility, DSC, FTIR, XRD, in vitro dissolution, and anticancer activities using MCF7 cell lines. The molecular docking suggested a salt/ion-pair complex between curcumin and L-arginine. Curcumin solubility was increased 335- and 440-fold by curcumin in L-arginine, physical, and co-precipitated mixtures, respectively. Thermal and spectral analyses supported the molecular docking and formation of a salt/ion pair between curcumin and L-arginine. The cytotoxicity of curcumin L-arginine salt significantly improved (p < 0.05) by 1.4-fold, as evidenced by the calculated IC50%, which was comparable to Taxol (the standard anticancer drug but with common side effects).

PH-Switchable Phase-Transfer Agents for Host Cell Protein Rejection in the Cascaded Biocatalytic Synthesis of an Active Pharmaceutical Ingredient

Given the increased reliance on multienzymatic cascades to synthesize biologically inspired small molecule drug candidates, the pharmaceutical industry requires efficient strategies to control immunogenic protein impurities in active pharmaceutical ingredients. Despite advances in directed enzyme evolution and biocatalysis, it is still economical to use unpurified enzymes overexpressed in host cell lysates in manufacturing processes. Due to similar solubility profiles between protein impurities and the pharmaceutical, separation strategies relying on solubility differences (direct crystallization from water or extraction) often fail. Leveraging the rapid generation of modular acid–base pairs, we describe a pH-responsive extraction strategy using tertiary ammonium phase-transfer agents for efficient purification and isolation of hydrophilic immuno-oncology drug candidate MK-1454 from an aqueous biocatalytic cascade containing crude E. coli cell lysates. We demonstrate this technique to isolate several hundred grams of MK-1454 with undetectable protein impurities for human administration in clinical trials. We further show that this extraction technique is driven by reverse micellization, and may be generally applied to the purification of other hydrophilic small molecules.

Anti-hardening effect and mechanism of silkworm sericin peptide in high protein nutrition bars during early storage

Hardening presents an inevitable challenge during the storage of high protein nutrition bars. Sericin peptide is the product of hydrolysis of sericin, a protein from the silkworm cocoon. Here in, the effects of sericin peptide addition on the hardening of high protein nutrition bars during 72 h of storage were investigated. The addition of sericin peptide to high protein nutrition bars reduced the hardening of the sample during the early storage, The main mechanism was to improve the mobility of water and small hydrophilic molecules, which slowed down the phase separation. As well, after sericin peptide addition, the ζ- potential, the content of secondary structure, and the surface hydrophobicity of the samples were also changed, which prevented the self-aggregation of proteins. These results indicate that SRP can be used as a promising anti-hardening ingredient in the food industry to improve the texture of food products.

Optimized Data Set and Feature Construction for Substrate Prediction of Membrane Transporters.

α-Helical transmembrane proteins termed membrane transporters mediate the passage of small hydrophilic substrate molecules across biological lipid bilayer membranes. Annotating the specific substrates of the dozens to hundreds of individual transporters of an organism is an important task. In the past, machine learning classifiers have been successfully trained on pan-organism data sets to predict putative substrates of transporters. Here, we critically examine the selection of an optimal data set of protein sequence features for the classification task. We focus on membrane transporters of the three model organisms Escherichia coli, Arabidopsis thaliana, and Saccharomyces cerevisiae, as well as human. We show that organism-specific classifiers can be robustly trained if at least 20 samples are available for each substrate class. If information from position-specific scoring matrices is included, such classifiers have F1 scores between 0.85 and 1.00. For the largest data set (A. thaliana), a 4-class classifier yielded an F-score of 0.97. On a pan-organism data set composed of transporters of all four organisms, amino acid and sugar transporters were predicted with an F1 score of 0.91.

In silico identification and synthesis of a multi-drug loaded MOF for treating tuberculosis.

Conventional drug delivery systems have been applied to a myriad of active ingredients but may be difficult to tailor for a given drug. Herein, we put forth a new strategy, which designs and selects the drug delivery material by considering the properties of encapsulated drugs (even multiple drugs, simultaneously). Specifically, through an in-silico screening process of 5109 MOFs using grand canonical Monte Carlo simulations, a customized MOF (referred as BIO-MOF-100) was selected and experimentally verified to be biologically stable, and capable of loading 3 anti-Tuberculosis drugs Rifampicin+Isoniazid+Pyrazinamide at 10%+28%+23% wt/wt (total>50% by weight). Notably, the customized BIO-MOF-100 delivery system cleared naturally Pyrazinamide-resistant Bacillus Calmette-Guérin, reduced growth of virulent Erdman infection in macaque macrophages 10-100-fold compared to soluble drugs in vitro and was also significantly reduced Erdman growth in mice. These data suggest that the methodology of identifying-synthesizing materials can be used to generate solutions for challenging applications such as simultaneous delivery of multiple, small hydrophilic and hydrophobic molecules in the same molecular framework.

Progress and challenges of lyotropic liquid crystalline nanoparticles for innovative therapies

Since the late 20th century, we have witnessed a growing and substantial advance in nanomedicine, in part due to the development of multifunctional and multimodal nanoplatforms that have enabled improved efficacy, biocompatibility, and novel therapeutic applications. Non-lamellar liquid-crystalline nanoparticles, especially, reverse hexagonal and cubic bicontinuous mesophases, have gained the attention of the scientific-academic community due to their intriguing and functional characteristics, such as self-organization into two- and three-dimensional supramolecular structures, high symmetry, and ability to accommodate hydrophobic and hydrophilic small molecules, peptides, proteins, nucleic acids, and imaging agents. Furthermore, these particles can be easily modified with specific and/or bioresponsive molecules allowing targeting and improved therapeutic performance. In this contribution we provide an overview of advances in the design and architecture of LCNPs, strategies to overcome biological barriers and main findings about interactions with different types of interfaces. We highlight recent applications in topical, oral, pulmonary and intravenous drug delivery in preclinical in vivo studies. We discussed the current scenario and translational obstacles faced for clinical translation, as well as our perspectives.

Cephalosporin translocation across enterobacterial OmpF and OmpC channels, a filter across the outer membrane

Gram-negative porins are the main entry for small hydrophilic molecules. We studied translocation of structurally related cephalosporins, ceftazidime (CAZ), cefotaxime (CTX) and cefepime (FEP). CAZ is highly active on E. coli producing OmpF (Outer membrane protein F) but less efficient on cells expressing OmpC (Outer membrane protein C), whereas FEP and CTX kill bacteria regardless of the porin expressed. This matches with the different capacity of CAZ and FEP to accumulate into bacterial cells as quantified by LC-MS/MS (Liquid Chromatography Tandem Mass Spectrometry). Furthermore, porin reconstitution into planar lipid bilayer and zero current assays suggest permeation of ≈1,000 molecules of CAZ per sec and per channel through OmpF versus ≈500 through OmpC. Here, the instant killing is directly correlated to internal drug concentration. We propose that the net negative charge of CAZ represents a key advantage for permeation through OmpF porins that are less cation-selective than OmpC. These data could explain the decreased susceptibility to some cephalosporins of enterobacteria that exclusively express OmpC porins.

Granulation and encapsulation of N-Acetylcysteine (NAC) by internal phase separation

The encapsulation of small hydrophilic molecules offers advantages for the delivery of hydrophilic food supplements and food ingredients. These advantages include: masking off-flavors, reducing astringency , and controlling release profiles. We used an internal phase separation (IPS) method for simultaneous granulation and encapsulation of a small hydrophilic food supplement in corn oil. To prove that IPS is a suitable method for the encapsulation of small hydrophilic molecules with controllable release times, we used the nutritional supplement N -acetylcysteine (NAC) as our model molecule. NAC has a variety of health and pharmaceutical benefits however it suffers from poor organoleptic properties such as a bitter aftertaste, astringency, and a strong sulfur smell. We used IPS to produce free-flowing granules containing 56–78% NAC with diameters ranging from 100 to 1000 μm. All the materials used in this process were either food safe or generally recognized as safe (GRAS). The granulated/encapsulated samples were characterized using FTIR, NMR, SEM, and LC-MS methods to confirm the presence of unreacted NAC in high loading concentrations. The kinetics of NAC release was then monitored using conductometry. The combination of hydrophilic carboxymethyl cellulose sodium salt (CMC) and hydroxypropyl methylcellulose (HPMC) gums with a hydrophobic coating such as ethyl cellulose (EC), provided an effective barrier to ensure a 5 min time window for NAC dispersion into water and drinks before the capsule starts to dissolve in water and loses more than 30 wt % of its total NAC loading. • Internal phase separation was used for NAC granulation and encapsulation. • Using all GRAS materials, NAC was granulated and encapsulated in high loading. • The encapsulated samples, showed delayed NAC release in water. • The spherical granules of encapsulated NAC can be used in ready-to-mix product.

LC-HRMS toxicological screening and biological sample preparation: Evaluation of extraction by turbulent flow chromatography and protein precipitation procedures

LC-HRMS has nowadays become the “gold standard” for identification of xenobiotics in clinical and forensic toxicology providing both high selectivity and specificity. Sample preparation and introduction nevertheless remain a key step and should be simple, universal, reproducible and sensitive for a broad range of analytes. Two types of sample introduction were tested for drugs spiked in biological samples: on-line extraction by turbulent flow chromatography (TurboFlow®) versus direct injection. Then, parameters of protein precipitation such as organic solvent, sample/organic solvent ratio and evaporation after precipitation were evaluated. In all, 62 compounds (drugs of abuse, pharmaceuticals and metabolites) were selected according to several criteria such as polarity, log P, retention time, occurrence in drug screening and divided in two groups regarding their therapeutic (A: n = 20) or regulatory (B: n = 42) concentrations. Plasma, blood and urine samples were prepared by spiking different concentrations (0.25 to 1000 μg/L, n = 6) into blank matrices. TurboFlow® and LC analyses were performed using a Transcend II TLX-1 and Vanquish systems. Data were acquired on an Exploris 120 Orbitrap Mass spectrometer and processed with TraceFinder 5.1 software (ThermoFisher Scientific®, San Jose, USA). The analytical workflow was based on previous publication (Helfer, J Chrom B, 2017, 1043, 138–149) with minor modifications: Phenyl replaced by Fluoro Turboflow column. 400 μL of sample was mixed with 600 μL of acetonitrile for precipitation. After shaking and centrifugation, the supernatant was injected. Several parameters were assessed for protein precipitation: – Sample/organic solvent ratio: 1:1,5; 1:2 and 1:3; – Organic solvent: acetonitrile, methanol, acetonitrile plus 0.1% formic acid, acetonitrile:methanol (50:50, v:v), methanol plus 0.2 M ZnSO4 (80:20, v:v) and acetonitrile with 80 mg QuEChERS extraction salts; – Concentration: direct injection or evaporation after precipitation. The performances were evaluated regarding the limits of identification (LOI), reproducibility at the respective LOI of each compound and linearity. Reproducibility gave satisfactory results for 59 and 56 compounds in direct injection and in TurboFlow® in all matrices (CV < 15%). Linearity was acceptable in both injection modes with no exception (R2 > 0.95). Finally, sensitivity appeared to be the most differentiating factor, direct injection showing lower limits for 29 (plasma), 31 (blood) and 28 (urine) compounds. For example, identification was possible at 10 μg/L in plasma for gabapentin, metformin and baclofen in direct injection and at 100 μg/L in TurboFlow®. For protein precipitation, 1:1.5 and 1:2 ratios were statistically equivalent in term of sensitivity regardless of the matrix, whereas 1:3 ratio showed an average decrease of 48%, 35% and 22% of pic areas in plasma, blood and urine samples respectively. The LOI obtained with different organic solvent were identical except for few compounds. Acetonitrile provided lower LOI for sufentanyl (0.25 vs. 1 μg/L for other solvents) whereas methanol in urine showed better results for zopiclone (1 μg/L vs. 5 μg/L). Finally, the addition of QuEChERS salt and evaporation after precipitation doubled at least the average signal intensity for all drugs. Despite some benefits such as large injection volume and automation, the sensitivity of the method including TurboFlow® extraction did not show improvements compared with direct injection. Indeed, small hydrophilic molecules had generally less retention on TurboFlow® column resulting in lower recovery and signal suppression can occur due to many matrix components. The addition of formic acid for protein precipitation does not improve the sensitivity, neither ZnSO4 nor mix of solvents. Simple and providing adequate sensitivity, the use of acetonitrile as organic solvent for protein precipitation with a ratio of 1:2 was chosen for the 3 matrices for routine. The addition of QuEChERS salts and evaporation after precipitation could be dedicated to specific cases where very low LOI has to be reached.

Synthesis, biological evaluation, and molecular docking analysis of novel 1, 3, 4-thiadiazole -based kojic acid derivatives as tyrosinase inhibitors

Kojic acid is a natural tyrosinase inhibitor that has been clinically used to cure hyperpigmentation in humans. However, kojic acid as a hydrophilic small-molecule has deficient inhibitory activity and stability. In this current work, a new series of kojic acid derivatives, 4a–h bearing 2-amino-5-mercapto-1,3,4-thiadiazole, were synthesized using TBTU as the catalyst, and their chemical structures were characterized by spectroscopic methods. The inhibitory activities of synthesized compounds against Mushroom tyrosinase were evaluated in vitro, and some derivatives showed potent anti-tyrosinase activity. In particular, compounds 4g (IC50=10.71 ± 2.47 µM) and 4h (IC50=18.62 ± 3.05 µM) were comparable with the reference compound, kojic acid (IC50=23.14 µM). The Docking study was in good agreement with experimental results and indicated that compound 4g interacted entirely with the active site of tyrosinase with proper binding energy and mode.

Development of Carboxymethyl Chitosan Nanoparticles Prepared by Ultrasound-Assisted Technique for a Clindamycin HCl Carrier.

Polymeric nanoparticles are one method to modify the drug release of small hydrophilic molecules. In this study, clindamycin HCl was used as a model drug loaded in carboxymethyl chitosan nanoparticles cross-linked with Ca2+ ions (CMCS-Ca2+). The ultrasonication with experimental design was used to produce CMCS-Ca2+ nanoparticles loading clindamycin HCl. The model showed that the size of nanoparticles decreased when amplitude and time increased. The nanoparticle size of 318.40 ± 7.56 nm, decreased significantly from 543.63 ± 55.07 nm (p < 0.05), was obtained from 75% of amplitude and 180 s of time, which was one of the optimal conditions. The clindamycin loading content in this condition was 34.68 ± 2.54%. The drug content in nanoparticles showed an inverse relationship with the size of the nanoparticles. The sodium carboxymethylcellulose film loading clindamycin HCl nanoparticles exhibited extended release with 69.88 ± 2.03% drug release at 60 min and a gradual increase to 94.99 ± 4.70% at 24 h, and demonstrated good antibacterial activity against S. aureus and C. acne with 40.72 ± 1.23 and 48.70 ± 1.99 mm of the zone of inhibition at 24 h, respectively. Thus, CMCS-Ca2+ nanoparticles produced by the ultrasound-assisted technique could be a potential delivery system to modify the drug release of small hydrophilic antibiotics.

Optimization of Biocompatibility for a Hydrophilic Biological Molecule Encapsulation System.

Despite considerable advances in recent years, challenges in delivery and storage of biological drugs persist and may delay or prohibit their clinical application. Though nanoparticle-based approaches for small molecule drug encapsulation are mature, encapsulation of proteins remains problematic due to destabilization of the protein. Reverse micelles composed of decylmonoacyl glycerol (10MAG) and lauryldimethylamino-N-oxide (LDAO) in low-viscosity alkanes have been shown to preserve the structure and stability of a wide range of biological macromolecules. Here, we present a first step on developing this system as a future platform for storage and delivery of biological drugs by replacing the non-biocompatible alkane solvent with solvents currently used in small molecule delivery systems. Using a novel screening approach, we performed a comprehensive evaluation of the 10MAG/LDAO system using two preparation methods across seven biocompatible solvents with analysis of toxicity and encapsulation efficiency for each solvent. By using an inexpensive hydrophilic small molecule to test a wide range of conditions, we identify optimal solvent properties for further development. We validate the predictions from this screen with preliminary protein encapsulation tests. The insight provided lays the foundation for further development of this system toward long-term room-temperature storage of biologics or toward water-in-oil-in-water biologic delivery systems.

6-Glycosylaminoquinoline-assisted LDI MS for detection and imaging of small molecules with enhanced detection selectivity and sensitivity

Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) is a powerful tool in the analysis and imaging of small molecules. However, MALDI MS analysis is easily subjected to poor signal reproducibility and selectivity, especially for complex samples. In this study, a matrix glycosylation strategy was proposed to synthesize glycosylated matrices with excellent performances by enhancing the interaction of the matrix with small molecules. A series of glycosylated matrices including 3-glycosylaminoquinoline (3-GAQ), 6-glycosylaminoquinoline (6-GAQ), and 1-amino-5-glycosylaminoquinoline (GDAN) were synthesized by connecting glucose with the existing amine matrices. Compared with their parent matrices and the existing matrix (1,5-diaminonaphathelene, 1,5-DAN), the glycosylated matrices exhibited remarkably-improved sensitivity, higher signal reproducibility (RSD < 9%) in detecting metabolites, demonstrating the effectiveness of the glycosylation strategy. Among them, 6-GAQ exhibited the best performance. Using 6-GAQ, the detection limit of citric acid reached the low fmol range, and the calibration curve of citric acid had ideal linearity (R2 > 0.99), proving that 6-GAQ was capable of accurate quantitative analysis of metabolites. Furthermore, 6-GAQ was used for the imaging of metabolites in the mouse kidney section, showing higher sensitivity and lower background noise than the commonly-used matrices, 9-aminoquinoline (9-AA), and 1,5-DAN. More importantly, 6-GAQ can selectively detect the hydrophilic metabolites, especially the hydrophilic lipids in the mouse kidney. Overall, 6-GAQ is an ideal matrix potentially applied in the imaging and quantitative analysis of hydrophilic small molecules in complex samples.

Kinetic Study of Encapsulated β-Carotene Degradation in Dried Systems: A Review.

β-Carotene serves as a precursor of vitamin A and provides relevant health benefits. To overcome the low bioavailability of β-carotene from natural sources, technologies have been designed for its encapsulation in micro- and nano-structures followed by freeze-drying, spray-drying, supercritical fluid-enhanced dispersion and electrospraying. A technological challenge is also to increase β-carotene stability, since due to its multiple conjugated double bonds, it is particularly prone to oxidation. This review analyzes the stability of β-carotene encapsulated in different dried micro- and nano-structures by comparing rate constants and activation energies of degradation. The complex effect of water activity and glass transition temperature on degradation kinetics is also addressed, since the oxidation process is remarkably dependent on the glassy or collapsed state of the matrix. The approaches to improve β-carotene stability, such as the development of inclusion complexes, the improvement of the performance of the interface between air and oil phase in which β-carotene was dissolved by application of biopolymer combinations or functionalization of natural biopolymers, the addition of hydrophilic small molecular weight molecules that reduce air entrapped in the powder and the co-encapsulation of antioxidants of various polarities are discussed and compared, in order to provide a rational basis for further development of the encapsulation technologies.

Self‐Assembled PSMA‐Targeted Nanoparticles Enhanced Photodynamic Therapy in Prostate Cancer

Prostate cancer greatly threatens human health. Many treatment strategies including surgery and radiotherapy are applied to treat prostate cancer. However, these treatment strategies will bring great side effects. Herein, we developed a functionalized Prostate‐Specific Membrane Antigen‐Chlorin e6 (PSMA‐Ce6) NPs to enhance photodynamic therapy to reduce the side effects of prostate cancer treatments. We linked the hydrophobic photosensitizer Ce6 with the small hydrophilic molecule PSMA ligand through a covalent bond to form a functionalized PSMA‐Ce6. In aqueous phase, the PSMA‐Ce6 self‐assembled into nanoparticles with a hydrophobic core and a hydrophilic shell, which significantly enhanced the accumulation of PSMA‐Ce6 in the tumor. The molecularly targeted drug PSMA inhibitor can not only specifically inhibit PSMA but also increase the accumulation of the photosensitizer Ce6 in the tumor and achieve a completed tumor ablation under the near infrared (NIR) irradiation. PSMA‐Ce6 NPs have great potential for translation into clinical applications.

Efficient Delivery of Hydrophilic Small Molecules to Retinal Cell Lines Using Gel Core-Containing Solid Lipid Nanoparticles.

In this study, we developed a novel solid lipid nanoparticle (SLN) formulation for drug delivery of small hydrophilic cargos to the retina. The new formulation, based on a gel core and composite shell, allowed up to two-fold increase in the encapsulation efficiency. The type of hydrophobic polyester used in the composite shell mixture affected the particle surface charge, colloidal stability, and cell internalization profile. We validated SLNs as a drug delivery system by performing the encapsulation of a hydrophilic neuroprotective cyclic guanosine monophosphate analog, previously demonstrated to hold retinoprotective properties, and the best formulation resulted in particles with a size of ±250 nm, anionic charge > −20 mV, and an encapsulation efficiency of ±60%, criteria that are suitable for retinal delivery. In vitro studies using the ARPE-19 and 661W retinal cell lines revealed the relatively low toxicity of SLNs, even when a high particle concentration was used. More importantly, SLN could be taken up by the cells and the release of the hydrophilic cargo in the cytoplasm was visually demonstrated. These findings suggest that the newly developed SLN with a gel core and composite polymer/lipid shell holds all the characteristics suitable for the drug delivery of small hydrophilic active molecules into retinal cells.

Beyond hydrophilic polymers in amphiphilic polymer-based self-assembled NanoCarriers: Small hydrophilic carboxylate-capped disulfide drug delivery system and its multifunctionality and multispatial targetability

Due to increasing safety and intracellular delivery concerns about hydrophilic polymers in amphiphilic polymer-based nanoparticles (NPs), this study investigates small hydrophilic molecule-stabilized NPs for effective intracellular delivery with multiorganelle targetability and dual responsiveness to acidic pH/glutathione (GSH). In the construction of small hydrophilic molecule-stabilized NP (MSPCL-NP), the A-B-A-type amphiphilic polymer (MSPCL-P) is composed of two short hydrophilic carboxylate-capped disulfide derivatives (A) that replace hydrophilic polymers and assist in providing colloidal stability and preventing antibody (e.g., at least anti-PEG antibody)-mediated specific interactions and complement activation in the plasma and a hydrophobic multiple disulfide-containing poly(ε-caprolactone) block (B) that carries hydrophobic drugs. The carboxylates on the surface of MSPCL-NP target the acidic extratumoral/endolysosomal milieu by sensing and buffering acidic pH values, and the hydrophobic carboxylic acids improve adsorptive endocytosis and effective endosomal escape. Multiple disulfide linkages selectively target cytosolic GSH, resulting in rapid drug release from the destroyed MSPCL-NP via the cleavage of disulfide bonds in MSPCL-P. Doxorubicin (DOX)-loaded NP (DOX@MSPCL-NP) exerts strong effects on killing cells in vitro and inhibits tumor growth in HCT116 xenograft tumor-bearing mice. In conclusion, the multifunctionality and multispatial targetability of MSPCL-NP might effectively overcome various sequential drug delivery hurdles, ranging from blood circulation to drug release. Furthermore, the introduction of small hydrophilic molecules represents a potential strategy to make self-assembled NPs without the use of hydrophilic polymers.

Tumor-dilated polymersome nanofactories for enhanced enzyme prodrug chemo-immunotherapy

Combination chemo-immunotherapy of cancers has attracted great attention due to its significant synergistic antitumor effect. The response rates and therapeutic efficacy of immunotherapy can be enhanced significantly after proper combination with chemotherapy. However, chemo-immunotherapy is frequently limited by severe immune-related adverse events and systemic side toxicity. In this report, efficient nanofactory-directed enzyme prodrug chemo-immunotherapy is demonstrated based on enzyme-loaded tumor-dilatable polymersomes with optimized membrane cross-linking density. Upon intravenous injection of the nanofactories, they can passively accumulate at the tumor site. The tumor pH-responsive nanofactories can swell from ~100 nm to ~200 nm under the trigger of tumor acidity, leading to prolonged retention of up to one week inside tumor tissues. Simultaneously, the membrane permeability of the nanofactories has improved significantly, which allows hydrophilic small molecules to pass across the membranes while keeping the enzymes in the inner cavities. Subsequently, the non-toxic prodrug mixtures of chemo-immunotherapy are administrated three times within 6 days, which are in situ activated by the nanofactories selectively at tumor sites. Activated chemotherapeutic drugs kill cancer cells and generate tumor-associated antigens to promote the maturation of dendritic cells. Activated indoleamine 2, 3-dioxygenase 1 inhibitors reverse the immunosuppressive tumor microenvironment. Finally, primary tumors can be effectively suppressed while causing minimal systemic toxicity. The distant tumors that are established after treatment can also be inhibited completely via activation of antitumor immunity in mice. Thus, the tumor-dilatable polymersome nanofactories with long-term intratumoral retention offer a promising paradigm for enhanced enzyme prodrug chemo-immunotherapy.

Enhancing the Pharmacokinetic Profile of Interleukin 2 through Site-Specific Conjugation to a Selective Small-Molecule Transthyretin Ligand.

Protein drugs hold great promise as therapeutics for a wide range of diseases. Unfortunately, one of the greatest challenges to be addressed during clinical development of protein therapeutics is their short circulation half-life. Several protein conjugation strategies have been developed for half-life extension. However, these strategies have limitations and there remains room for improvement. Here, we report a novel nature-inspired strategy for enhancing the in vivo half-life of proteins. Our strategy involves conjugating proteins to a hydrophilic small molecule that binds reversibly to the plasma protein, transthyretin. We show here that our strategy is effective in enhancing the pharmacokinetic and pharmacodynamic properties of human interleukin 2 in rats, potentially opening the door for more effective and safer cancer immunotherapies. To our knowledge, this is the first example of successful use of a small-molecule that not only extends the half-life but also maintains the smaller size, binding potency, and hydrophilicity of proteins.