SNAC for Enhanced Oral Bioavailability: An Updated Review.

The delivery of proteins and peptides via an oral route poses numerous challenges to improve the oral bioavailability and patient compliance. To overcome these challenges, as well as to improve the permeation of proteins and peptides via intestinal mucosa, several chemicals have been studied such as surfactants, fatty acids, bile salts, pH modifiers, and chelating agents, amongst these medium chain fatty acid like C10 (sodium caprate) and Sodium N-[8-(2-hydroxybenzoyl) amino] caprylate (SNAC) and its derivatives that have been well studied from a clinical perspective. This current review enumerates the challenges involved in protein and peptide delivery via the oral route, i.e., non-invasive routes of protein and peptide administration. This review also covers the chemistry behind SNAC and toxicity as well as mechanisms to enhance the oral delivery of clinically proven molecules like simaglutide and other small molecules under clinical development, as well as other permeation enhancers for efficient delivery of proteins and peptides.

Drug Reprofiling to Identify Potential HIV-1 Protease Inhibitors.

The use of protease inhibitors in human immunodeficiency virus type 1 (HIV-1) treatment is limited by adverse effects, including metabolic complications. To address these challenges, efforts are underway in the pursuit of more potent and less toxic HIV-1 protease inhibitors. Repurposing existing drugs offers a promising avenue to expedite the drug discovery process, saving both time and costs compared to conventional de novo drug development. This study screened FDA-approved and investigational drugs in the DrugBank database for their potential as HIV-1 protease inhibitors. Molecular docking studies and cell-based assays, including anti-HIV-1 in vitro assays and XTT cell viability tests, were conducted to evaluate their efficacy. The study findings revealed that CBR003PS, an antibiotic currently in clinical use, and CBR013PS, an investigational drug for treating endometriosis and uterine fibroids, exhibited significant binding affinity to the HIV-1 protease with high stability. Their EC50 values, measured at 100% cell viability, were 9.4 nM and 36.6 nM, respectively. Furthermore, cell-based assays demonstrated that these two compounds showed promising results, with therapeutic indexes higher than 32. In summary, based on their favorable therapeutic indexes, CBR003PS and CBR013PS show potential for repurposing as HIV-1 protease inhibitors.

Discovery of Novel HIV Protease Inhibitors Using Modern Computational Techniques.

The human immunodeficiency virus type 1 (HIV-1) has continued to be a global concern. With the new HIV incidence, the emergence of multi-drug resistance and the untoward side effects of currently used anti-HIV drugs, there is an urgent need to discover more efficient anti-HIV drugs. Modern computational tools have played vital roles in facilitating the drug discovery process. This research focuses on a pharmacophore-based similarity search to screen 111,566,735 unique compounds in the PubChem database to discover novel HIV-1 protease inhibitors (PIs). We used an in silico approach involving a 3D-similarity search, physicochemical and ADMET evaluations, HIV protease-inhibitor prediction (IC50/percent inhibition), rigid receptor-molecular docking studies, binding free energy calculations and molecular dynamics (MD) simulations. The 10 FDA-approved HIV PIs (saquinavir, lopinavir, ritonavir, amprenavir, fosamprenavir, atazanavir, nelfinavir, darunavir, tipranavir and indinavir) were used as reference. The in silico analysis revealed that fourteen out of the twenty-eight selected optimized hit molecules were within the acceptable range of all the parameters investigated. The hit molecules demonstrated significant binding affinity to the HIV protease (PR) when compared to the reference drugs. The important amino acid residues involved in hydrogen bonding and п-п stacked interactions include ASP25, GLY27, ASP29, ASP30 and ILE50. These interactions help to stabilize the optimized hit molecules in the active binding site of the HIV-1 PR (PDB ID: 2Q5K). HPS/002 and HPS/004 have been found to be most promising in terms of IC50/percent inhibition (90.15%) of HIV-1 PR, in addition to their drug metabolism and safety profile. These hit candidates should be investigated further as possible HIV-1 PIs with improved efficacy and low toxicity through in vitro experiments and clinical trial investigations.

Biomimetic Targeted Theranostic Nanoparticles for Breast Cancer Treatment.

The development of biomimetic drug delivery systems for biomedical applications has attracted significant research attention. As the use of cell membrane as a surface coating has shown to be a promising platform for several disease treatments. Cell-membrane-coated nanoparticles exhibit enhanced immunocompatibility and prolonged circulation time. Herein, human red blood cell (RBC) membrane-cloaked nanoparticles with enhanced targeting functionality were designed as a targeted nanotheranostic against cancer. Naturally, derived human RBC membrane modified with targeting ligands coated onto polymeric nanoparticle cores containing both chemotherapy and imaging agent. Using epithelial cell adhesion molecule (EpCAM)-positive MCF-7 breast cancer cells as a disease model, the nature-inspired targeted theranostic human red blood cell membrane-coated polymeric nanoparticles (TT-RBC-NPs) platform was capable of not only specifically binding to targeted cancer cells, effectively delivering doxorubicin (DOX), but also visualizing the targeted cancer cells. The TT-RBC-NPs achieved an extended-release profile, with the majority of the drug release occurring within 5 days. The TT-RBC-NPs enabled enhanced cytotoxic efficacy against EpCAM positive MCF-7 breast cancer over the non-targeted NPs. Additionally, fluorescence images of the targeted cancer cells incubated with the TT-RBC-NPs visually indicated the increased cellular uptake of TT-RBC-NPs inside the breast cancer cells. Taken together, this TT-RBC-NP platform sets the foundation for the next-generation stealth theranostic platforms for systemic cargo delivery for treatment and diagnostic of cancer.

Nanoparticle biointerfacing by platelet membrane cloaking.

Development of functional nanoparticles can be encumbered by unanticipated material properties and biological events, which can affect nanoparticle effectiveness in complex, physiologically relevant systems. Despite the advances in bottom-up nanoengineering and surface chemistry, reductionist functionalization approaches remain inadequate in replicating the complex interfaces present in nature and cannot avoid exposure of foreign materials. Here we report on the preparation of polymeric nanoparticles enclosed in the plasma membrane of human platelets, which are a unique population of cellular fragments that adhere to a variety of disease-relevant substrates. The resulting nanoparticles possess a right-side-out unilamellar membrane coating functionalized with immunomodulatory and adhesion antigens associated with platelets. Compared to uncoated particles, the platelet membrane-cloaked nanoparticles have reduced cellular uptake by macrophage-like cells and lack particle-induced complement activation in autologous human plasma. The cloaked nanoparticles also display platelet-mimicking properties such as selective adhesion to damaged human and rodent vasculatures as well as enhanced binding to platelet-adhering pathogens. In an experimental rat model of coronary restenosis and a mouse model of systemic bacterial infection, docetaxel and vancomycin, respectively, show enhanced therapeutic efficacy when delivered by the platelet-mimetic nanoparticles. The multifaceted biointerfacing enabled by the platelet membrane cloaking method provides a new approach in developing functional nanoparticles for disease-targeted delivery.

Macrophage-like nanoparticles concurrently absorbing endotoxins and proinflammatory cytokines for sepsis management.

Sepsis, resulting from uncontrolled inflammatory responses to bacterial infections, continues to cause high morbidity and mortality worldwide. Currently, effective sepsis treatments are lacking in the clinic, and care remains primarily supportive. Here we report the development of macrophage biomimetic nanoparticles for the management of sepsis. The nanoparticles, made by wrapping polymeric cores with cell membrane derived from macrophages, possess an antigenic exterior the same as the source cells. By acting as macrophage decoys, these nanoparticles bind and neutralize endotoxins that would otherwise trigger immune activation. In addition, these macrophage-like nanoparticles sequester proinflammatory cytokines and inhibit their ability to potentiate the sepsis cascade. In a mouse Escherichia coli bacteremia model, treatment with macrophage mimicking nanoparticles, termed MΦ-NPs, reduced proinflammatory cytokine levels, inhibited bacterial dissemination, and ultimately conferred a significant survival advantage to infected mice. Employing MΦ-NPs as a biomimetic detoxification strategy shows promise for improving patient outcomes, potentially shifting the current paradigm of sepsis management.

Biomimetic Micromotor Enables Active Delivery of Antigens for Oral Vaccination.

Vaccination represents one of the most effective means of preventing infectious disease. In order to maximize the utility of vaccines, highly potent formulations that are easy to administer and promote high patient compliance are desired. In the present work, a biomimetic self-propelling micromotor formulation is developed for use as an oral antivirulence vaccine. The propulsion is provided by a magnesium-based core, and a biomimetic cell membrane coating is used to detain and neutralize a toxic antigenic payload. The resulting motor toxoids leverage their propulsion properties in order to more effectively elicit mucosal immune responses. After demonstrating the successful fabrication of the motor toxoids, their uptake properties are shown in vitro. When delivered to mice via an oral route, it is then confirmed that the propulsion greatly improves retention and uptake of the antigenic material in the small intestine in vivo. Ultimately, this translates into markedly elevated generation of antibody titers against a model toxin. This work provides a proof-of-concept highlighting the benefits of active oral delivery for vaccine development, opening the door for a new set of applications, in which biomimetic motor technology can provide significant benefits.

Inhibition of Pathogen Adhesion by Bacterial Outer Membrane-Coated Nanoparticles.

Anti-adhesion therapies interfere with the bacterial adhesion to the host and thus avoid direct disruption of bacterial cycles for killing, which may alleviate resistance development. Herein, an anti-adhesion nanomedicine platform is made by wrapping synthetic polymeric cores with bacterial outer membranes. The resulting bacterium-mimicking nanoparticles (denoted "OM-NPs") compete with source bacteria for binding to the host. The "top-down" fabrication of OM-NPs avoids the identification of the adhesins and bypasses the design of agonists targeting these adhesins. In this study, OM-NPs are made with the membrane of Helicobacter pylori and shown to bind with gastric epithelial cells (AGS cells). Treatment of AGS cells with OM-NPs reduces H. pylori adhesion and such anti-adhesion efficacy is dependent on OM-NP concentration and its dosing sequence.

Toxoid Vaccination against Bacterial Infection Using Cell Membrane-Coated Nanoparticles.

As nanoparticles exhibit unique properties attractive for vaccine development, they have been progressively implemented as antigen delivery platforms and immune potentiators. Recently, cell membrane-coated nanoparticles have provided a novel approach for intercepting and neutralizing bacterial toxins by leveraging their natural affinity to cellular membranes. Such toxin-nanoparticle assemblies, termed nanotoxoids, allow rapid loading of different types of toxins and have been investigated for their ability to effectively confer protection against bacterial infection. This topical review will cover the current progress in antibacterial vaccine nanoformulations and highlight the nanotoxoid platform as a novel class of nanoparticulate vaccine. We aim to provide insights into the potential of nanotoxoids as a platform that is facile to implement and can be broadly applied to help address the rising threat of super pathogens.

Active Intracellular Delivery of a Cas9/sgRNA Complex Using Ultrasound-Propelled Nanomotors.

Direct and rapid intracellular delivery of a functional Cas9/sgRNA complex using ultrasound-powered nanomotors is reported. The Cas9/sgRNA complex is loaded onto the nanomotor surface through a reversible disulfide linkage. A 5 min ultrasound treatment enables the Cas9/sgRNA-loaded nanomotors to directly penetrate through the plasma membrane of GFP-expressing B16F10 cells. The Cas9/sgRNA is released inside the cells to achieve highly effective GFP gene knockout. The acoustic Cas9/sgRNA-loaded nanomotors display more than 80 % GFP knockout within 2 h of cell incubation compared to 30 % knockout using static nanowires. More impressively, the nanomotors enable highly efficient knockout with just 0.6 nm of the Cas9/sgRNA complex. This nanomotor-based intracellular delivery method thus offers an attractive route to overcome physiological barriers for intracellular delivery of functional proteins and RNAs, thus indicating considerable promise for highly efficient therapeutic applications.

Micromotors Spontaneously Neutralize Gastric Acid for pH-Responsive Payload Release.

The highly acidic gastric environment creates a physiological barrier for using therapeutic drugs in the stomach. While proton pump inhibitors have been widely used for blocking acid-producing enzymes, this approach can cause various adverse effects. Reported herein is a new microdevice, consisting of magnesium-based micromotors which can autonomously and temporally neutralize gastric acid through efficient chemical propulsion in the gastric fluid by rapidly depleting the localized protons. Coating these micromotors with a cargo-containing pH-responsive polymer layer leads to autonomous release of the encapsulated payload upon gastric-acid neutralization by the motors. Testing in a mouse model demonstrate that these motors can safely and rapidly neutralize gastric acid and simultaneously release payload without causing noticeable acute toxicity or affecting the stomach function, and the normal stomach pH is restored within 24 h post motor administration.

Remote Loading of Small-Molecule Therapeutics into Cholesterol-Enriched Cell-Membrane-Derived Vesicles.

The increasing popularity of biomimetic design principles in nanomedicine has led to therapeutic platforms with enhanced performance and biocompatibility. This includes the use of naturally derived cell membranes, which can bestow nanocarriers with cell-specific functionalities. Herein, we report on a strategy enabling efficient encapsulation of drugs via remote loading into membrane vesicles derived from red blood cells. This is accomplished by supplementing the membrane with additional cholesterol, stabilizing the nanostructure and facilitating the retention of a pH gradient. We demonstrate the loading of two model drugs: the chemotherapeutic doxorubicin and the antibiotic vancomycin. The therapeutic implications of these natural, remote-loaded nanoformulations are studied both in vitro and in vivo using animal disease models. Ultimately, this approach could be used to design new biomimetic nanoformulations with higher efficacy and improved safety profiles.

Nanoparticle-Based Antivirulence Vaccine for the Management of Methicillin-Resistant Staphylococcus aureus Skin Infection.

With the rising threat of antibiotic-resistant bacteria, vaccination is becoming an increasingly important strategy to prevent and manage bacterial infections. Made from deactivated bacterial toxins, toxoid vaccines are widely used in the clinic as they help to combat the virulence mechanisms employed by different pathogens. Herein, the efficacy of a biomimetic nanoparticle-based anti-virulence vaccine is examined in a mouse model of methicillin-resistant Staphylococcus aureus (MRSA) skin infection. Vaccination with nanoparticle-detained staphylococcal α-hemolysin (Hla) effectively triggers the formation of germinal centers and induces high anti-Hla titers. Compared to mice vaccinated with control samples, those vaccinated with the nanoparticle toxoid show superior protective immunity against MRSA skin infection. The vaccination not only inhibits lesion formation at the site of bacterial challenge, but also reduces the invasiveness of MRSA, preventing dissemination into other organs. Overall, this biomimetic nanoparticle-based toxin detainment strategy is a promising method for the design of potent anti-virulence vaccines for managing bacterial infections.

Modulating antibacterial immunity via bacterial membrane-coated nanoparticles.

Synthetic nanoparticles coated with cellular membranes have been increasingly explored to harness natural cell functions toward the development of novel therapeutic strategies. Herein, we report on a unique bacterial membrane-coated nanoparticle system as a new and exciting antibacterial vaccine. Using Escherichia coli as a model pathogen, we collect bacterial outer membrane vesicles (OMVs) and successfully coat them onto small gold nanoparticles (AuNPs) with a diameter of 30 nm. The resulting bacterial membrane-coated AuNPs (BM-AuNPs) show markedly enhanced stability in biological buffer solutions. When injected subcutaneously, the BM-AuNPs induce rapid activation and maturation of dendritic cells in the lymph nodes of the vaccinated mice. In addition, vaccination with BM-AuNPs generates antibody responses that are durable and of higher avidity than those elicited by OMVs only. The BM-AuNPs also induce an elevated production of interferon gamma (INFγ) and interleukin-17 (IL-17), but not interleukin-4 (IL-4), indicating its capability of generating strong Th1 and Th17 biased cell responses against the source bacteria. These observed results demonstrate that using natural bacterial membranes to coat synthetic nanoparticles holds great promise for designing effective antibacterial vaccines.

Deep Eutectic Solvents for Subcutaneous Delivery of Protein Therapeutics.

Proteins are among the most common therapeutics for the treatment of diabetes, autoimmune diseases, cancer, and metabolic diseases, among others. Despite their common use, current protein therapies, most of which are injectables, have several limitations. Large proteins such as monoclonal antibodies (mAbs) suffer from poor absorption after subcutaneous injections, thus forcing their administration by intravenous injections. Even small proteins such as insulin suffer from slow pharmacokinetics which poses limitations in effective management of diabetes. Here, a deep eutectic-based delivery strategy is used to offer a generalized approach for improving protein absorption after subcutaneous injections. The lead formulation enhances absorption of mAbs after subcutaneous injections by ≈200%. The same composition also improves systemic absorption of subcutaneously injected insulin faster than Humalog, the current gold-standard of rapid acting insulin. Mechanistic studies reveal that the beneficial effect of deep eutectics on subcutaneous absorption is mediated by their ability to reduce the interactions of proteins with the subcutaneous matrix, especially collagen. Studies also confirm that these deep eutectics are safe for subcutaneous injections. Deep eutectic-based formulations described here open new possibilities for subcutaneous injections of therapeutic proteins.

Development of Mucoadhesive Electrospun Scaffolds for Intravaginal Delivery of Lactobacilli spp., a Tenside, and Metronidazole for the Management of Bacterial Vaginosis.

Bacterial vaginosis (BV) is an infection of the vagina associated with thriving anaerobes, such as Gardnerella vaginitis and other associated pathogens. These pathogens form a biofilm responsible for the recurrence of infection after antibiotic therapy. The aim of this study was to develop a novel mucoadhesive polyvinyl alcohol and polycaprolactone electrospun nanofibrous scaffolds for vaginal delivery, incorporating metronidazole, a tenside, and Lactobacilli. This approach to drug delivery sought to combine an antibiotic for bacterial clearance, a tenside biofilm disruptor, and a lactic acid producer to restore healthy vaginal flora and prevent the recurrence of bacterial vaginosis. F7 and F8 had the least ductility at 29.25% and 28.39%, respectively, and this could be attributed to the clustering of particles that prevented the mobility of the crazes. F2 had the highest at 93.83% due to the addition of a surfactant that increased the affinity of the components. The scaffolds exhibited mucoadhesion between 31.54 ± 0.83% and 57.86 ± 0.95%, where an increased sodium cocoamphoacetate concentration led to increased mucoadhesion. F6 showed the highest mucoadhesion at 57.86 ± 0.95%, as compared to 42.67 ± 1.22% and 50.89 ± 1.01% for the F8 and F7 scaffolds, respectively. The release of metronidazole via a non-Fickian diffusion-release mechanism indicated both swelling and diffusion. The anomalous transport within the drug-release profile pointed to a drug-discharge mechanism that combined both diffusion and erosion. The viability studies showed a growth of Lactobacilli fermentum in both the polymer blend and the nanofiber formulation that was retained post-storage at 25 °C for 30 days. The developed electrospun scaffolds for the intravaginal delivery of Lactobacilli spp., along with a tenside and metronidazole for the management of bacterial vaginosis, provide a novel tool for the treatment and management of recurrent vaginal infection.

Freeze-drying for the preservation of immunoengineering products.

Immunoengineering technologies harness the power of immune system modulators such as monoclonal antibodies, cytokines, and vaccines to treat myriad diseases. Immunoengineering innovations have showed great promise in various practices including oncology, infectious disease, autoimmune diseases, and transplantation. Despite the countless successes, the majority of immunoengineering products contain active moieties that are prone to instability. The current review aims to feature freeze-drying as a robust and scalable solution to the inherent stability challenges in immunoengineering products by preventing the active moiety from degradation. Furthermore, this review describes the stability issues related to immunoengineering products and the utility of the lyophilization process to preserve the integrity and efficacy of immunoengineering tools ranging from biologics to nanoparticle-based vaccines. The concept of the freeze-drying process is described highlighting the quality by design (QbD) for robust process optimization. Case studies of lyophilized immunoengineering technologies and relevant clinical studies using immunoengineering products are discussed.

Modulation of Gastrointestinal Mucus Properties with Ionic Liquids for Drug Delivery.

The mucus barrier lining the gastrointestinal tract poses a significant barrier to the oral delivery of macromolecular drugs. Successful approaches to overcoming this barrier have primarily focused on reducing drug and carrier interactions with mucus or disrupting the mucus layer directly. Choline-based ionic liquids (ILs) such as choline geranate and choline glycolate (CGLY) have recently been shown to be effective in enhancing the intestinal absorption of macromolecules such as insulin and immunoglobulin (IgG), respectively. Herein, the use of choline-based ILs as mucus-modulating agents for safely improving drug penetration through mucus is described. Choline-based ILs significantly increase the diffusion rates of cationic dextrans through mucin solution. Choline-maleic acid (CMLC 2:1) enhances the diffusion of 4 kDa cationic dextran in mucin solution by more than fourfold when compared to phosphate-buffered saline control. Choline-based ILs also reduce mucus viscosity without significantly impacting the native mucus gel structure. In vitro studies in a mucus-secreting coculture model with Caco-2 and HT29MTX-E12 cells further demonstrate the effectiveness of ILs in improving transport of cationic molecules in the presence of secreted mucus. This work demonstrates the potential for choline-based ionic liquids to be used as nondestructive mucus-modulating agents for enabling enhanced oral delivery of macromolecular drugs.

Amphiphilic Polyacrylamide Excipients Lead to a Record-Breaking Fast-Acting Insulin.

Fast-acting insulins are central to the regulation of prandial glucose in diabetic patients. Current fast-acting insulins require 20-30 min for the onset and longer for the peak blood concentrations. The recent work by Mann et al. used high-throughput synthesis and screening of polyacrylamide-based excipients to yield a formulation with pharmacokinetics that is faster than the currently available fast-acting insulins.

Ionic-Liquid-Based Safe Adjuvants.

Adjuvants play a critical role in the design and development of novel vaccines. Despite extensive research, only a handful of vaccine adjuvants have been approved for human use. Currently used adjuvants are mostly composed of components that are non-native to the human body, such as aluminum salt, bacterial lipids, or foreign genomic material. Here, a new ionic-liquid-based adjuvant is explored, synthesized using two metabolites of the body, choline and lactic acid (ChoLa). ChoLa distributes the antigen efficiently upon injection, maintains antigen integrity, enhances immune infiltration at the injection site, and leads to a potent immune response against the antigen. Thus, it can serve as a promising safe adjuvant platform that can help to protect against pandemics and future infectious threats.