Delivery Of Peptides Research Articles

Strategies for transportation of peptides across the skin for treatment of multiple diseases.

An established view in genetic engineering dictates an increase in the discovery of therapeutic peptides to enable the treatment of multiple diseases. The use of hypodermic needle for delivery of proteins and peptides occurs due to the hydrophilic nature, sensitivity toward proteolytic enzymes and high molecular weight. The non-invasive nature of the transdermal delivery technique offers multiple advantages over the invasive route to release drugs directly into the systemic circulation to enhance bioavailability, better patient compliance, reduced toxicity and local irritability. The transdermal route seems highly desirable from the pharmaco-therapeutic and patient compliance point of view, however, the lipophilic barrier of skin restricts the application. The use of several techniques like electrical methods (iontophoresis, sonophoresis etc.), chemical penetration enhancers (e.g. protease inhibitors, penetration enhancers, etc.) and nanocarriers (dendrimers, lipid nanocapsules, etc.) are utilized to improve the passage of drug molecules across the biomembranes. Additionally, such clinical interventions facilitate the physicochemical characteristics of peptides, to enable effective preservation, conveyance and release of therapeutic agents. Moreover, strategies ensure the attainment of the intended targets and enhance treatment outcomes for multiple diseases. This review article focuses on the techniques of peptide transportation across the skin to advance the delivery approaches and therapeutic efficiency.

Advances in Mucoadhesive drug Delivery System: Enhancing Efficacy and Patient Compliance

The goal of mucoadhesive drug delivery systems (MDDS) is to improve drug delivery effectiveness by adhering to the body's mucosal tissues. The material used in this approach adheres to mucosal surfaces, including those in the mouth, gastrointestinal system, nasal passages, and vaginal walls. The most important goal is to extend the medication's residence duration at the absorption site in order to improve drug absorption and provide a more regulated and prolonged release. Mucoadhesion refers to the adhesion between two materials, one of which is biological, through interfacial forces over an extended period. This characteristic provides an effective solution to challenges in traditional drug delivery systems, such as avoiding first-pass metabolism and facilitating localized delivery of biomolecules, including proteins, peptides, and oligonucleotides. It holds significant promise for delivering compounds via routes like ocular, nasal, vaginal, and buccal administration This study addresses the introduction, benefits, drawbacks, mechanisms, and theories of mucoadhesion in addition to polymer categorization, mucoadhesive dosage forms, assessment, and factors affecting mucoadhesion and several kinds of mucoadhesive dosage forms. Keywords: Mucoadhesion, Theories, Mucoadhesive Dosage Form, bio adhesion

Tumor-specific antigen delivery for T-cell therapy via a pH-sensitive peptide conjugate.

Identifying an optimal antigen for targeted cancer therapy is challenging as the antigen landscape on cancerous tissues mimics that of healthy tissues, with few unique tumor-specific antigens identified in individual patients. pH low insertion peptides (pHLIPs) act as a unique delivery platform that can specifically target the acidic microenvironment of tumors, sparing healthy tissue in the process. We developed a pHLIP-peptide conjugate to deliver the SIINFEKL peptide, an immunogenic fragment of ovalbumin, to tumor cells in vivo. When processed intracellularly, SIINFEKL is presented for immune recognition through the major histocompatibility complex (MHC) class I pathway. We observed selective delivery of pHLIP-SIINFEKL both in vitro and in vivo using fluorescently labeled constructs. In vitro, treatment of melanoma tumor cells with pHLIP-SIINFEKL resulted in recognition by SIINFEKL-specific T cells (OT1), leading to T cell activation and effector function. Mechanistically, we show that this recognition by OT1 cells was abrogated by siRNA/shRNA knockdown of multiple components within the MHC class I pathway in the target tumor cells, indicating that an intact antigen processing pathway in the cancer cells is necessary to mediate the effect of pHLIP-directed SIINFEKL delivery. In vivo, pHLIP-SIINFEKL treatment of tumor-bearing mice resulted in recruitment of OT1 T cells and suppression of tumor growth in two syngeneic tumor models in immunocompetent mice, with no effect when mutating either the pHLIP or SIINFEKL components of the conjugate. These results suggest that pHLIP-mediated peptide delivery can be used to deliver novel artificial antigens that can be targeted by cell-based therapies.

Antiaging synergistic effect in noninvasive transdermal delivery of peptide loaded liposomes by low energy/frequency radiofrequency

Low energy/frequency radiofrequency (LRF) combined with the transdermal delivery of liposome (L) encapsulated antiaging peptides technology is a remarkable, newly developed physical noninvasive transdermal penetration technique; it is considered a highly efficient, comprehensive and safe technology. In this study, our objective was to evaluate the physical and chemical mechanisms underlying the efficacy of this innovative technique involving a combination of LRF and L, termed LLRF, that exerts a synergistic anti-aging effect on human skin, via an animal experiment. Physical and chemical analyses indicated that a relatively stable liposome with a uniform nano-size, which was formed, possessed good transdermal permeability that was 2.74 folds higher than that of the free peptide (F). LLRF exhibited a higher transdermal permeation performance that was of 3.65 folds higher than that of the free one, which was substantiated via confocal laser scanning fluorescence microscopy. The mouse UVB photoaging model trial confirmed that the LLRF technology exerted a significant synergistic effect compared to liposome technology, or free peptide, by downregulating inflammatory factors (IL-6, TNF-α), inhibiting the mRNA and protein expression of matrix metalloproteinases (MMP1, MMP3), promoting the mRNA and protein expression of related collagens (Procollagen, Col1α1 and Col3α1), and repairing the stratum corneum barrier function, as evidenced by trans-epidermal water loss (TEWL), skin cuticle hydration (SCH), and decreased expression of β-gal, an aging marker. These findings indicated that photoaging skin can be effectively and comprehensively rejuvenated, and that even photodamage can be reversed, thereby restoring the original physiological characteristics of healthy skin. Clinical tests have confirmed that although liposome technology is an effective antiaging method which helps exert tightening and anti-wrinkle effects on human skin, LLRF is an even more effective anti-aging technique. This study reveals a highly effective technique involving a combination physical and chemical therapy that may be utilized for antiaging purposes as well as repairing lightly damaged skin, and can be made readily available in the future.

Chitosan-melanin complex microsphere: A potential colonic delivery system for protein drugs

The characteristics and performance of chitosan-based colon delivery systems are significantly influenced by the method of preparation. Insect chitosan-melanin complex (CMC) may offer superior attributes over traditional shrimp and crab chitosan (CS) for colon-targeted administration. This study used dung beetle CMC as the carrier matrix and comprehensively examined the impact of various crosslinking techniques on the colonic drug delivery efficacy of microspheres, encompassing drug loading, swelling, drug release behavior, adhesion, enzymatic degradation, and absorption enhancement. The results indicate that F-TPPLC microspheres, crosslinked with a combination of formaldehyde and TPP, exhibit superior drug loading capabilities, optimal swelling behavior, and controlled in vitro drug release profiles in the colonic environment, along with excellent adhesion and enzymatic degradation properties within intestinal tract. Notably, these F-TPPLC microspheres increase paracellular permeability, possibly by disrupting the calcium-dependent adhesion junctions. In comparison to commercial CS, CMC demonstrates superior drug encapsulation efficiency, enhanced colonic drug release, adhesion, and absorption promotion, rendering it a favorable candidate as a carrier in colon-targeted drug delivery systems. Consequently, F-TPPLC microspheres derived from CMC are highly suitable for colon drug delivery applications and show promising potential for the oral delivery of peptide and protein-based therapeutics to the colon.

Recombinant ferritin-based nanoparticles as neoantigen carriers significantly inhibit tumor growth and metastasis

BackgroundTumor neoantigen peptide-based vaccines, systemic immunotherapies that enhance antitumor immunity by activating and expanding antigen-specific T cells, have achieved remarkable results in the treatment of a variety of solid tumors. However, how to effectively deliver neoantigens to induce robust antitumor immune responses remains a major obstacle.ResultsHere, we developed a safe and effective neoantigen peptide delivery system (neoantigen-ferritin nanoparticles, neoantigen-FNs) that successfully achieved effective lymph node targeting and induced robust antitumor immune responses. The genetically engineered self-assembled particles neoantigen-FNs with a size of 12 nm were obtained by fusing a neoantigen with optimized ferritin, which rapidly drainage to and continuously accumulate in lymph nodes. The neoantigen-FNs vaccine induced a greater quantity and quality of antigen-specific CD8+ T cells and resulted in significant growth control of multiple tumors, dramatic inhibition of melanoma metastasis and regression of established tumors. In addition, no obvious toxic side effects were detected in the various models, indicating the high safety of optimized ferritin as a vaccine carrier.ConclusionsHomogeneous and safe neoantigen-FNs could be a very promising system for neoantigen peptide delivery because of their ability to efficiently drainage to lymph nodes and induce efficient antitumor immune responses.Graphical

Multi-site esterification: a tunable, reversible strategy to tailor therapeutic peptides for delivery.

Peptides are naturally potent and selective therapeutics with massive potential; however, low cell membrane permeability limits their clinical implementation, particularly for hydrophilic, anionic peptides with intracellular targets. To overcome this limitation, esterification of anionic carboxylic acids on therapeutic peptides can simultaneously increase hydrophobicity and net charge to facilitate cell internalization, whereafter installed esters can be cleaved hydrolytically to restore activity. To date, however, most esterified therapeutics contain either a single esterification site or multiple esters randomly incorporated on multiple sites. This investigation provides molecular engineering insight into how the number and position of esters installed onto the therapeutic peptide α carboxyl terminus 11 (αCT11, RPRPDDLEI) with 4 esterification sites affect hydrophobicity and the hydrolysis process that reverts the peptide to its original form. After installing methyl esters onto αCT11 using Fischer esterification, we isolated 5 distinct products and used 2D nuclear magnetic resonance spectroscopy, reverse-phase high performance liquid chromatography, and mass spectrometry to determine which residues were esterified in each and the resulting increase in hydrophobicity. We found esterifying the C-terminal isoleucine to impart the largest increase in hydrophobicity. Monitoring ester hydrolysis showed the C-terminal isoleucine ester to be the most hydrolytically stable, followed by the glutamic acid, whereas esters on aspartic acids hydrolyze rapidly. LC-MS revealed the formation of transient intramolecular aspartimides prior to hydrolysis to carboxylic acids. In vitro proof-of-concept experiments showed esterifying αCT11 to increase cell migration into a scratch, highlighting the potential of multi-site esterification as a tunable, reversible strategy to enable the delivery of therapeutic peptides.

Enhanced Tumor-Targeted Delivery of Arginine-Rich Peptides via a Positive Feedback Loop Orchestrated by Piezo1/integrin β1 Signaling Axis.

Peptide-based drugs hold great potential for cancer treatment, and their effectiveness is driven by mechanisms on how peptides target cancer cells and escape from potential lysosomal entrapment post-endocytosis. Yet, the mechanisms remain elusive, which hinder the design of peptide-based drugs. Here hendeca-arginine peptides (R11) are synthesized for targeted delivery in bladder carcinoma (BC), investigated the targeting efficiency and elucidated the mechanism of peptide-based delivery, with the aim of refining the design and efficacy of peptide-based therapeutics. It is demonstrated that the over-activated Piezo1/integrin β1 (ITGB1) signaling axis significantly facilitates tumor-targeted delivery of R11 peptides via macropinocytosis. Furthermore, R11 peptides formed hydrogen bonds with integrin β1, facilitating targeting and penetration into tumor cells. Additionally, R11 peptides protected integrin β1 from lysosome degradation, promoting its recycling from cytoplasm to membrane. Moreover, this findings establish a positive feedback loop wherein R11 peptides activate Piezo1 by increasing membrane fusion, promoting Ca2+ releasing and resulting in enhanced integrin β1-mediated endocytosis in both orthotopic models and clinical tissues, demonstrating effective tumor-targeted delivery. Eventually, the Piezo1/integrin β1 signaling axis promoted cellular uptake and transport of peptides, establishing a positive feedback loop, promoting mechanical delivery to cancer and offering possibilities for drug modification in cancer therapy.

Microneedles for Proteins and Peptides Delivery: Current Aspect and Future Perspective

Microneedles for Proteins and Peptides Delivery: Current Aspect and Future Perspective

Biomimetic Supramolecular Assembly with IGF-1C Delivery Ameliorates Inflammatory Bowel Disease (IBD) by Restoring Intestinal Barrier Integrity.

The management of dysfunctional intestinal epithelium by promoting mucosal healing and modulating the gut microbiota represents a novel therapeutic strategy for inflammatory bowel disease (IBD). As a convenient and well-tolerated method of drug delivery, intrarectal administration may represent a viable alternative to oral administration for the treatment of IBD. Here, a biomimetic supramolecular assembly of hyaluronic acid (HA) and β-cyclodextrin (HA-β-CD) for the delivery of the C domain peptide of insulin-like growth factor-1 (IGF-1C), which gradually releases IGF-1C, is developed. It is identified that the supramolecular assembly of HA-β-CD enhances the stability and prolongs the release of IGF-1C. Furthermore, this biomimetic supramolecular assembly potently inhibits the inflammatory response, thereby restoring intestinal barrier integrity. Following HA-β-CD-IGF-1C administration, 16S rDNA sequencing reveals a significant increase in the abundance of the probiotic Akkermansia, suggesting enhanced intestinal microbiome homeostasis. In conclusion, the findings demonstrate the promise of the HA-based mimicking peptide delivery platform as a therapeutic approach for IBD. This biomimetic supramolecular assembly effectively ameliorates intestinal barrier function and intestinal microbiome homeostasis, suggesting its potential for treating IBD.

Development and Characterization of a Peptide-Bisphosphonate Nanoparticle for the Treatment of Breast Cancer.

In women, breast cancer (BC) is the most common cancer, and despite advancements in diagnosis and treatment, 20-30% of early stage BC patients develop metastatic disease. Metastatic BC is deemed an incurable disease, which accounts for 90% of BC related deaths, with only 26% of metastatic patients reaching a 5 year survival rate. Therefore, there is an unmet need for the prevention or treatment of metastasis in early stage breast cancer patients. Bisphosphonates (BPs) are potent inhibitors of bone resorption and are extensively used for the prevention of osteoporosis and other skeletal disorders, as well as for the treatment of secondary bone cancer in BC patients. Furthermore, the direct anticancer activity of BPs has been established in primary tumor models. However, these studies were limited by the need for dosages far above the clinical range to overcome BPs' high affinity for bones and poor accumulation in the tumor itself, which leads to toxicity, including osteonecrosis of the jaw. To decrease BP dosage, increase bioavailability, and direct anticancer activity, we used the RALA (R-) peptide delivery system to form highly stable NPs with the nitrogen containing BP, risedronate (R-RIS). In vitro studies showed that, in comparison to RIS, R-RIS nanoparticles increased cytotoxicity and reduced metastatic features such as proliferation, migration, invasion, and adhesion of metastatic BC cells to bones. Furthermore, in an in vivo model, R-RIS had increased tumor accumulation while still maintaining similar bone accumulation to RIS alone. This increase in tumor accumulation corresponded with decreased tumor volume and lungs metastasis. R-RIS has great potential to be used in combination with standard of care chemotherapy for the treatment of primary BC and its metastasis while still having its bone resorption inhibiting properties.

Transglutaminase-catalyzed covalent anti-myostatin peptide depots

Nature realizes protein and peptide depots by catalyzing covalent bonds with the extracellular matrix (ECM) of tissues. We are translating this natural blueprint for the sustained delivery of a myostatin-inhibiting peptide (Anti-Myo), resulting in an enzyme depot established from injectable solutions. For that, we fused Anti-Myo to the D-domain of insulin-like growth factor I, a transglutaminase (TG) substrate. TG catalyzed the covalent binding of the D-domain to ECM proteins, such as laminin and fibronectin, on bioengineered ECM and in mice. ECM decorated with Anti-Myo suppressed myostatin activity and pathway activation and reduced the differentiation of preconditioned bone marrow-derived macrophages into osteoclasts in vitro.

Caveolin Regulates the Transport Mechanism of the Walnut-Derived Peptide EVSGPGYSPN to Penetrate the Blood-Brain Barrier.

Bioactive peptides, derived from short protein fragments, are recognized for their neuroprotective properties and potential therapeutic applications in treating central nervous system (CNS) diseases. However, a significant challenge for these peptides is their ability to penetrate the blood-brain barrier (BBB). EVSGPGYSPN (EV-10) peptide, a walnut-derived peptide, has demonstrated promising neuroprotective effects in vivo. This study aimed to investigate the transportability of EV-10 across the BBB, explore its capacity to penetrate this barrier, and elucidate the regulatory mechanisms underlying peptide-induced cellular internalization and transport pathways within the BBB. The results indicated that at a concentration of 100 μM and osmotic time of 4 h, the apparent permeability coefficient of EV-10 was Papp = 8.52166 ± 0.58 × 10-6 cm/s. The penetration efficiency of EV-10 was influenced by time, concentration, and temperature. Utilizing Western blot analysis, immunofluorescence, and flow cytometry, in conjunction with the caveolin (Cav)-specific inhibitor M-β-CD, we confirmed that EV-10 undergoes transcellular transport through a Cav-dependent endocytosis pathway. Notably, the tight junction proteins ZO-1, occludin, and claudin-5 were not disrupted by EV-10. Throughout its transport, EV-10 was localized within the mitochondria, Golgi apparatus, endoplasmic reticulum, lysosomes, endosomes, and cell membranes. Moreover, Cav-1 overexpression facilitated the release of EV-10 from lysosomes. Evidence of EV-10 accumulation was observed in mouse brains using brain slice scans. This study is the first to demonstrate that Cav-1 can facilitate the targeted delivery of walnut-derived peptide to the brain, laying a foundation for the development of functional foods aimed at CNS disease intervention.

Intravascular delivery of an MK2 inhibitory peptide to prevent restenosis after angioplasty

Peripheral artery disease is commonly treated with balloon angioplasty, a procedure involving minimally invasive, transluminal insertion of a catheter to the site of stenosis, where a balloon is inflated to open the blockage, restoring blood flow. However, peripheral angioplasty has a high rate of restenosis, limiting long-term patency. Therefore, angioplasty is sometimes paired with delivery of cytotoxic drugs like paclitaxel to reduce neointimal tissue formation. We pursue intravascular drug delivery strategies that target the underlying cause of restenosis - intimal hyperplasia resulting from stress-induced vascular smooth muscle cell switching from the healthy contractile into a pathological synthetic phenotype. We have established MAPKAP kinase 2 (MK2) as a driver of this phenotype switch and seek to establish convective and contact transfer (coated balloon) methods for MK2 inhibitory peptide delivery to sites of angioplasty. Using a flow loop bioreactor, we showed MK2 inhibition in ex vivo arteries suppresses smooth muscle cell phenotype switching while preserving vessel contractility. A rat carotid artery balloon injury model demonstrated inhibition of intimal hyperplasia following MK2i coated balloon treatment in vivo. These studies establish both convective and drug coated balloon strategies as promising approaches for intravascular delivery of MK2 inhibitory formulations to improve efficacy of balloon angioplasty.

Thermosensitive and mucoadhesive Xanthan gum-based hydrogel for local release of anti-Candida peptide

A thermoresponsive and mucoadhesive hydrogel has been developed for the local delivery of a novel anti-Candida peptide. This antimicrobial peptide was custom-designed and synthesized, utilizing a natural peptide identified in the hemolymph of the freshwater crayfish Procambarus clarkii as a molecular scaffold. The hydrogel, fabricated from a xanthan gum/poly-N-isopropylacrylamide graft copolymer, demonstrates temperature-dependent viscoelastic properties and a pseudoplastic behaviour, making it suitable for potential administration in various tissues. Moreover, the high stability of the hydrogel (about 9 % weight loss after 24 h of incubation) in physiological fluids as well as its mucoadhesive properties indicate that it could withstand in the application site long enough to perform its intended function. Furthermore, the good cytocompatibility of the hydrogel and the peptide's release profile (approximately 90 % release within the first 24 h), coupled with its efficacy in inhibiting fungal growth (logarithmic reduction of 1.08 compared to the control), validates the prospective application of the formulation in managing mucosal and superficial skin C. albicans infections. This not only addresses concerns related to drug resistance but also establishes the hydrogel as a versatile platform for advanced drug delivery systems aimed at circumventing systemic administration of antifungal drugs for the treatment of superficial skin and mucosal candidiasis.

Targeted degradation of undruggable proteins using a novel heterobifunctional proteomimetic platform.

47 Background: Of the ~20,000 proteins in the human genome, only a subset can be modulated via traditional pharmacological approaches. “Undruggable” targets currently include proteins with pivotal roles in cancer pathogenesis. Our work aims to introduce a new paradigm for engaging these clinically relevant proteins using a modular platform we term the HYbrid DegRAding Copolymers (HYDRACs). HYDRACs multiplex targeting warheads with degradation inducers and have long circulation times plus high cell uptake. Preliminary studies on MYC and RAS, targets of great pharmaceutical interest, highlight the potential of this platform to revolutionize drug design. Methods: HYDRACs containing MYC or RAS targeting peptides randomly copolymerized with degrons were synthesized using ROMP. Fluorescent and biotin-tags were incorporated for use in uptake and pull-down assays. Viability following treatment with MYC-HYDRACs was performed in multiple cell lines with both cellular (MYC-independent lines) and polymer composition controls. Target engagement was assessed via immunoprecipitation and circular dichroism with protein degradation monitored by WB. Unbiased whole proteome analysis and RNAseq were done to confirm observed effects were selectively on-target. In vivo efficacy and biodistribution was assessed in tumor-bearing mice. Results: HYDRACs show high levels of cell uptake and antiproliferative effects at sub-micromolar concentrations in a formulation- and target-dependent manner. Cells treated with MYC-HYDRACs displayed reduced MYC protein levels, which were rescued by proteasome or neddylation inhibition. Swapping out the degron for various validated E3 ligase recruiters maintained MYC degradation, highlighting the “plug-and-play” nature of this approach. On-target activity was confirmed via RNAseq. Biophysical analysis showed strong HYDRAC-protein interactions, orthogonally supported by the presence of target protein following pull-down of biotin-terminated HYDRACs. Target protein degradation only occurred following treatment with an intact HYDRAC compound. Mice bearing MYC-CaP tumors showed delayed tumor growth following treatment with MYC-HYDRACs, with compound accumulation in the tumor for up to 72 h following a single injection. Generalizability was demonstrated against RAS, with RAS-HYDRACs capable of degrading KRAS across multiple different alleles, acting as a pan-KRAS degrader. Conclusions: We present a novel platform technology that addresses the challenges inherent to peptide delivery approaches. HYDRACs have the potential to dramatically alter the drug discovery landscape, allowing for the development of cancer-relevant target modulators for which there are no current viable therapies. We envision the HYDRAC platform as a generalizable approach to designing degraders of proteins of interest, greatly expanding the therapeutic armamentarium for TPD.

Milk Exosome-Liposome Hybrid Vesicles with Self-Adapting Surface Properties Overcome the Sequential Absorption Barriers for Oral Delivery of Peptides.

Milk exosomes (mExos) have demonstrated significant promise as vehicles for the oral administration of protein and peptide drugs owing to their superior capacity to traverse epithelial barriers. Nevertheless, certain challenges persist due to their intrinsic characteristics, including suboptimal drug loading efficiency, inadequate mucus penetration capability, and susceptibility to membrane protein loss. Herein, a hybrid vesicle with self-adaptive surface properties (mExos@DSPE-Hyd-PMPC) was designed by fusing functionalized liposomes with natural mExos, aiming to overcome the limitations associated with mExos and unlock their full potential in oral peptide delivery. The surface property transformation of mExos@DSPE-Hyd-PMPC was achieved by introducing a pH-sensitive hydrazone bond between the highly hydrophilic zwitterionic polymer and the phospholipids, utilizing the pH microenvironment on the jejunum surface. In comparison to natural mExos, hybrid vesicles exhibited a 2.4-fold enhancement in the encapsulation efficiency of the semaglutide (SET). The hydrophilic and neutrally charged surfaces of mExos@DSPE-Hyd-PMPC in the jejunal lumen exhibited improved preservation of membrane proteins and efficient traversal of the mucus barrier. Upon reaching the surface of jejunal epithelial cells, the highly retained membrane proteins and positively charged surfaces of the hybrid vesicle efficiently overcame the apical barrier, the intracellular transport barrier, and the basolateral exocytosis barrier. The self-adaptive surface properties of the hybrid vesicle resulted in an oral bioavailability of 8.7% and notably enhanced the pharmacological therapeutic effects. This study successfully addresses some limitations of natural mExos and holds promise for overcoming the sequential absorption barriers associated with the oral delivery of peptides.

Cubosome lipid nanocarriers for delivery of ultra-short antimicrobial peptides

HypothesisAlthough antimicrobial peptides (AMPs) are a promising class of new antibiotics, their inherent susceptibility to degradation requires nanocarrier-mediated delivery. While cubosome nanocarriers have been extensively studied for delivery of AMPs, we do not currently understand why cubosome encapsulation improves antimicrobial efficacy for some compounds but not others. This study therefore aims to investigate the link between the mechanism of action and permeation efficiency of the peptides, their encapsulation efficacy, and the antimicrobial activity of these systems. ExperimentsEncapsulation and delivery of Indolicidin, and its ultra-short derivative, Priscilicidin, were investigated using SAXS, cryo-TEM and circular dichroism. Molecular dynamics simulations were used to understand the loading of these peptides within cubosomes. The antimicrobial efficacy was assessed against gram-negative (E. coli) and gram-positive (MRSA) bacteria. FindingsA high ionic strength solution was required to facilitate high loading of the cationic AMPs, with bilayer encapsulation driven by tryptophan and Fmoc moieties. Cubosome encapsulation did not improve the antimicrobial efficacy of the AMPs consistent with their high permeation, as explained by a recent ’diffusion to capture model’. This suggests that cubosome encapsulation may not be an effective strategy for all antimicrobial compounds, paving the way for improved selection of nanocarriers for AMPs, and other antimicrobial compounds.

Enhancing peptide and PMO delivery to mouse airway epithelia by chemical conjugation with the amphiphilic peptide S10

Delivery of antisense oligonucleotides (ASOs) to airway epithelial cells is arduous due to the physiological barriers that protect the lungs, and the endosomal entrapment phenomenon which prevents ASOs from reaching their intracellular targets. Various delivery strategies involving peptide-, lipid-, and polymer-based carriers are being investigated, yet the challenge remains. S10 is a peptide-based delivery agent that enables the intracellular delivery of biomolecules such as GFP, CRISPR-associated nuclease ribonucleoprotein (RNP), base editor RNP, and a fluorescent peptide into lung cells after intranasal or intratracheal administrations to mice, ferrets, and rhesus monkeys. Herein, we demonstrate that covalently attaching S10 to a fluorescently labeled peptide or a functional splice-switching phosphorodiamidate morpholino oligomer (PMO) improves their intracellular delivery to airway epithelia in mice after a single intranasal instillation. Data reveal a homogenous delivery from the trachea to the distal region of the lungs, specifically into the cells lining the airway. Quantitative measurements further highlight that conjugation via a disulfide bond through a PEG linker was the most beneficial strategy compared to direct conjugation (without the PEG linker) or conjugation via a permanent thiol-maleimide bond. We believe that S10-based conjugation provides a great strategy to achieve intracellular delivery of peptides and ASOs with therapeutic properties in lungs.

Oral delivery of therapeutic peptides by milk-derived extracellular vesicles

Oral delivery of therapeutic peptides by milk-derived extracellular vesicles