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PEGylated PAMAM dendrimers (PEG‐PAMAM) are well‐characterized biomaterials with still unexplored applications as carriers of drugs acting via membrane receptors, such as octreotide. This work confirmed the safety and negligible internalization capacity of fourth‐generation 50%‐PEG‐PAMAM in HEK‐293 cells, to then assessed their supramolecular binding to octreotide through tryptophan quenching experiments and Gaussian‐accelerated molecular dynamics (GaMD) simulations. Tryptophan quenching showed that PEG‐PAMAM binds octreotide with a Kbind of 6 × 106 M−1 and a complex stoichiometry of 1:1.4, unlike native PAMAM. GaMD simulations revealed that octreotide binds at the outer PEG shell of PEG‐PAMAM, potentially hindering the drug from proteolytic degradation and enabling its release at a membrane level. Viability experiments on HeLa, PC‐12, and HEK‐293 cells incubated with increasing concentrations of octreotide in free drug solutions and equimolar mixtures with PEG‐PAMAM confirmed that the PEGylated dendrimer acts as an efficient supramolecular carrier for octreotide and enhances the antiproliferative effects of the drug. Our findings highlight a novel facet for PEG‐PAMAM dendrimers as macromolecular vehicles for peptide or non‐peptide drugs acting via membrane receptor sites. [ABSTRACT FROM AUTHOR]

Purpose: Sodium N-[8-(2-hydroxybenzoyl)amino]caprylate (SNAC) is a well-known penetration enhancer widely used in commercial applications. This study aims to broaden its properties through a novel strategy of converting it into its phenolate salts. The objective is to investigate the synthesis of SNAC phenolate salts, specifically SNAC–choline (SNAC-CH), SNAC–sodium (SNAC-Na), and SNAC–phosphatidylcholine (SNAC-PC), and to explore their potential application in improving the oral absorption of semaglutide. Methods: The synthesis of SNAC phenolate salts was confirmed through 1H-NMR, FTIR, and an elemental analysis of C, H, N, and O. In vivo testing was conducted to assess the oral delivery of semaglutide using these synthesized SNAC phenolate salts. Pharmacokinetic (PK) values were measured to evaluate the impact on drug absorption. Results: The synthesis of SNAC phenolate salts (SNAC-CH, SNAC-Na, and SNAC-PC) was successfully achieved under appropriate conditions, and their structures were confirmed using analytical techniques such as IR, NMR, and CHN elemental analysis. The paradigm of their use was evaluated through an oral pharmacokinetic (PK) in vivo study using SNAC phenolate salts, which did not impair the original SNAC PK values. This suggests that this strategy holds promise as a potential new effective enhancer for oral absorption. Conclusions: The utilization of SNAC phenolate salts presents a novel and promising strategy for extending the verity of penetration enhancers' molecules and properties. Synthesizing phenolate salts represents a new chemical strategy that may open new avenues in molecular development. This approach holds future potential to enhance the oral delivery of peptide drugs like semaglutide without compromising therapeutic efficacy. Overall, it offers significant advancements in the field by providing a potential alternative to injectable peptides through oral delivery systems. [ABSTRACT FROM AUTHOR]

Motivation There has been a burgeoning interest in cyclic peptide therapeutics due to their various outstanding advantages and strong potential for drug formation. However, it is undoubtedly costly and inefficient to use traditional wet lab methods to clarify their biological activities. Using artificial intelligence instead is a more energy-efficient and faster approach. MuCoCP aims to build a complete pre-trained model for extracting potential features of cyclic peptides, which can be fine-tuned to accurately predict cyclic peptide bioactivity on various downstream tasks. To maximize its effectiveness, we use a novel data augmentation method based on a priori chemical knowledge and multiple unsupervised training objective functions to greatly improve the information-grabbing ability of the model. Results To assay the efficacy of the model, we conducted validation on the membrane-permeability of cyclic peptides which achieved an accuracy of 0.87 and R-squared of 0.503 on CycPeptMPDB using semi-supervised training and obtained an accuracy of 0.84 and R-squared of 0.384 using a model with frozen parameters on an external dataset. This result has achieved state-of-the-art, which substantiates the stability and generalization capability of MuCoCP. It means that MuCoCP can fully explore the high-dimensional information of cyclic peptides and make accurate predictions on downstream bioactivity tasks, which will serve as a guide for the future de novo design of cyclic peptide drugs and promote the development of cyclic peptide drugs. Availability and implementation All code used in our proposed method can be found at https://github.com/lennonyu11234/MuCoCP. [ABSTRACT FROM AUTHOR]

Currently, cancer is the second leading cause of death worldwide, and potential targeted drugs and molecular pathways for cancer development and progression have been a hot research topic worldwide. In recent years, the importance of the kinase superfamily in diseases has been well demonstrated by studies on various molecular mechanisms of kinases and the successful application of their inhibitors in diseases. Pseudokinases are members of the kinase superfamily, which have been increasingly documented to play a crucial role in cancers year after year. As a member of pseudokinases, tribbles homolog 3 (TRIB3) also exerts diverse functions in different cancers through different interacting proteins and molecular pathways, especially in tumor immunity, stemness, drug resistance, metabolism, and autophagy. In addition, peptide drugs targeting TRIB3 have high specificity in preclinical studies, which shows great promise for TRIB3 application in diseases including cancers. In this review, we dissect diverse functions played by TRIB3 in different cancers, describing the underlying mechanisms in detail. Notably, inhibitors and agonists currently available for TRIB3 are discussed, indicating the potential for TRIB3 as a therapeutic target. [ABSTRACT FROM AUTHOR]

Objective: In this work, in order to optimize the structure of histidine-containing antifungal peptides with a "linear" type of amphipathicity (LTA), we carried out the synthesis and comparative screening of the antifungal activity of a series of histidine-containing SAMPs (F2H7, F2H10, F2H13, F2H16, H10, H10F2) with a "linear" type of amphipathicity that we constructed de novo. We also aimed to study their hemolytic properties, to assess their therapeutic index, and to compare the obtained results with similar data for the antifungal histidine-rich 12-mer peptide P113 (AKRHHGYKRKFH), a drug candidate known as PAC-113, one of the shortest and most active analogues of the natural antifungal histidine-rich peptide Hst 5. Methods: Peptides were synthesized by a solid phase method using a Fmoc-based strategy. To determine the antifungal activity of the peptides, we used the opportunistic fungus strain Candida albicans EMTK 34. The number of microorganisms (titer) in the suspension was determined by optical density at a wavelength of 595 nm. To determine the antifungal activity of the peptides, cell cultures were co-incubated with the studied peptides in 96-well plates. The cells were incubated with the drug for 24 h at 37°C. To assess the antifungal activity of the studied peptides, their minimum inhibitory concentrations (MIC50) were determined, i.e. such peptide concentration at which the growth of microorganisms was suppressed by 50%. To assess the toxicity of the peptides, their hemolytic activity was determined, which was tested against fresh human erythrocytes using a photometric method. To assess the selectivity of the action of peptides, the values of the minimum hemolytic concentration (MHC) of the peptides were determined and the values of their therapeutic index (TI) were calculated as the ratio of the minimum hemolytic concentration (MHC) to the MIC50 value. Results and Discussion: It was shown that the antifungal activity of the studied SAMPLTA increases with an increase in the number of histidine residues in their composition, reaching the maximum value for the histidine-containing peptide F2H16 (MIC50 = 1.0 µM). It has been shown that as the chain length of peptides increases, their hemolytic toxicity also increases. The TI values for these peptides were 233, 247, 79, and 60, respectively. It has been shown that, against C. albicans, histidine-containing SAMP-LTA derivatives with phenylalanine residues at the N-terminus of the peptide (F2H10) are less effective compared to similar peptides (H10F2) containing phenylalanine residues at the C-terminus. Among the studied peptides, the most active was the H10 peptide (MIC50 = 0.7 µM), which does not contain phenylalanine residues, which in its antifungal activity is not only more effective than all other histidine-containing peptides, including the F2H16 peptide with 16 histidine residues, but also 4– 5 times more effective than the antifungal peptide P113 (MIC50 = 3.4 µM), a short active fragment of natural Hst 5, well known from the literature. Among all the studied peptides, H10 and P113 have minimal (almost zero) values of hemolytic activity. However, due to its higher antifungal activity, the selectivity of the H10 peptide (TI > 1400) exceeds the selectivity of the P113 peptide (TI > 340) by more than 4 times. Conclusions: Thus, peptide H10, due to its high antifungal activity, low hemolytic toxicity and, accordingly, high therapeutic significance, can be used as a promising antifungal peptide drug. [ABSTRACT FROM AUTHOR]

Proton-coupled oligopeptide transporters (POTs) are of great pharmaceutical interest owing to their promiscuous substrate binding site that has been linked to improved oral bioavail ability of several classes of drugs. Members of the POT family are conserved across all phylogenetic kingdoms and function by coupling peptide uptake to the proton electrochemical gradient. Cryo-EM structures and alphafold models have recently provided new insights into different conformational states of two mammalian POTs, SLC15A1, and SLC15A2. Nevertheless, these studies leave open important questions regarding the mechanism of proton and substrate coupling, while simultaneously providing a unique opportunity to investigate these processes using molecular dynamics (MD) simulations. Here, we employ extensive unbiased and enhanced-sampling MD to map out the full SLC15A2 conformational cycle and its thermodynamic driving forces. By computing conformational free energy landscapes in different protonation states and in the absence or presence of peptide substrate, we identify a likely sequence of intermediate protonation steps that drive inward-directed alternating access. These simulations identify key differences in the extracellular gate between mammalian and bacterial POTs, which we validate experimentally in cell-based transport assays. Our results from constant-PH MD and absolute binding free energy (ABFE) calculations also establish a mechanistic link between proton binding and peptide recognition, revealing key details underpining secondary active transport in POTs. This study provides a vital step forward in understanding proton-coupled peptide and drug transport in mammals and pave the way to integrate knowledge of solute carrier structural biology with enhanced drug design to target tissue and organ bioavailability. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is an age‐related neurodegenerative disease with amyloid‐β (Aβ) deposition as the main pathological feature. It's an important challenge to find new ways to clear Aβ from the brain. The soluble amyloid precursor protein α (sAPPα) is a neuroprotective protein and can attenuate neuronal damage, including toxic Aβ. However, the regulatory role of sAPPα in non‐neuronal cells, such as microglia, is less reported and controversial. Here, we showed that sAPPα promoted the phagocytosis and degradation of Aβ in both normal and damaged microglia. Moreover, the function of damaged microglia was improved by the sAPPα through normalizing mitochondrial function. Furthermore, the results of molecular docking simulation showed that sAPPα had a good affinity with Aβ. We preliminarily reveal that sAPPα is similar to antibodies and can participate in the regulation of microglia phagocytosis and degradation of Aβ after binding to Aβ. sAPPα is expected to be a mild and safe peptide drug or drug carrier for AD. [ABSTRACT FROM AUTHOR]

As the least abundant residue in proteins, tryptophan widely exists in peptide drugs and bioactive natural products and contributes to drug-target interactions in multiple ways. We report here a clickable tryptophan modification for late-stage diversification of native peptides, via catalyst-free C2-sulfenylation with 8-quinoline thiosulfonate reagents in trifluoroacetic acid (TFA). A wide range of groups including trifluoromethylthio (SCF3), difluoromethylthio (SCF2H), (ethoxycarbonyl)difluoromethylthio (SCF2CO2Et), alkylthio, and arylthio were readily incorporated. The rapid reaction kinetics of Trp modification and full tolerance with other 19 proteinogenic amino acids, as well as the super dissolving capability of TFA, render this method suitable for all kinds of Trp-containing peptides without limitations from sequences, hydrophobicity, and aggregation propensity. The late-stage modification of 15 therapeutic peptides (1.0 to 7.6 kilodaltons) and the improved bioactivity and serum stability of SCF3- and SCF2Hmodified melittin analogs illustrated the effectiveness of this method and its potential in pharmacokinetic property improvement. [ABSTRACT FROM AUTHOR]

Peptide drugs are becoming star drug agents with high efficiency and selectivity which open up new therapeutic avenues for various diseases. However, the sensitivity to hydrolase and the relatively short half-life have severely hindered their development. In this study, a new generation artificial intelligence-based system for accurate prediction of peptide half-life was proposed, which realized the half-life prediction of both natural and modified peptides and successfully bridged the evaluation possibility between two important species (human, mouse) and two organs (blood, intestine). To achieve this, enzymatic cleavage descriptors were integrated with traditional peptide descriptors to construct a better representation. Then, robust models with accurate performance were established by comparing traditional machine learning and transfer learning, systematically. Results indicated that enzymatic cleavage features could certainly enhance model performance. The deep learning model integrating transfer learning significantly improved predictive accuracy, achieving remarkable R 2 values: 0.84 for natural peptides and 0.90 for modified peptides in human blood, 0.984 for natural peptides and 0.93 for modified peptides in mouse blood, and 0.94 for modified peptides in mouse intestine on the test set, respectively. These models not only successfully composed the above-mentioned system but also improved by approximately 15% in terms of correlation compared to related works. This study is expected to provide powerful solutions for peptide half-life evaluation and boost peptide drug development. [ABSTRACT FROM AUTHOR]

Among all health-related issues, the rising concerns about drug resistance led to the look for alternative pharmaceutical drugs that are effective against both infectious and noninfectious diseases. Antimicrobial peptides (AMPs) are small molecular peptides that play a crucial role in the innate immunity of various organisms. They have amphiphilic structure and net positive charge, allowing them to interact with membranes and hydrophobic surfaces, showing strong broad-spectrum activity against different microorganisms, including bacteria, fungi, and viruses. They also exhibit other host-beneficial activities, including immunomodulation, anti-inflammatory, tissue regeneration, etc. AMPs exhibit antimicrobial activity through wide mechanisms of action, particularly by focusing on intracellular targets to inhibit the synthesis of nucleic acids and proteins. These wide ranges of mechanisms of action of AMPs have contributed to the slow development of resistance against microorganisms. The increasing pathogen resistance is a major global public health threat, and AMPs are constantly being explored and developed as another treatment for viral diseases such as HIV infection and (COVID-19). This review analyzes the potential of AMPs to combat antimicrobial resistance developed by several antimicrobial-resistant (AMR) microorganisms against existing antibiotics. This review focuses on the highlights of the sources, synthesis, mode, and mechanism of action, the evaluation of several benefits, and the outline of various hurdles. The review has also included the possible solution to the limitations associated with the clinical applications of AMPs, along with its future perspectives and development needed in drug discovery against AMR pathogens. [ABSTRACT FROM AUTHOR]

Covering: up to 2024 Ribosomally synthesised and post-translationally modified peptides (RiPPs) comprise a substantial group of peptide natural products exhibiting noteworthy bioactivities ranging from antiinfective to anticancer and analgesic effects. Furthermore, RiPP biosynthetic pathways represent promising production routes for complex peptide drugs, and the RiPP technology is well-suited for peptide engineering to produce derivatives with specific functions. Thus, RiPP natural products possess features that render them potentially ideal candidates for drug discovery and development. Nonetheless, only a small number of RiPP-derived compounds have successfully reached the market thus far. This review initially outlines the therapeutic opportunities that RiPP-based compounds can offer, whilst subsequently discussing the limitations that require resolution in order to fully exploit the potential of RiPPs towards the development of innovative drugs. [ABSTRACT FROM AUTHOR]

There is a lack of effective therapeutic drugs for pulmonary arterial hypertension. Previous studies have demonstrated the positive cardiovascular system protective effects of the new peptide ACTY116. However, its stability in ordinary aqueous solution injections is poor and its half‐life in the body is short, which has hindered the development of preparations. This study aimed to prepare in situ forming implants (ISFIs) of the peptide ACTY116 and investigate its impact on pulmonary arterial hypertension. We prepared ISFIs using NMP/TA as a solvent and PLGA as a polymer. These ISFIs exhibited low viscosity, low toxicity and sustained release properties. In a mouse model of pulmonary hypertension induced by SU5416/hypoxia, both ISFIs and ACTY116 peptides effectively reduced pulmonary hypertension, cardiac hypertrophy and pulmonary blood vessel wall thickness. In conclusion, this study highlights the potential of ACTY116 as a treatment for pulmonary arterial hypertension and suggests that incorporating it into an in‐situ gel implant could be a promising option. [ABSTRACT FROM AUTHOR]

This study aimed to evaluate Zn/Ag-electrode-printed patches for the transdermal delivery of small molecules through iontophoresis. The Zn/Ag-electrode-printed patches interact with biological liquid electrolytes and generate suitable microcurrents for the iontophoretic delivery of small molecules across the skin. In fluorescein permeation studies, Zn/Ag-electrode-printed patches increased the transdermal depth of fluorescein into the dermis, while the permeation of fluorescein was limited when Zn/C-electrode-printed patches were tested. Further permeation experiments were conducted with 3D skin models, which showed a similar trend to the above, indicating that Zn/Ag-electrode-printed patches had a higher penetration rate compared to the blank. Studies using acetyl hexapeptide-8 as a peptide drug model and sodium ascorbyl phosphate (SAP) as a hydrophilic derivative of ascorbic acid showed that the iontophoretic patch with Zn/Ag electrodes promoted more penetration of drugs than unprinted patches. The permeation of SAP exhibited a two-phase profile with a relatively rapid permeation followed by a sustained, slower permeation. The permeation of acetyl hexapeptide-8 was slower due to its higher molecular weight, but the iontophoretic patch increased the permeation up to 1.5 times more than the unprinted patch. The microcurrent generated by the patch drives the transport of small molecule components through the skin, for the controlled and efficient delivery of therapeutic agents. The flexible design, efficient microcurrent generation, and stable electrodes make the Zn/Ag-electrode-printed patch a promising tool for transdermal drug delivery. [ABSTRACT FROM AUTHOR]

Breast cancer persistently ranks as the second most prevalent cause of cancer fatalities among the female population worldwide, underscoring the imperative need for the invention of groundbreaking therapeutic strategies. In this context, our study delves into the potentialities offered by Anticancer Peptides (ACPs), an emergent class of therapeutics that function by selectively intercalating into and perturbing the membranes of neoplastic cells. Operating under the conjecture that their inherent characteristics considerably modulate the anticancer efficacy of ACPs, we conducted a statistical examination of over 60 ACP sequences and cytotoxicity profiles. We applied multiple regression models to pinpoint the attributes wielding the most predictive power. Our investigation revealed that the peptides' net charge and the isoelectric point emerged as the most consequential parameters. Our research endeavors to craft a predictive model as a preliminary screening instrument for pinpointing prospective peptides with high therapeutic potential for breast cancer treatment. This tool could potentially economize time and resources by circumventing the investigation of peptides that, due to their intrinsic properties, have a low likelihood of demonstrating efficacious results. Ultimately, our research aims to contribute to the progress in breast cancer research to improve outcomes for the millions of people impacted each year. [ABSTRACT FROM AUTHOR]

With the discovery of the therapeutic activity of peptides, they have emerged as a promising class of anti-cancer agents due to their specific targeting, low toxicity, and potential for high selectivity. In particular, as peptide-drug conjugates enter clinical, the coupling of targeted peptides with traditional chemotherapy drugs or cytotoxic agents will become a new direction in cancer treatment. To facilitate the drug development of cancer therapy peptides, we have constructed DCTPep, a novel, open, and comprehensive database for cancer therapy peptides. In addition to traditional anticancer peptides (ACPs), the peptide library also includes peptides related to cancer therapy. These data were collected manually from published research articles, patents, and other protein or peptide databases. Data on drug library include clinically investigated and/or approved peptide drugs related to cancer therapy, which mainly come from the portal websites of drug regulatory authorities and organisations in different countries and regions. DCTPep has a total of 6214 entries, we believe that DCTPep will contribute to the design and screening of future cancer therapy peptides. [ABSTRACT FROM AUTHOR]

Motivation Peptides are promising agents for the treatment of a variety of diseases due to their specificity and efficacy. However, the development of peptide-based drugs is often hindered by the potential toxicity of peptides, which poses a significant barrier to their clinical application. Traditional experimental methods for evaluating peptide toxicity are time-consuming and costly, making the development process inefficient. Therefore, there is an urgent need for computational tools specifically designed to predict peptide toxicity accurately and rapidly, facilitating the identification of safe peptide candidates for drug development. Results We provide here a novel computational approach, CAPTP, which leverages the power of convolutional and self-attention to enhance the prediction of peptide toxicity from amino acid sequences. CAPTP demonstrates outstanding performance, achieving a Matthews correlation coefficient of approximately 0.82 in both cross-validation settings and on independent test datasets. This performance surpasses that of existing state-of-the-art peptide toxicity predictors. Importantly, CAPTP maintains its robustness and generalizability even when dealing with data imbalances. Further analysis by CAPTP reveals that certain sequential patterns, particularly in the head and central regions of peptides, are crucial in determining their toxicity. This insight can significantly inform and guide the design of safer peptide drugs. Availability and implementation The source code for CAPTP is freely available at https://github.com/jiaoshihu/CAPTP. [ABSTRACT FROM AUTHOR]

In situ depot gel is a type of polymeric long-acting injectable (pLAI) drug delivery system; compared to microsphere technology, its preparation process is simpler and more conducive to industrialization. To ensure the chemical stability of peptide ACTY116, we avoided the use of harsh conditions such as high temperatures, high shear mixing, or homogenization; maintaining a water-free and oxygen-free environment was also critical to prevent hydrolysis and oxidation. Molecular dynamics (MDs) simulations were employed to assess the stability mechanism between ACTY116 and the pLAI system. The initial structure of ACTY116 with an alpha helix conformation was constructed using SYBYL-X, and the copolymer PLGA was generated by AMBER 16; results showed that PLGA-based in situ depot gel improved conformational stability of ACTY116 through hydrogen bonds formed between peptide ACTY116 and the components of the pLAI formulation, while PLGA (Poly(DL-lactide-co-glycolide)) also created steric hindrance and shielding effects to prevent conformational changes. As a result, the chemical and conformational stability and in vivo long-acting characteristics of ACTY116 ensure its enhanced efficacy. In summary, we successfully achieved our objective of developing a highly stable peptide-loaded long-acting injectable (LAI) in situ depot gel formulation that is stable for at least 3 months under harsh conditions (40 °C, above body temperature), elucidating the underlying stabilisation mechanism, and the high stability of the ACTY116 pLAI formulation creates favourable conditions for its in vivo pharmacological activity lasting for weeks or even months. [ABSTRACT FROM AUTHOR]

Peptides show high promise in the targeting and intracellular delivery of next-generation biotherapeutics. The main limitation is peptides' susceptibility to proteolysis in biological systems. Numerous strategies have been developed to overcome this challenge by chemically enhancing the resistance to proteolysis. In nature, amino acids, except glycine, are found in L- and D-enantiomers. The change from one form to the other will change the primary structure of polypeptides and proteins and may affect their function and biological activity. Given the inherent chiral nature of biological systems and their high enantiomeric selectivity, there is rising interest in manipulating the chirality of polypeptides to enhance their biomolecular interactions. In this review, we discuss the first examples of up-and-down homeostasis regulation by two enantiomeric drugs: immunostimulant Thymogen (L-Glu-L-Trp) and immunosuppressor Thymodepressin (D-Glu(D-Trp)). This study shows the perspective of exploring chirality to remove the chiral wall between L- and D-biomolecules. The selected clinical result will be discussed. [ABSTRACT FROM AUTHOR]

Ganirelix, a peptide‐based drug used to treat female infertility, has been in high market demand, which attracted generic formulation. A hitherto unknown impurity of ganirelix was observed in our formulation process, which reached ~0.3% in 6 months and led to a detailed investigation of its structure. In‐depth analysis of ESI‐MS/MS data of this impurity coupled with an artificial intelligence prediction tool led to a highly unusual putative structure, that is, N‐(2‐carboxyethyl)‐ganirelix (NCE‐GA), which was authenticated by chemical synthesis from ganirelix and NMR analysis and via corroborated HPLC and MS/MS data with the formulation‐derived impurity. [ABSTRACT FROM AUTHOR]

Pure shift NMR experiments greatly enhance spectral resolution by collapsing multiplet structures into singlets and, with water suppression, can be used for aqueous samples. Here, we combine ultra-clean pure-shift NMR (SAPPHIRE) with two different internally encoded water suppression schemes to achieve optimal performance for small molecule and macrocyclic peptide pharmaceuticals in water and acetonitrile–water mixtures. [ABSTRACT FROM AUTHOR]

Rationale: Disulfide bridges (DSB) play an important role in stabilizing threedimensional structures of biopharmaceuticals, single purified proteins, and various cyclic peptide drugs that contain disulfide in their structures. Incorrect cross-linking known as DSB scrambling results in misfolded structures that can be inactive, immunogenic, and susceptible to aggregation. Very few articles have been published on the experimental annotation of DSBs in proteins and cyclic peptide drugs. Accurate characterization of the disulfide bond is essential for understanding protein confirmation. Methods: Characterizing DSBs using mass spectrometry (MS) involves the chemical and enzymatic digestion of samples to obtain smaller peptide fragments, in both reduced and nonreduced forms. Subsequently, these samples are analyzed using MS to locate the DSB, either through interpretation or by employing various software tools. Results: The main challenge in DSB analysis methods using sample preparation is to obtain a sample solution in which nonnative DSBs are not formed due to high pH, temperature, and presence of free sulfhydryl groups. Formation of nonnative DSBs can lead to erroneous annotation of disulfide bond. Sample preparation techniques, fragmentation methods for DSB analysis, and contemporary approaches for DSB mapping using this fragmentation were discussed. Conclusions: This review presents the latest advancement in MS-based characterization; also a critical perspective is presented for further annotation of DSBs using MS, primarily for single purified proteins or peptides that are densely connected and rich in cysteine. Despite significant breakthroughs resulting from advancements in MS, the analysis of disulfide bonds is not straightforward; it necessitates expertise in sample preparation and interpretation. [ABSTRACT FROM AUTHOR]

Poor selectivity to tumor cells is a major drawback in the clinical application of the antitumor drug doxorubicin (DOX). Peptide–drug conjugates (PDCs) constructed by modifying antitumor drugs with peptide ligands that have high affinity to certain overexpressed receptors in tumor cells are increasingly assessed for their possibility of tumor-selective drug delivery. However, peptide ligands composed of natural L-configuration amino acids have the defects of easy enzymatic degradation and insufficient biological stability. In this study, two new PDCs (LT7-SS-DOX and DT7-SS-DOX) were designed and synthesized by conjugating a transferrin receptor (TfR) peptide ligand LT7 (HAIYPRH) and its retro-inverso analog DT7 (hrpyiah), respectively, with DOX via a disulfide bond linker. Both conjugates exhibited targeted antiproliferative effects on TfR overexpressed tumor cells and little toxicity to TfR low-expressed normal cells compared with free DOX. Moreover, the DT7-SS-DOX conjugate possessed higher serum stability, more sustained reduction-triggered drug release characteristics, and stronger in vitro antiproliferative activity as compared to LT7-SS-DOX. In conclusion, the coupling of antitumor drugs with the DT7 peptide ligand can be used as a promising strategy for the further development of stable and efficient PDCs with the potential to facilitate TfR-targeted drug delivery. [ABSTRACT FROM AUTHOR]

Teriparatide is an anabolic peptide drug indicated for the treatment of osteoporosis. Recombinant teriparatide was first approved in 2002 and has since been followed by patent-free alternatives under biosimilar or hybrid regulatory application. The aim of this study is to demonstrate the essential similarity between synthetic teriparatide BGW and the reference medicinal product (RMP), and thus to ensure the development of the first generic teriparatide drug. Hence, an extensive side-by-side comparative exercise, focusing on structural and biological activity, was performed using a wide range of state-of-the-art orthogonal methods. Nuclear magnetic resonance (NMR), ion mobility–mass spectrometry (IM–MS), UV, circular dichroism (CD) and Fourier transform infrared (FTIR) demonstrated the structural similarity between teriparatide BGW and the RMP. Comparative cell-based bioassays showed that the synthetic and recombinant peptides have identical behaviors. Teriparatide BGW, as a generic drug, provides an available treatment option for patients with osteoporosis and offers clinical benefits identical to those provided by the RMP. [ABSTRACT FROM AUTHOR]

This study was tasked with the design of mucoadhesive buccal films (MBFs) containing a peptide drug, leuprolide (LEU), or its diverse nanoparticles (NPs), for enhanced membrane permeability via self-assembled nanonization and deformable behavior. An LEU-oleic acid conjugate (LOC) and its self-assembled NPs (LON) were developed. Additionally, a deformable variant of LON (d-LON) was originally developed by incorporating l-α-phosphatidylcholine into LON as an edge activator. The physicochemical properties of LON and d-LON, encompassing particle size, zeta potential, and deformability index (DI), were evaluated. MBFs containing LEU, LOC, and NPs (LON, d-LON) were prepared using the solvent casting method by varying the ratio of Eudragit RLPO and hydroxypropyl methylcellulose, with propylene glycol used as a plasticizer. The optimization of MBF formulations was based on their physicochemical properties, including in vitro residence time, dissolution, and permeability. The dissolution results demonstrated that the conjugation of oleic acid to LEU exhibited a more sustained LEU release pattern by cleaving the ester bond of the conjugate, as compared to the native LEU, with reduced variability. Moreover, the LOC and its self-assembled NPs (LON, d-LON), equivalent to 1 mg LEU doses in MBF, exhibited an amorphous state and demonstrated better permeability through the nanonization process than LEU alone, regardless of membrane types. The incorporation of lauroyl-L-carnitine into the films as a permeation enhancer synergistically augmented drug permeability. Most importantly, the d-LON-loaded buccal films showed the highest permeability, due to the deformability of NPs. Overall, MBF-containing peptide NPs and permeation enhancers have the potential to replace parenteral LEU administration by improving LEU druggability and patient compliance. [ABSTRACT FROM AUTHOR]

Biodegradable polyesters are widely employed in the development of controlled release systems for peptide drugs. However, one of the challenges in developing a polyester-based delivery system for peptides is the acylation reaction between peptides and polymers. Peptide acylation is an important factor that affects formulation stability and can occur during storage, in vitro release, and after drug administration. This review focuses on the mechanisms and parameters that influence the rate of peptide acylation within polyesters. Furthermore, it discusses reported strategies to minimize the acylation reaction. [ABSTRACT FROM AUTHOR]

Ovarian cancer is the most deadly female gynaecological malignancy in developed countries and new treatments are urgently needed. The luteinising hormone releasing hormone (LHRH) peptide drug conjugate Zoptarelin doxorubicin is one such potential new drug modality that entered clinical trials for treating LHRH receptor‐positive gynaecological cancers. However, development stopped after disappointing Phase 3 results in 2017. We believe the lack of efficacy was due to linker instability and payload potency. In this work, we replaced its linker‐toxin with vedotin (MC‐VC‐PABC‐MMAE), yielding the novel peptide drug conjugate D‐Cys6‐LHRH vedotin. A GI50 and cell specificity comparison against cancerous and non‐cancerous ovarian cell lines showed significantly superior bioactivity and selectivity over Zoptarelin doxorubicin (GI50 4 vs. 453 nM) and other chemotherapeutic drugs used for treating ovarian cancers. Our results suggest D‐Cys6‐LHRH vedotin can potentially be used as a treatment for ovarian cancer. [ABSTRACT FROM AUTHOR]

The central dogma of modern biology underscores the pivotal roles proteins play in diverse biological processes, the study of which necessitates advanced methods to produce proteins with precision and versatility. Chemical protein synthesis, a powerful approach utilizing chemical reactions for the de novo construction of structurally accurate proteins, has emerged as a transformative tool for studying proteins and generating protein derivatives/mimics inaccessible by natural biological machinery, including post-translationally modified proteins, proteins comprised of unnatural amino acids, as well as mirror-image proteins. This review summarizes recent strides in synthetic method developments for chemical protein synthesis, including innovative techniques in solid-phase peptide synthesis, the challenges presented by difficult sequences in either synthesis or folding and the exploration of novel ligation reactions using both chemical and enzymatic methods. Furthermore, the review also delves into newly developed protocols for site-selective protein modifications and the generation of stapled or macrocyclized peptides/mini-proteins, highlighting the power of chemical methods to make structurally diverse proteins. Recent applications of synthetic proteins in investigating post-translational modifications (phosphorylation, lipidation, glycosylation, ubiquitination, etc.), mirror-image biological processes and drug development are further discussed. Together, these topics provide a comprehensive overview of the current landscape of chemical protein synthesis. [ABSTRACT FROM AUTHOR]

This article explores the potential of peptide drugs in cancer immunotherapy. Peptides offer advantages such as selectivity, infiltration into solid tumors, and easy synthesis, making them promising candidates for immune checkpoint inhibition. The article discusses the development of peptide inhibitors that target immune checkpoints like PD-1/PD-L1, TIGIT/PVR, and CD47/SIRPα. It also explores the use of dual-function peptides that can target multiple signaling pathways simultaneously. Additionally, the article mentions the conjugation of peptides with small molecule drugs to create peptide-drug conjugates and self-assembly nanoparticles for enhanced therapeutic effects. The article emphasizes the need for further research in oral peptide delivery and the combination of immune checkpoint blockade with peptide vaccines for a synergistic anti-tumor response. [Extracted from the article]

Mast cells (MCs) are primary effector cells involved in immediate allergic reactions. Mas-related G protein-coupled receptor-X2 (MrgX2), which is highly expressed on MCs, is involved in receptor-mediated drug-induced pseudo-anaphylaxis. Many small-molecule drugs and peptides activate MrgX2, resulting in MC activation and allergic reactions. Although small-molecule drugs can be identified using existing MrgX2 ligand-screening systems, there is still a lack of effective means to screen peptide ligands. In this study, to screen for peptide drugs, the MrgX2 high-affinity endogenous peptide ligand substance P (SP) was used as a recognition group to design a fluorescent peptide probe. Spectroscopic properties and fluorescence imaging of the probe were assessed. The probe was then used to screen for MrgX2 agonists among peptide antibiotics. In addition, the effects of peptide antibiotics on MrgX2 activation were investigated in vivo and in vitro. The environment-sensitive property of the probe was revealed by the dramatic increase in fluorescence intensity after binding to the hydrophobic ligand-binding domain of MrgX2. Based on these characteristics, it can be used for in situ selective visualization of MrgX2 in live cells. The probe was used to screen ten types of peptide antibiotics, and we found that caspofungin and bacitracin could compete with the probe and are hence potential ligands of MrgX2. Pharmacological experiments confirmed this hypothesis; caspofungin and bacitracin activated MCs via MrgX2 in vitro and induced local anaphylaxis in mice. Our research can be expected to provide new ideas for screening MrgX2 peptide ligands and reveal the mechanisms of adverse reactions caused by peptide drugs, thereby laying the foundation for improving their clinical safety. [Display omitted] • A fluorescent peptide probe for MrgX2 were designed and synthesized. • Fluorescent intensity of probe increased dramatically after binding to MrgX2. • The probe can be used for selective visualization of MrgX2 in living cells. • The probe was established to screen MrgX2 ligands in peptide antibiotics. • Caspofungin and bacitracin was found as MrgX2 agonists to activate MCs. [ABSTRACT FROM AUTHOR]

Therapeutic peptides are a major class of pharmaceutical drugs owing to their target‐binding specificity as well as their versatility in inhibiting aberrant protein‐protein interactions associated with human pathologies. Within the realm of amyloid diseases, the use of peptides and peptidomimetics tailor‐designed to overcome amyloidogenesis has been an active research endeavor since the late 90s. In more recent years, incorporating nanoparticles for enhancing the biocirculation and delivery of peptide drugs has emerged as a frontier in nanomedicine, and nanoparticles have further demonstrated a potency against amyloid aggregation and cellular inflammation to rival strategies employing small molecules, peptides, and antibodies. Despite these efforts, however, a fundamental understanding of the chemistry, characteristics and function of peptido‐nanocomposites is lacking, and a systematic analysis of such strategy for combating a range of amyloid pathogeneses is missing. Here we review the history, principles and evolving chemistry of constructing peptido‐nanocomposites from bottom up and discuss their future application against amyloid diseases that debilitate a significant portion of the global population. [ABSTRACT FROM AUTHOR]

We employed a structural bioinformatics approach to develop novel peptides with predicted affinity to the binding site for negative allosteric modulators (NAMs) of metabotropic glutamate receptor 5 (mGluR5). Primary screening in zebrafish (Danio rerio) revealed a stimulatory effect of two peptides, LCGM-10 and LCGM-15. Target validation studies using calcium ion flux imaging and a luciferase reporter assay confirmed mGluR5 as the target. LCGM-10 showed greater potency than LCGM-15; it was comparable to that of the mGluR5 NAM 2-methyl-6-(phenylethynyl) pyridine (MPEP). Rodent behavioral screening in the open field and elevated plus maze revealed increased locomotor activity in both tests after acute LCGM-10 treatment, supported by further analysis of home cage spontaneous locomotor activity (SLA). The stimulating effect of a single LCGM-10 administration on SLA was evident up to 60 min after administration and was not accompanied by hypokinetic rebound observed for caffeine. According to our results, LCGM-10 has therapeutic potential to treat hypoand dyskinesias of various etiologies. Further investigation of LCGM-10 effects in the delay discounting model of impulsive choice in rats revealed reduced trait impulsivity after single and chronic administrations, suggesting potential implication for attention deficit hyperactivity disorder, obsessive compulsive disorder, and addictions. [ABSTRACT FROM AUTHOR]

D-amino acid-containing peptides (DAACPs) occur in biological and artificial environments. Since the importance of DAACPs has been recognized, various mass spectrometry-based analytical approaches have been developed. However, the capability of higher-energy collisional dissociation (HCD) fragmentation to characterize DAACP sites has not been evaluated. In this study, we compared the normalized spectra intensity under different conditions of HCD and used liraglutide along with its DAACPs as examples. Our results indicated that the difference in the intensity of y ions between DAACPs and all-L liraglutide could not only distinguish them but also localize the sites of D-amino acids in the DAACPs. Our data demonstrate the potential of using HCD for the site characterization of DAACPs, which may have great impact in biological studies and peptide drug development. [ABSTRACT FROM AUTHOR]

This research theoretically modeled the binding interaction of the Sars-Cov-2 (Spike protein) utilizing molecular docking with some potential repurposed antiviral medications and two botanical products (Curcumin and Quercetin). Molecular docking between the drugs and the Sars-Cov-2 proteins reflecting the pure electrostatic forces and H-bond formation is complemented with the DFT results that shed light on the electronic nature of the interactions. Moreover, DFT computations provide invaluable information about the drug reactivity indices calculated from the energies of the frontier orbitals. The DFT results indicate intermolecular electron donor-acceptor interaction besides the H-bond formation. Most of the considered medication molecules act as electron-sink candidates except EIDD-2801 (molnupiravir), the electron donor. The theoretical results show the high possibility of blocking the human cellular entry against Sare-Cov-2 via strong interaction of the drugs with its peptide sequence (β6) measured by the high binding energy of docking or weakening its activity due to the electronic donor-acceptor interactions indicated by reactivity indices computed from the frontier molecular orbitals. [ABSTRACT FROM AUTHOR]

Although peptides notoriously have poor intrinsic pharmacokinetic properties, it is well-known that nanostructures with excellent pharmacokinetic properties can be designed. Noticing that peptide inhibitors are generally nonpolar, here, we consolidate the peptide inhibitor targeting intracellular protein–protein interactions (PPIs) as an integral part of biodegradable self-assembled depsipeptide nanostructures (SdPNs). Because the peptide inhibitor has the dual role of PPI inhibition and self-assembly in this design, problems associated with the poor pharmacokinetics of peptides and encapsulation/entrapment processes can be overcome. Optimized SdPNs displayed better tumor targeting and PPI inhibition properties than the comparable small molecule inhibitor in vivo. Kinetics of PPI inhibition for SdPNs were gradual and controllable in contrast to the rapid inhibition kinetics of the small molecule. Because SdPN is modular, any appropriate peptide inhibitor can be incorporated into the platform without concern for the poor pharmacokinetic properties of the peptide. [Display omitted] • Consolidation of peptide drugs with biodegradable peptide assemblies. • Self-assembled depsipeptide nanostructure (SdPN) targets protein–protein interaction. • Poor pharmacokinetics of peptides can be overcome with SdPNs. • Peptide drugs are released from SdPNs in a controlled manner. • Modular SdPNs enables incorporation of any peptide drugs targeting PPIs [ABSTRACT FROM AUTHOR]

Peptides are a particular molecule class with inherent attributes of some small-molecule drugs and macromolecular biologics, thereby inspiring continuous searches for peptides with therapeutic and/or agrochemical potentials. However, the success rate is decreasing, presumably because many interesting but less-abundant peptides are so scarce or labile that they are likely 'overlooked' during the characterization effort. Here, we present the biochemical characterization and druggability improvement of an unprecedented minor fungal RiPP (ribosomally synthesized and post-translationally modified peptide), named acalitide, by taking the relevant advantages of metabolomics approach and disulfide-bridged substructure which is more frequently imprinted in the marketed peptide drug molecules. Acalitide is biosynthetically unique in the macrotricyclization via two disulfide bridges and a protease (AcaB)-catalyzed lactamization of AcaA, an unprecedented precursor peptide. Such a biosynthetic logic was successfully re-edited for its sample supply renewal to facilitate the identification of the in vitro and in vivo antiparkinsonian efficacy of acalitide which was further confirmed safe and rendered brain-targetable by the liposome encapsulation strategy. Taken together, the work updates the mining strategy and biosynthetic complexity of RiPPs to unravel an antiparkinsonian drug candidate valuable for combating Parkinson's disease that is globally prevailing in an alarming manner. Acalitide is an anti-parkinsonian minor fungal RiPP with its macrotricyclic molecular framework sculpted by the unique biosynthetic pathway which is re-edited for its sample supply renewal. [Display omitted] [ABSTRACT FROM AUTHOR]

Cyclic peptides are fascinating molecules abundantly found in nature and exploited as molecular format for drug development as well as other applications, ranging from research tools to food additives. Advances in peptide technologies made over many years through improved methods for synthesis and drug development have resulted in a steady stream of new drugs, with an average of around one cyclic peptide drug approved per year. Powerful technologies for screening random peptide libraries, and de novo generating ligands, have enabled the development of cyclic peptide drugs independent of naturally derived molecules and now offer virtually unlimited development opportunities. In this review, we feature therapeutically relevant cyclic peptides derived from nature and discuss the unique properties of cyclic peptides, the enormous technological advances in peptide ligand development in recent years, and current challenges and opportunities for developing cyclic peptides that address unmet medical needs. [ABSTRACT FROM AUTHOR]

Drugs based on peptides and proteins (PPs) have been widely used in medicine, beginning with insulin therapy in patients with diabetes mellitus over a century ago. Although the oral route of drug administration is the preferred one by the vast majority of patients and improves compliance, medications of this kind due to their specific chemical structure are typically delivered parenterally, which ensures optimal bioavailability. In order to overcome issues connected with oral absorption of PPs such as their instability depending on digestive enzymes and pH changes in the gastrointestinal (GI) system on the one hand, but also their limited permeability across physiological barriers (mucus and epithelium) on the other hand, scientists have been strenuously searching for novel delivery methods enabling peptide and protein drugs (PPDs) to be administered enterally. These include utilization of different nanoparticles, transport channels, substances enhancing permeation, chemical modifications, hydrogels, microneedles, microemulsion, proteolytic enzyme inhibitors, and cell-penetrating peptides, all of which are extensively discussed in this review. Furthermore, this article highlights oral PP therapeutics both previously used in therapy and currently available on the medical market. [ABSTRACT FROM AUTHOR]

Peptides with a molecular mass between 2 and 10 kDa that are prohibited in elite sports usually require dedicated sample preparation and mass spectrometric detection that commonly cannot be combined with other (lower molecular mass) substances. In most instances, the physicochemical differences are too significant to allow for a generic analytical procedure. A simplification of established and comparably complex analytical approaches is therefore desirable and has been accomplished in the context of this study. With urine samples representing still the most frequently collected doping control specimens, efficient extraction of peptidic analytes from this matrix was a major goal of this method, as demonstrated for the included compounds such as insulins (human, lispro, aspart, glulisine, tresiba, glargine metabolite, bovine insulin, porcine insulin), growth hormone‐releasing hormones (sermorelin, CJC‐1295, tesamorelin) incl. their respective metabolites, insulin‐like‐growth factors (long‐R3‐IGF‐I, R3‐IGF‐I, des1–3‐IGF‐I), synacthen, gonadorelin and mechano growth factors (human MGF, MGF‐Goldspink). Sample preparation and detection are controlled by five internal standards, covering all five included peptide drug categories. Nearly all requirements of the recent technical documents from the World Anti‐Doping Agency (WADA) considering their minimum required performance levels (MRPL) are fulfilled, and the method was validated for its utilisation as initial testing procedure in doping controls. Finally, the approach was applied to authentic post‐administration study urine samples (for insulins and gonadorelin) in order to provide proof of principle. [ABSTRACT FROM AUTHOR]

Purpose: Proteins and peptides have secured a place as excellent therapeutic moieties on account of their high selectivity and efficacy. However due to oral absorption limitations, current formulations are mostly delivered parenterally. Oral delivery of peptides and proteins (PPs) can be considered the need of the hour due to the immense benefits of this route. This review aims to critically examine and summarize the innovations and mechanisms involved in oral delivery of peptide and protein drugs. Methods: Comprehensive literature search was undertaken, spanning the early development to the current state of the art, using online search tools (PubMed, Google Scholar, ScienceDirect and Scopus). Results: Research in oral delivery of proteins and peptides has a rich history and the development of biologics has encouraged additional research effort in recent decades. Enzyme hydrolysis and inadequate permeation into intestinal mucosa are the major causes that result in limited oral absorption of biologics. Pharmaceutical and technological strategies including use of absorption enhancers, enzyme inhibition, chemical modification (PEGylation, pro-drug approach, peptidomimetics, glycosylation), particulate delivery (polymeric nanoparticles, liposomes, micelles, microspheres), site-specific delivery in the gastrointestinal tract (GIT), membrane transporters, novel approaches (self-nanoemulsifying drug delivery systems, Eligen technology, Peptelligence, self-assembling bubble carrier approach, luminal unfolding microneedle injector, microneedles) and lymphatic targeting, are discussed. Limitations of these strategies and possible innovations for improving oral bioavailability of protein and peptide drugs are discussed. Conclusion: This review underlines the application of oral route for peptide and protein delivery, which can direct the formulation scientist for better exploitation of this route. [ABSTRACT FROM AUTHOR]

Peptide‐based drugs offer high specificity, potency, and selectivity. However, their inherent flexibility and differences in conformational preferences between their free and bound states create unique challenges that have hindered progress in effective drug discovery pipelines. The emergence of AlphaFold (AF) and Artificial Intelligence (AI) presents new opportunities for enhancing peptide‐based drug discovery. We explore recent advancements that facilitate a successful peptide drug discovery pipeline, considering peptides' attractive therapeutic properties and strategies to enhance their stability and bioavailability. AF enables efficient and accurate prediction of peptide‐protein structures, addressing a critical requirement in computational drug discovery pipelines. In the post‐AF era, we are witnessing rapid progress with the potential to revolutionize peptide‐based drug discovery such as the ability to rank peptide binders or classify them as binders/non‐binders and the ability to design novel peptide sequences. However, AI‐based methods are struggling due to the lack of well‐curated datasets, for example to accommodate modified amino acids or unconventional cyclization. Thus, physics‐based methods, such as docking or molecular dynamics simulations, continue to hold a complementary role in peptide drug discovery pipelines. Moreover, MD‐based tools offer valuable insights into binding mechanisms, as well as the thermodynamic and kinetic properties of complexes. As we navigate this evolving landscape, a synergistic integration of AI and physics‐based methods holds the promise of reshaping the landscape of peptide‐based drug discovery. This article is categorized under:Structure and Mechanism > Computational Biochemistry and BiophysicsMolecular and Statistical Mechanics > Molecular InteractionsSoftware > Molecular Modeling [ABSTRACT FROM AUTHOR]

Abstract Background Middle size peptides (MSPs) have emerged as a promising new pharmaceutical modality. We are seeking the best way to assess the non-clinical safety of MSPs. Consideration The requirements for assessing the genotoxicity of pharmaceuticals differ between small molecule drugs and biotherapeutics. Genotoxicity tests are necessary for small molecule drugs but not for biotherapeutics. MSPs, however, share similarities with both small molecule drugs and biotherapeutics. Here, we describe important points to consider in assessing the genotoxicity of MSP drugs. The current standard of genotoxicity assessment for small molecules may not be entirely appropriate for MSP drugs. MSP drugs need genotoxicity assessment mostly according to the current standard of small molecule drugs. Conclusion We propose a few modifications to the standard test battery of genotoxicity tests, specifically, the inclusion of an in vitro gene mutation test using mammalian cells, and exclusion of (Q)SAR assessment on MSP-related impurities.

Background: Middle size peptides (MSPs) have emerged as a promising new pharmaceutical modality. We are seeking the best way to assess the non-clinical safety of MSPs. Consideration: The requirements for assessing the genotoxicity of pharmaceuticals differ between small molecule drugs and biotherapeutics. Genotoxicity tests are necessary for small molecule drugs but not for biotherapeutics. MSPs, however, share similarities with both small molecule drugs and biotherapeutics. Here, we describe important points to consider in assessing the genotoxicity of MSP drugs. The current standard of genotoxicity assessment for small molecules may not be entirely appropriate for MSP drugs. MSP drugs need genotoxicity assessment mostly according to the current standard of small molecule drugs. Conclusion: We propose a few modifications to the standard test battery of genotoxicity tests, specifically, the inclusion of an in vitro gene mutation test using mammalian cells, and exclusion of (Q)SAR assessment on MSP-related impurities. [ABSTRACT FROM AUTHOR]

Herein, we designed alkylated lysine-dendron oxytocin amphiphiles (ALOAs) 1G-OTK and 2G-OTK, which were self-assembled into spherical nanoparticles and nanostrips, respectively, and showed superior stability compared to native oxytocin. We found similar trends in the functional activity of ALOAs and native OT for human oxytocin receptor. This work may inspire the development of peptide drugs for clinical applications. [ABSTRACT FROM AUTHOR]

Targeted therapy has attracted more and more attention in cancer treatment in recent years. However, due to the diversity of tumor types and the mutation of target sites on the tumor surface, some existing targets are no longer suitable for tumor therapy. In addition, the long‐term administration of a single targeted drug can also lead to drug resistance and attenuate drug potency, so it is important to develop new targets for tumor therapy. The expression of Type III transforming growth factor β receptor (TGFBR3) is upregulated in colon, breast, and prostate cancer cells, and plays an important role in the occurrence and development of these cancers, so TGFBR3 may be developed as a novel target for tumor therapy, but so far there is no report on this research. In this study, the structure of bone morphogenetic protein 4 (BMP4), one of the ligands of TGFBR3 was analyzed through the docking analysis with TGFBR3 and sequence charge characteristic analysis, and a functional tumor‐targeting penetrating peptide T3BP was identified. The results of fluorescent labeling experiments showed that T3BP could target and efficiently enter tumor cells with high expression of TGFBR3, especially A549 cells. When the expression of TGFBR3 on the surface of tumor cells (HeLa) was knocked down by RNA interference, the high delivery efficiency of T3BP was correspondingly reduced by 40%, indicating that the delivery was TGFBR3‐dependent. Trichosanthin (TCS, a plant‐derived ribosome inactivating protein) fused with T3BP can enhance the inhibitory activity of the fusion protein on A549 cells by more than 200 times that of TCS alone. These results indicated that T3BP, as a novel targeting peptide that can efficiently bind TGFBR3 and be used for targeted therapy of tumors with high expression of TGFBR3. This study enriches the supply of tumor‐targeting peptides and provides a new potential application option for the treatment of tumors with high expression of TGFBR3. [ABSTRACT FROM AUTHOR]

Proteins and peptides are rapidly developing pharmaceutical products and are expected to continue growing in the future. However, due to their nature, their delivery is often limited to injection, with drawbacks such as pain and needle waste. To overcome these limitations, microneedles technology is developed to deliver protein and peptide drugs through the skin. One type of microneedles, known as dissolving microneedles, has been extensively studied for delivering various proteins and peptides, including ovalbumin, insulin, bovine serum albumin, polymyxin B, vancomycin, and bevacizumab. This article discusses polymer materials used for fabricating dissolving microneedles, which are poly(vinylpyrrolidone), hyaluronic acid, poly(vinyl alcohol), carboxymethylcellulose, GantrezTM, as well as other biopolymers like pullulan and ulvan. The paper is focused solely on solvent casting micromoulding method for fabricating dissolving microneedles containing proteins and peptides, which will be divided into one-step and two-step casting micromoulding. Additionally, future considerations in the market plan for dissolving microneedles are discussed in this article. [ABSTRACT FROM AUTHOR]

Organoids are three‐dimensional cell aggregates with near‐physiologic cell behaviors and can undergo long‐term expansion in vitro. They are amenable to high‐throughput drug screening processes, which renders them a viable preclinical model for drug development. The procedure of organoid‐based high‐throughput screening has been extensively employed to discover small‐molecule drugs, encompassing the steps of generating organoids, examining efficient drugs in organoid cultures, and data assessment. Compared to small molecules, peptides are more straightforward to synthesize, can be modified chemically, and demonstrate high target specificity and low cytotoxicity. Therefore, they have emerged as promising carriers to deliver drugs to disease‐associated targets and could be efficient therapeutic drugs for various diseases. To date, organoids have been used to evaluate the efficacy of certain peptide agents; however, no organoid‐based high‐throughput screening of peptide drugs has been reported. Given the advantages of peptide drugs, there is an urgent need to establish organoid‐based peptide high‐throughput screening platforms. In this review, we discuss the typical approach of screening small‐molecular drugs with the use of organoid cultures, as well as provide an overview of the studies that have incorporated organoids in peptide research. Drawing on the knowledge from small molecular screens, we explore the difficulties and potential avenues for creating new platforms to identify peptide agents using organoid models. [ABSTRACT FROM AUTHOR]

The pharmacokinetics of peptide drugs are strongly affected by their aggregation properties and the morphology of the nanostructures they form in their native state as well as in their therapeutic formulation. In this contribution, we analyze the aggregation properties of a Liraglutide analogue (LG18), a leading drug against diabetes type 2. LG18 is a lipopeptide characterized by the functionalization of a lysine residue (K26) with an 18C lipid chain. To this end, spectroscopic experiments, dynamic light scattering measurements, and molecular dynamics simulations were carried out, following the evolution of the aggregation process from the small LG18 clusters formed at sub-micromolar concentrations to the mesoscopic aggregates formed by aged micromolar solutions. The critical aggregation concentration of LG18 in water (pH = 8) was found to amount to 4.3 μM, as assessed by the pyrene fluorescence assay. MD simulations showed that the LG18 nanostructures are formed by tetramer building blocks that, at longer times, self-assemble to form micrometric supramolecular architectures. [ABSTRACT FROM AUTHOR]

[Display omitted] Due to the lack of pharmacopeia guidelines for injectable microspheres based on poly (D, L-lactide-co-glycolide) (PLGA), an internal method validation is a critical prerequisite for quality assurance. One of the essential issues of developing peptide-based drugs loaded PLGA microspheres is the precise determination of the amount of peptide drug entrapped in the microspheres. The aim of this study is the development and optimization of a method for measuring the drug content loading of PLGA microspheres using exenatide as a model peptide drug. Exenatide-loaded PLGA microspheres were prepared by a double emulsion solvent evaporation method. The extraction method to determine exenatide content in microspheres was optimized using Design of Experiments (DoE) approach. After the initial screening of six factors, using Fractional Factorial design (FFD), four of them, including type of organic solvent, buffer/organic solvent ratio (v/v), shaking time and pH, exhibited significant effects on the response, namely the exenatide loading, and a Box-Behnken design (BBD) was subsequently applied to obtain its optimum level. The optimum level for organic solvent volume, buffer/organic solvent ratio, shaking time, and pH were 4 ml, 1, 5.6 hrs, and pH 6, respectively. The exenatide content in microspheres under these conditions was 6.4 ± 0.0 (%w/w), whereas a value of 6.1% was predicted by the derived equation. This excellent agreement between the actual and the predicted value demonstrates that the fitted model can thus be used to determine the exenatide content. [ABSTRACT FROM AUTHOR]