Synthesis and anti-pseudomonal activity of new ß-Ala modified analogues of the antimicrobial peptide anoplin.

Due to the rise of antibiotic-resistant bacteria around the world, AMPs (antimicrobial peptides), depending on non-specific membrane mechanism and low tendency to develop bacterial resistance, attract widespread attentions as novel antimicrobial alternatives for treating bacterial infections. In this study, a series of new ß-Ala modified-antimicrobial peptide analogues of anoplin were designed and synthesized, and their biological activities were described. Most of the new peptides showed perfect antimicrobial activities against two antibiotic-sensitive Pseudomonas aeruginosa strains and three clinical isolates of multidrug-resistant P. aeruginosa strains without significant hemolysis or cytotoxicity. More significantly, Ano-1ß and Ano-8ß (substituting positions 1 and 8 of anoplin with ß-Ala, respectively) exhibited the best antimicrobial potency. Additionally, the two new peptides were stable under physiological conditions and displayed preferable in vivo antimicrobial activity with less acute toxicity. Notably, Ano-1ß and Ano-8ß hardly generated resistance in contrast to conventional antibiotics rifampicin and gentamicin, and they exhibited better anti-biofilm activity and synergistic or additive effects in combination with conventional antibiotics. What's more, Ano-1ß and Ano-8ß had strong membrane disruption as evidenced by outer membrane permeabilization and cytoplasmic membrane depolarization assays. Confocal laser scanning microscopy and scanning electron microscopy further demonstrated that the two new peptides could destroy the bacterial membrane integrity. Collectively, the incorporation of ß-Ala was a reasonable approach for new antimicrobial peptides design, and the new peptides Ano-1ß and Ano-8ß might be promising antimicrobial candidates in combating the increasing antibiotic-resistant bacteria.

Study on the effects of different dimerization positions on biological activity of partial d-Amino acid substitution analogues of Anoplin.

Antimicrobial peptides have recently attracted much attention due to their broad-spectrum antimicrobial activity, rapid microbial effects, and minimal tendency toward resistance development. In this study, a series of new C-C terminals and C-N terminals dimer peptides were designed and synthesized by intermolecular dimerization of the partial d-amino acid substitution analogues of Anoplin, and the effects of different dimerization positions on biological activity were researched. The antimicrobial activity and stability of the new C-C terminals and C-N terminals dimer peptides were significantly improved compared with their parent peptide Anoplin. They displayed no obvious hemolytic activity and lower cytotoxicity, with a higher therapeutic index. Furthermore, the new dimer peptides not only enabled to rapidly disrupt bacterial membrane and damage its integrity which was different from conventional antibiotics but also penetrated bacterial membrane into binding to intracellular genomic DNA. More importantly, the new dimer peptides showed excellent antimicrobial activity against multidrug-resistant strains isolated from clinics in contrast to conventional antibiotics with low tendency to develop the bacterial resistance, besides they exhibited better anti-biofilm activity than antibiotic Rifampicin. Interestingly, the C-N terminals dimer peptides were superior to C-C terminals ones in antimicrobial and anti-biofilm activity, therapeutic index, outer membrane permeability, and DNA binding ability, whereas there were no noteworthy effects in different dimerization positions on stability. Thus, these data suggested that dimerization in different positions represented a potent strategy to develop novel antimicrobial agents for fighting against increasing bacterial resistance.

Amphiphilic Endomorphin-1 Derivative Functions as Self-assembling Nanomedicine for Effective Brain Delivery.

Endomorphin-1 (Tyr-Pro-Trp-Phe-NH2, EM-1), an endogenous µ-opioid receptor ligand with strong antinociceptive activity, is not in clinical use because of its limited metabolic stability and membrane permeability. In this study, we develop a short-peptide self-delivery system for brain targets with the capability to deliver EM-1 without vehicle. Two amphiphilic EM-1 derivatives, C18-SS-EM1 and C18-CONH-EM1, were synthesized by attaching a stearyl moiety to EM-1 via a disulfide and amide bond, respectively. The amphiphilicity of EM-1 derivatives enabled self-assembling into nanoparticles for brain delivery. The study assessed morphology, circular dichroism, and metabolic stability of the formulations, as well as their pharmacodynamics and in vivo distribution, directly monitored by near-IR fluorescence imaging in mouse brains. In aqueous solution, the C18-SS-EM1 derivative self-assembled into spherical nanostructures with a diameter of 10-20 nm. Near-IR fluorescence analysis visualized the accumulation of the peptides in the brain. Importantly, the analgesic effect of C18-SS-EM1 nanoparticles was significantly stronger as compared to that of unmodified EM-1 or C18-CONH-EM1 nanoparticles. An in vitro release study demonstrated that self-assembled C18-SS-EM1 nanoparticles possessed reduction-responsive behavior. In summary, self-assembling C18-SS-EM1 nanoparticles, which integrate the advantages of lipidization, nanoscale characteristics and, labile disulfide bonds, represent a promising strategy for brain delivery of short peptides.

Intramolecular cyclization of the antimicrobial peptide Polybia-MPI with triazole stapling: influence on stability and bioactivity.

Cationic antimicrobial peptides have attracted increasing attention as a novel class of antibiotics to treat infectious diseases caused by pathogenic bacteria. However, susceptibility to protease is a shortcoming in their development. Cyclization is one approach to increase the proteolytic resistance of peptides. Therefore, to improve the proteolytic resistance of Polybia-MPI, we have synthesized the MPI cyclic analogs C-MPI-1 (i-to-i+4) and C-MPI-2 (i-to-i+6) by copper(I)-catalyzed azide-alkyne cycloaddition. Compared with MPI, C-MPI-1 displayed sustained antimicrobial activity and had enhanced anti-trypsin resistance, while C-MPI-2 displayed no antimicrobial activity. The relationship between peptide structure and bioactivity was further investigated by probing the secondary structure of the peptides by circular dichroism. This showed that C-MPI-1 adopted an α-helical structure in aqueous solution and, interestingly, had increased α-helical conformation in 30 mM sodium dodecyl sulfate and 50% trifluoroethyl alcohol compared with MPI. C-MPI-2 that was not α-helical in structure, suggesting that the propensity for α-helix conformation may play an important role in cyclic peptide design. In addition, scanning electron microscopy, propidium iodide uptake, and membrane permeabilization assays indicated that MPI and the optimized analog C-MPI-1 had membrane-active action modes, indicating that the peptides would not be susceptible to conventional resistance mechanisms. Our study provides additional insight into the influence of intramolecular cyclization at various positions on peptide structure and biological activity. In conclusion, the design and synthesis of cyclic analogs via click chemistry offer a new strategy for the development of stable antimicrobial agents. Copyright © 2017 European Peptide Society and John Wiley & Sons, Ltd.

[Advances of tumor targeting peptides drug delivery system with pH-sensitive activities].

The pH-sensitive peptides drug delivery systems, which target to acidic extracellular environment of tumor tissue, have many advantages in drug delivery. They exhibit a high specificity to tumor and low cytotoxicity, which significantly increase the efficacy of traditional anti-cancer drugs. In recent years the systems have received a great attention. The pH-sensitive peptides drug delivery systems can be divided into five types according to the difference in pH-responsive mechanism,type of peptides and carrier materials. This paper summarizes the recent progresses in the field with a focus on the five types of pH-sensitive peptides in drug delivery systems. This may provide a guideline to design and application of tumor targeting drugs.

Simultaneous determination of oleanolic acid, ursolic acid, quercetin and apigenin in Swertia mussotii Franch by capillary zone electrophoresis with running buffer modifier.

The method of capillary zone electrophoresis (CZE) with direct UV detection was developed for the determination of oleanolic acid, ursolic acid, quercetin and apigenin. and then for the first time successfully applied to the analysis of four analytes in Swertia mussotii Franch and its preparations. Various factors affecting the CZE procedure were investigated and optimized, and the optimal conditions were: 50 × 10(-3) mol/L borate-phosphate buffer (pH 9.5) with 5.0 × 10(-3) mol/L ß-cyclodextrin, 15 kV separation voltage, 20 °C column temperature, 250 nm detection wavelength and 5 s electrokinetic injection time (voltage 20 psi). Under the conditions, oleanolic acid, ursolic acid, quercetin and apigenin could be determined within the test ranges with a good correlation coefficient (r(2) > 0.9991). The limits of detection for conditions, oleanolic acid, ursolic acid, quercetin and apigenin were 0.3415, 0.2003, 0.0062 and 0.2538 µg/mL, respectively, and the intra- and inter-day relative standard deviations were no more than 4.72%. This procedure provided a convenient, sensitive and accurate method for simultaneous determination of oleanolic acid, ursolic acid, quercetin and apigenin in S. mussotii Franch.

Simultaneous determination of phenylethanoid glycosides and aglycones by capillary zone electrophoresis with running buffer modifier.

Although the separation efficiency of capillary electrophoresis (CE) is much higher than that of other chromatographic methods, it is sometimes difficult to adequately separate the complex ingredients in biological samples. This article describes how one effective and simple way to develop the separation efficiency in CE is to add some modifiers to the running buffer. The suitable running buffer modifier ß-cyclodextrin (ß-CD) was explored to fast and completely separate four phenylethanoid glycosides and aglycones (homovanillyl alcohol, hydroxytyrosol, 3,4-dimethoxycinnamic acid, and caffeic acid) in Lamiophlomis rotata (Lr) and Cistanche by capillary zone electrophoresis with ultraviolet (UV) detection. It was found that when ß-CD was used as running buffer modifier, a baseline separation of the four analytes could be accomplished in less than 20 min and the detection limits were as low as 10(-3) mg L(-1). Other factors affecting the CE separation, such as working potential, pH value and ionic strength of running buffer, separation voltage, and sample injection time, were investigated extensively. Under the optimal conditions, a successful practical application on the determination of Lr and Cistanche samples confirmed the validity and practicability of this method.

Functional properties of a novel hybrid antimicrobial peptide NS: potent antitumor activity and efficient plasmid delivery.

Antimicrobial peptides have been widely recognized as potential candidates for treating tumor, especially for defending against multidrug-resistant cells. Previously, based on the structure of substance P, we have designed a novel class of hybrid antimicrobial peptide NS, which possesses potent antimicrobial activity against a broad spectrum of bacterial pathogens. In this study, we evaluated its cytotoxicity to tumor cells and studied the possible mechanism of action. We showed that NS could efficiently kill tumor cells by rapidly disrupting the tumor cell membrane and inhibiting the DNA synthesis. In addition, we also found that NS could efficiently deliver plasmids into cells and exhibit high transfection efficiency after the introduction of a stearyl moiety to its N-terminus, like many reported cell-penetrating peptides. Taken together, this study revealed the potential multiple functions of NS, providing fundamental support for further therapeutic application as potential antitumor agent.

Organoid as a promising tool for primary liver cancer research: a comprehensive review.

Primary liver cancer (PLC) is one of the most common malignant gastrointestinal tumors worldwide. Limited by the shortage of liver transplantation donors and the heterogeneity of tumors, patients with liver cancer lack effective treatment options, which leads to rapid progression and metastasis. Currently, preclinical models of PLC fall short of clinical reality and are limited in their response to disease progression and the effectiveness of drug therapy. Organoids are in vitro three-dimensional cultured preclinical models with a high degree of heterogeneity that preserve the histomorphological and genomic features of primary tumors. Liver cancer organoids have been widely used for drug screening, new target discovery, and precision medicine; thus representing a promising tool to study PLC. Here, we summarize the progress of research on liver cancer organoids and their potential application as disease models. This review provides a comprehensive introduction to this emerging technology and offers new ideas for researchers to explore in the field of precision medicine.

Structure modification of anoplin for fighting resistant bacteria.

The emergence of bacterial resistance has posed a significant challenge to clinical antimicrobial treatment, rendering commonly used antibiotics ineffective. The development of novel antimicrobial agents and strategies is imperative for the treatment of resistant bacterial infections. Antimicrobial peptides (AMPs) are considered a promising class of antimicrobial agents due to their low propensity for resistance and broad-spectrum activity. Anoplin is a small linear α-helical natural antimicrobial peptide that was isolated from the venom of the solitary wasp Anplius samariensis. It exhibits rich biological activity, particularly broad-spectrum antimicrobial activity and low hemolytic activity. Over the past three decades, more than 40 research publications on anoplin have been made available online. This review focuses on the advancements of anoplin in antimicrobial research, encompassing its sources, characterization, antimicrobial activity, influencing factors and structural modifications. The aim is to provide assistances for the development of new antimicrobial agents that can combat bacterial resistance.

Novel ß-Hairpin Antimicrobial Peptide Containing the ß-Turn Sequence of -NG- and the Tryptophan Zippers Facilitate Self-Assembly into Nanofibers, Exhibiting Excellent Antimicrobial Performance.

Antimicrobial peptides (AMPs) have emerged as promising agents to combat the antibiotic resistance crisis due to their rapid bactericidal activity and low propensity for drug resistance. However, AMPs face challenges in terms of balancing enhanced antimicrobial efficacy with increased toxicity during modification processes. In this study, de novo d-type ß-hairpin AMPs are designed. The conformational transformation of self-assembling peptide W-4 in the environment of the bacterial membrane and the erythrocyte membrane affected its antibacterial activity and hemolytic activity and finally showed a high antibacterial effect and low toxicity. Furthermore, W-4 displays remarkable stability, minimal occurrence of drug resistance, and synergistic effects when combined with antibiotics. The in vivo studies confirm its high safety and potent wound-healing properties at the sites infected by bacteria. This study substantiates that nanostructured AMPs possess enhanced biocompatibility. These advances reveal the superiority of self-assembled AMPs and contribute to the development of nanoantibacterial materials.

Arginine and tryptophan-rich dendritic antimicrobial peptides that disrupt membranes for bacterial infection in vivo.

The potent antibacterial activity and low resistance of antimicrobial peptides (AMPs) render them potential candidates for treating multidrug-resistant bacterial infections. Herein, a minimalist design strategy was proposed employing the "golden partner" combination of arginine (R) and tryptophan (W), along with a dendritic structure to design AMPs. By extension, the α/ε-amino group and the carboxyl group of lysine (K) were utilized to link R and W, forming dendritic peptide templates αRn(εRn)KWm-NH2 and αWn(εWn)KRm-NH2, respectively. The corresponding linear peptide templates R2nKWm-NH2 and W2nKRm-NH2 were used as controls. Their physicochemical properties, activity, toxicity, and stability were compared. Among these new peptides, the dendritic peptide R2(R2)KW4 was screened as a prospective candidate owing to its preferable antibacterial properties, biocompatibility, and stability. Additionally, R2(R2)KW4 not only effectively restrained the progression of antibiotic resistance, but also demonstrated synergistic utility when combined with conventional antibiotics due to its unique membrane-disruptive mechanism. Furthermore, R2(R2)KW4 possessed low toxicity (LD50 = 109.31 mg/kg) in vivo, while efficiently clearing E. coli in pulmonary-infected mice. In conclusion, R2(R2)KW4 has the potential to become an antimicrobial regent or adjuvant, and the minimalist design strategy of dendritic peptides provides innovative and encouraging thoughts in designing AMPs.

A novel antimicrobial peptide with broad-spectrum and exceptional stability derived from the natural peptide Brevicidine.

The global issue of antibiotic resistance is increasingly severe, highlighting the urgent necessity for the development of new antibiotics. Brevicidine, a natural cyclic lipopeptide, exhibits remarkable antimicrobial activity against Gram-negative bacteria. In this study, a comprehensive structure-activity relationship of Brevicidine was investigated through 20 newly synthesized cyclic lipopeptide analogs, resulting in the identification of an optimal linear analog 22. The sequence of analog 22 consisted of five d-amino acids and four non-natural amino acid 2,5-diaminovaleric acid (Orn) and conjugated with decanoic acid at N-terminal. Compared to Brevicidine, analog 22 was easier to synthesize, and exerted broad spectrum antimicrobial activity and excellent stability (t1/2 = 40.98 h). Additionally, analog 22 demonstrated a rapid bactericidal effect by permeating non-specifically through the bacterial membranes, thereby minimizing the likelihood of inducing resistance. Moreover, it exhibited remarkable efficacy in combating bacterial biofilms and reversing bacterial resistance to conventional antibiotics. Furthermore, it effectively suppressed the growth of bacteria in vital organs of mice infected with S. aureus ATCC 25923. In conclusion, analog 22 may represent a potential antimicrobial peptide for further optimization.

Rational design of a new short anticancer peptide with good potential for cancer treatment.

Anticancer peptides (ACPs) have regarded as a new generation of promising antitumor drugs due to the unique mode of action. The main challenge is to develop potential anticancer peptides with satisfied antitumor activity and low toxicity. Here, a series of new α-helical anticancer peptides were designed and synthesized based on the regular repeat motif KLLK. The optimal peptides 14E and 14Aad were successfully derived from the new short α-helical peptide KL-8. Our results demonstrated that 14E and 14Aad had good antitumor activity and low toxicity, exhibiting excellent selectivity index. This result highlighted that the desirable modification position and appropriate hydrophobic side-chain structure of acidic amino acids played critical roles in regulating the antitumor activity/toxicity of new peptides. Further studies indicated that they could induce tumor cell death via the multiple actions of efficient membrane disruption and intracellular mechanisms, displaying apparent superiority in combination with PTX. In addition, the new peptides 14E and 14Aad showed excellent antitumor efficacy in vivo and low toxicity in mice compared to KL-8 and PTX. Particularly, 14Aad with the longer side chain at the 14th site exhibited the best therapeutic performance. In conclusion, our work provided a new avenue to develop promising anticancer peptides with good selectivity for tumor therapy.

High Therapeutic Index α-Helical AMPs and Their Therapeutic Potential on Bacterial Lung and Skin Wound Infections.

Antimicrobial peptides (AMPs) possess strong antibacterial activity and low drug resistance, making them ideal candidates for bactericidal drugs for addressing the issue of traditional antibiotic resistance. In this study, a template (G(XXKK)nI, G = Gly; X = Leu, Ile, Phe, or Trp; n = 2, 3, or 4; K = Lys; I = Ile.) was employed for the devised of a variety of novel α-helical AMPs with a high therapeutic index. The AMP with the highest therapeutic index, WK2, was ultimately chosen following a thorough screening process. It demonstrates broad-spectrum and potent activity against both standard and multidrug-resistant bacteria, while also showing low hemolysis and rapid and efficient time-kill kinetics. Additionally, WK2 exhibits excellent efficacy in treating mouse models of Klebsiella pneumonia-induced lung infections and methicillin-resistant Staphylococcus aureus (MRSA)-induced skin wound infections while demonstrating good safety profiles in vivo. In conclusion, the template-based design methodology for novel AMPs with high therapeutic indices offers new insights into addressing antibiotic resistance problems. WK2 represents a promising antimicrobial agent.

Phosphorylation as an Effective Tool to Improve Stability and Reduce Toxicity of Antimicrobial Peptides.

Developing a straightforward and effective strategy to modify antimicrobial peptides (AMPs) is crucial in overcoming the challenges posed by their instability and toxicity. Phosphorylation can reduce toxicity and improve the stability of AMPs. Based on these, we designed a series of peptides and their corresponding phosphorylated forms. The results showed that all phosphorylated peptides displayed reduced toxicity and enhanced stability compared to their unphosphorylated counterparts. Among them, W3BipY8-P stood out as the most promising peptide, exhibiting similar antibacterial activity as its unphosphorylated analog W3BipY8 but with significantly reduced hemolytic activity (19-fold decrease), cytotoxicity (3.3-fold decrease), and an extended serum half-life 6.3 times longer than W3BipY8. W3BipY8-P exerted bactericidal effects by disrupting bacterial membranes. Notably, W3BipY8-P significantly prolonged the survival of bacteria-infected animals while its LD50 was 4.2 times higher than that of W3BipY8. These findings highlight phosphorylation as an effective strategy for improving the antimicrobial properties of AMPs.

Design of an acid-activated antimicrobial peptide for tumor therapy.

Antimicrobial peptides have received increasing attention as potential antitumor drugs due to their new mode of action. However, the systemic toxicity at high concentration always hampers their successful utilization for tumor therapy. Here, we designed a new type of acid-activated antimicrobial peptide AMitP by conjugating antimicrobial peptide MitP to its anionic binding partner MitPE via a disulfide linker. Compared with MitP, AMitP displayed significant antitumor activity at acidic pH and low cytotoxicity at normal pH. The results of MD simulations demonstrate that the changes of structure and membrane binding tendency of AMitP at different pH values played an important role in its pH-dependent antitumor activity. In addition, AMitP showed significant enzymatic stability compared with MitP, suggesting a potential for in vivo application. In short, our work opens a new avenue to develop antimicrobial peptides as potential antitumor drugs with high selectivity.

[Carboxyl nanodiamond as intracellular transporters of anticancer drug--podophyllotoxin].

The purpose of this study is to investigate the intracellular transporters effect and the cytotoxicity of carboxyl nanodiamond (CND) - podophyllotoxin (PPT). Nanodiamond (ND) was treated with mixed carboxylic acid and finally got 64 nm CND by centrifugation, and then it was reacted with PPT to form CND-PPT. UV spectrophotometry was used to calculate the content of PPT in CND-PPT, the particle size distribution and zeta potential were measured by Dynamic laser scattering instrument. CND, PPT, CND-PPT and CND + PPT (physical mixture of CND and PPT) were characterized by Fourier transform infrared spectroscopy, at the same time, thermal analysis and element analysis were used to estimate the content of the PPT in CND-PPT. The affect of CND, PPT, CND-PPT on HeLa cell was measured with MTT assay. The results showed that content of PPT combined with CND accounted for about 10%. MTT assay showed that CND has low cytotoxicity and CND-PPT can increase the water soluble of PPT. As a conclusion, CND as a hydrophilic pharmaceutical carrier combined with PPT is able to increase the water solubility of PPT, at low concentration, CND-PPT can enhance the antitumor activity in comparison with PPT, so CND can be used as a potential anticancer drug carrier.

Structure-Activity Relationship Study of Antimicrobial Peptide PE2 Delivered Novel Linear Derivatives with Potential of Eradicating Biofilms and Low Incidence of Drug Resistance.

The ongoing emergence of antibiotic-resistant pathogens had been dramatically stimulating and accelerating the need for new drugs. PE2 is a kind of cyclic lipopeptide with broad-spectrum antimicrobial activity. Herein, its structure-activity relationship was systematically investigated by employing 4 cyclic analogues and 23 linear analogues for the first time. The screened linear analogues 26 and 27 bearing different fatty acyls at N-termini and a Tyr residue at the 9th position had superior potency compared to the cyclic analogues and showed equivalent antimicrobial activity compared with PE2. Notably, 26 and 27 exhibited significant ability against multidrug-resistant bacteria, favorable resistance to protease, excellent performance against biofilm, low drug resistance, and high effectiveness against the mice pneumonia model. The antibacterial mechanisms of PE2 and linear derivatives 26 and 27 were also preliminarily explored in this study. As described above, 26 and 27 are promising antimicrobial candidates for the treatment of infections associated with drug-resistant bacteria.

One New Acid-Activated Hybrid Anticancer Peptide by Coupling with a Desirable pH-Sensitive Anionic Partner Peptide.

Anticancer peptides (ACPs) are promising antitumor resources, and developing acid-activated ACPs as more effective and selective antitumor drugs would represent new progress in cancer therapy. In this study, we designed a new class of acid-activated hybrid peptides LK-LE by altering the charge shielding position of the anionic binding partner LE based on the cationic ACP LK and investigated their pH response, cytotoxic activity, and serum stability, in hoping to achieve a desirable acid-activatable ACP. As expected, the obtained hybrid peptides could be activated and exhibit a remarkable antitumor activity by rapid membrane disruption at acidic pH, whereas its killing activity could be alleviated at normal pH, showing a significant pH response compared with LK. Importantly, this study found that the peptide LK-LE3 with the charge shielding in the N-terminal of LK displayed notably low cytotoxicity and more stability, demonstrating that the position of charge masking is extremely important for the improvement of peptide toxicity and stability. In short, our work opens a new avenue to design promising acid-activated ACPs as potential targeting agents for cancer treatment.