Degradation of Polo-like Kinase 1 by the Novel Poly-Arginine N-Degron Pathway PROTAC Regulates Tumor Growth in Nonsmall Cell Lung Cancer.

Polo-like kinase 1 (PLK1), which is crucial in cell cycle regulation, is considered a promising anticancer drug target. Herein, we present the N-degron pathway-based proteolysis targeting chimera (PROTAC) for PLK1 degradation, targeting the Polo-box domain (PBD). We identified DD-2 as the most potent PROTAC that selectively induces PLK1 degradation in cancer cells, including HeLa and nonsmall cell lung cancer (NSCLC), through the N-degron pathway. DD-2 exhibited significant in vitro anticancer effects, inducing G2/M arrest and apoptosis in HeLa and NSCLC cell lines. DD-2 showed significant tumor growth inhibition in a xenograft mouse model using HeLa and NSCLC cell lines, highlighting its potential in cancer treatment. Furthermore, the combination of DD-2 with tyrosine kinase inhibitor (TKI), osimertinib, effectively suppressed tumor growth in double-mutated H1975 cell lines, emphasizing DD-2's potential in combination cancer therapies. Collectively, this study demonstrates the potential of the N-degron pathway, especially using DD-2, for targeted cancer therapies.

Transcriptomic profiling and the first spatial expression analysis of candidate genes in the salivary gland of the East Asian medicinal leech, Hirudo nipponia.

Hirudo nipponia, a blood-sucking leech native to East Asia, possesses a rich repertoire of active ingredients in its saliva, showcasing significant medical potential due to its anticoagulant, anti-inflammatory, and antibacterial effects against human diseases. Despite previous studies on the transcriptomic and proteomic characteristics of leech saliva, which have identified medicinal compounds, our knowledge of tissue-specific transcriptomes and their spatial expression patterns remains incomplete. In this study, we conducted an extensive transcriptomic profiling of the salivary gland tissue in H. nipponia based on de novo assemblies of tissue-specific transcriptomes from the salivary gland, teeth, and general head region. Through gene ontology (GO) analysis and hierarchical clustering, we discovered a novel set of anti-coagulant factors-i.e., Hni-Antistasin, Hni-Ghilanten, Hni-Bdellin, Hni-Hirudin-as well as a previously unrecognized immune-related gene, Hni-GLIPR1 and uncharacterized salivary gland specific transcripts. By employing in situ hybridization, we provided the first visualization of gene expression sites within the salivary gland of H. nipponia. Our findings expand on our understanding of transcripts specifically expressed in the salivary gland of blood-sucking leeches, offering valuable resources for the exploration of previously unidentified substances with medicinal applications.

Multifunctional Properties of BMAP-18 and Its Aliphatic Analog against Drug-Resistant Bacteria.

BMAP-18, derived from the N-terminal region of bovine myeloid antimicrobial peptide BMAP-27, demonstrates potent antimicrobial activity without cytotoxicity. This study aimed to compare the antibacterial, antibiofilm, and anti-inflammatory properties of BMAP-18, rich in aromatic phenylalanine residues, with its aliphatic analog, BMAP-18-FL. Both aromatic BMAP-18 and aliphatic BMAP-18-FL exhibited equally potent antimicrobial activities against Gram-positive and Gram-negative bacteria, particularly methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant Pseudomonas aeruginosa (MDRPA). Mechanistic investigations employing SYTOX green uptake, DNA binding, and FACScan analysis revealed that both peptides acted by inducing membrane permeabilization and subsequent intracellular targeting. Moreover, both BMAP-18 and BMAP-18-FL effectively prevented biofilm formation and eradicated existing biofilms of MRSA and MDRPA. Notably, BMAP-18-FL displayed a superior anti-inflammatory activity compared to BMAP-18, significantly reducing the expression levels of pro-inflammatory cytokines in lipopolysaccharide-stimulated macrophages. This study emphasizes the similarities and differences in the antimicrobial, antibiofilm, and anti-inflammatory properties between aromatic BMAP-18 and aliphatic BMAP-18-FL, providing valuable insights for the development of multifunctional antimicrobial peptides against drug-resistant bacteria.

Fusarium solani infection disrupts metabolism during the germination of roselle (Hibiscus sabdariffa L.) seeds.

Fungal infections adversely influence the production and quality of seeds. Previously, Fusarium solani was reported as the causal agent of roselle (Hibiscus sabdariffa L.) seed rot. This study was designed to evaluate the effect of F. solani infection on the germination, biochemical composition, energy reserves, and antioxidant activity of roselle seeds because there is currently a lack of information on the relationship between seed metabolism and infection with F. solani. The results showed that roselle seeds infected with F. solani exhibited a ca. 55% reduction in overall germination. Additionally, the fungal infection decreased antioxidant activity, total phenolic content, protein, sugar (sucrose, fructose, and glucose), and some amino acid (glutamine, serine, and arginine) contents. In contrast, some metabolites were more abundant in infected seeds, including alanine (2.1-fold) and some fatty acids (palmitic acid and heptadecanoic acid by 1.1- and 1.4-fold, respectively). The infection-associated changes in fatty acid profile resulted in the ratio of unsaturated/saturated fatty acids being 2.1-fold higher in infected seeds. Therefore, our results reveal that F. solani infection remarkably altered the biochemical composition of roselle seeds, which may have contributed to the loss of germination and quality of roselle seeds.

Evaluation of deoxythymidine-based cationic amphiphiles as antimicrobial, antibiofilm, and anti-inflammatory agents.

OBJECTIVES: We recently designed a series of cationic deoxythymidine-based amphiphiles that mimic the cationic amphipathic structure of antimicrobial peptides (AMPs). Among these amphiphiles, ADG-2e and ADL-3e displayed the highest selectivity against bacterial cells. In this study, ADG-2e and ADL-3e were evaluated for their potential as novel classes of antimicrobial, antibiofilm, and anti-inflammatory agents. METHODS: Minimum inhibitory concentrations of ADG-2e and ADL-3e against bacteria were determined using the broth microdilution method. Proteolytic resistance against pepsin, trypsin, α-chymotrypsin, and proteinase K was determined by radial diffusion and HPLC analysis. Biofilm activity was investigated using the broth microdilution and confocal microscopy. The antimicrobial mechanism was investigated by membrane depolarization, cell membrane integrity analysis, scanning electron microscopy (SEM), genomic DNA influence and genomic DNA binding assay. Synergistic activity was evaluated using checkerboard method. Anti-inflammatory activity was investigated using ELISA and RT-PCR. RESULTS: ADG-2e and ADL-3e showed good resistance to physiological salts and human serum, and a low incidence of drug resistance. Moreover, they exhibit proteolytic resistance against pepsin, trypsin, α-chymotrypsin, and proteinase K. ADG-2e and ADL-3e were found to kill bacteria by an intracellular target mechanism and bacterial cell membrane-disrupting mechanism, respectively. Furthermore, ADG-2e and ADL-3e showed effective synergistic effects when combined with several conventional antibiotics against methicillin-resistant Staphylococcus aureus (MRSA) and multidrug-resistant Pseudomonas aeruginosa (MDRPA). Importantly, ADG-2e and ADL-3e not only suppressed MDRPA biofilm formation but also effectively eradicated mature MDRPA biofilms. Furthermore, ADG-2e and ADL-3e drastically decreased tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) gene expression and protein secretion in lipopolysaccharide (LPS)-stimulated macrophages, implying potent anti-inflammatory activity in LPS-induced inflammation. CONCLUSION: Our findings suggest that ADG-2e and ADL-3e could be further developed as novel antimicrobial, antibiofilm, and anti-inflammatory agents to combat bacterial infections.

Cell selectivity and antibiofilm and anti-inflammatory activities and antibacterial mechanism of symmetric-end antimicrobial peptide centered on D-Pro-Pro.

This study aimed to develop a new symmetric-end antimicrobial peptide (AMP) with cell selectivity, antibiofilm, and anti-inflammatory activities. Two symmetric-end AMPs, Lf6-pP and Lf6-GG, were designed based on the sequence RRWQWRzzRWQWRR, which contains two symmetric repeat sequences connected by a ß-turn-promoting sequence (zz) that can be a rigid turn by D-Pro-Pro (pP) or a flexible turn by Gly-Gly (GG). Both Lf6-pP and Lf6-GG exhibited potent antibacterial activity without causing hemolysis, but Lf6-pP exhibited better cell selectivity, likely due to the more significant impact of the rigid pP turn. Compared to Lf6-GG, Lf6-pP demonstrated approximately three times higher antimicrobial activity against drug-resistant bacteria, had a low incidence of drug resistance, and maintained its activity in the presence of physiological salts and human serum. Additionally, Lf6-pP was more effective than Lf6-GG in inhibiting biofilm formation and eradicating mature biofilms. The BODIPY-cadaverine assay indicated that the potent anti-inflammatory activity of Lf6-pP may be attributed to its direct interaction with LPS, resulting in decreased TNF-α and IL-6 levels in LPS-stimulated macrophages. Mechanistic studies, including membrane depolarization, outer/inner membrane permeation, and membrane integrity change, demonstrated that Lf6-pP exerts its antibacterial action through an intracellular-target mechanism. Overall, we propose that Lf6-pP has potential as a novel antibacterial, antibiofilm, and anti-inflammatory agent against drug-resistant bacterial infections.

Brain compartmentalization based on transcriptome analyses and its gene expression in Octopus minor.

Coleoid cephalopods have a high intelligence, complex structures, and large brain. The cephalopod brain is divided into supraesophageal mass, subesophageal mass and optic lobe. Although much is known about the structural organization and connections of various lobes of octopus brain, there are few studies on the brain of cephalopod at the molecular level. In this study, we demonstrated the structure of an adult Octopus minor brain by histomorphological analyses. Through visualization of neuronal and proliferation markers, we found that adult neurogenesis occurred in the vL and posterior svL. We also obtained specific 1015 genes by transcriptome of O. minor brain and selected OLFM3, NPY, GnRH, and GDF8 genes. The expression of genes in the central brain showed the possibility of using NPY and GDF8 as molecular marker of compartmentation in the central brain. This study will provide useful information for establishing a molecular atlas of cephalopod brain.

Discovery of structural and functional transition sites for membrane-penetrating activity of sheep myeloid antimicrobial peptide-18.

Cathelicidin antimicrobial peptides have an extended and/or unstructured conformation in aqueous solutions but fold into ordered conformations, such as the α-helical structure, when interacting with cellular membranes. These structural transitions can be directly correlated to their antimicrobial activity and its underlying mechanisms. SMAP-18, the N-terminal segment (residues 1-18) of sheep cathelicidin (SMAP-29), is known to kill microorganisms by translocating across membranes and interacting with their nucleic acids. The amino acid sequence of SMAP-18 contains three Gly residues (at positions 2, 7, and 13) that significantly affect the flexibility of its peptide structure. This study investigated the role of Gly residues in the structure, membrane interaction, membrane translocation, and antimicrobial mechanisms of SMAP-18. Five analogs were designed and synthesized through Gly → Ala substitution (i.e., G2A, G7A, G13A, G7,13A, and G2,7,13A); these substitutions altered the helical content of SMAP-18 peptides. We found that G7,13A and G2,7,13A changed their mode of action, with circular dichroism and nuclear magnetic resonance studies revealing that these analogs changed the structure of SMAP-18 from a random coil to an α-helical structure. The results of this experiment suggest that the Gly residues at positions 7 and 13 in SMAP-18 are the structural and functional determinants that control its three-dimensional structure, strain-specific activity, and antimicrobial mechanism of action. These results provide valuable information for the design of novel peptide-based antibiotics.

Cationic, amphipathic small molecules based on a triazine-piperazine-triazine scaffold as a new class of antimicrobial agents.

Poor proteolytic resistance, toxicity and salt/serum sensitivity of antimicrobial peptides (AMPs) limits their practical clinical application. Here, to overcome these drawbacks of AMPs and develop novel antimicrobial agents, a series of small molecules based on a triazine-piperazine-triazine scaffold that mimic the cationic amphipathic structure of AMPs were synthesized and evaluated their potential as a new class of antimicrobial agents. All designed compounds showed strong antimicrobial activity and negligible hemolytic activity. Particularly, five compounds (9, 11, 12, 15, and 16) presented excellent cell selectivity with proteolytic resistance, salt/serum stability and anti-inflammatory activity against lipopolysaccharide (LPS)-induced inflammation. These five compounds exhibited similar or 2-4 fold higher antimicrobial activity than melittin against six antibiotic-resistant bacteria tested. Similar to the intracellular-targeting AMP, buforin-2, these compounds displayed an intracellular mode of antimicrobial action. These compounds showed potent biofilm inhibitory and eradicating activities against multidrug-resistant Pseudomonas aeruginosa (MDRPA). Additionally, these compounds displayed synergistic or additive effects when combined with selected clinically used antibiotics. Furthermore, these compounds have been proven to inhibit pro-inflammatory cytokine release by directly binding to LPS and blocking the interaction between LPS and CD14/TLR4 receptor in LPS-stimulated RAW264.7 macrophage cells. Overall, our results demonstrate the potential of the designed compounds as a novel class of multifunctional antimicrobial agents to combat bacterial infection.

A novel hybrid peptide composed of LfcinB6 and KR-12-a4 with enhanced antimicrobial, anti-inflammatory and anti-biofilm activities.

Hybridizing two known antimicrobial peptides (AMPs) is a simple and effective strategy for designing antimicrobial agents with enhanced cell selectivity against bacterial cells. Here, we generated a hybrid peptide Lf-KR in which LfcinB6 and KR-12-a4 were linked with a Pro hinge to obtain a novel AMP with potent antimicrobial, anti-inflammatory, and anti-biofilm activities. Lf-KR exerted superior cell selectivity for bacterial cells over sheep red blood cells. Lf-KR showed broad-spectrum antimicrobial activities (MIC: 4-8 µM) against tested 12 bacterial strains and retained its antimicrobial activity in the presence of salts at physiological concentrations. Membrane depolarization and dye leakage assays showed that the enhanced antimicrobial activity of Lf-KR was due to increased permeabilization and depolarization of microbial membranes. Lf-KR significantly inhibited the expression and production of pro-inflammatory cytokines (nitric oxide and tumor necrosis factor-α) in LPS-stimulated mouse macrophage RAW264.7 cells. In addition, Lf-KR showed a powerful eradication effect on preformed multidrug-resistant Pseudomonas aeruginosa (MDRPA) biofilms. We confirmed using confocal laser scanning microscopy that a large portion of the preformed MDRPA biofilm structure was perturbed by the addition of Lf-KR. Collectively, our results suggest that Lf-KR can be an antimicrobial, anti-inflammatory, and anti-biofilm candidate as a pharmaceutical agent.

The Central PXXP Motif Is Crucial for PMAP-23 Translocation across the Lipid Bilayer.

PMAP-23, a cathelicidin-derived host defense peptide, does not cause severe membrane permeabilization, but exerts strong and broad-spectrum bactericidal activity. We have previously shown that it forms an amphipathic α-helical structure with a central hinge induced by the PXXP motif, which is implicated in the interaction of PMAP-23 with negatively charged bacterial membranes. Here, we studied the potential roles of the PXXP motif in PMAP-23 translocation across the lipid bilayer by replacing Pro residues with either α-helix former Ala (PMAP-PA) or α-helix breaker Gly (PMAP-PG). Although both PMAP-PA and PMAP-PG led to effective membrane depolarization and permeabilization, they showed less antimicrobial activity than wild-type PMAP-23. Interestingly, we observed that PMAP-23 crossed lipid bilayers much more efficiently than its Pro-substituted derivatives. The fact that the Gly-induced hinge was unable to replace the PXXP motif in PMAP-23 translocation suggests that the PXXP motif has unique structural properties other than the central hinge. Surface plasmon resonance sensorgrams showed that the running buffer almost entirely dissociated PMAP-23 from the membrane surface, while its Pro-substituted derivatives remained significantly bound to the membrane. In addition, kinetic analysis of the sensorgrams revealed that the central PXXP motif allows PMAP-23 to rapidly translocate at the interface between the hydrophilic and hydrophobic phases. Taken together, we propose that the structural and kinetic understanding of the PXXP motif in peptide translocation could greatly aid the development of novel antimicrobial peptides with intracellular targets by promoting peptide entry into bacterial cells.

Helicity Modulation Improves the Selectivity of Antimicrobial Peptoids.

The modulation of conformational flexibility in antimicrobial peptides (AMPs) has been investigated as a strategy to improve their efficacy against bacterial pathogens while reducing their toxicity. Here, we synthesized a library of helicity-modulated antimicrobial peptoids by the position-specific incorporation of helix-inducing monomers. The peptoids displayed minimal variations in hydrophobicity, which permitted the specific assessment of the effect of conformational differences on antimicrobial activity and selectivity. Among the moderately helical peptoids, the most dramatic increase in selectivity was observed in peptoid 17, providing more than a 20-fold increase compared to fully helical peptoid 1. Peptoid 17 had potent broad-spectrum antimicrobial activity that included clinically isolated multi-drug-resistant pathogens. Compared to pexiganan AMP, 17 showed superior metabolic stability, which could potentially reduce the dosage needed, alleviating toxicity. Dye-uptake assays and high-resolution imaging revealed that the antimicrobial activity of 17 was, as with many AMPs, mainly due to membrane disruption. However, the high selectivity of 17 reflected its unique conformational characteristics, with differential interactions between bacterial and erythrocyte membranes. Our results suggest a way to distinguish different membrane compositions solely by helicity modulation, thereby improving the selectivity toward bacterial cells with the maintenance of potent and broad-spectrum activity.

Antimicrobial and anti-inflammatory activities of short dodecapeptides derived from duck cathelicidin: Plausible mechanism of bactericidal action and endotoxin neutralization.

Antimicrobial peptides (AMPs) have gained increasing attention to combat antibiotic-resistant pathogens. dCATH (duck cathelicidin) is a 20-residue avian cathelicidin with potent bactericidal activity. However, its therapeutic application is limited due to high mammalian cell cytotoxicity. To develop therapeutically useful AMPs with enhanced antimicrobial and cell-selective property, we designed a series of 12-meric (dodeca) short amphiphilic peptides based on dCATH. Among these, Trp and Lys-rich dCATH 12-4 and dCATH 12-5 exhibited higher selectivity towards bacterial cells than erythrocytes and macrophages. Additionally, these AMPs significantly reduced NO and TNF-α secretion in LPS-stimulated macrophage cells, suggesting their anti-inflammatory properties. Various fluorophore-based studies and confocal microscopic observations demonstrated that dCATH 12-4 and dCATH 12-5 could penetrate the bacterial cell membrane and accumulate in the cytoplasm, without disrupting membrane integrity. Results from the microscopic examination and gel-retardation DNA binding assay suggested that both the designed AMPs could bind with bacterial DNA, subsequently leading to cell death via arrest of DNA synthesis. Fluorescence spectroscopy and flow cytometry analysis revealed that the designed AMPs induced strong binding to LPS oligomers which resulted in dissociation of LPS aggregates, thereby preventing LPS from binding to the carrier protein lipopolysaccharide-binding protein (LBP) or alternatively to CD14 receptors of macrophage cells. Additionally, both dCATH 12-4 and dCATH 12-5 demonstrated synergistic actions with various conventional antibiotics against antibiotic resistant pathogens, thus indicating their ability as promising adjuncts to combination therapy. In summary, these findings contribute to the design of short AMPs with bactericidal and immunomodulatory properties for combating bacterial infection and sepsis.

Mechanisms of antimicrobial and antiendotoxin activities of a triazine-based amphipathic polymer.

TZP4 is a triazine-based amphipathic polymer designed to mimic the amphipathic structure found in antimicrobial peptides. TZP4 showed potent antimicrobial activity comparable to melittin against antibiotic-resistant bacteria, such as methicillin-resistant Staphylococcus aureus and multidrug-resistant Pseudomonas aeruginosa. TZP4 showed high resistance to proteolytic degradation and low tendency to develop drug resistance. The results from membrane depolarization, SYTOX Green uptake, flow cytometry, and gel retardation revealed that the mechanism of antimicrobial action of TZP4 involved an intracellular target rather than the bacterial cell membrane. Furthermore, TZP4 suppressed the messenger RNA levels of inducible nitric oxide synthase and tumor necrosis factor-α (TNF-α) and inhibited the release of nitric oxide and TNF-α in lipopolysaccharide (LPS)-stimulated RAW264.7 cells. BODIPY-TR-cadaverine displacement and dissociation of fluorescein isothiocyanate (FITC)-labeled LPS assays revealed that TZP4 strongly bound to LPS and disaggregated the LPS oligomers. Flow cytometric analysis demonstrated that TZP4 inhibits the binding of FITC-conjugated LPS to RAW264.7 cells. These observations indicate that TZP4 may exert its antiendotoxic activity by directly binding with LPS and inhibiting the interaction between LPS and CD14+ cells. Collectively, TZP4 is a promising drug candidate for the treatment of endotoxic shock and sepsis caused by Gram-negative bacterial infections.

Proadrenomedullin N-terminal 20 peptide (PAMP) and its C-terminal 12-residue peptide, PAMP(9-20): Cell selectivity and antimicrobial mechanism.

Proadrenomedullin N-terminal 20 peptide (PAMP) is a regulatory peptide that is found in various cell types. It is involved in many biological activities and is rich in basic and hydrophobic amino acids, a common feature of antimicrobial peptides (AMPs). In this study, the cell selectivity and antimicrobial mechanism of PAMP and its C-terminal peptide, PAMP(9-20), were investigated. PAMP and PAMP(9-20) displayed potent antimicrobial activity (minimum inhibitory concentration: 4-32 µM) against standard bacterial strains, but showed no hemolytic activity even at the highest tested concentration of 256 µM. PAMP(9-20) showed 2- to 4-fold increase in antimicrobial activity against gram-negative bacteria compared to PAMP. Cytoplasmic membrane depolarization, leakage of calcein dye from membrane mimic liposomes, SYTOX Green uptake, membrane permeabilization, and flow cytometry studies indicated that the major target of PAMP and PAMP(9-20) is not the microbial cell membrane. Interestingly, laser-scanning confocal microscopy demonstrated that FITC-labeled PAMP and PAMP(9-20) enter the cytoplasm of Escherichia coli similar to buforin-2, and gel retardation assay indicated that PAMP and PAMP(9-20) effectively bind to bacterial DNA. These results suggest that the intracellular target mechanism is responsible for the antimicrobial action of PAMP and PAMP(9-20). Collectively, PAMP and PAMP(9-20) could be considered promising candidates for the development of new antimicrobial agents.

Synthesis of Fmoc-Triazine Amino Acids and Its Application in the Synthesis of Short Antibacterial Peptidomimetics.

To combat the escalating rise of antibacterial resistance, the development of antimicrobial peptides (AMPs) with a unique mode of action is considered an attractive strategy. However, proteolytic degradation of AMPs remains the greatest challenge in their transformation into therapeutics. Herein, we synthesized Fmoc-triazine amino acids that differ from each other by anchoring either cationic or hydrophobic residues. These unnatural amino acids were adopted for solid-phase peptide synthesis (SPPS) to synthesize a series of amphipathic antimicrobial peptidomimetics. From the antimicrobial screening, we found that the trimer, BJK-4 is the most potent short antimicrobial peptidomimetic without showing hemolytic activity and it displayed enhanced proteolytic stability. Moreover, the mechanism of action to kill bacteria was found to be an intracellular targeting.

Antibacterial AZT derivative regulates metastasis of breast cancer cells.

Antimicrobial peptides (AMP) with anticancer activity have drawn remarkable attention in modern treatments. However, long peptide length and protease instability are the most addressing factors, which hampers their further development as therapeutic agents. In view of this, herein, we designed and synthesized a series of AZT-based cationic small molecule incorporating a variety of hydrophobic groups and cationic charges, including amine and guanidine groups to mimic the amphipathic structure of AMPs. These compounds were evaluated for their antibacterial activity against Gram-positive and Gram-negative bacteria. Through an extensive structure activity relationship study (SAR), we identified ADG-2e as the most potent antibacterial agent, which exhibited remarkable potency against drug resistant bacterial strains such as MRSA and MDRPA. Further, ADG-2e was examined for their anti-metastatic ability by investigating the cancer cell migration and invasiveness through scratch wound-healing assay and transwell invasive assay, respectively. In addition, time-lapse cell tracking analysis also performed for analyzing the cell movement pattern. Treatment of ADG-2e against metastatic breast cancer cells (MDA-MB-231) suppressed tumor cell migration by multi-directional lamellipodium formation, indicating their anti-metastatic potential. Thus, our cationic AZT based small molecules may evolve as an appealing class of antibacterial agents with anti-metastasis potential.

Amphiphilic Triazine Polymer Derivatives as Antibacterial And Anti-atopic Agents in Mice Model.

Considering the emergence of bacterial resistance and low proteolytic stability of antimicrobial peptides (AMPs), herein we developed a series of ultra-short triazine based amphipathic polymers (TZP) that are connected with ethylene diamine linkers instead of protease sensitive amide bond. The most potent oligomers, TZP3 and TZP5 not only displayed potent antibacterial action on various drug-resistant pathogens but also exhibited a strong synergic antibacterial activity in combination with chloramphenicol against multidrug-resistant Pseudomonas aeruginosa (MDRPA). Since most of atopic dermatitis (AD) infections are caused by bacterial colonization, we evaluated the potency of TZP3 and TZP5 on AD in vitro and in vivo. In vitro AD analysis of these two polymers showed significant inhibition against the release of ß-hexosaminidase and tumor necrosis factor (TNF-α) from RBL-2H3 cells. In AD-like skin lesions in BALB/c mice model, these two polymers displayed significant potency in suppressing dermal and epidermal thickness, mast cell infiltration and pro-inflammatory cytokines expression. Moreover, these polymers exhibited remarkable efficacy over the allergies caused by the imbalance of Th1/Th2 by regulating total IgE and IgG2a. Finally, the impact of treatment effects of these polymers was examined through analyzing the weights and sizes of spleen and lymph node of AD-induced mice.

The design of a cell-selective fowlicidin-1-derived peptide with both antimicrobial and anti-inflammatory activities.

Fowlicidin-1 (Fowl-1), a cathelicidin expressed in chicken intestine, is known to have both antimicrobial and anti-inflammatory properties. However, its pharmaceutical development has been ultimately compromised by its high host cytotoxicity. In this study, a series of N- and C-terminal-truncated 19-meric Fowl-1 peptides were synthesized. Among these truncated peptides, Fowl-1 (8-26) exhibited broad-spectrum antimicrobial activity without human erythrocyte cytotoxicity while reducing anti-inflammatory activity. Further, Fowl-1 (8-26)-WRK was designed via Thr5→Trp, Ile7→Arg, and Asn11→Lys substitutions in Fowl-1 (8-26) to exhibit more amphipathicity. The results revealed that it exhibited both antimicrobial and anti-inflammatory properties. This study also demonstrated that the inhibitory activity of Fowl-1 (8-26)-WRK against LPS-induced inflammation was mainly due to the binding of LPS to the peptide. Interestingly, compared with human cathelicidin LL-37 and melittin, Fowl-1 (8-26)-WRK showed more potent activity against drug-resistant bacteria. It was also resistant to physiological salts and human serum and acted synergistically in combination with conventional antibiotics, such as chloramphenicol, ciprofloxacin, and oxacillin, suggesting that combined with conventional antibiotics, it is a promising adjuvant. Furthermore, membrane depolarization, SYTOX Green uptake, and flow cytometry revealed that it kills bacteria by damaging their membrane integrity. Therefore, this study suggests that Fowl-1 (8-26)-WRK has considerable potential for future development as an antimicrobial and anti-inflammatory agent for treating antibiotic-resistant infections.

Structural analysis and mode of action of BMAP-27, a cathelicidin-derived antimicrobial peptide.

BMAP-27, a member of cathelicidin family, plays an important role against microorganisms, including bacteria and fungi. BMAP-27 may exert antimicrobial effects through membrane integrity disruption, but the exact molecular mechanism remains unclear. To identify the structural features important for antimicrobial activity and propose a mechanism underlying antibacterial effects, we determined the nuclear magnetic resonance structure of BMAP-27 in a membrane-mimetic environment and investigated its interactions with lipid membranes. BMAP-27 exhibited a long N-terminal α-helix with faces patterned into aromatic and cationic regions, central kink, and short hydrophobic C-terminal helix. While the N-terminal 18-residue peptide (BMAP-18) exerted only antibacterial activity, BMAP-27 showed potent activity against bacteria and cancer cells. Both peptides inhibited bacterial growth, but BMAP-18 showed delayed bactericidal activity and BMAP-27 completely killed bacteria within 20 min. The differences in antimicrobial activities and microbicidal kinetics may be associated with membrane permeabilisation; BMAP-27 rapidly and largely disrupted membrane integrity, whereas BMAP-18 showed low membrane disruption activity. Thus, the N-terminal helix is sufficient to inhibit bacterial growth and the C-terminal helix is involved in membrane permeabilisation for rapid bactericidal and efficient anticancer activities. The structural and functional characterisation of BMAP-27 may encourage the development of novel antimicrobial/anticancer agents.