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Prior studies examined Acidocin 4356's antibacterial and antivirulence effects against Pseudomonas aeruginosa, including cell membrane penetration abilities. Building on prior research, an in-vitro co-culture of human cells was established to evaluate the selectivity of Acidocin (ACD) by concurrently cultivating human cells and bacterial pathogens. This study evaluated the antibacterial effectiveness of ACD against Acinetobacter baumannii and Pseudomonas aeruginosa. Laser scanning confocal microscopy (LSCM) and scanning electron microscopy (SEM) revealed significant biofilm dispersion at ACD concentrations as low as 1/2 MIC. The cytotoxicity of ACD was evaluated on two human cell lines, Calu-6 and THP-1, using the MTT assay. The IC50 values were 114 µg/mL and 24 µg/mL after a 12-hour treatment duration. In a co-culture model, the IC50 increased to 118 µg/mL, showing greater resilience of THP-1 cells under these settings, mimicking in-vivo conditions. Fluorescent microscopy and flow cytometry analysis confirmed the MTT results, showing ACD's potent antimicrobial effects and minimal toxicity to human cells, even after 12 h of treatment. Transmission electron microscopy (TEM) study revealed that normal Calu-6 cells included papillary outgrowths and microvilli, while infected cells displayed secretory vesicles, indicating an active response to P. aeruginosa infection. The present study thus serves as a critical step toward the development of an innovative therapeutic strategy targeting biofilm-associated infections. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) have attracted considerable attention as potential substitutes for traditional antibiotics. In our previous research, a novel antimicrobial peptide YS12 derived from the Bacillus velezensis strain showed broad-spectrum antimicrobial activity against Gram-positive and Gram-negative bacteria. In this study, the fractional inhibitory concentration index (FICI) indicated that combining YS12 with commercial antibiotics produced a synergistic effect. Following these findings, the combination of YS12 with an antibiotic resulted in a faster killing effect against bacterial strains compared to the treatment with the peptide YS12 or antibiotic alone. The peptide YS12 maintained its antimicrobial activity under different physiological salts (Na+, Mg2+, and Fe3+). Most importantly, YS12 exhibited no cytotoxicity towards Raw 264.7 cells and showed low hemolytic activity, whereas positive control melittin indicated extremely high toxicity. In terms of mode of action, we found that peptide YS12 was able to bind with LPS through electrostatic interaction. The results from fluorescent measurement revealed that peptide YS12 damaged the integrity of the bacterial membrane. Confocal laser microscopy further confirmed that the localization of peptide YS12 was almost in the cytoplasm of the cells. Peptide YS12 also exhibited anti-inflammatory activity by reducing the release of LPS-induced pro-inflammatory mediators such as TNF-α, IL-1β, and NO. Collectively, these properties strongly suggest that the antimicrobial peptide YS12 may be a promising candidate for treating microbial infections and inflammation. [ABSTRACT FROM AUTHOR]

Abstract BP100 is a short cationic antimicrobial peptide (AMP) designed using a combinatorial chemistry approach based on the cecropin A-melittin hybrid. It displays potent antimicrobial activity against gram-negative bacteria and low toxicity toward eukaryotic cells. To develop a short AMP with potent cell selectivity, antibiofilm and anti-inflammatory activities, we designed a newly BP100 analog, BP100-W, in which Leu-3 at the hydrophobic face of BP100 was replaced by Trp. BP100-W possessed better cell selectivity, with a 1.7-fold higher therapeutic index than BP100. BP100-W displayed more effective synergistic activity when combined with several antibiotics, such as chloramphenicol, ciprofloxacin and oxacillin, compared to BP-100. BP100-W also exhibited stronger antibiofilm activity than BP100 in inhibiting biofilm formation by multidrug-resistant Pseudomonas aeruginosa (MDRPA) and eradicating the preformed biofilms of MDRPA. Moreover, unlike BP100, BP100-W significantly suppressed the production and expression of lipopolysaccharide (LPS)-induced pro-inflammatory cytokines, such as the tumor necrosis factor-α and nitric oxide. Boron-dipyrromethene-TR-cadaverine displacement assay demonstrated that the inhibitory activity of BP100-W on LPS-induced inflammation in RAW 264.7 cells may be due to increased direct interaction with LPS. Our results suggest that BP100-W exhibits potential for future use as an antimicrobial, antibiofilm and anti-inflammatory agent.

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 β-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. [Display omitted] • Cell selectivity of Lf6-pP was better than that of Lf6-GG. • Lf6-pP shows potent antibiofilm and anti-inflammatory activities. • Lf6-pP exerts its antibacterial action by the intracellular target mechanism. • Lf6-pP is resistant to physiological salts and human serum. • Lf6-pP has a low incidence of drug resistance. [ABSTRACT FROM AUTHOR]

The active form of vitamin D3, 1α,25-dihydroxyvitamin D3 [1,25(OH)2D3], is a major regulator of calcium homeostasis through activation of the vitamin D receptor (VDR). We have previously synthesized vitamin D derivatives with large adamantane (AD) rings at position 24, 25, or 26 of the side chain to study VDR agonist and/or antagonist properties. One of them—ADTK1, with an AD ring and 23,24-triple bond—shows a high VDR affinity and cell-selective VDR activity. In this study, we synthesized novel vitamin D derivatives (ADKM1-6) with an alkyl group substituted at position 25 of ADTK1 to develop more cell-selective VDR ligands. ADKM2, ADKM4, and ADKM6 had VDR transcriptional activity comparable to 1,25(OH)2D3 and ADTK1, although their VDR affinities were weaker. Interestingly, ADKM2 has selective VDR activity in kidney- and skin-derived cells—a unique phenotype that differs from ADTK1. Furthermore, ADKM2, ADKM4, and ADKM6 induced osteoblast differentiation in human dedifferentiated fat cells more effectively than ADTK1. The development of vitamin D derivatives with bulky modifications such as AD at position 24, 25, or 26 of the side chain is useful for increased stability and tissue selectivity in VDR-targeting therapy. [ABSTRACT FROM AUTHOR]

I 4 K 2 Y*, co-localizing with tyrosinase abundantly expressed in melanoma cells, is catalyzed by tyrosinase to self-assemble into amyloid-like aggregates around the nucleus. Such aggregates prevent the exchange of substances between nucleus and cytoplasm, leading to cell apoptosis. [Display omitted] With the rising global incidence of melanoma, new anti-melanoma drugs with low-inducing drug resistance and high selectivity are in urgent need. Inspired by the physiological events in which fibrillar aggregates formed by amyloid proteins are toxic to normal tissues, we here rationally design a tyrosinase responsive peptide, I 4 K 2 Y* (Ac-IIIIKKDopa-NH 2). Such peptide self-assembled into long nanofibers outside the cells, while it was catalyzed into amyloid-like aggregates by tyrosinase which was rich in melanoma cells. The newly formed aggregates concentrated around the nucleus of melanoma cells, blocking the exchange of biomolecules between the nucleus and cytoplasm and finally leading to cell apoptosis via the S phase arrest in cell cycle distribution and dysfunction of mitochondria. Furthermore, I 4 K 2 Y* effectively inhibited B16 melanoma growth in a mouse model but with minimal side effects. We believe that the strategy of combining the usage of toxic amyloid-like aggregates and in-situ enzymatic reactions by specific enzymes in tumor cells will bring profound implications for designing new anti-tumor drugs with high selectivity. [ABSTRACT FROM AUTHOR]

BP100 is a short cationic antimicrobial peptide (AMP) designed using a combinatorial chemistry approach based on the cecropin A-melittin hybrid. It displays potent antimicrobial activity against gram-negative bacteria and low toxicity toward eukaryotic cells. To develop a short AMP with potent cell selectivity, antibiofilm and anti-inflammatory activities, we designed a newly BP100 analog, BP100-W, in which Leu-3 at the hydrophobic face of BP100 was replaced by Trp. BP100-W possessed better cell selectivity, with a 1.7-fold higher therapeutic index than BP100. BP100-W displayed more effective synergistic activity when combined with several antibiotics, such as chloramphenicol, ciprofloxacin and oxacillin, compared to BP-100. BP100-W also exhibited stronger antibiofilm activity than BP100 in inhibiting biofilm formation by multidrug-resistant Pseudomonas aeruginosa (MDRPA) and eradicating the preformed biofilms of MDRPA. Moreover, unlike BP100, BP100-W significantly suppressed the production and expression of lipopolysaccharide (LPS)-induced pro-inflammatory cytokines, such as the tumor necrosis factor-α and nitric oxide. Boron-dipyrromethene-TR-cadaverine displacement assay demonstrated that the inhibitory activity of BP100-W on LPS-induced inflammation in RAW 264.7 cells may be due to increased direct interaction with LPS. Our results suggest that BP100-W exhibits potential for future use as an antimicrobial, antibiofilm and anti-inflammatory agent. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are considered to be powerful weapons in the fight against traditional antibiotic resistance due to their unique membrane-disruptive mechanism. The combination of traditional and classical hydrophobic tryptophan (W) residues and hydrophilic charged arginine (R) residues is considered as the first choice for the minimalist design of AMPs due to its potent performance in antibacterial activity. However, some W- and R-rich AMPs that are not rationally designed and contain excessive repeats of W and R residues may cause severe cytotoxicity and hemolysis. To address this issue, we designed the (WRX) n (where X = hydrophilic uncharged amino residues; n = number of repeat units) series engineered peptides with high cell selectivity by introducing hydrophilic uncharged threonine (T), serine (S), glutamine (Q) or asparagine (N) residues into the minimalist design of W- and R-rich AMPs. The results showed that the introduction of these hydrophilic uncharged amino residues, especially T residues, significantly improved the cell selectivity of the W- and R-rich engineered peptides. Among (WRX) n series engineered peptides, T6 presents a mixture structure of β-turn and α-helix. It has broad spectrum and potent antibacterial activity (no activity against probiotics), good biocompatibility, high selectivity index, strong tolerance (physiological salts, serum acid, alkali, and heat conditions), rapid and efficient time-kill kinetics, and no tendency of resistance. Studies on antibacterial mechanism show that T6 exert antibacterial activity mainly by disrupting bacterial cell membrane and inducing the accumulation of reactive oxygen species in bacterial cells. Furthermore, T6 exhibited potent antibacterial and antiinflammatory capabilities in vivo in a mouse peritonitis-sepsis model infected with Escherichia coli. In conclusion, our study confirms an effective strategy for the minimalist design of highly cell selective W- and R-rich AMPs by introducing hydrophilic uncharged T residues, which may trigger widespread attention to hydrophilic uncharged amino acid residues, including T residues, and provide new insights into the design of peptide-based antibacterial biomaterials. We have introduced hydrophilic uncharged T, S, Q or N residues into the minimalist design of W- and R-rich engineered peptides and found that the introduction of these hydrophilic uncharged amino residues, especially the T residues, can significantly improve the cell selectivity of W- and R-rich engineered peptides. The target compound T6 showed potent antibacterial activity, high cell selectivity, strong tolerance, good in vivo efficacy and killed bacteria through multiple mechanisms mainly membrane-disruptive. These findings may spark widespread interest in hydrophilic uncharged amino acid residues, and provide new insights into the design of peptide-based antimicrobial biomaterials. [Display omitted] [ABSTRACT FROM AUTHOR]

Distinctively directing endothelial cells (ECs) and smooth muscle cells (SMCs), potentially by surface topography cue, is of central importance for enhancing bioefficacy of vascular implants. For the first time, surface gradients with a broad range of nano-micrometer roughness are developed on Mg, a promising next-generation biodegradable metal, to carry out a systematic study on the response of ECs and SMCs. Cell adhesion, spreading, and proliferation are quantified along gradients by high-throughput imaging, illustrating drastic divergence between ECs and SMCs, especially in highly rough regions. The profound role of surface topography overcoming the biochemical cue of released Mg2+ is unraveled at different roughness ranges for ECs and SMCs. Further insights into the underlying regulatory mechanism are gained at subcellular and gene levels. Our work enables high-efficient exploration of optimized surface morphology for modulating favored cell selectivity of promoting ECs and suppressing SMCs, providing a potential strategy to achieve rapid endothelialization for Mg.

The dynamic change of cell-surface glycans is involved in diverse biological and pathological events such as oncogenesis and metastasis. Despite tremendous efforts, it remains a great challenge to selectively distinguish and label glycans of different cancer cells or cancer subtypes. Inspired by biomimetic cell membrane-coating technology, herein, we construct pH-responsive azidosugar liposomes camouflaged with natural cancer-cell membrane for tumor cell-selective glycan engineering. With cancer cell-membrane camouflage, the biomimetic liposomes can prevent protein corona formation and evade phagocytosis of macrophages, facilitating metabolic glycans labeling in vivo. More importantly, due to multiple membrane receptors, the biomimetic liposomes have prominent cell selectivity to homotypic cancer cells, showing higher glycanlabeling efficacy than a single-ligand targeting strategy. Further in vitro and in vivo experiments indicate that cancer cell membrane-camouflaged azidosugar liposomes not only realize cell-selective glycan imaging of different cancer cells and triple-negative breast cancer subtypes but also do well in labeling metastatic tumors. Meanwhile, the strategy is also applicable to the use of tumor tissue-derived cell membranes, which shows the prospect for individual diagnosis and treatment. This work may pave a way for efficient cancer cell-selective engineering and visualization of glycans in vivo. [ABSTRACT FROM AUTHOR]

Metabolic glycan labeling (MGL) followed by bioorthogonal chemistry provides a powerful tool for tumor imaging and therapy. However, selectively metabolic labeling of cells or tissues of interest remains a challenge. Particularly, owing to tumor heterogeneity including tumor subtypes and interpatient heterogeneity, it is far more difficult to realize tumor‐cell‐selective metabolic labeling for precise diagnosis. Inspired by nature, we designed azidosugar‐functionalized metal–organic frameworks camouflaged with cancer cell membranes to accomplish cancer‐cell‐selective MGL in vivo. With abundant receptors, this biomimetic platform not only selectively targets homotypic cells but also realizes different breast cancer subtype‐selective MGL. Moreover, the endo/lysosomal‐escaped ZIF‐8 can make azidosugar escape from lysosomes and accelerate its metabolic incorporation. This strategy also takes advantage of cancer‐tissue‐derived cell membranes, which may have huge potential for personalized diagnosis and therapy. [ABSTRACT FROM AUTHOR]

The surfaces and interfaces of biomaterials interact with the biological systems in multi-scale levels, and thereby influence the biological functions and comprehensive performance in vitro and in vivo. In particular, a surface promoting the selective adhesion and directional migration of desired types of cells in complex environment is extremely important in the repair and regeneration of tissues such as peripheral nerve and blood vessel, and long-term application of intracorporal devices such as intravascular implants. Therefore, surface modification of biomaterials is a facile and effective method to achieve the desired cell-biomaterials interactions. In this short review, recent advances on the surface modification of biomaterials to regulate selective cell adhesion and migration are briefly summarized. In particular, the surface properties of biomaterials are manipulated via the convenient introduction of amino groups to the ester-based polymers, the formation of polyelectrolyte multilayers, and the fabrication of topology and gradient cues, etc., followed by the association of chemical and biological signals such as collagen, heparin, hyaluronic acid, peptides and cell growth factors. The selective adhesion and directional migration of various types of cells such as endothelial cells (ECs), smooth muscle cells (SMCs), hepatocytes and Schwann cells (SCs) are achieved over the competitive counterpart cells by the use of cell-resisting substances and cell-selective motifs on gradient substrates in most cases. Recent advances on cell behaviors in three-dimensional (3D) cell-extracellular matrix (ECM)-mimicking substrates are also reviewed. [ABSTRACT FROM AUTHOR]

The rise of antibiotic-resistant strains of bacterial pathogens has necessitated the development of new therapeutics. Antimicrobial peptides (AMPs) are a class of compounds with potentially attractive therapeutic properties, including the ability to target specific groups of bacteria. In nature, AMPs exhibit remarkable structural and functional diversity, which may be further enhanced through genetic engineering, high-throughput screening, and chemical modification strategies. In this review, we discuss the molecular mechanisms underlying AMP selectivity and highlight recent computational and experimental efforts to design selectively targeting AMPs. While there has been an extensive effort to find broadly active and highly potent AMPs, it remains challenging to design targeting peptides to discriminate between different bacteria on the basis of physicochemical properties. We also review approaches for measuring AMP activity, point out the challenges faced in assaying for selectivity, and discuss the potential for increasing AMP diversity through chemical modifications. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) have attracted more attention for their potential candidates for new antibiotic drugs. As a novel identified cathelicidin AMP from duck, dCATH owns broad-spectrum antimicrobial activities but with a noticeable toxicity. To explore dCATH-derived AMPs with reduced cell toxicity and improved cell selectivity, a series of truncated and tryptophan-replaced peptides of dCATH were designed. Two truncated peptides containing one of the two tryptophan (Trp) residues at the positions of 4 and 17 (W4 and W17) of dCATH, dCATH(1–16) and dCATH(5–20), showed strong antibacterial activity, but didn’t show obvious hemolytic activity and cytotoxicity. The derived peptides not containing Trp didn’t possess obvious antimicrobial activity, and their hemolytic and cytotoxic effect was also diminished. Also as evidence by Trp fluorescence experiment that existence of W4 and W17 was crucially important to the antimicrobial activity, hemolysis and cytotoxicity of dCATH, and one of the two Trp residues was competent and necessary to retain its antimicrobial activity. Antibacterial mechanism analysis showed that dCATH(1–16) and dCATH(5–20) killed bacterial cells by increasing permeability and causing a loss of membrane integrity. dCATH(1–16) and dCATH(5–20) possessed insignificant inhibitory activity against levels of IL-6, TNF-α, and NO in RAW 264.7 cells treated with LPS. In vivo, intraperitoneal administration of the two peptides significantly decreased mortality and provided protection against LPS-induced inflammation in mice challenged with lethal dose of LPS. The two peptides, dCATH(1–16) and dCATH(5–20), which possessed high antibacterial activity and cell selectivity, may herald development prospects as new antibacterial agents in the future.

Although Trp‐rich antimicrobial peptides (TRAMPs) such as tritrpticin and indolicidin exert potent antimicrobial activity, they have several drawbacks in therapeutic application, such as low cell selectivity and strong hemolytic activity. Here, we designed α‐helical TRAMPs with symmetric amino acid sequences based on pseudo‐symmetric tritrpticin and indolicidin. Compared with the parental pseudo‐symmetric TRAMPs, the symmetric α‐helical TRAMPs were highly effective against Gram‐positive bacteria, including clinically isolated methicillin‐resistant Staphylococcus aureus and vancomycin‐resistant Enterococcus faecium. The symmetric α‐helical TRAMPs exhibited negligible or very low toxicity against mammalian cells, including human erythrocytes. The symmetric peptides caused a rapid collapse of bacterial membrane potential and destabilized negatively charged lipid membranes, indicating that they act on the bacterial cytoplasmic membrane as a primary mode of action. Taken together, these findings imply that symmetric TRAMPs with an α‐helical structure offer significant potential for development as novel therapeutics to overcome the problem of antibiotic resistance in Gram‐positive pathogens. [ABSTRACT FROM AUTHOR]

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. Image 1 • PAMP and PAMP (9–20) acts as an antimicrobial peptide. • Compared with melittin, PAMP and PAMP(9–20) showed increased cell selectivity of 24.9- and 39.4-fold, respectively.. • PAMP(9–20) exhibits more potent antimicrobial activity against gram-negative bacteria compared to PAMP.. • Antimicrobial action of PAMP and PAMP(9–20) seem to exert via an intracellular target. • PAMP and PAMP(9–20) could be considered new promising candidates for antimicrobial drugs.. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) have attracted more attention for their potential candidates for new antibiotic drugs. As a novel identified cathelicidin AMP from duck, dCATH owns broad-spectrum antimicrobial activities but with a noticeable toxicity. To explore dCATH-derived AMPs with reduced cell toxicity and improved cell selectivity, a series of truncated and tryptophan-replaced peptides of dCATH were designed. Two truncated peptides containing one of the two tryptophan (Trp) residues at the positions of 4 and 17 (W4 and W17) of dCATH, dCATH(1–16) and dCATH(5–20), showed strong antibacterial activity, but didn't show obvious hemolytic activity and cytotoxicity. The derived peptides not containing Trp didn't possess obvious antimicrobial activity, and their hemolytic and cytotoxic effect was also diminished. Also as evidence by Trp fluorescence experiment that existence of W4 and W17 was crucially important to the antimicrobial activity, hemolysis and cytotoxicity of dCATH, and one of the two Trp residues was competent and necessary to retain its antimicrobial activity. Antibacterial mechanism analysis showed that dCATH(1–16) and dCATH(5–20) killed bacterial cells by increasing permeability and causing a loss of membrane integrity. dCATH(1–16) and dCATH(5–20) possessed insignificant inhibitory activity against levels of IL-6, TNF-α, and NO in RAW 264.7 cells treated with LPS. In vivo , intraperitoneal administration of the two peptides significantly decreased mortality and provided protection against LPS-induced inflammation in mice challenged with lethal dose of LPS. The two peptides, dCATH(1–16) and dCATH(5–20), which possessed high antibacterial activity and cell selectivity, may herald development prospects as new antibacterial agents in the future. [ABSTRACT FROM AUTHOR]

Selectively targeting the membrane‐perturbing potential of peptides towards a distinct cellular phenotype allows one to target distinct populations of cells. We report the de novo design of a new class of peptide whose ability to perturb cellular membranes is coupled to an enzyme‐mediated shift in the folding potential of the peptide into its bioactive conformation. Cells rich in negatively charged surface components that also highly express alkaline phosphatase, for example many cancers, are susceptible to the action of the peptide. The unfolded, inactive peptide is dephosphorylated, shifting its conformational bias towards cell‐surface‐induced folding to form a facially amphiphilic membrane‐active conformer. The fate of the peptide can be further tuned by peptide concentration to affect either lytic or cell‐penetrating properties, which are useful for selective drug delivery. This is a new design strategy to afford peptides that are selective in their membrane‐perturbing activity. [ABSTRACT FROM AUTHOR]

The cRGD-conjugated Aib-containing amphipathic helical peptide, MAP(Aib) derivative (PI), has been reported to be a useful carrier for siRNA delivery into cells. We have conducted a series of structure-activity relationship studies of the influence of the balance between hydrophobicity and basicity on the amphipathicity of PI , and synthesized peptides having a larger number of Lys residues than PI. Increasing the number of basic residues in the amphipathic helix suppressed the ability to deliver siRNA into cells. It was concluded that the balance between hydrophobicity and basicity in the PI helix was important for siRNA delivery into cells. Furthermore, the siRNA delivering ability of PI was specific to cancer cells, such as A549, U-87 MG, and WiDr cells, and was low in normal cells, namely, NIH3T3 cells. Next, we examined the potential of PI as a carrier for the delivery of microRNA-133b (miR-133b), which is known to be an anti-oncomiR. PI enhanced the delivery of miR-133b into WiDr cells, which resulted in the suppression of endogenous protein expression. [ABSTRACT FROM AUTHOR]

Fowlicidin‐1 (Fowl‐1) is a cathelicidin antimicrobial peptide from chicken that has potent antimicrobial activity and considerable cytotoxicity for human erythrocytes and mammalian cells. To improve the selectivity for bacterial cells, a Fowl‐1 analog ([Y7,P20]‐Fowl‐1) was designed by swapping the amino acids between Pro‐7 and Tyr‐20 in Fowl‐1. [Y7,P20]‐Fowl‐1 maintained relatively good antimicrobial activity (MIC 4 to 8 μM) but was much less toxic to human erythrocytes and RAW 264.7 murine macrophage cells, as compared to Fowl‐1. Tryptophan fluorescence quenching suggested that the bacterial selectivity of [Y7, P20]‐Fowl‐1 is associated with a preferential interaction with anionic phospholipids. The three‐dimensional structure of the peptides predicted by the ITASSER server and reverse phase‐high performance liquid chromatography (RP‐HPLC) retention time revealed that the significant reduction in cytotoxicity of [Y7,P20]‐Fowl‐1 was correlated with increased structural flexibility and decreased hydrophobicity. The data increase the understanding of the antimicrobial activity of Fowl‐1 and the factors that influence bacterial selectivity. [ABSTRACT FROM AUTHOR]

The estrogen receptor-positive (ER+) breast cancers constitute more than 50 % of breast cancers, seriously threatening the health of women. Unfortunately, the detection and targeted therapy of ER+ breast cancers remain a challenge. Here, a novel nucleic acid aptamer S1–4 was developed to specifically target ER+ breast cancer MCF-7 cells by using Cell-SELEX and nucleic acid truncation strategies. The affinity dissociation constant of the binding of aptamer S1–4 to MCF-7 cells was 97.6 ± 7.5 nM in vitro. Compared with HER2+ breast cells SK-BR-3 and triple-negative breast cancer cells MDA-MB-231, MCF-7 cells were selectively recognized and targeted by aptamer S1–4. Fluorescence tracing in vivo results also indicated that aptamer S1–4 selectively targeted the cell membrane of tumor tissues in MCF-7- but not in SK-BR3 or MDB-MA-231-bearing mice. This selectively developed novel aptamer probe S1–4 with high affinity could be used for the diagnosis and treatment of ER+ breast cancers. [ABSTRACT FROM AUTHOR]

Several antimicrobial peptides (AMPs) have been discovered, developed, and purified from natural sources and peptide engineering; however, the clinical applications of these AMPs are limited owing to their lack of abundance and side effects related to cytotoxicity, immunogenicity, and hemolytic activity. Accordingly, to improve cell selectivity for pseudin-2, an AMP from Pseudis paradoxa skin, in mammalian cells and pathogenic fungi, the sequence of pseudin-2 was modified by alanine or lysine at each position of two amino acids within the leucine-zipper motif. Alanine-substituted variants were highly selective toward fungi over HaCaT and erythrocytes and maintained their antifungal activities and mode of action (membranolysis). However, the antifungal activities of lysine-substituted peptides were reduced, and the compound could penetrate into fungal cells, followed by induction of mitochondrial reactive oxygen species and cell death. In vivo antifungal assays of analogous peptide showed excellent antifungal efficiency in a Candida tropicalis skin infection mouse model. Our results demonstrated the usefulness of selective amino acid substitution in the repeated sequence of the leucine-zipper motif for the design of AMPs with potent antimicrobial activities and low toxicity.

Antimicrobial peptides (AMPs) are excellent candidates to combat the increasing number of multi- or pan-resistant pathogens worldwide based on their mechanism of action, which is different from that of antibiotics. In this study, we designed short peptides by fusing an α-helix and β-turn sequence-motif in a symmetric-end template to promote the higher cell selectivity, antibacterial activity and salt-resistance of these structures. The results showed that the designed peptides PQ and PP tended to form an α-helical structure upon interacting with a membrane-mimicking environment. They displayed high cell selectivity toward bacterial cells over eukaryotic cells. Their activities were mostly maintained in the presence of different conditions (salts, serum, heat, and pH), which indicated their stability in vivo. Fluorescence spectroscopy and electron microscopy analyses indicated that PP and PQ killed bacterial cells through membrane pore formation, thereby damaging membrane integrity. This study revealed the potential application of these designed peptides as new candidate antimicrobial agents.

Abstract CXCL14 is a CXC chemokine family that exhibits antimicrobial activity and contains an amphipathic cationic α-helical region in the C-terminus, a characteristic structure of antimicrobial peptides (AMPs). In this study, we designed three analogs of CXCL14 59–75 (named CXCL14-C17) corresponding to the C-terminal α-helix of CXCL14, which displayed potential antimicrobial activity against a wide variety of gram-negative and gram-positive bacteria with minimum inhibitory concentrations of 4−16 μM without mammalian cell toxicity. Furthermore, two CXCL14-C17 analogs (CXCL14-C17-a1 and CXCL14-C17-a3) with improved cell selectivity were engineered by introducing Lys, Arg, or Trp in CXCL14-C17. Additionally, CXCL14-C17 analogs showed much greater synergistic effect (FICI: 0.3125–0.375) with chloramphenicol and ciprofloxacin against multidrug-resistant Pseudomonas aeruginosa (MDRPA) than LL-37 did (FICI: 0.75–1.125). CXCL14-C17 analogs were more active against antibiotic-resistant bacteria including methicillin-resistant Staphylococcus aureus (MRSA), MDRPA, and vancomycin-resistant Enterococcus faecium (VREF) than LL-37 and melittin. In particular, CXCL14-C17-a2 and CXCL14-C17-a3 completely inhibited the biofilm formation at sub-MIC and all of the peptides were able to eliminate pre-formed biofilm as well. Membrane depolarization, flow cytometry, sytox green uptake, ONPG hydrolysis and confocal microscopy revealed the possible target of the native peptide (CXCL14-C17) to likely be intracellular, and the amphipathic designed analogs targeted the bacterial membrane. CXCL14-C17 also showed DNA binding characteristic activity similar to buforin-2. Interestingly, CXCL14-C17-a2 and CXCL14-C17-a3 effectively inhibited the production and expression of nitric oxide (NO), tumor necrosis factor (TNF)-α, interleukin (IL)-6, and monocyte chemoattractant protein (MCP)-1 from lipopolysaccharide (LPS)-stimulated RAW264.7 cells, suggesting that these peptides could be promising anti-inflammatory and antimicrobial agents. Graphical abstract Unlabelled Image Highlights • CXCL14-C17 corresponding to the C-terminal α-helix of a CXC chemokine family CXCL14 and its analogs were designed and synthesized. • CXCL14-C17 displayed potent antimicrobial activity (MIC range: 4−16μM) without mammalian cell toxicity. • The main target of CXCL14-C17 and its analogs is intracellular and bacterial membrane, respectively. • CXCL14-C17 analogs were more active against antibiotic-resistant bacteria including MRSA, MDRPA and VREF than LL-37 and melittin. • CXCL14-C17-a2 and CXCL14-C17-a3 could be promising antimicrobial and anti-inflammatory agents. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are excellent candidates to combat the increasing number of multi- or pan-resistant pathogens worldwide based on their mechanism of action, which is different from that of antibiotics. In this study, we designed short peptides by fusing an α-helix and β-turn sequence-motif in a symmetric-end template to promote the higher cell selectivity, antibacterial activity and salt-resistance of these structures. The results showed that the designed peptides PQ and PP tended to form an α-helical structure upon interacting with a membrane-mimicking environment. They displayed high cell selectivity toward bacterial cells over eukaryotic cells. Their activities were mostly maintained in the presence of different conditions (salts, serum, heat, and pH), which indicated their stability in vivo. Fluorescence spectroscopy and electron microscopy analyses indicated that PP and PQ killed bacterial cells through membrane pore formation, thereby damaging membrane integrity. This study revealed the potential application of these designed peptides as new candidate antimicrobial agents. [ABSTRACT FROM AUTHOR]

High manufacturing costs and weak cell selectivity have limited the clinical application of naturally occurring peptides when faced with an outbreak of drug resistance. To overcome these limitations, a set of antimicrobial peptides was synthesized with the general sequence of (WL)n, where n = 1, 2, 3, and WL was truncated from the N‐terminus of Cecropin P1 without initial serine residues. The antimicrobial peptide WL3 exhibited stronger antimicrobial activity against both Gram‐negative and Gram‐positive microbes than the parental peptide CP‐1. WL3 showed no hemolysis even at the highest test concentrations compared to the parental peptide CP‐1. The condition sensitivity assays (salts, serum, and trypsin) demonstrated that WL3 had high stability in vitro. Fluorescence spectroscopy and electron microscopy indicated that WL3 killed microbes by means of penetrating the membrane and causing cell lysis. In a mouse model, WL3 was able to significantly reduce the bacteria load in major organs and cytokines (TNF‐α, IL‐6, and IL‐1β) levels in serum. In summary, these findings suggest that WL3, which was modified from a natural antimicrobial peptide, has enormous potential for application as a novel antibacterial agent. [ABSTRACT FROM AUTHOR]

Facing rising global antibiotics resistance, physical membrane-damaging antimicrobial peptides (AMPs) represent promising antimicrobial agents. Various strategies to design effective hybrid peptides offer many advantages in overcoming the adverse effects of natural AMPs. In this study, hybrid peptides from different species were investigated, and three hybrid antimicrobial peptides, LI, LN, and LC, were designed by combining the typical fragment of human cathelicidin-derived LL37 with either indolicidin, pig nematode cecropin P1 (CP-1) or rat neutrophil peptide-1 (NP-1). In an aqueous solution, all hybrid peptides had an unordered conformation. In simulated membrane conditions, the hybrid peptide LI displayed more β-turn and β-hairpin structures, whereas LN and LC folded into α-helix structures. The three interspecific hybrid peptides LI, LN, and LC exhibited different levels of antimicrobial activity against Gram-positive and Gram-negative bacteria. LI demonstrated the highest antimicrobial activity and cell selectivity. The results of the swimming motility indicated that LI repressed bacterial motility in a concentration-dependent method. Endotoxin binding assay demonstrated that hybrid peptide LI conserved the binding ability to LPS (polyanionic lipopolysaccharides) of its parental peptides. Fluorescence assays, flow cytometry, and SEM further revealed that hybrid peptide LI acted through different bacteriostatic mechanisms than LL37 and indolicidin and that LI killed bacterial cells via membrane damage. In summary, this study demonstrated that hybrid peptide LI produced by interspecific hybrid synthesis possessed strong cell selectivity and is a promising therapeutic candidate for drug-resistant bacteria infection. [ABSTRACT FROM AUTHOR]

Fowlicidin‐3 (Fow‐3), a chicken‐derived cathelicidin, is an antimicrobial peptide with considerable antimicrobial activity and mammalian cell toxicity. To reduce toxicity toward mammalian cells and improve selectivity for bacteria, a Fow‐3 analog was synthesized by shifting its PXXP motif from the N‐terminus to a central position. This analog was more selective against bacteria and was less toxic to mammalian cells, including human erythrocytes, than Fow‐3. Circular dichroism and retention time in reversed‐phase high‐performance chromatography demonstrated that the drastic decrease in mammalian cell toxicity of the Fow‐3 analog correlated with increased conformational flexibility and decreased hydrophobicity. This analog had potent antimicrobial activity against antibiotic‐resistant bacteria, including methicillin‐resistant Staphylococcus aureus, multidrug‐resistant Pseudomonas aeruginosa, and vancomycin‐resistant Enterococcus faecium. Furthermore, this analog was more synergistic with chloramphenicol and oxacillin compared to LL‐37, against a multidrug‐resistant strain of P. aeruginosa. Thus, repositioning the proline residues to the center of the α‐helical antimicrobial peptides improved cell selectivity. [ABSTRACT FROM AUTHOR]

Cellular mobility is one of the key factors that must be studied to understand the intracellular environment and intracellular mechanisms. To achieve this, various methods using micro/nano fabrication technology have been attempted. However, there are many restrictions, depending on materials, substrates, and situations. In this study, we modified surfaces readily by exploiting the mechanical fragility of nanostructures. This method is applicable to various substrates for bioengineering applications. It can also be used to modify a surface while growing cells, allowing for in situ patterning. The surface will affect cellular co-culture and tissue engineering by controlling cellular mobility. [ABSTRACT FROM AUTHOR]

Papiliocin is a 37-residue antimicrobial peptide, with Trp2 and Phe5 previously reported as key residues necessary for its antibacterial activity. This study determined the essential length of the N-terminal fragment of papiliocin necessary to retain its biological activity. We designed and synthesized an array of seven peptides from the N-terminal helix (PapN), with longest peptide with 22 residues and the shortest peptide consisting of the first 10 residues. The minimum inhibitory concentration (MIC) values and cytotoxicity measurement revealed that a PapN-12mer containing a three-turn, amphipathic helix was the shortest peptide exhibiting antibacterial activity without cytotoxicity. Additionally, PapN-20mer peptide containing two isoleucines at the C-terminus represented the shortest peptide exhibiting potent anti-inflammatory activities by inhibiting nitric oxide production and inflammatory cytokine production in lipopolysaccharide-stimulated mouse macrophage RAW264.7 cells. These results provided valuable insights into the design of short, potent peptide analogs of papiliocin. [ABSTRACT FROM AUTHOR]

KR-12-a5 is a 12-meric α-helical antimicrobial peptide (AMP) with dual antimicrobial and anti-inflammatory activities designed from human cathelicidin LL-37. We designed and synthesized a series of d -amino acid-substituted analogs of KR-12-a5 with the aim of developing novel α-helical AMPs that possess higher cell selectivity than KR-12-a5, while maintaining the anti-inflammatory activity. d -amino acid incorporation into KR-12-a5 induced a significant improvement in the cell selectivity by 2.6- to 13.6-fold as compared to KR-12-a5, while maintaining the anti-inflammatory activity. Among the three analogs, KR-12-a5 (6- D L) with d -amino acid in the polar-nonpolar interface (Leu 6 ) showed the highest cell selectivity (therapeutic index: 61.2). Similar to LL-37, KR-12-a5 and its analogs significantly inhibited the expression and secretion of NO, TNF-α, IL-6 and MCP-1 from LPS-stimulated RAW264.7 cells. KR-12-a5 and its analogs showed a more potent antimicrobial activity against antibiotic-resistant bacteria, including clinically isolated MRSA, MDRPA, and VREF than LL-37 and melittin. Furthermore, compared to LL-37, KR-12-a5 and its analogs showed greater synergistic effects with conventional antibiotics, such as chloramphenicol, ciprofloxacin, and oxacillin against MDRPA; KR-12-a5 and its analogs had a FICI range between 0.25 and 0.5, and LL-37 had a range between 0.75 and 1.5. KR-12-a5 and its analogs were found to be more effective anti-biofilm agents against MDRPA than LL-37. In addition, KR-12-a5 and its analogs maintained antimicrobial activity in physiological salts and human serum. SYTOX Green uptake and membrane depolarization studies revealed that KR-12-a5 and its analogs kills microbial cells by permeabilizing the cell membrane and damaging membrane integrity. Taken together, our results suggest that KR-12-a5 and its analogs can be developed further as novel antimicrobial/anti-inflammatory agents to treat antibiotic-resistant infections. [ABSTRACT FROM AUTHOR]

Although the human-derived antimicrobial peptide (AMP) LL-37 has potent antimicrobial and anti-inflammatory activities, its therapeutic application is limited by its low cell selectivity and high production cost due to its large size. To overcome these problems, we tried to develop novel LL-37-derived short α-helical AMPs with improved cell selectivity and without a significant loss of anti-inflammatory activity relative to that of parental LL-37. Using amino acid substitution, we designed and synthesized a series of FK13 analogs based on the sequence of the 13-meric short FK13 peptide (residues 17–29 of LL-37) that has been identified as the region responsible for the antimicrobial activity of LL-37. Among the designed FK13 analogs, FK-13-a1 and FK-13-a7 showed high cell selectivity and retained the anti-inflammatory activity. The therapeutic index (a measure of cell selectivity) of FK-13-a1 and FK-13-a7 was 6.3- and 2.3-fold that of parental LL-37, respectively. Furthermore, FK-13-a1 and FK-13-a7 displayed more potent antimicrobial activity against antibiotic-resistant bacteria including MRSA, MDRPA, and VREF, than did LL-37. In addition, FK-13-a1 and FK-13-a7 exhibited greater synergistic effects with chloramphenicol against MRSA and MDRPA and were more effective anti-biofilm agents against MDRPA than LL-37 was. Moreover, FK-13-a1 and FK-13-a7 maintained their activities in the presence of physiological salts and human serum. SYTOX green uptake, membrane depolarization and killing kinetics revealed that FK13-a1 and FK13-a7 kills microbial cells by permeabilizing the cell membrane and damaging membrane integrity. Taken together, our results suggest that FK13-a1 and FK13-a7 can be developed as novel antimicrobial/anti-inflammatory agents. [ABSTRACT FROM AUTHOR]

[Display omitted] • N-terminal modified peptides preferentially bind with anionic lipid bilayer membrane, which could be used for bio-analysis. • The anti-cancer activities of both peptides were evaluated against the four cancer cells and normal cells namely, Hep-2, HepG2, MCF-7, A375, and Vero. • The peptide is largely unordered to ordered structure and non-toxic towards normal cell. • In-silico studies suggested that the peptides have good binding interactions with the proteins (PDB ID: 2W3L, 5FC4, and 5GWK). The short peptides, containing the amino acid sequence asparagine-glycine-arginine (NGR) and arginine-glycine-aspartic acid (RGD), possess the strong binding ability to N (APN/CD13) aminopeptidase receptor and integrin proteins involved in antitumor properties are overexpressed. A novel short N-terminal modified hexapeptides P1 and P2 was designed and synthesized using the Fmoc-chemistry solid phase peptide synthesis protocol. Notably, the cytotoxicity of the MTT assay demonstrated the viability of normal and cancer cells up to lower peptide concentrations. Interestingly, both peptides show good anticancer activities against the four cancer cells and normal cells namely, Hep-2, HepG2, MCF-7, A375, and Vero and compared with standard drugs, doxorubicin and paclitaxel. Additionally, in silico studies were applied to predict the binding sites and binding orientation of the peptides for potential anticancer targets. Steady-state fluorescence measurements showed that peptide P1 exhibits preferential interactions with POPC/POPG anionic bilayers rather than the zwitterionic POPC lipid bilayers and peptide P2 , did not show any preferential interaction with lipids bilayers. But impressively, peptide P2 shows anticancer activity due to the NGR/RGD motif. Circular dichroism studies demonstrated that the peptide's secondary structure changes only minimally upon binding to the anionic lipid bilayers. [ABSTRACT FROM AUTHOR]

Photodynamic molecular beacons (PMBs) are highly appealing for activatable photodynamic therapy (PDT), but their applications are hindered by limited therapeutic efficacy. Here, by molecular engineering of enzyme-responsive units in the loop region of DNA-based PMBs, we present for the first time the modular design of an enzyme/microRNA dual-regulated PMB (D-PMB) to achieve cancer-cell-selective amplification of PDT efficacy. In the design, the "inert" photosensitizers in D-PMB could be repeatedly activated in the presence of both tumor-specific enzyme and miRNA, leading to amplified generation of cytotoxic singlet oxygen species and therefore enhanced PDT efficacy in vitro and in vivo. By contrast, low photodynamic activity could be observed in healthy cells, as D-PMB activation has been largely avoided by the dual-regulatable design. This work presents a cooperatively activated PDT strategy, which enables enhanced therapeutic efficacy with improved tumor-specificity and thus conceptualizes an approach to expand the repertoire of designing smart tumor treatment modality.

Trigger-activatable antisense oligonucleotides have been widely applied to regulate gene function. Among them, caged cyclic antisense oligonucleotides (cASOs) maintain a specific topology that temporarily inhibits their interaction with target genes. By inserting linkers that respond to cell-specific endogenous stimuli, they can be powerful tools and potential therapeutic agents for specific types of cancer cells with low off-target effects on normal cells. Here, we developed enzyme-activatable cASOs by tethering two terminals of linear antisense oligonucleotides through a cathepsin B (CB) substrate peptide (Gly-Phe-Leu-Gly [GFLG]), which could be efficiently uncaged by CB. CB-activatable cASOs were used to successfully knock down two disease-related endogenous genes in CB-abundant PC-3 tumor cells at the mRNA and protein levels but had much less effect on gene knockdown in CB-deficient human umbilical vein endothelial cell (HUVECs). In addition, reduced nonspecific immunostimulation was found using cASOs compared with their linear counterparts. Further in vivo studies indicated that CB-activatable cASOs showed effective tumor inhibition in PC-3 tumor model mice through downregulation of translationally controlled tumor protein (TCTP) protein in tumors. This study applies endogenous enzyme-activatable cASOs for antitumor therapy in tumor model mice, which demonstrates a promising stimulus-responsive cASO strategy for cell-specific gene knockdown upon endogenous activation and ASO prodrug development., Competing Interests: The authors declare no competing interests., (© 2023 The Author(s).)

Thirty-five types of novel pyridocarbazoles (5-(N-alkyl)carbamoyl-11-methyl-6H-pyrido[4,3-b]carbazoles and 5-(N-alkyl)carbamoyl-2,11-dimethyl-6H-pyrido[4,3-b]carbazol-2-ium chloride derivatives), that were conjugated with amine, N-methylurea, and N-methyl-N-nitrosourea moieties through alkyl-, oxyalkyl-, and iminoalkylcarbamoyl linkers, were synthesized by a series of reactions of methyl 11-methyl-6H-pyrido[4,3-b]carbazole-5-carboxylates with polymethylenediamine (n=2-5), p-nitrophenyl N-methylcarbamate, and N-methyl-N-nitrosocarbamate in high yields, and their cytotoxic activities were evaluated against Sarcoma-180, NIH3T3, HeLa S-3, and L1210 cell lines. These compounds exhibited potent cytotoxic activity (IC50=1.6-50 µM) and odd-even alternation effect. 9-Methoxy-2,11-dimethyl-5-((2-(3-methyl-3-nitrosoureido)ethyl)carbamoyl)-6H-pyrido[4,3-b]carbazol-2-ium chloride exhibited the most potent cytotoxic activity (IC50=0.15 µM) and cell selectivity against HeLa S-3. Some of the un-charged 5-(N-alkyl)carbamoyl-6H-pyrido[4,3-b]carbazole derivatives, which were conjugated with the amine and N-methyl-N-nitrosourea moieties through a dimethylene spacer, also exhibited potent cytotoxic activity (IC50=0.43-2.4 µM) and remarkable cell selectivity as well as ellipticine, 9-hydroxyellipticine, and the starting methyl ester of the pyridocarbazole-5-carboxylate (IC50=0.30-2.2 µM). [ABSTRACT FROM AUTHOR]

Multifunctional constructs providing a proper environment for adhesion and growth of selected cell types are needed for most tissue engineering and regenerative medicine applications. In this context, vinylsulfone (VS)-modified dextran was proposed as a matrix featuring low-fouling properties as well as multiple versatile moieties. The displayed VS groups could indeed react with thiol, amine or hydroxyl groups, be it for surface grafting, crosslinking or subsequent tethering of biomolecules. In the present study, a library of dextran-VS was produced, grafted to aminated substrates and characterized in terms of degree of VS modification (%VS), cell-repelling properties and potential for the oriented grafting of cysteine-tagged peptides. As a bioactive coating of vascular implants, ECM peptides (e.g. RGD) as well as vascular endothelial growth factor (VEGF) were co-immobilized on one of the most suitable dextran-VS coating (%VS = ca. 50% of saccharides units). Both RGD and VEGF were efficiently tethered at high densities ( ca. 1 nmol/cm 2 and 50 fmol/cm 2 , respectively), and were able to promote endothelial cell adhesion as well as proliferation. The latter was enhanced to the same extent as with soluble VEGF and proved selective to endothelial cells over smooth muscle cells. Altogether, multiple biomolecules could be efficiently incorporated into a dextran-VS construct, while maintaining their respective biological activity. Statement of Significance This work addresses the need for multifunctional coatings and selective cell response inherent to many tissue engineering and regenerative medicine applications, for instance, vascular graft. More specifically, a library of dextrans was first generated through vinylsulfone (VS) modification. Thoroughly selected dextran-VS provided an ideal platform for unbiased study of cell response to covalently grafted biomolecules. Considering that processes such as healing and angiogenesis require multiple factors acting synergistically, vascular endothelial growth factor (VEGF) was then co-immobilized with the cell adhesive RGD peptide within our dextran coating through a relevant strategy featuring orientation and specificity. Altogether, both adhesive and proliferative cues could be incorporated into our construct with additive, if not synergetic, effects. [ABSTRACT FROM AUTHOR]

Messenger RNA (mRNA) is being used as part of an emerging class of biotherapeutics with great promise for preventing and treating a wide range of diseases, as well as encoding programmable nucleases for genome editing. However, mRNA's low stability and immunogenicity, as well as the impermeability of the cell membrane to mRNA greatly limit mRNA's potential for therapeutic use. Lipid nanoparticles (LNPs) are currently one of the most extensively studied nanocarriers for mRNA delivery and have recently been clinically approved for developing mRNA-based vaccines to prevent COVID-19. In this review, we summarize the latest advances in designing ionizable lipids and formulating LNPs for intracellular and tissue-targeted mRNA delivery. Furthermore, we discuss the progress of intracellular mRNA delivery for spatiotemporally controlled CRISPR/Cas9 genome editing by using LNPs. Finally, we provide a perspective on the future of LNP-based mRNA delivery for CRISPR/Cas9 genome editing and the treatment of genetic disorders., (© 2023 Wiley-VCH GmbH.)

To generate effective antimicrobial peptides (AMPs) with good antimicrobial activities and cell selectivity, many synthetic strategies have been implemented to facilitate the development of AMPs. However, these synthetic strategies represent only a small proportion of the methods used for the development of AMPs and are not optimal with the requirements needed for the design of AMPs. In this investigation, we designed a mirror-like structure with a lower charge and a higher number of hydrophobic amino acids. The amino acid sequence of the designed mirror-like peptides was XXYXXXY XXXYXX [X represents L (Leu) and/or A (Ala); Y represents K (Lys)]. These mirror-like peptides displayed antimicrobial activity against both Gram-positive and Gram-negative bacteria. Hemolysis activity and cytotoxicity, detected by using human red blood cells (hRBCs) and human embryonic kidney cells (HEK293), respectively, demonstrated that the frequency of Ala residues in this structure had a regulatory effect on the high hydrophobic region. In particular, KLA showed a greater antimicrobial potency than the other three mirror-like peptides, folded into an α-helical structure, and displayed the highest therapeutic index, suggesting its good cell selectivity. Observations from fluorescence spectroscopy, flow cytometry, and electron microscopy experiments indicated that KLA exhibited good membrane penetration potential by inducing membrane blebbing, disruption and lysis. Therefore, generating mirror-like peptides is a promising strategy for designing effective AMPs with regions of high hydrophobicity. [ABSTRACT FROM AUTHOR]

The β-hairpin structure has been proposed to exhibit potent antimicrobial properties with low cytotoxicity, thus, multiple β-hairpin structures have been proved to be highly stable in structures containing tightly packed hydrophobic cores. The aim of this study was to develop peptide-based synthetic strategies for generating short, but effective AMPs as inexpensive antimicrobial agents. Multiple-stranded β-hairpin peptides with the same β-hairpin unit, (WRXxRW) n where n = 1, 2, 3, or 4 and Xx represent the turn sequence, were synthesized, and their potential as antimicrobial agents was evaluated. Owning to the tightly packed hydrophobic core and paired Trp of this multiple-stranded β-hairpin structure, all the 12-residues peptides exhibited high cell selectivity towards bacterial cells over human red blood cells (hRBCs), and the peptide W2 exhibited stronger antimicrobial activities with the MIC values of 2–8 μM against various tested bacteria. Not only that, but W2 also showed obvious synergy with streptomycin and chloramphenicol against Escherichia coli , and displayed synergy with ciprofloxacin against Staphylococcus aureus with the FICI values ⩽0.5. Fluorescence spectroscopy and electron microscopy analyses indicated that W2 kills microbial cells by permeabilizing the cell membrane and damaging membrane integrity. Collectively, based on the multiple β-hairpin peptides, the ability to develop libraries of short and effective peptides will be a powerful approach to the discovery of novel antimicrobial agents. Statement of significance We successfully screened a peptide W2 ((WRPGRW)2) from a series of multiple-stranded β-hairpin antimicrobial peptides based on the “S-shaped” motif that induced the formation of a globular structure, and Trp zipper was used to replace the disulfide bonds to reduce the cost of production. This novel structure applied to AMPs improved cell selectivity and salt stability. The findings of this study will promote the development of peptide-based antimicrobial biomaterials. Further exploration of these AMPs will allow for diverse biotechnological and clinical applications such as biomedical coating, food storaging, and animal feeding. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) with amphipathic β-hairpin structures have been demonstrated to possess potent antimicrobial activities and great cell selectivities. However, our understanding of β-hairpin antimicrobial peptides lags behind that of α-helices, mainly because it is difficult for short peptides to form robust β-hairpin structures. Tryptophan zipper (trpzip) peptides are among the most stable β-hairpin peptides known to fold spontaneously without requiring covalent disulfide constraint or metal binding. To develop model β-hairpin AMPs with small size and remarkable stability, a series of amphiphilic linear peptides were designed based on the trpzip motif. The sequence of designed peptides is (WK)PG(KW)-NH ( n = 1, 2, 3, 4, 5), and the antimicrobial activity and membrane interaction mechanism of the peptides were evaluated. The results showed that these peptides readily fold into β-hairpin structures in aqueous and membrane-mimicking environments and exhibit broad-spectrum antimicrobial activities against both gram-positive and gram-negative bacteria. The antibacterial potency of the peptides initially increased and then decreased with increasing chain length. WK3, a 14-residue peptide, displayed excellent antimicrobial activity with minimal hemolytic activity and cytotoxicity, suggesting that it possesses great cell selectivity. Scanning electron microscopy (SEM), transmission electron microscopy (TEM), fluorescence spectroscopy, and flow cytometry indicated that representative peptides WK3 and WK4 exert their activities by permeabilizing the microbial membrane and damaging cell membrane integrity. This study reveals the application potential of the designed peptides as promising antimicrobial agents for the control of infectious diseases, and it also provides new insights into the design and optimization of highly stable β-hairpin AMPs with great antimicrobial activities and cell selectivities. [ABSTRACT FROM AUTHOR]

In the solid state each of three binuclear zinc dithiocarbamates bearing hydroxyethyl groups, {Zn[S 2 CN(R)CH 2 CH 2 OH] 2 } 2 for R = iPr ( 1 ), CH 2 CH 2 OH ( 2 ), and Me ( 3 ), and an all alkyl species, [Zn(S 2 CNEt 2 ) 2 ] 2 ( 4 ), features a centrosymmetric {ZnSCS} 2 core with a step topology; both 1 and 3 were isolated as monohydrates. All compounds were broadly cytotoxic, specifically against human cancer cell lines compared with normal cells, with greater potency than cisplatin. Notably, some selectivity were indicated with 2 being the most potent against human ovarian carcinoma cells (cisA2780), and 4 being more cytotoxic toward multidrug resistant human breast carcinoma cells (MCF-7R), human colon adenocarcinoma cells (HT-29), and human lung adenocarcinoma epithelial cells (A549). Based on human apoptosis PCR-array analysis, caspase activities, DNA fragmentation, cell apoptotic assays, intracellular reactive oxygen species (ROS) measurements and human topoisomerase I inhibition, induction of apoptosis in HT-29 cells is demonstrated via both extrinsic and intrinsic pathways. Compounds 2 – 4 activate the p53 gene while 1 activates both p53 and p73. Cell cycle arrest at the S and G 2 /M phases correlates with inhibition of HT-29 cell growth. Cell invasion is also inhibited by 1 – 4 which is correlated with down-regulation of NF -κ B. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) are considered as potential antibiotic substitutes because of their potent activities. Previous studies mainly focused on the effects of peptide charges and secondary structures, but the self-assembly of AMPs was neglected. As more and more researchers notice the roles of peptide self-assembly in AMPs, it has been considered as another important property. In this review, we will discuss the influences of peptide self-assembly on the activity and mode of action, and some specific features it introduces to the AMPs, such as particular responsiveness, improved cell selectivity and stability and sustained release. In addition, some methods to design self-assembling AMPs are primarily discussed. With further understanding about the self-assembling regularity, design of particular self-assembling AMPs will be very helpful for their applications, especially in the fields of drug delivery and biomedical engineering. Copyright © 2015 European Peptide Society and John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Antimicrobial peptides (AMPs) serve as a defense mechanism within multicellular organisms and are attracting increasing attention because of their potential application in the treatment of multidrug-resistant infections. Amphipathicity has long been believed to be the most important consideration for the structural modification and design of both naturally occurring and synthetic AMPs. Previous studies indicated that disruptive amphipathicity formed by replacing the paired charged amino acid residues on the polar face of an amphipathic helix with tryptophan residues linked with hydrogen bonds on the basis of α-helical protein folding principles endowed the AMPs with increased cell selectivity. In an attempt to augment and hone this strategy further, we designed a series of imperfect amphipathic peptides by placing different types of amino acid residues at the hydrogen bond linked positions of α-helix structures to characterize their antimicrobial properties and mechanism of action. The d -Trp-substituted sequence (PRW4-d) showed greater antimicrobial potency than Cys-(C4), Asp-(D4), Ile-(I4), and Pro-(P4) substituted sequences, comparable to the l -Trp-substituted parent sequence (PRW4). Furthermore, the total replacement of Lys residues with Arg residues along the peptide sequence (PRW4-R) exhibited enhanced antimicrobial activity and cell selectivity. In addition, no cytotoxicity was observed among these synthetic peptides. PRW4-d and PRW4-R maintained their activities in the presence of physiological salts and human serum. The fluorescence spectroscopy, flow cytometry, and electron microscopy observations indicated that the optimized sequences exhibited excellent antimicrobial potency by inducing cytoplasmic membrane potential loss, membrane permeabilization and disruption. Collectively, the results could be useful for designing short AMPs with great antimicrobial activity and cell selectivity. [ABSTRACT FROM AUTHOR]