Transiently restricting individual amino acids protects Drosophila melanogaster against multiple stressors.

Nutrition and resilience are linked, though it is not yet clear how diet confers stress resistance or the breadth of stressors that it can protect against. We have previously shown that transiently restricting an essential amino acid can protect Drosophila melanogaster against nicotine poisoning. Here, we sought to characterize the nature of this dietary-mediated protection and determine whether it was sex, amino acid and/or nicotine specific. When we compared between sexes, we found that isoleucine deprivation increases female, but not male, nicotine resistance. Surprisingly, we found that this protection afforded to females was not replicated by dietary protein restriction and was instead specific to individual amino acid restriction. To understand whether these beneficial effects of diet were specific to nicotine or were generalizable across stressors, we pre-treated flies with amino acid restriction diets and exposed them to other types of stress. We found that some of the diets that protected against nicotine also protected against oxidative and starvation stress, and improved survival following cold shock. Interestingly, we found that a diet lacking isoleucine was the only diet to protect against all these stressors. These data point to isoleucine as a critical determinant of robustness in the face of environmental challenges.

Distinct Amino Acid-Based PROTACs Target Oncogenic Kinases for Degradation in Non-Small Cell Lung Cancer (NSCLC).

Proteolysis-targeting chimeras (PROTACs) selectively eliminate detrimental proteins by exploiting the ubiquitin-proteasome system (UPS), representing a promising therapeutic strategy against various diseases. Effective adaptations of degradation signal sequences and E3 ligases for PROTACs remain limited. Here, we employed three amino acids─Gly, Pro, and Lys─as the ligand to recruit the corresponding E3 ligases: CRL2ZYG11B/ZER1, GID4, and UBRs, to degrade EML4-ALK and mutant EGFR, two oncogenic drivers in NSCLC. We found that the extent of EML4-ALK and EGFR reduction can be easily fine-tuned by using different degradation signals. These amino acid-based PROTACs, termed AATacs, hindered proliferation and induced cell cycle arrest and apoptosis of NSCLC cells in vitro. Compared to other PROTACs, AATacs are small, interchangeable but with different degradation efficiency. Our study further expands the repertoire of E3 ligases and their ligands for PROTAC application, improving the versatility and utility of targeted protein degradation for therapeutic purposes.

Amino Acid-Conjugated Polymer-Silver Bromide Nanocomposites for Eradicating Polymicrobial Biofilms and Treating Burn Wound Infections.

The rise in antimicrobial resistance, the increasing occurrence of bacterial, and fungal infections, and the challenges posed by polymicrobial biofilms necessitate the exploration of innovative therapeutic strategies. Silver-based antimicrobials have garnered attention for their broad-spectrum activity and multimodal mechanisms of action. However, their effectiveness against single-species or polymicrobial biofilms remains limited. In this study, we present the fabrication of polymer-silver bromide nanocomposites using amino acid conjugated polymers (ACPs) through a green and water-based in situ technique. The nanocomposite architecture facilitated prolonged and controlled release of the active components. Remarkably, the nanocomposites exhibited broad-spectrum activity against multidrug-resistant (MDR) human pathogenic bacteria (MIC = 2-16 µg/mL) and fungi (MIC = 1-8 µg/mL), while displaying no detectable toxicity to human erythrocytes (HC50 > 1024 µg/mL). In contrast to existing antimicrobials and silver-based therapies, the nanocomposite effectively eradicated bacterial, fungal, and polymicrobial biofilms, and prevented the development of microbial resistance due to their membrane-active properties. Furthermore, the lead polymer-silver bromide nanocomposite demonstrated a 99% reduction in the drug-resistant Pseudomonas aeruginosa burden in a murine model of burn wound infection, along with excellent in vivo biocompatibility.

Intake of Special Amino Acids Mixture Leads to Blunted Murine Colon Cancer Growth In Vitro and In Vivo.

Cancer cells require substantial amounts of energy and substrates for their metabolic hyperactivity, enabling the synthesis of new cells at the expense of healthy ones. Preliminary in vitro data suggest that a mix of free essential amino acids (EAA-mix) can promote cancer cell apoptosis by enhancing autophagy. This study aimed to confirm, both in vitro and in vivo, whether EAA intake could influence the development of colon cancer in mice. We investigated changes in cancer proliferation in CT26 cells treated with EAA-mix and in mice fed with EAA-rich modified diets (EAARD) as compared to those on a standard laboratory diet (StD). CT26 cells were injected subcutaneously (s.c.) or intraperitoneally (i.p.). After 21 days, tumors were removed and measured. In vitro data corroborated that EAA-mix impairs cancer growth by inducing apoptosis. In vivo data revealed that mice on StD developed significantly larger (s.c.) and more numerous (i.p.) cancers than those on EAARD. EAA administration appears to influence cancer cell survival with notable antiproliferative properties.

Design, synthesis and evaluation of carbamate-bridged amino acid prodrugs of cycloicaritin with improved antitumor activity, aqueous solubility and phase II metabolic stability.

Cycloicaritin (CICT), a bioactive flavonoid derived from the genus Epimedium, exhibits a variety of beneficial biological activities, including promising anticancer effects. However, its poor oral bioavailability is attributed to its extremely low aqueous solubility and rapid elimination via phase II conjugative metabolism. To overcome these limitations, we designed and synthesized a series of carbamate-bridged prodrugs, protecting the hydroxyl group at the 3-position of cycloicaritin by binding with the N-terminus of a natural amino acid. The optimal prodrug 4b demonstrated a significant increase in aqueous solubility as compared to CICT, as well as improved stability in phase II metabolism, while allowing for a rapid release of CICT in the blood upon gastrointestinal absorption. The prodrug 4b also facilitated oral absorption through organic anion-transporting polypeptide 2B1-mediated transport and exhibited moderate cytotoxicity. Importantly, the prodrug enhanced the oral bioavailability of CICT and displayed dose-dependent antitumor activity with superior safety. In summary, the prodrug 4b is a novel potential antitumor drug candidate, and the carbamate-bridged amino acid prodrug approach is a promising strategy for the oral delivery of CICT.

Select amino acids recover cytokine-altered ENaC function in human bronchial epithelial cells.

The airway epithelium plays a pivotal role in regulating mucosal immunity and inflammation. Epithelial barrier function, homeostasis of luminal fluid, and mucociliary clearance are major components of mucosal defense mechanisms. The epithelial sodium channel (ENaC) is one of the key players in controlling airway fluid volume and composition, and characteristic cytokines cause ENaC and barrier dysfunctions following pulmonary infections or allergic reactions. Given the limited understanding of the requisite duration and magnitude of cytokines to affect ENaC and barrier function, available treatment options for restoring normal ENaC activity are limited. Previous studies have demonstrated that distinct amino acids can modulate epithelial ion channel activities and barrier function in intestines and airways. Here, we have investigated the time- and concentration-dependent effect of representative cytokines for Th1- (IFN-γ and TNF-α), Th2- (IL-4 and IL-13), and Treg-mediated (TGF-ß1) immune responses on ENaC activity and barrier function in human bronchial epithelial cells. When cells were exposed to Th1 and Treg cytokines, ENaC activity decreased gradually while barrier function remained largely unaffected. In contrast, Th2 cytokines had an immediate and profound inhibitory effect on ENaC activity that was subsequently followed by epithelial barrier disruption. These functional changes were associated with decreased membrane protein expression of α-, ß-, and γ-ENaC, and decreased mRNA levels of ß- and γ-ENaC. A proprietary blend of amino acids was developed based on their ability to prevent Th2 cytokine-induced ENaC dysfunction. Exposure to the select amino acids reversed the inhibitory effect of IL-13 on ENaC activity by increasing mRNA levels of ß- and γ-ENaC, and protein expression of γ-ENaC. This study indicates the beneficial effect of select amino acids on ENaC activity in an in vitro setting of Th2-mediated inflammation suggesting these amino acids as a novel therapeutic approach for correcting this condition.

Bioinformatic Approaches Identify Hybrid Antibiotics against Tuberculosis via d-Amino Acid-Activating Adenylation Domains from Cordyceps militaris.

The development of tuberculosis (TB) therapy has been marked by the discovery of natural-product-derived streptomycin, followed by the introduction of NP-derived rifampicin, representing a significant milestone in the history of TB management. However, TB remains a global challenge, with the emergence of multidrug-resistant Mycobacterium tuberculosis highlighting the need for novel therapeutic agents. In this study, a bioinformatic approach was employed to investigate d-amino acid-activating adenylation domains, leading to the identification of cordysetin A (1), a novel trans-decalin tetramic acid antibiotic from the ascomycete fungi Cordyceps militaris. Cordysetin A (1) exhibits considerable activity against M. tuberculosis in vitro and in vivo while maintaining low cytotoxicity. These results reveal that the d-configuration of the amino acid within this hybrid polyketide-nonribosomal antibiotic is crucial for preserving its anti-tuberculosis efficacy. These findings emphasize the significant translational potential of cordysetin A as a promising candidate for TB treatment, furthering our understanding of bioinformatic approaches in the development of effective anti-tuberculosis agents.

An Amino Acid Mixture to Counteract Skeletal Muscle Atrophy: Impact on Mitochondrial Bioenergetics.

Skeletal muscle atrophy (SMA) is caused by a rise in muscle breakdown and a decline in protein synthesis, with a consequent loss of mass and function. This study characterized the effect of an amino acid mixture (AA) in models of SMA, focusing on mitochondria. C57/Bl6 mice underwent immobilization of one hindlimb (I) or cardiotoxin-induced muscle injury (C) and were compared with controls (CTRL). Mice were then administered AA in drinking water for 10 days and compared to a placebo group. With respect to CTRL, I and C reduced running time and distance, along with grip strength; however, the reduction was prevented by AA. Tibialis anterior (TA) muscles were used for histology and mitochondria isolation. I and C resulted in TA atrophy, characterized by a reduction in both wet weight and TA/body weight ratio and smaller myofibers than those of CTRL. Interestingly, these alterations were lightly observed in mice treated with AA. The mitochondrial yield from the TA of I and C mice was lower than that of CTRL but not in AA-treated mice. AA also preserved mitochondrial bioenergetics in TA muscle from I and C mice. To conclude, this study demonstrates that AA prevents loss of muscle mass and function in SMA by protecting mitochondria.

Design, Synthesis, and Antitumor Activity of Isoliquiritigenin Amino Acid Ester Derivatives.

Isoliquiritigenin (ISL) is a chalcone that has shown great potential in the treatment of cancer. However, its relatively weak activity and low water solubility limit its clinical application. In this study, we designed and synthesized 21 amino acid ester derivatives of ISL and characterized the compounds using 1H NMR and 13C NMR. Among them, compound 9 (IC50 = 14.36 µM) had a better inhibitory effect on human cervical cancer (Hela) than ISL (IC50 = 126.5 µM), and it was superior to the positive drug 5-FU (IC50 = 33.59 µM). The mechanism of the action experiment showed that compound 9 could induce Hela cell apoptosis and autophagy through the PI3K/Akt/mTOR pathway.

Design of Proteolytic-Resistant Antifungal Peptides by Utilizing Minimum d-Amino Acid Ratios.

Antifungal peptides are an appealing alternative to standard antifungal medicines due to their unique mechanism of action and low-level resistance. However, their susceptibility to protease degradation keeps hindering their future development. Herein, a library was established to design peptides with protease resistance and high antifungal activity. The peptides were incorporated with minimal D-amino acids to further improve the protease stability. The most active peptide, IR3, demonstrated good antifungal activity and low toxicity, and its molecular integrity was maintained after protease hydrolysis for 8 h at 2 mg/mL. Furthermore, IR3 could permeate the fungal cell wall, disrupt the cell membrane, produce reactive oxygen species, and induce apoptosis in fungal cells. In vivo experiments confirmed that IR3 could effectively treat fungal keratitis. Collectively, these findings suggest that IR3 is a promising antifungal agent and may be beneficial in the design and development of protease-resistant antifungal peptides.

Synthesis and anti-SARS-CoV-2 activity of amino acid modified cephalotaxine derivatives.

The severe acute respiratory syndrome coronavirus (SARS-CoV-2) pandemic has triggered a significant impact on global public health security, it is urgent to develop effective antiviral drugs. Previous studies have found that binding to ACE2 is a key step in the invasion of SARS-CoV-2 into host cells, so virus invasion can be inhibited by blocking ACE2, but there are few reports on this kind of specific inhibitor. Our previous study found that Harringtonine (HT) can inhibit the entry of SARS-CoV-2 spike pseudovirus into ACE2h cells, but its relatively high cytotoxicity limits its further development. Amino acid modification of the active components can increase their solubility and reduce their cytotoxicity. Therefore, in this study, seven new derivatives were synthesized by amino acid modification of its core structure Cephalotaxine. The target compounds were evaluated by cell viability assay and the SARS-CoV-2 spike pseudovirus entry assay. Compound CET-1 significantly inhibited the entry of pseudovirus into ACE2h cells and showed less cytotoxicity than HT. Molecular docking results showed that CET-1 could bind TYR83, an important residue of ACE2, just like HT. In conclusion, our study provided a novel compound with more potential activity and lower toxicity than HT on inhibiting the SARS-CoV-2 spike pseudovirus infection, which makes it possible to be a lead compound as an antiviral drug in the future.

The administration of Cerebrolysin elicits neuroprotective and neurorepair effects in an animal model of type 1 diabetes mellitus.

Diabetes mellitus (DM) is a metabolic disorder impacting cerebral function. The administration of Streptozotocin (STZ) is a well-known animal model of insulinopenic type 1 DM in rats. STZ-induced DM results in a myriad of alteration in the periphery and central nervous system (CNS). Cerebrolysin (CBL) is a neuropeptide preparation that promotes synaptic and neuronal plasticity in various animal models. In all cases, CBL was administered when the model was established. This research aims to investigate the neuroprotective and neurorepair effect of CBL on the cytoarchitecture of neurons and spine density in pyramidal neurons of the prefrontal (PFC) and the CA1 region of the dorsal hippocampus, as well as spheroidal neurons of the dentate gyrus (DG), in STZ-induced DM. In the first experimental condition, STZ and CBL are administered at the same time to evaluate the potential preventive effect of CBL. In the second experimental condition, CBL was administered two months after establishing the DM model to measure the potential neurorepair effect of CBL. STZ-induced hyperglycemia remained unaltered by the administration of CBL in both experimental conditions. In the first experimental condition, CBL treatment preserved the neuronal morphology in PFC layer 3, PFC layer 5 and the DG of the hippocampus, while also maintaining spine density in the PFC-3, DG and CA1 hippocampus. Furthermore, CBL induced neurorepair in neurons within the PFC-3, PFC-5 and CA1 regions of the hippocampus, along with an increase in spine density in the PFC-3, DG and CA1 hippocampus. These findings suggest that CBL´s effects on neuroplasticity could be observed before or after the damage was evident.

Self-assembly of enzymes and prodrugs with clickable amino acids for nucleus-targeted cancer therapy.

A nucleus-targeted nanocomposite was prepared by clickable amino acid-tuned one-step co-assembly of proteins and chemotherapeutics. The nanocomposite with favorable pharmacokinetic behavior can effectively accumulate in the nucleus, thereby significantly enhancing the anticancer therapeutic effect both in vitro and in vivo.

Inverted amino acids reduce the adhesion and biofilm biomass of early oral colonizers.

BACKGROUND: Early colonizers adhere to the dental surface and facilitate the initial adhesion of secondary colonizers to form oral biofilms, which may cause oral infections. OBJECTIVES: This study aimed to determine the antimicrobial, anti-adhesion and antibiofilm potency of inverted amino acids on early colonizer streptococci and their mixed species. MATERIAL AND METHODS: The following test strains were used: Streptococcus gordonii (American Type Culture Collection (ATCC) 35105); Streptococcus mitis (ATCC 49456); Streptococcus oralis (ATCC 10557); Streptococcus salivarius (ATCC 7073); and Streptococcus sanguinis (ATCC BAA-1455). The concentration-dependent antimicrobial potency of d-alanine (d-ala), d-arginine (d-arg), d-leucine (d-leu), d-methionine (d-met), and d-tryptophan (d-try) was determined using the Clinical and Laboratory Standards Institute (CLSI) broth microdilution method with AlamarBlue modification. The adhesion of primary colonizers in the presence of 25-mM d-amino acids (dAAs) was assessed using the colony forming unit (CFU) assay. The CFU assay was conducted on 24-h flow cell bacterial biofilm models after exposure to 25-mM inverted dAAs. RESULTS: No minimum inhibitory concentration (MIC) point was detected at any concentration tested. The minimum bactericidal concentration (MBC) point was not observed. The adhesion of S. mitis, S. oralis and mixed species was reduced by all tested dAAs. No adverse effects were observed on S. gordonii with any of the tested dAAs. The biofilm biomass of test strains under flow conditions was significantly reduced after a 5-min exposure to all tested dAAs at 25-mM concentration. CONCLUSIONS: D-amino acids did not inhibit bacterial growth and did not show bactericidal or bacteriostatic effects on test strains at any concentration tested (ranging from 6.25 mM to 100 mM). However, dAAs effectively inhibit the adhesion of early colonizers, thereby preventing the formation of oral biofilm.

Amino acids inhibit the photodynamic inactivation effect by hindering cellular oxidative stress.

Photodynamic inactivation (PDI) could utilize light to activate reactive oxygen species (ROS) produced by photosensitizers to kill bacteria for preservation. To delve into the complex effects arising during the post-harvest PDI processing, we conducted experiments using Pseudomonas reinekei, a food spoilage bacteria extracted from rotten Pakchoi. Through analyzing the metabolomics results, we discovered that methionine (Met) and glutamate (Glu) exhibited significant inhibitory effects during the PDI process. The oxidative stress generated by light treatment resulted in a reduction of 30.31% and 36.37% in the levels of Met and Glu, respectively. The data also showed that exogenous Met and Glu reduced intracellular oxidative stress levels, increased peroxidase activity, and prevented the damage of intracellular material and cell membrane deformation. That amino acids could inhibit the effect of PDI by hindering oxidative stress. Therefore, the amino acid content should be considered when applying PDI to treat Met- or Glu-rich foods.

Glycyrrhizic acid conjugates with amino acid methyl esters target the main protease, exhibiting antiviral activity against wild-type and nirmatrelvir-resistant SARS-CoV-2 variants.

COVID-19 pandemic is predominantly caused by SARS-CoV-2, with its main protease, Mpro, playing a pivotal role in viral replication and serving as a potential target for inhibiting different variants. In this study, potent Mpro inhibitors were identified from glycyrrhizic acid (GL) derivatives with amino acid methyl/ethyl esters. Out of the 17 derivatives semisynthesized, Compounds 2, 6, 9, and 15, with methionine methyl esters, D-tyrosine methyl esters, glutamic acid methyl esters, and methionines in the carbohydrate moiety, respectively, significantly inhibited wild-type SARS-CoV-2 Mpro-mediated proteolysis, with IC50 values ranging from 0.06 µM to 0.84 µM. They also demonstrated efficacy in inhibiting trans-cleavage by mutant Mpro variants (Mpro_P132H, Mpro_E166V, Mpro_P168A, Mpro_Q189I), with IC50 values ranging from 0.05 to 0.92 µM, surpassing nirmatrelvir (IC50: 1.17-152.9 µM). Molecular modeling revealed stronger interactions with Valine166 in the structural complex of Mpro_E166V with the compounds compared to nirmatrelvir. Moreover, these compounds efficiently inhibited the post-entry viral processes of wild-type SARS-CoV-2 single-round infectious particles (SRIPs), mitigating viral cytopathic effects and reducing replicon-driven GFP reporter signals, as well as in vitro infectivity of wild-type, Mpro_E166V, and Mpro_Q189I SRIPs, with EC50 values ranging from 0.02 to 0.53 µM. However, nirmatrelvir showed a significant decrease in inhibiting the replication of mutant SARS-CoV-2 SRIPs carrying Mpro_E166V (EC50: >20 µM) and Mpro_Q189I (EC50: 13.2 µM) compared to wild-type SRIPs (EC50: 0.06 µM). Overall, this study identifies four GL derivatives as promising lead compounds for developing treatments against various SARS-CoV-2 strains, including Omicron, and nirmatrelvir-resistant variants.

Comparing the biological activity and composition of Cerebrolysin with other peptide preparations.

Neurological disorders, ranging from acute forms such as stroke and traumatic brain injury to neurodegenerative diseases like dementia, are the leading cause of disability-adjusted life years (DALYs) worldwide. A promising approach to address these conditions and promote nervous system regeneration is the use of the neuropeptide preparation Cerebrolysin, which has been shown to be effective in both clinical and preclinical studies. Despite claims of similar clinical efficacy and safety by several peptide preparations, concerns regarding their generic composition and efficacy have been previously raised. Based on these reports, we analyzed the peptide composition and neurotrophic activity of several peptide preparations allegedly similar to Cerebrolysin and approved in some countries for treating neurological diseases. Our results demonstrate that these preparations lack relevant biological activity and that the peptide composition is significantly different from Cerebrolysin. peptide.

Excluding alanine from minimum essential medium (MEM) nonessential amino acid supplementation of the culture medium facilitates post-fertilization events and early cleavages of bovine oocytes fertilized in vitro.

Embryonic transfer of bovine blastocysts produced by in vitro fertilization is widely utilized-despite a compromised conception rate. It has been suggested that a set of four evaluation criteria for judging the quality of embryos, based on the timing of early cleavages and proper morphologies of embryos, can effectively predict pregnancy success. These blastocysts are hereafter referred to as four-criteria-compliant blastocysts. The same criteria should be used to modify the culture media to improve embryo quality. For example, culture media is often supplemented with nonessential amino acids (NEAA) at a uniform concentration despite the major variation in their concentration in the oviductal fluid. In the present study, the effects of the embryo culture medium, namely CR1, supplemented with all seven MEM NEAA or six of them, excluding one at a time, were examined. All media, except for the medium that did not contain proline and serine, tended to improve the efficiency of producing four-criteria-compliant blastocysts, and excluding alanine was particularly effective. The absence of alanine resulted in the rapid occurrence of the first cleavage and pronuclear formation of fertilized oocytes in the alanine-free medium compared to that in the medium containing alanine. These results suggested that alanine hinders certain events involved in the progression of early embryogenesis, which is necessary to achieve the four criteria that provide a benchmark for pregnancy. Therefore, a significantly higher percentage of embryos satisfied the recommended criteria and developed into four-criteria-compliant blastocysts when developed in alanine-free medium than in alanine-containing medium.

Screening of Amino Acids as a Safe Energy Source for Isolated Rat Pancreatic Acini.

OBJECTIVES: Amino acids play an essential role in protein synthesis, metabolism, and survival of pancreatic acini. Adequate nutritional support is important for acute pancreatitis treatment. However, high concentrations of arginine and lysine may induce acute pancreatitis. The study aimed to identify the most suitable l -amino acids as safe energy sources for pancreatic acinar cells. MATERIALS AND METHODS: Pancreatic acini were isolated from male Wistar rats. Effects of amino acids (0.1-20 mM) on uncoupled respiration of isolated acini were studied with a Clark electrode. Cell death was evaluated with fluorescent microscopy and DNA gel electrophoresis. RESULTS: Among the tested amino acids, glutamate, glutamine, alanine, lysine, and aspartate were able to stimulate the uncoupled respiration rate of isolated pancreatic acini, whereas arginine, histidine, and asparagine were not. Lysine, arginine, and glutamine (20 mM) caused complete loss of plasma membrane integrity of acinar cells after 24 hours of incubation. Glutamine also caused early (2-4 hours) cell swelling and blebbing. Aspartate, asparagine, and glutamate only moderately decreased the number of viable cells, whereas alanine and histidine were not toxic. DNA fragmentation assay and microscopic analysis of nuclei showed no evidence of apoptosis in cells treated with amino acids. CONCLUSIONS: Alanine and glutamate are safe and effective energy sources for mitochondria of pancreatic acinar cells.