Discovery of low-molecular-weight phenylalanine derivatives as novel HIV capsid modulators with improved antiretroviral activity and metabolic stability.

The HIV capsid (CA) protein is a promising target for anti-AIDS treatment due to its critical involvement in viral replication. Herein, we utilized the well-documented CA inhibitor PF74 as our lead compound and designed a series of low-molecular-weight phenylalanine derivatives. Among them, compound 7t exhibited remarkable antiviral activity with a high selection index (EC50 = 0.040 µM, SI = 2815), surpassing that of PF74 (EC50 = 0.50 µM, SI = 258). Furthermore, when evaluated against the HIV-2 strain, 7t (EC50 = 0.13 µM) demonstrated approximately 14-fold higher potency than that of PF74 (EC50 = 1.76 µM). Insights obtained from surface plasmon resonance (SPR) revealed that 7t exhibited stronger target affinity to the CA hexamer and monomer in comparison to PF74. The potential interactions between 7t and the HIV-1 CA were further elucidated using molecular docking and molecular dynamics simulations, providing a plausible explanation for the enhanced target affinity with 7t over PF74. Moreover, the metabolic stability assay demonstrated that 7t (T1/2 = 77.0 min) significantly outperforms PF74 (T1/2 = 0.7 min) in human liver microsome, exhibiting an improvement factor of 110-fold. In conclusion, 7t emerges as a promising drug candidate warranting further investigation.

Evaluation of Pharmacokinetic and Pharmacodynamic (PK/PD) of Novel Fluorenylmethoxycarbonyl- Phenylalanine Antimicrobial Agent.

OBJECTIVE: To assess the pharmacokinetic profile, in-vivo toxicity, and efficacy of 9-Fluorenylmethoxycarbonyl-L-phenylalanine (Fmoc-F) as a potential antibacterial agent, with a focus on its suitability for clinical translation. METHODS: An RP-HPLC-based bio-analytical method was developed and qualified to quantify Fmoc-F levels in mouse plasma for pharmacokinetic analysis. Oral bioavailability was determined, and in-vivo toxicity was evaluated following intra-peritoneal administration. Efficacy was assessed by measuring the reduction in Staphylococcus aureus burden and survival rates in BALB/c mice. RESULTS: The RP-HPLC method is highly sensitive, detecting as low as 0.8 µg mL-1 (~ 2 µM) of Fmoc-F in blood plasma. This study revealed that Fmoc-F has an oral bioavailability of 65 ± 18% and suitable pharmacokinetic profile. Further, we showed that intra-peritoneal administration of Fmoc-F is well tolerated by BALB/c mice and Fmoc-F treatment (100 mg/kg, i.p.) significantly reduces Staphylococcus aureus burden from visceral organs in BALB/c mice but falls short in enhancing survival rates at higher bacterial loads. CONCLUSIONS: The study provides crucial insights into the pharmacokinetic and pharmacodynamic properties of Fmoc-F. The compound displayed favourable oral bioavailability and in-vivo tolerance. Its significant reduction of bacterial burden underscores its potential as a treatment for systemic infections. However, limited effectiveness for severe infections, short half-life, and inflammatory response at higher doses need to be addressed for its clinical application.

Design, Synthesis, and Target Identification of Novel Phenylalanine Derivatives by Drug Affinity Responsive Target Stability (DARTS) in Xanthomonas oryzae pv Oryzae.

The increasing resistance displayed by plant phytopathogenic bacteria to conventional pesticides has heightened the urgency for the exploration of novel antibacterial agents possessing distinct modes of action (MOAs). In this study, a series of novel phenylalanine derivatives with the unique structure of acylhydrazone dithioether have been designed and synthesized. Bioassay results demonstrated that most target compounds exhibited excellent in vitro antibacterial activity against Xanthomonas oryzae pv oryzae (Xoo) and Xanthomonas axonopodis pv citri (Xac). Among them, the EC50 values of L3, L4, L6, L21, and L22 against Xoo were 7.4, 9.3, 6.7, 8.9, and 5.1 µg/mL, respectively, superior to that of bismerthiazol (BT) and thiodiazole copper (TC) (41.5 and >100 µg/mL); the EC50 values of L3, L4, L5, L6, L7, L8, L20, L21, and L22 against Xac were 5.6, 2.5, 6.2, 4.1, 4.2, 6.4, 6.3, 3.6, and 5.2 µg/mL, respectively, superior to that of BT and TC (43.3 and >100 µg/mL). An unmodified drug affinity responsive target stability (DARTS) technology was used to investigate the antibacterial MOAs of active compound L22, and the 50S ribosomal protein L2 (RL2) as an unprecedented target protein in Xoo cells was first discovered. The target protein RL2 was then expressed and purified. Furthermore, the in vitro interactions by microscale thermophoresis (Kd = 0.050 µM) and fluorescence titration (Ka = 1.4 × 105 M-1) experiments also demonstrated a strong binding force between compound L22 and RL2. Overall, these results not only facilitate the development of novel antibacterial agents but also establish a reliable method for exploring the targets of bactericides.

Phenylalanine requirements using the direct amino acid oxidation technique, and the effects of dietary phenylalanine on food intake, gastric emptying, and macronutrient metabolism in adult cats.

Despite Phe being an indispensable amino acid for cats, the minimum Phe requirement for adult cats has not been empirically defined. The objective of study 1 was to determine the minimum Phe requirement, where Tyr is in excess, in adult cats using the direct amino acid oxidation (DAAO) technique. Four adult male cats were used in an 8 × 4 Latin rectangle design. Cats were adapted to a basal diet for 7 d, top dressed with Phe to meet 140% of the adequate intake (NRC, 2006. Nutrient requirements of dogs and cats. Washington, DC: Natl. Acad. Press). Cats were randomly assigned to one of eight experimental Phe diets (0.29%, 0.34%, 0.39%, 0.44%, 0.54%, 0.64%, 0.74%, and 0.84% Phe in the diet on a dry matter [DM] basis). Following 1 d of diet adaptation, individual DAAO studies were performed. During each DAAO study, cats were placed into individual indirect calorimetry chambers, and 75% of the cat's daily meal was divided into 13 equal meals supplied with a dose of L-[1-13C]-Phe. Oxidation of L-[1-13C]-Phe (F13CO2) during isotopic steady state was determined from the enrichment of 13CO2 in breath. Competing models were applied using the NLMIXED procedure in SAS to determine the effects of dietary Phe on 13CO2. The mean population minimum requirement for Phe was estimated at 0.32% DM and the upper 95% population confidence limit at 0.59% DM on an energy density of 4,200 kcal of metabolizable energy/kg DM calculated using the modified Atwater factors. In study 2, the effects of a bolus dose of Phe (44 mg kg-1 BW) on food intake, gastric emptying (GE), and macronutrient metabolism were assessed in a crossover design with 12 male cats. For food intake, cats were given Phe 15 min before 120% of their daily food was offered and food intake was measured. Treatment, day, and their interaction were evaluated using PROC GLIMMIX in SAS. Treatment did not affect any food intake parameters (P > 0.05). For GE and macronutrient metabolism, cats were placed into individual indirect calorimetry chambers, received the same bolus dose of Phe, and 15 min later received 13C-octanoic acid (5 mg kg-1 BW) on 50% of their daily food intake. Breath samples were collected to measure 13CO2. The effect of treatment was evaluated using PROC GLIMMIX in SAS. Treatment did not affect total GE (P > 0.05), but cats receiving Phe tended to delay time to peak enrichment (0.05 < P ≤ 0.10). Overall, Phe at a bolus dose of 44 mg kg-1 BW had no effect on food intake, GE, or macronutrient metabolism. Together, these results suggest that the bolus dose of Phe used may not be sufficient to elicit a GE response, but a study with a greater number of cats and greater food intake is warranted.

Two studies were conducted to evaluate 1) the minimum requirement for dietary Phe and 2) the effects of Phe on gastric emptying (GE) and food intake in adult cats. In study 1, the minimum Phe requirement was estimated using the direct amino acid oxidation (DAAO) technique. Four cats were used and received all diets in random order in a Latin rectangle design (0.29%, 0.34%, 0.39%, 0.44%, 0.54%, 0.64%, 0.74%, and 0.84% Phe in the diet on a dry matter [DM] basis). The minimum Phe requirement, in the presence of excess of Tyr, for adult cats was estimated to be 0.59% DM on an energy density of 4,200 kcal of metabolizable energy/kg DM calculated using the modified Atwater factors; higher than current recommendations set in place by the National Research Council and the American Association of Feed Control Officials. In study 2, we first validated the use of the 13C-octanoic acid breath test (13C-OABT) in cats. Then, the effects of an oral bolus of Phe on food intake, GE, and macronutrient metabolism were evaluated. Phe supplementation did not influence food intake, macronutrient metabolism, or total GE, but tended to delay the time to peak GE.

Preparation of healing-promoting and fibrosis-inhibiting asymmetric poly(ethylene glycol-b-L-phenylalanine)/cRGD-modified hyaluronate sponges and their applications in hemorrhage and nasal mucosa repair.

Acute or chromic bleeding, such as epistaxis, requires hemostatic materials to assist hemostasis. Even in complex cases, hemostatic materials must have other functions, including the promotion of healing and prevention of adhesion. Herein, a series of fibrosis-suppressive functional cRGD-modified crosslinking hyaluronic acid sponges were prepared. The in vitro hemostatic efficiency and mechanism were determined using blood clotting time, blood coagulation index, lactate dehydrogenase (LDH) and thromboxane B2 (TX-B2) ELISA, and proteomics. Among the prepared sponges, both poly(ethylene-b-L-Phe) (PEBP)-and cRGD contained SPN4 and exhibited the highest platelet concentration and activation efficiency as well as the most effective coagulative effect. In addition, no significant cytotoxicity was observed for the sponges in rat airway epithelial cells. The in vivo hemostatic and adhesion-preventive effects of the sponges were evaluated using rat models of liver injury and sidewall defect-cecum abrasion. PEBP-containing sponges effectively prevented postoperative adhesion and cRGD-modified sponges exhibited excellent hemostatic effects. Finally, the comprehensive repair effects of the sponges were evaluated using a rabbit maxillary sinus mucosal injury model, based on CT, MRI examination, and pathological staining. SPN4 exhibited the best comprehensive reparative effects, including the promotion of mucosal repair and infection inhibition. Thus, SPN4 is a promising multifunctional hemostatic material.

Phenylalanine-Arginine-ß-Naphthylamide Enhances the Photobactericidal Effect of Methylene Blue on Pseudomonas aeruginosa.

Objective: To investigate the effectiveness, dosing sequence, concentration, and mechanism of antimicrobial photodynamic inactivation (aPDI) using methylene blue (MB) plus phenylalanine-arginine-ß-naphthylamide (PAßN) against Pseudomonas aeruginosa. Methods: P. aeruginosa bacterial suspension was incubated with MB for different times (5-240 min), and then, 10 J/cm2 red light was irradiated. The efflux pump inhibitor (EPI) PAßN (10-100 µg/mL) was combined with MB (1-20 µM) in different sequences (PAßN-first, PAßN+MB, PAßN-after). Colony-forming units were then determined by serial dilution. Results: Using MB 10 µM plus 10 J/cm2, the killing effect of MB-aPDI on P. aeruginosa increased first and then decreased with longer incubation time. The killing effect of MB+PAßN-aPDI on P. aeruginosa was better than that of MB-aPDI (p < 0.05) by up to 2 logs. PAßN-first had the best killing effect, whereas PAßN-after had the worst killing effect. The killing effect increased with PAßN concentration and at 100 µg/mL reached 5.1 logs. Conclusions: The EPI PAßN enhanced the bactericidal effect of MB-aPDI on P. aeruginosa, especially when added before MB. It is proposed that MB is a substrate of the resistance-nodulation-division family efflux pump.

Peptidomimetics based on ammonium decasubstituted pillar[5]arenes: Influence of the alpha-amino acid residue nature on cholinesterase inhibition.

Cholinesterase inhibitors are a group of medicines that are widely used for the treatment of cognitive impairments accompanying Alzheimer's disease as well as for the treatment of pathological muscle weaknesses syndromes such as myasthenia gravis. The search for novel non-toxic and effective cholinesterase inhibitors for creating neuroprotective and neurotransmitter agents is an urgent interdisciplinary problem. For the first time, the application of water-soluble pillar[5]arenes containing amino acid residues as effective cholinesterase inhibitors was shown. The influence of the nature of aliphatic and aromatic alpha-amino acid residues (glycine, l-alanine, l-phenylalanine and l-tryptophan) on self-assembly, aggregate's stability, cytotoxicity on A549 and LEK cells and cholinesterase inhibition was studied. It was found that the studied compounds with aliphatic amino acid residues showed a low inhibitory ability against cholinesterases. It was established that the pillar[5]arene containing fragments of l-phenylalanine is the most promising inhibitor of butyrylcholinesterase (IC50 = 0.32 ± 0.01 µM), the pillar[5]arene with l-tryptophan residues is the most promising inhibitor of acetylcholinesterase (IC50 = 0.32 ± 0.01 µM). This study has shown a possible application of peptidomimetics based on pillar[5]arenes to inhibit cholinesterase, as well as control the binding affinity to a particular enzyme in a structure-dependent manner.

Equilibria in the aqueous system of cobalt(II) based on 2-picolinehydroxamic acid and N-(2-hydroxybenzyl)phenylalanine and its ability to inhibit the propagation of cancer cells.

Mixed-ligand complexes of cobalt(II) with two bioligands, viz. 2-picolinehydroxamic acid and the reduced Schiff base N-(2-hydroxybenzyl)phenylalanine, were studied in aqueous solution by potentiometry and UV-Vis spectroscopic analysis. The coordination mode of the complexes and their stability were determined and compared to their parent species. Stacking interactions between the rings present in the ligands influence the stability of the complexes. Also, UV-Vis spectroscopy revealed that the stacking interactions affected the intercalation of DNA and mixed-ligand complexes. The in vitro anticancer activity of the free ligand 2-picolinehydroxamic acid and the complexes was tested against cervical and gastric human adenocarcinoma epithelial cell lines. At concentrations of 0.06 and 0.11 mM, the mixed-ligand structures showed the ability to reduce gastric cancer cells with no inhibitory effect on mouse fibroblasts. The cytotoxic effect was accompanied by damage to the cell nuclei, which may confirm that the complexes demonstrate effective binding to DNA. No determination of minimal inhibitory and bactericidal/fungicidal concentrations against the test organisms was possible at higher complex concentrations due to precipitation.

Zn(II)-Coordination Complex(s) from Chiral Bisamides of L-Phenylalanine: Nanoscale-Based Biological Applications and Metallogelation.

Three 3-pyridyl-containing small organic bisamide molecules attached with innocent L-phenylalanine (PHE) side chain as building blocks and positional isomeric toluoyl terminals (PME, MME, and OME) have rationally been designed and synthesized for developing a new series of ZnII-coordination complexes. One of the unique molecular frameworks, having two hydrogen bond-equipped monodentate metal-coordinating sites and biologically potent chiral PHE moiety, was combined with ZnII halide salts under various conditions to produce the coordination complexes (CC1-CC7), thoroughly characterized by the single-crystal X-ray diffraction (SXRD) technique. Maintaining the similar component ratios of acquired CCs in 1:1 DMSO-water produced low-molecular weight metallogels (LMWGs) of PME/MME as envisaged from a rheology- and crystal engineering-based structural rationale. A structure-property correlation from the basis of PXRD of the bulk and xerogels and SXRD data of the isolated single crystals of reaction products clearly supports the crystal engineering-based design strategy based on which the metallogels are prepared. Hand-ground nanoscale ZnCl2-based coordination complex CC1 of PME was also studied for cytotoxicity (HEK-293 cell line) and anticancer activities (B16-F10 cell line) in the MTT assay.

Metabolic engineering strategies for naringenin production enhancement in Streptomyces albidoflavus J1074.

BACKGROUND: Naringenin is an industrially relevant compound due to its multiple pharmaceutical properties as well as its central role in flavonoid biosynthesis. RESULTS: On our way to develop Streptomyces albidoflavus J1074 as a microbial cell factory for naringenin production, we have significantly increased the yields of this flavanone by combining various metabolic engineering strategies, fermentation strategies and genome editing approaches in a stepwise manner. Specifically, we have screened different cultivation media to identify the optimal production conditions and have investigated how the additive feeding of naringenin precursors influences the production. Furthermore, we have employed genome editing strategies to remove biosynthetic gene clusters (BGCs) associated with pathways that might compete with naringenin biosynthesis for malonyl-CoA precursors. Moreover, we have expressed MatBC, coding for a malonate transporter and an enzyme responsible for the conversion of malonate into malonyl-CoA, respectively, and have duplicated the naringenin BGC, further contributing to the production improvement. By combining all of these strategies, we were able to achieve a remarkable 375-fold increase (from 0.06 mg/L to 22.47 mg/L) in naringenin titers. CONCLUSION: This work demonstrates the influence that fermentation conditions have over the final yield of a bioactive compound of interest and highlights various bottlenecks that affect production. Once such bottlenecks are identified, different strategies can be applied to overcome them, although the efficiencies of such strategies may vary and are difficult to predict.

Helical-Like Assembly of Nateglinide as Coating for Oral Delivery of Insulin and Their Synergistic Prevention of Diabetes Mellitus.

Oral delivery of antidiabetic active components promises to free millions of people from daily suffering who require routine injections. However, oral insulin (Ins) and other short-acting compounds such as nateglinide (NG) in harsh gastrointestinal tract still face great challenging, including low bioavailability, and rapid elimination. In this study, inspired by the self-assembly of phenylalanine-based peptides in nature, it is showed that NG a small phenylalanine derivative, assembles into left-handed helical nanofibers in the presence of Ca2+ . These helical NG nanofibers functioned as a coating layer on the surface of Ca2+ -linked alginate (Alg) microgels for the effective encapsulation of Ins. As expected, the sustained release and prolonged circulation of Ins and NG from the Ins-loading Alg/NG microgels (Ins@Alg/NG) in the intestinal tract synergistically maintain a relatively normal blood glucose level in streptozotocin-induced diabetic mice after oral administration of Ins@Alg/NG. This further confirms that Ins@Alg/NG ameliorated Ins resistance mainly through activating Insreceptor substrate 1 (IRS1), protein kinase B (AKT), and AMP-activated protein kinase (AMPK), as well as by repressing glycogen synthase kinase-3ß (GSK-3ß). The strategy of using the assembly of NG as a coating achieves the oral delivery of insulin and showcases a potential for the treatment of diabetes.

Antibacterial Effect of Melanoidins Derived From Xylose and Phenylalanine Against Bacillus cereus and Clostridium perfringens.

Melanoidins produced from the combination of D-xylose and L-phenylalanine have been reported to exhibit strong antibacterial effects. This study investigated the influence of environmental factors, such as temperatures (10, 15, 20, 25, 30, 35, 40, and 45°C), pH (5.5, 6.0, 6.5, 7.0, 7.5, and 8.0), and water activity (aw: 0.99, 0.96, and 0.93), on the antibacterial effect of the melanoidins produced from the combination of D-xylose with L-phenylalanine against Bacillus cereus and Clostridium perfringens in culture media. Furthermore, freeze-dried powdered melanoidin was used to determine the minimum concentration for growth inhibition, to compare the antibacterial effect of the melanoidin with conventional food preservatives. The liquid melanoidins significantly inhibited the growth of B. cereus (up to 4 log CFU/mL at the maximum) and C. perfringens (up to 6.5 log CFU/mL at the maximum) regardless of the incubation temperatures. However, the remarkable difference between the presence and absence of the melanoidins was demonstrated in the range of 20-35°C as 4 log-cycle lower in B. cereus and 2 log-cycle lower in C. perfringens than those without the melanoidins. The antibacterial effect of the melanoidin on B. cereus was not influenced by pH from 5.5 to 7.0, which exhibited 2-3 log-cycle lower viable counts than those without the melanoidin. Only one log-cycle difference between with and without the melanoidin was shown in C. perfringens growth under the pH range of 5.5-7.0. Although there was no significant difference in the growth of B. cereus between three aw conditions, the melanoidin exhibited a significant antibacterial effect at aw 0.99 demonstrating 4 log-cycle lower viable numbers than those without the melanoidin. Minimum inhibitory concentration of the melanoidin powder for B. cereus and C. perfringens was 7 mg/mL and 15 mg/mL, respectively, regardless of the kind of foods. Furthermore, the melanoidin exhibited comparable antibacterial effect on B. cereus and C. perfringens to potassium sorbate and sodium benzoate under the same concentration as the minimum inhibitory concentration of the melanoidin, demonstrating 2 log-cycle reduction during 3 days of incubation period at 25°C. The results presented here suggest that the xylose- and phenylalanine-based melanoidin demonstrates the possibility to be an alternative food preservative.

Anti-amyloidogenic amphipathic arginine-dehydrophenylalanine spheres capped selenium nanoparticles as potent therapeutic moieties for Alzheimer's disease.

Aggregation of both amyloid beta (Aß) peptide and hyperphosphorylated tau proteins is the major pathological hallmark of Alzheimer's disease (AD). Moieties that carry anti-amyloidogenic potency against both of the aggregating entities are considered to be promising drug candidatures for the disease. In the current work, we have synthesized amphipathic dipeptide vesicle-templated selenium nanoparticles (RΔF-SeNPs) as potential entities to combat AD. We have investigated and established their anti-amyloidogenic activity against different peptide-based amyloid models, such as the reductionist model based on the dipeptide phenylalanine-phenylalanine (FF) derived from Aß; a model based on the hexapeptide Ac-PHF6 (306VQIVYK311) derived from tau protein; and the full-length Aß42 polypeptide-based model. We also evaluated the neuroprotective characteristics of RΔF-SeNPs against FF, Ac-PHF6, and Aß42 fibril-induced toxicity in neuroblastoma, SH-SY5Y cells. RΔF-SeNPs further exhibited neuroprotective effects in streptozotocin (STZ) treated neuronal (N2a) cells carrying AD-like features. In addition, studies conducted in an intra-cerebroventricular STZ-instigated rat model of dementia revealed that RΔF-SeNP-treated animals showed improved cognitive activity and reduced Aß42 aggregate burden in brain tissues as compared with the STZ-treated group. Moreover, in vivo brain distribution studies conducted in animal models additionally demonstrated the brain-homing ability of RΔF-SeNPs. All together, these studies supported the potency of RΔF-SeNPs as efficient and propitious disease-modifying therapeutic agents for combating AD.

Synthesis, characterization, HSA binding, molecular docking, cytotoxicity study, and antimicrobial activity of new palladium(II) complexes with propylenediamine derivatives of phenylalanine.

The four new ligands, propylenediamine derivatives of phenylalanine (R2-S,S-pddbaˑ2HCl; L1-L4) and their palladium(II) complexes (C1-C4) were synthesized and characterized by elemental analysis, infrared, 1H and 13C NMR spectroscopy. The interactions of new palladium(II) complexes with human serum albumin (HSA) were studied by fluorescence spectroscopy. All investigated compounds can be transported to target cells by binding to HSA, but complex C4 interacts most strongly. Molecular docking simulations were applied to comprehend the binding of the complex to the molecular target of HSA. Obtained results are in good correlations with experimental data regarding binding affinity by HSA. In vitro cytotoxicity activities were investigated on four tumor cell lines (mouse mammary (4 T1) and colon (CT26), human mammary (MDA-MD-468) and colon (HCT116)) and mouse mesenchymal stem cells as non-tumor control cells. Cytotoxic capacity was determined by MTT test and according to obtained results ligand L4 stands out as the most active and selective compound and as a good candidate for future in vivo testing. Further examination of the ligand L4 and corresponding complex C4 led to the conclusion that both induced cell death mainly by apoptosis. Ligand L4 facilitated cycle arrest in G0/G1 phase and decreased proliferative capacity of tumor cells. In vitro antimicrobial activity for ligands and corresponding Pd(II) complexes was investigated against eleven microorganisms (eight strains of pathogenic bacteria and three yeast species) using microdilution method. The minimum inhibitory concentration and minimum microbicidal concentration were determined.

Boosting the Antibacterial Activity of Azithromycin on Multidrug-Resistant Escherichia coli by Efflux Pump Inhibition Coupled with Outer Membrane Permeabilization Induced by Phenylalanine-Arginine ß-Naphthylamide.

The global spread of multidrug-resistant (MDR) bacteria increases the demand for the discovery of new antibiotics and adjuvants. Phenylalanine-arginine ß-naphthylamide (PAßN) is an inhibitor of efflux pumps in Gram-negative bacteria, such as the AcrAB-TolC complex in Escherichia coli. We aimed to explore the synergistic effect and mechanism of action of PAßN combined with azithromycin (AZT) on a group of MDR E. coli strains. Antibiotic susceptibility was tested for 56 strains, which were screened for macrolide resistance genes. Then, 29 strains were tested for synergy using the checkerboard assay. PAßN significantly enhanced AZT activity in a dose-dependent manner in strains expressing the mphA gene and encoding macrolide phosphotransferase, but not in strains carrying the ermB gene and encoding macrolide methylase. Early bacterial killing (6 h) was observed in a colistin-resistant strain with the mcr-1 gene, leading to lipid remodeling, which caused outer membrane (OM) permeability defects. Clear OM damage was revealed by transmission electron microscopy in bacteria exposed to high doses of PAßN. Increased OM permeability was also proven by fluorometric assays, confirming the action of PAßN on OM. PAßN maintained its activity as an efflux pump inhibitor at low doses without permeabilizing OM. A non-significant increase in acrA, acrB, and tolC expression in response to prolonged exposure to PAßN was noted in cells treated with PAßN alone or with AZT, as a reflection of bacterial attempts to counteract pump inhibition. Thus, PAßN was found to be effective in potentiating the antibacterial activity of AZT on E. coli through dose-dependent action. This warrants further investigations of its effect combined with other antibiotics on multiple Gram-negative bacterial species. Synergetic combinations will help in the battle against MDR pathogens, adding new tools to the arsenal of existing medications.

The effect of silicon groups on the physicochemical property and bioactivity of L-phenylalanine derived poly(amide-imide).

Poly(amide-imide) (PAI), serving as a synthetic polymer, has been widely used in industry for excellent mechanical properties, chemical resistance and high thermal stability. However, lack of suitable cell niche and biological activity limited the further application of PAI in biomedical engineering. Herein, silicon modified L-phenylalanine derived poly(amide-imide) (PAIS) was synthesized by introducing silica to L-phenylalanine derived PAI to improve physicochemical and biological performances. The influence of silicon amount on physicochemical, immune, and angiogenic performances of PAIS were systemically studied. The results show that PAIS exerts excellent hydrophilic, mechanical, biological activity. PAIS shows no effects on the number of macrophages, but can regulate macrophage polarization and angiogenesis in a dose-dependent manner. This study advanced our understanding of silicon modification in PAI can modulate cell responses via initiating silicon concentration regulation. The acquired knowledge will provide a new strategy to design and optimize biomedical PAI in the future.

The effects of exogenous amino acids on production of microcystin variants in Microcystis aeruginosa.

Dissolved free amino acids are a significant component of dissolved organic nitrogen (DON) in natural waterbodies. The effects of four amino acids (glutamic acid, phenylalanine, leucine, and arginine) on the growth and microcystins (MCs) production of Microcystis aeruginosa were studied in batch culture. The profiles of five MCs variants and the expression levels of target genes involved in MCs biosynthesis and nitrogen metabolism were measured. When amino acids were used as the sole nitrogen source instead of nitrate at different levels (0.5, 2.0 and 8.0 mg/L based on N) in BG-11 medium, algal cell growth and intracellular MCs quotas were inhibited slightly by the treatments with glutamic acid and arginine. The treatments with phenylalanine and leucine, on the other hand, had a strong inhibitory effect on algal cell growth and MCs production. Moreover, the concentrations of Chlorophyll a, phycocyanin and allophycocyanin in cells cultured in glutamic acid, leucine and phenylalanine were lower than those in the control group with nitrate as nitrogen source. The existence of leucine or phenylalanine can lead to a significant increase in the relative abundance of MCs variants structured with the corresponding amino acids. The expression of microcystin-producing gene mcyD was downregulated while the gene pipX associated with nitrogen metabolism was upregulated during the cultivation of M. aeruginosa with amino acids as sole nitrogen source. M. aeruginosa undergoes significant alterations due to exogenous amino acids and exhibits advanced strategies for MCs production.

A rapid intracellular enrichment of alkylating payload is essential for melphalan flufenamide potency and mechanism of action.

Despite decades of development of treatments and the successful application of targeted therapies for multiple myeloma, clinical challenges remain for patients with relapsed/refractory disease. A drug designed for efficient delivery of an alkylating payload into tumor cells that yields a favorable therapeutic window can be an attractive choice. Herein we describe melphalan flufenamide (melflufen), a drug with a peptide carrier component conjugated to an alkylating payload, and its cellular metabolism. We further underline the fundamental role of enzymatic hydrolysis in the rapid and robust accumulation of alkylating metabolites in cancer cells and their importance for downstream effects. The formed alkylating metabolites were shown to cause DNA damage, both on purified DNA and on chromatin in cells, with both nuclear and mitochondrial DNA affected in the latter. Furthermore, the rapid intracellular enrichment of alkylating metabolites is shown to be essential for the rapid kinetics of the downstream intracellular effects such as DNA damage signaling and induction of apoptosis. To evaluate the importance of enzymatic hydrolysis for melflufen's efficacy, all four stereoisomers of the compound were studied in a systematic approach and shown to have a different pattern of metabolism. In comparison with melflufen, stereoisomers lacking intracellular accumulation of alkylating payloads showed cytotoxic activity only at significantly higher concentration, slower DNA damage kinetics, and different mechanisms of action to reach cellular apoptosis.

Enhanced inhibition of HEDP on SRB-mediated corrosion with D-phenylalanine.

Microbiologically influenced corrosion (MIC) caused by biofilm is a serious problem in many industries. D-amino acids could be a potential strategy to enhance traditional corrosion inhibitors due to their roles in biofilm reduction. However, the synergistic mechanism of D-amino acids and inhibitors remains unknown. In this study, D-Phenylalanine (D-Phe) and 1-hydroxyethane-1,1-diphosphonic acid (HEDP) were selected as the typical D-amino acid and corrosion inhibitor to evaluate their effect on the corrosion caused by Desulfovibrio vulgaris. The combination of HEDP and D-Phe obviously slowed down the corrosion process by 32.25%, decreased the corrosion pit depth and retarded cathodic reaction. SEM and CLSM analysis indicated that D-Phe reduced the content of extracellular protein and thus inhibited the biofilm formation. The molecular mechanism of D-Phe and HEDP on corrosion inhibition was further explored via transcriptome. The combination of HEDP and D-Phe down-regulated the gene expression of peptidoglycan, flagellum, electron transfer, ferredoxin and quorum sensing (QS) molecules, leading to less peptidoglycan synthesis, weaker electron transfer and stronger QS factor inhibition. This work provides a new strategy for improving traditional corrosion inhibitors, retarding MIC and mitigating subsequent water eutrophication.

A novel L-phenylalanine dipeptide inhibits prostate cancer cell proliferation by targeting TNFSF9.

In the present study, a series of novel L-phenylalanine dipeptides were designed and synthesized by a multi-step sequence of reactions, including carbodiimide-mediated condensation, hydrolysis, mixed anhydride condensation, and nucleophilic substitution. Among them, compound 7c exhibited potent antitumor activity against prostate cancer cell PC3 in vitro and in vivo via inducing apoptosis. We investigated the significantly differentially expressed proteins in the cells caused by the compound 7c to unravel the molecular mechanisms underlying the regulation of PCa cell growth, which indicated that 7c mainly regulated the protein expression of apoptosis-related transcription factors, including c-Jun, IL6, LAMB3, OSMR, STC1, OLR1, SDC4 and PLAU; and 7c also regulated the protein expression of inflammatory cytokines including IL6, CXCL8, TNFSF9, TNFRSF12A and OSMR, and the phosphorylation levels of RelA. The action target confirmed that TNFSF9 protein is the critical binding target of 7c. These findings suggested that 7c could regulate the apoptosis and inflammatory response related signaling pathways for the inhibition of the proliferation of PC3 cells, implying that 7c could be considered a promising therapeutic candidate for PCa therapy.