In vitro activity of cefiderocol against European Pseudomonas aeruginosa and Acinetobacter spp., including isolates resistant to meropenem and recent ß-lactam/ß-lactamase inhibitor combinations.

Carbapenem-resistant Pseudomonas aeruginosa and Acinetobacter spp. represent major threats and have few approved therapeutic options. Non-|fermenting Gram-negative isolates were collected from hospitalized inpatients from 49 sites in 6 European countries between 01 January 2020 and 31 December 2020 and underwent susceptibility testing against cefiderocol and ß-lactam/ß-lactamase inhibitor combinations. Meropenem-resistant (MIC >8 mg/L), cefiderocol-susceptible isolates were analyzed by PCR, and cefiderocol-resistant isolates were analyzed by whole-genome sequencing to identify resistance mechanisms. Overall, 1,451 (950 P. aeruginosa; 501 Acinetobacter spp.) isolates were collected, commonly from the respiratory tract (42.0% and 39.3%, respectively). Cefiderocol susceptibility was higher than |ß|-|l|a|c|t|a|m|/|ß|-|l|a|c|t|a|mase| inhibitor combinations against P. aeruginosa (98.9% vs 83.3%-91.4%), and P. |aeruginosa resistant to meropenem (n = 139; 97.8% vs 12.2%-59.7%), ß-lactam/ß-lactamase inhibitor combinations (93.6%-98.1% vs 10.7%-71.8%), and both meropenem and ceftazidime-avibactam (96.7% vs 5.0%-||45.0%) or |ceftolozane-tazobactam (98.4% vs 8.1%-54.8%), respectively. Cefiderocol and sulbactam-durlobactam susceptibilities were high against Acinetobacter spp. (92.4% and 97.0%) and meropenem-resistant Acineto|bacter |spp. (n = 227; 85.0% and 93.8%) but lower against sulbactam-durlobactam- (n |= 15; 13.3%) and cefiderocol- (n = 38; 65.8%) resistant isolates, respectively. Among meropenem-resistant P. aeruginosa and Acinetobacter spp., the most common ß-||lactamase genes were metallo-ß-lactamases [30/139; blaVIM-2 (15/139)] and oxacillinases [215/227; blaOXA-23 (194/227)], respectively. Acquired ß-lactamase genes were identified in 1/10 and 32/38 of cefiderocol-resistant P. aeruginosa and Acinetobacter spp., and pirA-like or piuA mutations in 10/10 and 37/38, respectively. Conclusion: cefiderocol susceptibility was high against P. aeruginosa and Acinetobacter spp., including meropenem-resistant isolates and those resistant to recent ß-lactam/ß-lactamase inhibitor combinations common in first-line treatment of European non-fermenters. IMPORTANCE: This was the first study in which the in vitro activity of cefiderocol and non-licensed ß-lactam/ß-lactamase inhibitor combinations were directly compared against Pseudomonas aeruginosa and Acinetobacter spp., including meropenem- and ß-lactam/ß-lactamase inhibitor combination-resistant isolates. A notably large number of European isolates were collected. Meropenem resistance was defined according to the MIC breakpoint for high-dose meropenem, ensuring that data reflect antibiotic activity against isolates that would remain meropenem resistant in the clinic. Cefiderocol susceptibility was high against non-fermenters, and there was no apparent cross resistance between cefiderocol and ß-lactam/ß-lactamase inhibitor combinations, with the exception of sulbactam-durlobactam. These results provide insights into therapeutic options for infections due to resistant P. aeruginosa and Acinetobacter spp. and indicate how early susceptibility testing of cefiderocol in parallel with ß-lactam/ß-lactamase inhibitor combinations will allow clinicians to choose the effective treatment(s) from all available options. This is particularly important as current treatment options against non-fermenters are limited.

Role of microsomal metabolism in bromfenac-induced cytotoxicity.

This study delves into the intricate mechanisms underlying drug-induced liver injury (DILI) with a specific focus on bromfenac, the withdrawn nonsteroidal anti-inflammatory drug. DILI is a pervasive concern in drug development, prompting market withdrawals and posing significant challenges to healthcare. Despite the withdrawal of bromfenac due to DILI, the exact role of its microsomal metabolism in inducing hepatotoxicity remains unclear. Herein, employing HepG2 cells with human liver microsomes and UDP-glucuronic acid (UDPGA), our investigation revealed a substantial increase in bromfenac-induced cytotoxicity in the presence of UDPGA, pointing to the significance of UDP-glucuronosyltransferase (UGT)-dependent metabolism in augmenting toxicity. Notably, among the recombinant UGTs examined, UGT2B7 emerged as a pivotal enzyme in the metabolic activation of bromfenac. Metabolite identification studies disclosed the formation of reactive intermediates, with bromfenac indolinone (lactam) identified as a potential mediator of hepatotoxic effects. Moreover, in cytotoxicity experiments, the toxicity of bromfenac lactam exhibited a 34-fold increase, relative to bromfenac. The toxicity of bromfenac lactam was mitigated by nicotinamide adenine dinucleotide phosphate-dependent metabolism. This finding underscores the role of UGT-dependent metabolism in generating reactive metabolites that contribute to the observed hepatotoxicity associated with bromfenac. Understanding these metabolic pathways and the involvement of specific enzymes, such as UGT2B7, provides crucial insights into the mechanisms of bromfenac-induced liver injury. In conclusion, this research sheds light on the metabolic intricacies leading to cytotoxicity induced by bromfenac, especially emphasizing the role of UGT-dependent metabolism and the formation of reactive intermediates like bromfenac lactam. These findings offer insight into the mechanistic basis of DILI and emphasize the importance of understanding metabolism-mediated toxicity.

Structural Basis for Coronaviral Main Proteases Inhibition by the 3CLpro Inhibitor GC376.

The main protease (Mpro) of coronaviruses participates in viral replication, serving as a hot target for drug design. GC376 is able to effectively inhibit the activity of Mpro, which is due to nucleophilic addition of GC376 by binding covalently with Cys145 in Mpro active site. Here, we used fluorescence resonance energy transfer (FRET) assay to analyze the IC50 values of GC376 against Mpros from six different coronaviruses (SARS-CoV-2, HCoV-229E, HCoV-HUK1, MERS-CoV, SARS-CoV, HCoV-NL63) and five Mpro mutants (G15S, M49I, K90R, P132H, S46F) from SARS-CoV-2 variants. The results showed that GC376 displays effective inhibition to various coronaviral Mpros and SARS-CoV-2 Mpro mutants. In addition, the crystal structures of SARS-CoV-2 Mpro (wide type)-GC376, SARS-CoV Mpro-GC376, MERS-CoV Mpro-GC376, and SARS-CoV-2 Mpro mutants (G15S, M49I, S46F, K90R, and P132H)-GC376 complexes were solved. We found that GC376 is able to fit into the active site of Mpros from different coronaviruses and different SARS-CoV-2 variants properly. Detailed structural analysis revealed key molecular determinants necessary for inhibition and illustrated the binding patterns of GC376 to these different Mpros. In conclusion, we not only proved the inhibitory activity of GC376 against different Mpros including SARS-CoV-2 Mpro mutants, but also revealed the molecular mechanism of inhibition by GC376, which will provide scientific guidance for the development of broad-spectrum drugs against SARS-CoV-2 as well as other coronaviruses.

An RNA replicon system to investigate promising inhibitors of feline coronavirus.

Feline infectious peritonitis (FIP) is a fatal feline disease, caused by a feline coronavirus (FCoV), namely feline infectious peritonitis virus (FIPV). We produced a baby hamster kidney 21 (BHK) cell line expressing a serotype I FCoV replicon RNA with a green fluorescent protein (GFP) reporter gene (BHK-F-Rep) and used it as an in vitro screening system to test different antiviral compounds. Two inhibitors of the FCoV main protease (Mpro), namely GC376 and Nirmatrelvir, as well as the nucleoside analog Remdesivir proved to be effective in inhibiting the replicon system. Different combinations of these compounds also proved to be potent inhibitors, having an additive effect when combined. Remdesivir, GC376, and Nirmatrelvir all have a 50% cytotoxic concentration (CC50) more than 200 times higher than their half-maximal inhibitory concentrations (IC50), making them important candidates for future in vivo studies as well as clinically implemented drug candidates. In addition, results were acquired with a virus infection system, where Felis catus whole fetus 4 (Fcwf-4) cells were infected with a previously described recombinant GFP-expressing FIPV (based on the laboratory-adapted serotype I FIPV strain Black) and treated with the most promising compounds. Results acquired with the replicon system were comparable to the results acquired with the virus infection system, demonstrating that we successfully implemented the FCoV replicon system for antiviral screening. We expect that this system will greatly facilitate future screens for anti-FIPV compounds and provide a non-infectious system to study and evaluate drug-resistant mutations that may emerge in the FIPV genome.IMPORTANCEFIPV is of great significance in the cat population around the world, causing 0.3%-1.4% of feline deaths in veterinary practices (2). As there are neither effective preventive measures nor approved treatment options available, there is an urgent need to identify antiviral drugs against FIPV. Our FCoV replicon system provides a valuable tool for drug discovery in vitro. Due to the lack of cell culture systems for serotype I FCoVs (the serotype most prevalent in the feline population) (2), a different system is needed to study these viruses. A viral replicon system is a valuable tool for studying FCoVs. Overall, our results demonstrate the utility of the serotype I feline coronavirus replicon system for antiviral screening as well as to study this virus in general. We propose several compounds representing promising candidates for future clinical trials and ultimately with the potential to save cats suffering from FIP.

Dissecting the role of ALK double mutations in drug resistance to lorlatinib with in-depth theoretical modeling and analysis.

Anaplastic lymphoma kinase (ALK) is implicated in the genesis of multiple malignant tumors. Lorlatinib stands out as the most advanced and effective inhibitor currently used in the clinic for the treatment of ALK-positive non-small cell lung cancer. However, resistance to lorlatinib has inevitably manifested over time, with double/triple mutations of G1202, L1196, L1198, C1156 and I1171 frequently observed in clinical practice, and tumors regrow within a short time after treatment with lorlatinib. Therefore, elucidating the mechanism of resistance to lorlatinib is paramount in paving the way for innovative therapeutic strategies and the development of next-generation drugs. In this study, we leveraged multiple computational methodologies to delve into the resistance mechanisms of three specific double mutations of ALKG1202R/L1196M, ALKG1202R/L1198F and ALKI1171N/L1198F to lorlatinib. We analyzed these mechanisms through qualitative (PCA, DCCM) and quantitative (MM/GBSA, US) kinetic analyses. The qualitative analysis shows that these mutations exert minimal perturbations on the conformational dynamics of the structural domains of ALK. The energetic and structural assessments show that the van der Waals interactions, formed by the conserved residue Leu1256 within the ATP-binding site and the residues Glu1197 and Met1199 in the hinge domain with lorlatinib, play integral roles in the occurrence of drug resistance. Furthermore, the US simulation results elucidate that the pathways through which lorlatinib dissociates vary across mutant systems, and the distinct environments during the dissociation process culminate in diverse resistance mechanisms. Collectively, these insights provide important clues for the design of novel inhibitors to combat resistance.

Biological evaluation of indolactams for in vitro bryostatin 1-like activity.

Small molecule activators of protein kinase C (PKC) have traditionally been classified as either tumor promoters or suppressors. Although bryostatin 1 has well established anti-cancer activity, most natural products that target the PKC regulator domain exhibit tumor promotion properties. In this study, we examine a focused library of indolactam analogues in cell-based assays to establish the structural features of the scaffold that enhance bryostatin 1-like activity. These systematic biological assessments identified specific indole substitution patterns that impart diminished tumor promotion behavior in vitro for indolactam analogues, while still maintaining nanomolar potency for PKC.

Fulvanines J-K, two rare lactam pyrrole alkaloids from Hemerocallis fulva.

Two rare jatropham lactam derivatives, named as fulvanines J-K (1-2), together with six known pyrrole alkaloids, 5,5'-oxydi(3-methyl-3-pyrrolin-2-one) (3), (-)-5-hydroxy-3-methyl-3-pyrrolin-2-one (jatropham) (4), (±)-5-O-methyljatropham (5), perlolyrine (6), butyl-2-formyl-5-(hydroxymethyl)-1H-pyrrole-1-butanoate (7), and hemerocallisamine II (8), were isolated from the flower of Hemerocallis fulva. Their structures were elucidated on the basis of spectroscopic methods and compared with the NMR spectra data in the literature. All compounds were evaluated for their anti-complementary activity in vitro, and compounds 1, 4, and 6 exhibited anti-complement effect with CH50 values from 0.61 to 1.42 mM.

Two New Lactam Derivatives from Micromelum falcatum (Lour.) Tan. with Brine Shrimp Larvae Toxicity.

Chemical investigation of the stems of Micromelum falcatum (Lour.) Tan. led to the isolation of two new lactam derivatives, named 3-(hydroxy(10-hydroxyphenyl)methyl)-4-(16-hydroxyphenyl)-1-methylpyrrolidin-2-one (1) and 3-(hydroxy(10-hydroxy-9-methoxyphenyl)methyl)-4-(16-hydroxyphenyl)-1-methylpyrrolidin-2-one (2), along with five known compounds, trans-4-hydroxycinnamic acid (3), 4-hydroxybenzaldehyde (4), m-hydroxybenzoic acid (5), p-hydroxybenzoic acid (6), and gallic acid (7). Their structures were determined on the basis of spectroscopic studies, including nuclear magnetic resonance (NMR) spectrum, mass spectrometry (MS) data, ultraviolet (UV) spectrum, infrared (IR) data, and comparison with the literature. All compounds were evaluated for toxicity against brine shrimp larvae and cytotoxicity to HeLa and HepG-2 cells. Compounds 1-2 exhibited moderate brine shrimp larvae toxicity with an LC50 value of 50.6 and 121.8 µg mL-1, respectively.

Synthesis by diastereomeric resolution, biochemical evaluation and molecular modelling of chiral 3-hydroxyl b-lactam microtubule-targeting agents for the treatment of triple negative breast and chemoresistant colorectal cancers.

The synthesis and biochemical activity of a series of chiral trans 3-hydroxyl ß-lactams targeting tubulin is described. Synthesis of the series of enantiopure ß-lactams was achieved using chiral derivatising reagent N-Boc-l-proline. The absolute configuration was determined as 3S,4S for (+) enantiomer 4EN1 and 3R,4R for (-) enantiomer 4EN2. Antiproliferative studies identified chiral 3S,4S b-lactams with subnanomolar IC50 values across a range of cancer cell lines, improving potency with respect to the corresponding racemates. Fluoro-substituted (+)-(3S,4S)-4-(3-fluoro-4-methoxyphenyl)-3-hydroxy-1-(3,4,5-trimethoxyphenyl)azetidin-2-one (27EN1) was determined as the lead eutomer with dual antiproliferative activity in triple negative breast cancer cells (TNBC), and combretastatin A-4 resistant HT-29 colorectal cancer cells. IC50 values were in the range of 0.26-0.7 nM across four cell lines. Tubulin polymerisation assays, confocal microscopy and molecular modelling studies indicated that 3S,4S eutomers are microtubule destabilisers, while 3R,4R distomers have lower potency as microtubule destabilisers. 27EN1 demonstrated anti-mitotic and pro-apoptotic activity in MDA-MB-231 and HT-29 cells in addition to selective toxicity toward MCF-7 breast cancer versus non-tumorigenic MCF-10-2A cells. The related 3S,4S ß-lactam eutomer 4EN1 downregulated expression of key cell survival anti-apoptotic proteins Bcl-2 and Mcl-1 in MDA-MB-231 cells while 27EN1 downregulated Mcl-1 in HT-29 cells. Chiral ß-lactam 27EN1 will be further developed for treatment of TNBC and CA-4 resistant colorectal cancers.

Haneummycin, a new 22-membered macrolide lactam antibiotic, produced by marine-derived Streptomyces sp. KM77-8.

A new antibiotic named haneummycin (1) was isolated from a culture broth of marine-derived Streptomyces sp. KM77-8 by solvent extraction and HPLC using a C4 column. The structure of 1 was elucidated including relative stereochemistry as a new 22-membered macrolide lactam associated with a cyclopentanone and three sugars by various spectroscopic analyses, such as MS and NMR. Compound 1 displayed significant antibacterial activities against Gram-positive bacteria including vancomycin-resistant Enterococcus faecium (VRE) and methicillin-resistant Staphylococcus aureus (MRSA) with both MIC values of 8.0 µg ml-1.

Novel D-modified heterocyclic androstane derivatives as potential anticancer agents: Synthesis, characterization, in vitro and in silico studies.

Cancer remains a major health concern worldwide. The most frequently diagnosed types of cancer are caused by abnormal production or action of steroid hormones. In the present study, the synthesis and structural characterization of new heterocyclic androstane derivatives with D-homo lactone, 17α-(pyridine-2''-ylmethyl) or 17(E)-(pyridine-2''-ylmethylidene) moiety are presented. All compounds were evaluated for their anti-proliferative activity against HeLa cervical cancer cell line and non-cancerous kidney MDCK cells, where A-homo lactam compound 9A showed the greatest selectivity. Based on in vitro binding assays, N-formyl lactam compound 18 appeared to be the strong and isoform-selective ligand for ERα, while compound 9A displayed binding affinity for the GR-LBD, but also inhibited aldo-keto reductase 1C4 enzyme. Out of four selected compounds, methylpyrazolo derivative 13 showed potential for aromatase binding, while in silico studies provided insight into experimentally confirmed protein-ligand interactions.

Determination of potential combination of non-ß-lactam, ß-lactam, and ß-lactamase inhibitors/ß-lactam enhancer against class D oxacillinases producing Acinetobacter baumannii: Evidence from in-vitro, molecular docking and dynamics simulation.

Carbapenem-resistant Acinetobacter baumannii, a predominant nosocomial pathogen in hospitals of intensive care units, is associated with bacteremia and ventilator-associated pneumonia with a high-risk mortality rate. To increase the effectiveness of the ß-lactam (BL) antibiotics, the use of ß-lactamase inhibitors (BLI) acts as a booster when given in combination with BL antibiotics. To this aspect, we selected BL antibiotics of cefiderocol, cefepime, non-BL antibiotic eravacycline, BLI of durlobactam, avibactam, and a ß-lactam enhancer (BLE) of zidebactam. To prove our hypothesis, we determined the minimum inhibitory concentration (MIC) of various BL or non-BL/BLI or BLE combinations using broth microdilution method followed by in silico analysis of molecular docking, molecular dynamics (MD) simulation, and molecular mechanics Poisson-Boltzmann surface area (MM-PBSA) identifies the potential combination. In MIC testing, eravacycline, cefepime/zidebactam, cefiderocol/zidebactam, and eravacycline in combination with zidebactam or durlobactam were found to be effective against oxacillinases (OXAs) (OXA-23/24/58 like) expressing A. baumannii isolates. The docking results of the selected ligands toward OXA-23, OXA-24, and OXA-58 had an excellent binding score ranging from -5.8 to -9.3 kcal/mol. Further, the docked complexes were subjected and evaluated using gromacs for molecular dynamics simulation of 50 ns toward selected class D OXAs. The binding energies obtained from MM-PBSA shed light on the binding efficiencies of each non-BL, BL, and BLI/BLE, thereby helping us to propose the drug combinations. Based on the MD trajectories scoring acquired, we propose using eravacycline, cefepime/zidebactam, cefiderocol/zidebactam, and eravacycline in combination with durlobactam or zidebactam would be promising for treating OXA-23, OXA-24, and OXA-58 like expressing A. baumannii infections.

Unexpected Cascade Sequence Forming the C(sp3)-N/C(sp2)-C(sp2) Bond: Direct Access to γ-Lactam-Fused Pyridones with Anticancer Activity.

An unexpected Ugi cascade reaction was developed for the facile construction of γ-lactam-fused pyridone derivatives with high tolerance of substrates. A C(sp3)-N bond and a C(sp2)-C(sp2) bond were formed together, accompanied by a chromone ring-opening in Ugi adducts, under the basic conditions without any metal catalyst for the whole process. Screening data of several difficult-to-inhibit cancer cell lines demonstrated that 7l displayed a high cytotoxicity against HCT116 cells (IC50 = 5.59 ± 0.78 µM). Taken together, our findings revealed new insights into the molecular mechanisms underlying compound 7l and provided potential usage of this scaffold for cancer therapeutics.

Synthesis of novel ciprofloxacin-avibactam conjugates for the development of second-generation non-ß-lactam-ß-lactamase inhibitors.

To overcome the antibiotic resistance challenge, we synthesized a novel class of conjugates based on ciprofloxacin and avibactam, covalently linked by diverse amino acids. In vitro studies of these conjugates have shown improved antibacterial efficacy of avibactam when used alone against some ESKAPE pathogens, i.e., S. aureus, E. coli, and A. baumannii. Further, ceftazidime was screened in combination with all conjugates and found to be less synergistically effective than avibactam-ceftazidime co-dosing against K. pneumoniae and E. coli bacterial strains. Subsequently, the top-ranked active conjugates were investigated against the commercially available ß-lactamase-II (Penicillinase from Bacillus cereus) through in vitro studies. These studies elucidated two conjugates i.e, 9 (IC50 = 1.69±0.35 nM) and 24b (IC50 = 57.37±5.39 nM), which have higher inhibition profile than avibactam (IC50 = 141.08±12.20 nM). These outcomes allude to avibactam integration with ciprofloxacin is a novel and fruitful approach to discovering clinically valuable next-generation non-ß-lactam-ß-lactamase inhibitors.

Phthalazinone-based lactams and cyclic ureas as ROCK2 selective inhibitors.

Derivatives of lactam, cyclic urea and carbamate were explored as aniline amide replacements in a series of phthalazinone-based ROCK inhibitors. Potent ROCK2 inhibitors such as 22 were identified with excellent overall kinase selectivity as well as good isoform selectivity over ROCK1.

Sulbactam-durlobactam: A novel ß-lactam-ß-lactamase inhibitor combination targeting carbapenem-resistant Acinetobacter baumannii infections.

Carbapenem-resistant Acinetobacter baumannii (CRAB) is a difficult-to-treat nosocomial pathogen responsible for significant morbidity and mortality. Sulbactam-durlobactam (SUL-DUR), formerly ETX2514SUL, is a novel ß-lactam-ß-lactamase inhibitor designed specifically for the treatment of CRAB infections. The United States Food and Drug Administration (FDA) fast-track approval of SUL-DUR for the treatment of CRAB infections is currently pending after completion of the phase III ATTACK trial, which compared SUL-DUR to colistin, both in combination with imipenem-cilastatin (IMI) for patients with CRAB-associated hospital-acquired bacterial pneumonia, ventilator-associated pneumonia, and bacteremia. The results of this trial demonstrated that SUL-DUR was non-inferior to colistin for CRAB while also possessing a much more favorable safety profile. SUL-DUR was well-tolerated with the most common side effects being headache, nausea, and injection-site phlebitis. With the current landscape of limited effective treatment options for CRAB infections, SUL-DUR represents a promising therapeutic option for the treatment of these severe infections. This review will discuss the pharmacology, spectrum of activity, pharmacokinetics/pharmacodynamics, in vitro and clinical studies, safety, dosing, administration, as well as the potential role in therapy for SUL-DUR.

Investigating the Antibacterial Effects of Synthetic Gamma-Lactam Heterocycles on Methicillin-Resistant Staphylococcus aureus Strains and Assessing the Safety and Effectiveness of Lead Compound MFM514.

Methicillin-resistant Staphylococcus aureus (MRSA) continues to be one of the main causes of hospital-acquired infections in all regions of the world, while linezolid is one of the only commercially available oral antibiotics available against this dangerous gram-positive pathogen. In this study, the antibacterial activity from 32 analogues of synthetic gamma-lactam heterocycles against MRSA was determined. Amongst screened analogues for the minimum inhibitory concentration (MIC) assay, compound MFM514 displayed good inhibitory activity with MIC values of 7.8-15.6 µg/mL against 30 MRSA and 12 methicillin-sensitive S. aureus (MSSA) clinical isolates, while cytotoxicity evaluations displayed a mean inhibitory concentration (IC50) value of > 625 µg/mL, displaying a potential to becoming as a lead compound. In subsequent animal studies for MFM514, a single-dose oral acute toxicity test revealed an estimated mean lethal dose (LD50) value of <5000 mg/kg, while in the mice infection test, a mean effective dose (ED50) value of 29.39 mg/kg was obtained via oral administration. These results suggest that gamma-lactam carbon skeleton, particularly MFM514, is highly recommended to be evaluated further as a new safe and efficacious orally delivered antibacterial agent against MRSA.

Impact of short chain fatty acids (SCFAs) on antimicrobial activity of new ß-lactam/ß-lactamase inhibitor combinations and on virulence of Escherichia coli isolates.

In a healthy gut microbiota, short chain fatty acids (SCFAs) are produced. The antibacterial action of SCFAs against intestinal pathogens makes them useful for ensuring the safety of food and human health. In this study, we aimed to assess the in vitro inhibitory activity of SCFAs, and to report, for the first time, their impact on the activity of new ß-lactam/ß-lactamase inhibitor combinations. The minimum inhibitory concentrations of acetic, propionic, and butyric acids were determined against E. coli clinical isolates recovered from gastrointestinal infections. Cefoperazone/sulbactam, ceftazidime/avibactam and cefepime/enmetazobactam are new ß-lactam/ß-lactamase inhibitor combinations that were studied for their combined therapeutic effects. Also, the effects of pH and concentration of SCFAs were evaluated on in vitro bacterial growth and expression of genes encoding for motility, adhesion, invasion, and biofilm formation. SCFAs were tested at concentrations of 12 mM at pH 7.4 (ileum-conditions), in addition to 60 mM and 123 mM, at pH 6.5 (colon-conditions). The tested SCFAs showed the same MIC (3750 µg ml-1 ≃ 60 mM) against all isolates. Furthermore, the addition of SCFAs to the tested ß-lactam/ß-lactamase inhibitor combinations greatly restored the susceptibility of the isolates. SCFAs had significant effect on bacterial growth and virulence in a pH and concentration-dependent manner; low ileal concentration potentiated E. coli growth, while higher colonic concentration significantly suppressed growth and down-regulated the expression of virulence genes (fliC, ipaH, FimH, BssS). Therefore, the significant inhibitory effect of colonic SCFAs on ß-lactam/ß-lactamase inhibitor combinations might lead to the development of promising treatment strategies.

Somalactams A-D: Anti-inflammatory Macrolide Lactams with Unique Ring Systems from an Arctic Actinomycete Strain.

Four new PKS-NRPS-derived macrolide lactams with three unique ring fusion types were discovered from the Arctic sponge associated actinomycete Streptomyces somaliensis 1107 using a genome mining strategy. Their structures were elucidated by a combination of MS, NMR spectroscopic analysis, and single-crystal X-ray diffraction. Biosynthetically, a novel gene cluster sml consisting of three polyketide synthases and one hybrid polyketide synthase-nonribosomal peptide synthetase together with cytochrome P450s and flavin-containing monooxygenases and oxidoreductases was demonstrated to assemble the unique skeleton. Pharmacological studies revealed that compound 1 displayed a potent anti-inflammatory effect without cytotoxicity. It inhibited IL-6 and TNF-α release in the serum of LPS-stimulated RAW264.7 macrophage cells with IC50 values of 5.76 and 0.18 µM, respectively, and modulated the MAPK pathway. Moreover, compound 1 alleviated LPS-induced systemic inflammation in our transgenic fluorescent zebrafish model.

Synthesis and antiproliferative activities of steroidal lactam conjugates bearing a new nitrogen mustard.

Alkylating agents are potent anticancer compounds that exert their anticancer properties through the inhibition of cell replication and transcription leading to cell death. Despite the numerous benefits, these agents also have serious drawbacks such as their high toxicity and low specificity towards cancer cells. As previously reported by our group, conjugation of alkylating agents with azasteroids can reduce their systemic toxicity and enhance their anticancer activity. In this work, novel steroidal alkylating agents bearing POPAM-OH were synthesized and their anticancer efficacy was evaluated in vitro and in vivo. All the novel hybrids demonstrated high antiproliferative effects against 5 different cancer cell lines in the low micromolar range. Treatment of SCID mice bearing SKOV-3 or PC-3 tumor xenografts with the most potent hybrid 19 led to significant reduction of tumor size (tumor inhibition TI = 95% in SKOV3 models and TI = 85.2% in PC3 models). Importantly, the acute toxicity of hybrid 19 (LD10 = 36 µΜ, LD50 = 62 µΜ) in CB17 SCID mice exhibited three-fold decrease compared to the acute toxicity of previously reported hybrids of POPAM-NH2. This is an important finding since systemic cytotoxicity is a critical limitation of alkylating agents. Collectively, the steroidal conjugates of POPAM-OH displayed significant anticancer efficacy and reduced toxicity in vitro and in vivo rendering them as good candidates for cancer therapy.