Accraspiroketides A-B, Phenylnaphthacenoid-Derived Polyketides with Unprecedented [6 + 6+6 + 6] + [5 + 5] Spiro-Architecture.

Two novel polyketides, accraspiroketides A (1) and B (2), which feature unprecedented [6 + 6+6 + 6] + [5 + 5] spiro chemical architectures, were isolated from Streptomyces sp. MA37 ΔaccJ mutant strain. Compounds 1-2 exhibit excellent activity against Gram-positive bacteria (MIC = 1.5-6.3 µg/mL). Notably, 1 and 2 have superior activity against clinically isolated Enterococcus faecium K60-39 (MIC = 4.0 µg/mL and 4.7 µg/mL, respectively) than ampicillin (MIC = 25 µg/mL).

Design, synthesis, apoptotic, and antiproliferative effects of 5-chloro-3- (2-methoxyvinyl)-indole-2-carboxamides and pyrido[3,4-b]indol-1-ones as potent EGFRWT/EGFRT790M inhibitors.

A new series of indole-2-carboxamides 5a-g, 6a-f and pyrido[3,4-b]indol-1-ones 7a and 7b have been developed as new antiproliferative agents that target both wild and mutant type EGFR. The antiproliferative effect of the new compounds was studied. 5c, 5d, 5f, 5 g, 6e, and 6f have the highest antiproliferative activity with GI50 values ranging from 29 nM to 47 nM in comparison to the reference erlotinib (GI50 = 33 nM). Compounds 5d, 5f, and 5 g inhibited EGFRWT with IC50 values ranging from 68 to 85 nM while the GI50 of erlotinib is 80 nM. Moreover, compounds 5f and 5 g had the most potent inhibitory activity against EGFRT790M with IC50 values of 9.5 ± 2 and 11.9 ± 3 nM, respectively, being equivalent to the reference osimertinib (IC50 = 8 ± 2 nM). Compounds 5f and 5 g demonstrated excellent caspase-3 protein overexpression levels of 560.2 ± 5.0 and 542.5 ± 5.0 pg/mL, respectively, being more active than the reference staurosporine (503.2 ± 4.0 pg/mL). they also increase the level of caspase 8, and Bax while decreasing the levels of anti-apoptotic Bcl2 protein. Computational docking studies supported the enzyme inhibition results and provided favourable dual binding modes for both compounds 5f and 5 g within EGFRWT and EGFRT790M active sites. Finally, in silico ADME/pharmacokinetic studies predict good safety and pharmacokinetic profile of the most active compounds.

Design, Synthesis, and Antiproliferative Activity of New 5-Chloro-indole-2-carboxylate and Pyrrolo[3,4-b]indol-3-one Derivatives as Potent Inhibitors of EGFRT790M/BRAFV600E Pathways.

Mutant EGFR/BRAF pathways are thought to be crucial targets for the development of anticancer drugs since they are over-activated in several malignancies. We present here the development of a novel series of 5-chloro-indole-2-carboxylate 3a-e, 4a-c and pyrrolo[3,4-b]indol-3-ones 5a-c derivatives as potent inhibitors of mutant EGFR/BRAF pathways with antiproliferative activity. The cell viability assay results of 3a-e, 4a-c, and 5a-c revealed that none of the compounds tested were cytotoxic, and that the majority of those tested at 50 µM had cell viability levels greater than 87%. Compounds 3a-e, 4a-c, and 5a-c had significant antiproliferative activity with GI50 values ranging from 29 nM to 78 nM, with 3a-e outperforming 4a-c and 5a-c in their inhibitory actions against the tested cancer cell lines. Compounds 3a-e were tested for EGFR inhibition, with IC50 values ranging from 68 nM to 89 nM. The most potent derivative was found to be the m-piperidinyl derivative 3e (R = m-piperidin-1-yl), with an IC50 value of 68 nM, which was 1.2-fold more potent than erlotinib (IC50 = 80 nM). Interestingly, all the tested compounds 3a-e had higher anti-BRAFV600E activity than the reference erlotinib but were less potent than vemurafenib, with compound 3e having the most potent activity. Moreover, compounds 3b and 3e showed an 8-fold selectivity index toward EGFRT790M protein over wild-type. Additionally, molecular docking of 3a and 3b against BRAFV600E and EGFRT790M enzymes revealed high binding affinity and active site interactions compared to the co-crystalized ligands. The pharmacokinetics properties (ADME) of 3a-e revealed safety and good pharmacokinetic profile.

Optimization and SAR investigation of novel 2,3-dihydropyrazino[1,2-a]indole-1,4-dione derivatives as EGFR and BRAFV600E dual inhibitors with potent antiproliferative and antioxidant activities.

Using a single drug to treat cancer with dual-targeting is an unusual approach when compared to other drug combinations. Dual-targeting agents were developed as a result of insufficient efficacy and drug resistance when single-targeting agents were used. As a result, the 2,3-dihydropyrazino[1,2-a]indole-1,4-dione derivatives 13-22 have been developed as dual EGFR and BRAFV600E inhibitors. The target compounds were synthesized and tested in vitro against four cancer cell lines, with compounds 15, and 19-22 demonstrating potent antiproliferative activity. In vitro studies revealed that these compounds have dual inhibitory effect on EGFR and BRAFV600E. Compounds 15, and 19-22 exhibited inhibitions of EGFR with IC50 ranging from 32 nM to 63 nM which were superior to erlotinib (IC50 = 80 ± 10 nM). Compounds 20, 21 and 22 showed promising inhibitory activity of BRAFV600E (IC50 = 55, 45 and 51 nM, respectively) and were found to be potent inhibitors of cancer cell proliferation (GI50 = 51, 35 and 44 nM, respectively). Compounds 20, 21 and 22 showed good antioxidant activity comparable to the reference Trolox. Lastly, the best active dual inhibitors were docked inside EGFR and BRAFV600E active sites to clarify their binding modes.

Novel 1,5-diaryl pyrazole-3-carboxamides as selective COX-2/sEH inhibitors with analgesic, anti-inflammatory, and lower cardiotoxicity effects.

COX-2 selective drugs have been withdrawn from the market due to cardiovascular side effects, just a few years after their discovery. As a result, a new series of 1,5-diaryl pyrazole carboxamides 19-31 was synthesized as selective COX-2/sEH inhibitors with analgesic, anti-inflammatory, and lower cardiotoxic properties. The target compounds were synthesized and tested in vitro against COX-1, COX-2, and sEH enzymes. Compounds 20, 22 and 29 exhibited the most substantial COX-2 inhibitory activity (IC50 values: 0.82-1.12 µM) and had SIs of 13, 18, and 16, respectively, (c.f. celecoxib; SI = 8). Moreover, compounds 20, 22, and 29 were the most potent dual COX-2/sEH inhibitors, with IC50 values of 0.95, 0.80, and 0.85 nM against sEH, respectively, and were more potent than the standard AUDA (IC50 = 1.2 nM). Furthermore, in vivo studies revealed that these compounds were the most active as analgesic/anti-inflammatory derivatives with a good cardioprotective profile against cardiac biomarkers and inflammatory cytokines. Finally, the most active dual inhibitors were docked inside COX-2/sEH active sites to explain their binding modes.

Design, synthesis, and biological evaluation of novel EGFR inhibitors containing 5-chloro-3-hydroxymethyl-indole-2-carboxamide scaffold with apoptotic antiproliferative activity.

New EGFR inhibitor series of fifteen 5-chloro-3-hydroxymethyl-indole-2-carboxamide derivatives has been designed, synthesized, and tested for antiproliferative activity against a panel of cancer cell lines. The results showed that p-substituted phenethyl derivatives 10, 11, 13, 15 and 17-19 showed superior antiproliferative activity compared to their m-substituted counterparts 12, 14, 16 and 20. Compounds 15, 16, 19 and 20 displayed promising EGFR inhibitory activity as well as an increase in caspase 3 levels. Compounds 15 and 19 increased caspase-8 and 9 levels, as well as inducing Bax and decreasing Bcl-2 protein levels. Compound 19 demonstrated cell cycle arrest at pre-G1 and G2/M phases. The results of the docking study into the active site of EGFR revealed strong fitting of the new compounds with higher binding affinities compared to erlotinib.

Discovery of novel oxazole-based macrocycles as anti-coronaviral agents targeting SARS-CoV-2 main protease.

We have discovered a family of synthetic oxazole-based macrocycles to be active against SARS-CoV-2. The synthesis, pharmacological properties, and docking studies of the compounds are reported in this study. The structure of the new macrocycles was confirmed by NMR spectroscopy and mass spectrometry. Compounds 13, 14, and 15a-c were evaluated for their anti-SARS-CoV-2 activity on SARS-COV-2 (NRC-03-nhCoV) virus in Vero-E6 cells. Isopropyl triester 13 and triacid 14 demonstrated superior inhibitory activities against SARS-CoV-2 compared to carboxamides 15a-c. MTT cytotoxicity assays showed that the CC50 (50% cytotoxicity concentration) of 13, 14, and 15a-c ranged from 159.1 to 741.8 µM and their safety indices ranged from 2.50 to 39.1. Study of the viral inhibition via different mechanisms of action (viral adsorption, replication, or virucidal property) showed that 14 had mild virucidal (60%) and inhibitory effects on virus adsorption (66%) at 20 µM concentrations. Compound 13 displayed several inhibitory effects at three levels, but the potency of its action is primarily virucidal. The inhibitory activity of compounds 13, 14, and 15a-c against the enzyme SARS-CoV-2 Mpro was evaluated. Isopropyl triester 13 had a significant inhibition activity against SARS-CoV-2 Mpro with an IC50 of 2.58 µM. Large substituents on the macrocyclic template significantly reduced the inhibitory effects of the compounds. Study of the docking of the compounds in the SARS CoV-2-Mpro active site showed that the most potent macrocycles 13 and 14 exhibited the best fit and highest affinity for the active site binding pocket. Taken together, the present study shows that the new macrocyclic compounds constitute a new family of SARS CoV-2-Mpro inhibitors that are worth being further optimized and developed.

Discovery and Biosynthetic Investigation of a New Antibacterial Dehydrated Non-Ribosomal Tripeptide.

Dehydroalanine (Dha) and dehydrobutyrine (Dhb) display considerable flexibility in a variety of chemical and biological reactions. Natural products containing Dha and/or Dhb residues are often found to display diverse biological activities. While the (Z) geometry is predominant in nature, only a handful of metabolites containing (E)-Dhb have been found thus far. Here we report discovery of a new antimicrobial peptide, albopeptide, through NMR analysis and chemical synthesis, which contains two contiguous unsaturated residues, Dha-(E)-Dhb. It displays narrow-spectrum activity against vancomycin-resistant Enterococcus faecium. In-vitro biochemical assays show that albopeptide originates from a noncanonical NRPS pathway featuring dehydration processes and catalysed by unusual condensation domains. Finally, we provide evidence of the occurrence of a previously untapped group of short unsaturated peptides in the bacterial kingdom, suggesting an important biological function in bacteria.

Productive Syntheses of Privileged Scaffolds Inspired by the Recognition of a Diels-Alder Pattern Common to Three Classes of Natural Products.

Identification of a common Diels-Alder pattern in three classes of bioactive natural products led us to study the synthesis and cycloaddition of a new class of cyclic dienes readily available from ß,γ-unsaturated lactams. A practical and readily scalable route to the parent p-methoxybenzyl-protected 6- and 7-membered ß,γ-unsaturated lactams was developed. These were readily transformed into the corresponding O-silylated dienes, which were reacted with dimethyl and diethyl fumarate to yield stereoselectively highly functionalized bicyclic adducts. These exhibited unexpected and versatile transformations upon acid hydrolysis depending on the nature of the dienophile substituents and the acid catalyst. All reactions have been performed on multigram quantities. These transformations provide a convenient, economical, and easily scalable pathway for the rapid construction of functionally and stereochemically dense privileged scaffolds for the construction of libraries of natural products-inspired molecules of pharmacological relevance.

Characterization of the promiscuous N-acyl CoA transferase, LgoC, in legonoxamine biosynthesis.

More than 500 siderophores are known to date, but only three were identified to be aryl-containing hydroxamate siderophores, legonoxamines A and B from Streptomyces sp. MA37, and aryl ferrioxamine 2 from Micrococcus luteus KLE1011. Siderophores are produced by microorganisms to scavenge iron from the environment, thereby making this essential metal nutrient available to the microbe. We demonstrate here that LgoC from MA37 is responsible for the key aryl-hydroxamate forming step in legonoxamine biosynthesis. Biochemical characterization established that LgoC displays considerable promiscuity for the acylation between N-hydroxy-cadaverine and SNAC (N-acetylcysteamines) thioester derivatives.

Design and synthesis of novel 2,3-dihydropyrazino[1,2-a]indole-1,4-dione derivatives as antiproliferative EGFR and BRAFV600E dual inhibitors.

Recent studies have shown additive and synergistic effects associated with the combination of kinase inhibitors. BRAFV600E and EGFR are attractive targets for many diseases treatments and have been studied extensively. In keeping with our interest in developing anticancer targeting EGFR and BRAFV600E, a novel series of 2,3-dihydropyrazino[1,2-a]indole-1,4-dione has been rationally designed, synthesized and evaluated for their antiproliferative activity against a panel of four human cancer cell lines. Compounds 20-23, 28-31, and 33 showed promising antiproliferative activities. These compounds were further tested for their inhibitory potencies against EGFR and BRAFV600E kinases with erlotinib as a reference drug. Compounds 23 and 33 exhibited equipotency to doxorubicin against the four cell lines and efficiently inhibited both EGFR (IC50 = 0.08 and 0.09 µM, respectively) and BRAFV600E (IC50 = 0.1 and 0.29 µM, respectively). In cell cycle study of MCF-7 cell line, compounds 23 and 33 induced apoptosis and exhibited cell cycle arrest in both Pre-G1 and G2/M phases. Molecular docking analyses revealed that the new compounds can fit snugly into the active sites of EGFR, and BRAFV600E kinases. Compound 23, 31 and 33 adopted similar binding orientations and interactions to those of erlotinib and vemurafenib.

Structure-Based Design, Synthesis and Bioactivity of a New Anti-TNFα Cyclopeptide.

As opposed to small molecules, macrocyclic peptides possess a large surface area and are recognised as promising candidates to selectively treat diseases by disrupting specific protein-protein interactions (PPIs). Due to the difficulty in predicting cyclopeptide conformations in solution, the de novo design of bioactive cyclopeptides remains significantly challenging. In this study, we used the combination of conformational analyses and molecular docking studies to design a new cyclopeptide inhibitor of the interaction between the human tumour necrosis factor alpha (TNFα) and its receptor TNFR-1. This interaction is a key in mediating the inflammatory response to tissue injury and infection in humans, and it is also an important causative factor of rheumatoid arthritis, psoriasis and inflammatory bowel disease. The solution state NMR structure of the cyclopeptide was determined, which helped to deduce its mode of interaction with TNFα. TNFα sensor cells were used to evaluate the biological activity of the peptide.

Enzymatic Reconstitution and Biosynthetic Investigation of the Bacterial Carbazole Neocarazostatin A.

Tricyclic carbazole is an important scaffold in many naturally occurring metabolites, as well as valuable building blocks. Here we report the reconstitution of the ring A formation of the bacterial neocarazostatin A carbazole metabolite. We provide evidence of the involvement of two unusual aromatic polyketide proteins. This finding suggests how new enzymatic activities can be recruited to specific pathways to expand biosynthetic capacities. Finally, we leveraged our bioinformatics survey to identify the untapped capacity of carbazole biosynthesis.

Development of indole sulfonamides as cannabinoid receptor negative allosteric modulators.

Existing CB1 negative allosteric modulators (NAMs) fall into a limited range of structural classes. In spite of the theoretical potential of CB1 NAMs, published in vivo studies have generally not been able to demonstrate the expected therapeutically-relevant CB1-mediated effects. Thus, a greater range of molecular tools are required to allow definitive elucidation of the effects of CB1 allosteric modulation. In this study, we show a novel series of indole sulfonamides. Compounds 5e and 6c (ABD1075) had potencies of 4 and 3nM respectively, and showed good oral exposure and CNS penetration, making them highly versatile tools for investigating the therapeutic potential of allosteric modulation of the cannabinoid system.

A Unique Tryptophan C-Prenyltransferase from the Kawaguchipeptin Biosynthetic Pathway.

Cyanobactins are a rapidly growing family of linear and cyclic peptides produced by cyanobacteria. Kawaguchipeptins A and B, two macrocyclic undecapeptides reported earlier from Microcystis aeruginosa NIES-88, are shown to be products of the cyanobactin biosynthetic pathway. The 9 kb kawaguchipeptin (kgp) gene cluster was identified in a 5.26 Mb draft genome of Microcystis aeruginosa NIES-88. We verified that this gene cluster is responsible for the production of the kawaguchipeptins through heterologous expression of the kgp gene cluster in Escherichia coli. The KgpF prenyltransferase was overexpressed and was shown to prenylate C-3 of Trp residues in both linear and cyclic peptides in vitro. Our findings serve to further enhance the structural diversity of cyanobactins to include tryptophan-prenylated cyclic peptides.

Discovery of a Single Monooxygenase that Catalyzes Carbamate Formation and Ring Contraction in the Biosynthesis of the Legonmycins.

Pyrrolizidine alkaloids (PAs) are a group of natural products with important biological activities. The discovery and characterization of the multifunctional FAD-dependent enzyme LgnC is now described. The enzyme is shown to convert indolizidine intermediates into pyrrolizidines through an unusual ring expansion/contraction mechanism, and catalyze the biosynthesis of new bacterial PAs, the so-called legonmycins. By genome-driven analysis, heterologous expression, and gene inactivation, the legonmycins were also shown to originate from non-ribosomal peptide synthetases (NRPSs). The biosynthetic origin of bacterial PAs has thus been disclosed for the first time.

In-vivo pharmacological evaluation of the CB1-receptor allosteric modulator Org-27569.

Several allosteric modulators (AMs) of the CB1 receptor have been characterized in vitro, including Org27569, which enhances CB1-specific binding of [H]CP55,940, but behaves as an insurmountable CB1-receptor antagonist in several biochemical assays. Although a growing body of research has investigated the molecular actions of this unusual AM, it is unknown whether these actions translate to the whole animal. The purpose of the present study was to determine whether Org27569 would produce effects in well-established mouse behavioral assays sensitive to CB1 orthosteric agonists and antagonists. Similar to the orthosteric CB1 antagonist/inverse agonist rimonabant, Org27569 reduced food intake; however, this anorectic effect occurred independently of the CB1 receptor. Org27569 did not elicit CB1-mediated effects alone and lacked efficacy in altering antinociceptive, cataleptic, and hypothermic actions of the orthosteric agonists anandamide, CP55,940, and Δ-tetrahydrocannabinol. Moreover, it did not alter the discriminative stimulus effects of anandamide in FAAH-deficient mice or Δ-tetrahydrocannabinol in wild-type mice in the drug discrimination paradigm. These findings question the utility of Org27569 as a 'gold standard' CB1 AM and underscore the need for the development of CB1 AMs with pharmacology that translates from the molecular level to the whole animal.

Isolation and synthesis of pulmonarins A and B, acetylcholinesterase inhibitors from the colonial ascidian Synoicum pulmonaria.

Pulmonarins A and B are two new dibrominated marine acetylcholinesterase inhibitors that were isolated and characterized from the sub-Arctic ascidian Synoicum pulmonaria collected off the Norwegian coast. The structures of natural pulmonarins A and B were tentatively elucidated by spectroscopic methods and later verified by comparison with synthetically prepared material. Both pulmonarins A and B displayed reversible, noncompetitive acetylcholinesterase inhibition comparable to several known natural acetylcholinesterase inhibitiors. Pulmonarin B was the strongest inhibitor, with an inhibition constant (Ki) of 20 µM. In addition to reversible, noncompetitive acetylcholinesterase inhibition, the compounds displayed weak antibacterial activity but no cytotoxicity or other investigated bioactivities.

Design, Synthesis, and Biological Evaluation of Indole-2-carboxamides as Potential Multi-Target Antiproliferative Agents.

A small set of indole-based derivatives, IV and Va-I, was designed and synthesized. Compounds Va-i demonstrated promising antiproliferative activity, with GI50 values ranging from 26 nM to 86 nM compared to erlotinib's 33 nM. The most potent antiproliferative derivatives-Va, Ve, Vf, Vg, and Vh-were tested for EGFR inhibitory activity. Compound Va demonstrated the highest inhibitory activity against EGFR with an IC50 value of 71 ± 06 nM, which is higher than the reference erlotinib (IC50 = 80 ± 05 nM). Compounds Va, Ve, Vf, Vg, and Vh were further tested for BRAFV600E inhibitory activity. The tested compounds inhibited BRAFV600E with IC50 values ranging from 77 nM to 107 nM compared to erlotinib's IC50 value of 60 nM. The inhibitory activity of compounds Va, Ve, Vf, Vg, and Vh against VEGFR-2 was also determined. Finally, in silico docking experiments attempted to investigate the binding mode of compounds within the active sites of EGFR, BRAFV600E, and VEGFR-2.

Discovery of new 5-substituted-indole-2-carboxamides as dual epidermal growth factor receptor (EGFR)/cyclin dependent kinase-2 (CDK2) inhibitors with potent antiproliferative action.

A new series of 5-substituted-3-ethylindole-2-carboxamides 5a-k and 6a-c was designed and synthesised in an attempt to develop a dual targeted antiproliferative agent. Various spectroscopic methods of analysis were used to confirm the structures of the new compounds. The antiproliferative effect of compounds 5a-k and 6a-c against four cancer cell lines was investigated. Compounds 5a-k and 6a-c had significant antiproliferative activity against the four cancer cell lines tested, with mean GI50 values ranging from 37 nM to 193 nM. The most powerful derivatives were compounds 5g, 5i, and 5j, with GI50 values of 55 nM, 49 nM, and 37 nM, respectively, in comparison to the reference erlotinib, which had a GI50 of 33 nM. The four most potent compounds, 5c, 5g, 5i, and 5j, were then investigated for their efficacy as EGFR inhibitors, and the findings showed that the tested compounds inhibited EGFR with IC50 values ranging from 85 nM to 124 nM when compared to the reference erlotinib (IC50 = 80 nM). Moreover, compounds 5c and 5g inhibited CDK2 with IC50 values of 46 ± 05 nM and 33 ± 04 nM, respectively. The EGFR and CDK2 assays revealed that compounds 5i and 5j displayed potent antiproliferative activity and can be considered as potential dual EGFR and CDK2 inhibitors.