Synthetic Peroxides Promote Apoptosis of Cancer Cells by Inhibiting P-Glycoprotein ABCB5

This article discloses a new horizon for the application of peroxides in medical chemistry. Stable cyclic peroxides are demonstrated to have cytotoxic activity against cancer cells; in addition a mechanism of cytotoxic action is proposed. Synthetic bridged 1,2,4,5-tetraoxanes and ozonides were effective against HepG2 cancer cells and some ozonides selectively targeted liver cancer cells (the selectivity indexes for compounds 11 b and 12 a are 8 and 5, respectively). In some cases, tetraoxanes and ozonides were more selective than paclitaxel, artemisinin, and artesunic acid. Annexin V flow-cytometry analysis revealed that the active ozonides 22 a and 23 a induced cell death of HepG2 by apoptosis. Further study showed that compounds 22 a and 23 a exhibited a strong inhibitory effect on P-glycoprotein (P-gp/ABCB5)-overexpressing HepG2 cancer cells. ABCB5 is a key player in the multidrug-resistant phenotype of liver cancer. Peroxides failed to demonstrate a direct correlation between oxidative potential and their biological activity. To our knowledge this is the first time that peroxide diastereoisomers have been found to show stereospecific antimalarial action against the chloroquine-sensitive 3D7 strain of Plasmodium falciparum. Stereoisomeric ozonide 12 b is 11 times more active than stereoisomeric ozonide 12 a (IC₅₀=5.81 vs 65.18 μm ). Current findings mean that ozonides merit further investigation as potential therapeutic agents for drug-resistant hepatocellular carcinoma.     

Hit to Leads with Cytotoxic Effect in Leukemic Cells: Total Synthesis Intermediates as a Molecule Treasure Chest

A previously designed and developed 12-step total synthesis that includes [1,1′-biphenyl]-2-amine and carbazole intermediates and that ultimately produces the carbazole alkaloid carbazomycin G was exploited as a screening compound library with the goal of identifying potential lead compound(s) with cytotoxic effect. These compounds were investigated by using in-vitro tests involving the two human cell lines HL-60 and MOLM-13, which both model acute myeloid leukaemia (AML). The in-vitro biological test results were used together with the molecular structures of the various intermediates in a concise SAR analysis. Several of the intermediates revealed cytotoxicity (IC₅₀<10⁻⁴ M), although the final natural product carbazomycin G did not reveal cytotoxicity versus the two said human cell lines.     

Reexamination of the Ergothioneine Biosynthetic Methyltransferase EgtD from Mycobacterium tuberculosis as a Protein Kinase Substrate

Ergothioneine has emerged as a crucial cytoprotectant in the pathogenic lifestyle of Mycobacterium tuberculosis. Production of this antioxidant from primary metabolites may be regulated by phosphorylation of Thr213 in the active site of the methyltransferase EgtD. The structure of mycobacterial EgtD suggests that this post-translational modification would require a large-scale change in conformation to make the active-site residue accessible to a protein kinase. In this report, we show that, under in vitro conditions, EgtD is not a substrate of protein kinase PknD.     

Discovery of Affinity-Based Probes for Btk Occupancy Assays

Bruton's tyrosine kinase (Btk) is an attractive target for the treatment of a wide array of B-cell malignancies and autoimmune diseases. Small-molecule covalent irreversible Btk inhibitors targeting Cys481 have been developed for the treatment of such diseases. In clinical trials, probe molecules are required in occupancy studies to measure the level of engagement of the protein by these covalent irreversible inhibitors. The result of this pharmacodynamic (PD) activity provides guidance for appropriate dosage selection to optimize inhibition of the drug target and correlation of target inhibition with disease treatment efficacy. This information is crucial for successful evaluation of drug candidates in clinical trials. Based on the pyridine carboxamide scaffold of a novel solvent-accessible pocket (SAP) series of covalent irreversible Btk inhibitors, we successfully developed a potent and selective affinity-based biotinylated probe 12 (2-[(4-{4-[5-(1-{5-[(3aS,4S,6aR)-2-oxo-hexahydro-1H-thieno[3,4-d]imidazol-4-yl]pentanamido}-3,6,9,12-tetraoxapentadecan-15-amido)pentanoyl]piperazine-1-carbonyl}phenyl)amino]-6-[1-(prop-2-enoyl)piperidin-4-yl]pyridine-3-carboxamide). Compound 12 has been used in Btk occupancy assays for preclinical studies to determine the therapeutic efficacy of Btk inhibition in two mouse lupus models driven by TLR7 activation and type I interferon.     

Intersecting Xenobiology and Neometabolism To Bring Novel Chemistries to Life

The diversity of life relies on a handful of chemical elements (carbon, oxygen, hydrogen, nitrogen, sulfur and phosphorus) as part of essential building blocks; some other atoms are needed to a lesser extent, but most of the remaining elements are excluded from biology. This circumstance limits the scope of biochemical reactions in extant metabolism – yet it offers a phenomenal playground for synthetic biology. Xenobiology aims to bring novel bricks to life that could be exploited for (xeno)metabolite synthesis. In particular, the assembly of novel pathways engineered to handle nonbiological elements (neometabolism) will broaden chemical space beyond the reach of natural evolution. In this review, xeno-elements that could be blended into nature's biosynthetic portfolio are discussed together with their physicochemical properties and tools and strategies to incorporate them into biochemistry. We argue that current bioproduction methods can be revolutionized by bridging xenobiology and neometabolism for the synthesis of new-to-nature molecules, such as organohalides.     

Dynamic Stabilization of DNA Assembly by Using Pyrrole-Imidazole Polyamide

We used N-methylpyrrole (Py)-N-methylimidazole-(Im) polyamide as an exogenous agent to modulate the formation of DNA assemblies at specific double-stranded sequences. The concept was demonstrated on the hybridization chain reaction that forms linear DNA. Through a series of melting curve analyses, we demonstrated that the binding of Py−Im polyamide positively influenced both the HCR initiation and elongation steps. In particular, Py−Im polyamide was found to drastically stabilize the DNA duplex such that its thermal stability approached that of an equivalent hairpin structure. Also, the polyamide served as an anchor between hairpin pairs in the HCR assembly, thus improving the originally weak interstrand stability. We hope that these proof-of-concept results can inspire future use of Py−Im polyamide as a molecular tool to modulate the formation of DNA assemblies.     

Proton-Detected Solid-State NMR of the Cell-Free Synthesized α-Helical Transmembrane Protein NS4B from Hepatitis C Virus

Proton-detected 100 kHz magic-angle-spinning (MAS) solid-state NMR is an emerging analysis method for proteins with only hundreds of microgram quantities, and thus allows structural investigation of eukaryotic membrane proteins. This is the case for the cell-free synthesized hepatitis C virus (HCV) nonstructural membrane protein 4B (NS4B). We demonstrate NS4B sample optimization using fast reconstitution schemes that enable lipid-environment screening directly by NMR. 2D spectra and relaxation properties guide the choice of the best sample preparation to record 2D ¹H-detected ¹H,¹⁵N and 3D ¹H,¹³C,¹⁵N correlation experiments with linewidths and sensitivity suitable to initiate sequential assignments. Amino-acid-selectively labeled NS4B can be readily obtained using cell-free synthesis, opening the door to combinatorial labeling approaches which should enable structural studies.     

Application of Mono‐ and Disaccharides in Drug Targeting and Efficacy

Carbohydrates and their conjugates play important roles in many biological processes including fertilization, differentiation, development, immune response, and infection. Their activities are largely dependent on the properties of terminal mono‐ or disaccharides. Galactose, mannose, fucose, glucose, sialic acid, etc., are commonly used as powerful scaffolds installed on drug molecules for targeting specific tissues including brain, liver, and cancers, and as epitopes for enhancing the targeting of various vaccines. This review focuses on the influence of their structural variations, including changes in sugar type, substituent groups and their positions, as well as length of linker portion, on the targeting of drugs or their efficacy. Particular attention is paid to the targeting properties of mono‐ and disaccharides applied in drug design and discovery.