Synthesis, biological evaluation and structure-activity relationship of novel dichloroacetophenones targeting pyruvate dehydrogenase kinases with potent anticancer activity.

Pyruvate dehydrogenase kinases (PDKs) are promising therapeutic targets that have received increasing attentions in cancer metabolism. In this paper, we report the synthesis and biological evaluation of a series of novel dichloroacetophenones as potent PDKs inhibitors. Structure-activity relationship analysis enabled us to identify a potent compound 6u, which inhibited PDKs with an EC50 value of 0.09 µM, and reduced various cancer cells proliferation with IC50 values ranging from 1.1 to 3.8 µM, while show weak effect against non-cancerous L02 cell (IC50 > 10 µM). In the A375 xenograft model, 6u displayed an obvious antitumor activity at a dose of 5 mg/kg, but with no negative effect to the mice weight. Molecular docking suggested that 6u formed direct hydrogen bond interactions with Ser75 and Gln61 in PDK1, and meanwhile the aniline skeleton in 6u was sandwiched by the conserved hydrophobic residues Phe78 and Phe65, which contribute to the biochemical activity improvement. Moreover, 6u induced A375 cell apoptosis and cell arrest in G1 phase, and inhibited cancer cell migration. In addition, 6u altered glucose metabolic pathway in A375 cell by decreasing lactate formation and increasing ROS production and OCR consumption, which could serve as a potential modulator to reprogram the glycolysis pathway in cancer cell.

Transmammary modulation of xenobiotic metabolizing enzymes in liver of mouse pups by mace (Myristica fragrans Houtt.).

The present study examines the possible transfer of the active principle(s) of mace (aril of the plant Myristica fragrans) through the transmammary route and its ability to modulate hepatic xenobiotic metabolizing enzymes in the F1 progeny of mice. An aqueous suspension of mace at the dose levels of 0.025 or 0.1 g/animal/day was administered by oral gavage to dams from day 1 of lactation and continued daily for 14 or 21 days. Dams receiving mace treatment and their F1 pups showed significantly elevated hepatic sulfhydryl content, glutathione S-transferase and glutathione reductase activities and cytochrome b5 content. Hepatic cytochrome P450 content decreased in dams (P < 0.05) receiving the lower mace dose for 21 days and the F1 pups (P < 0.001), but increased in dams receiving the higher dose for both time periods (P < 0.001) and the lower dose for 14 days (P < 0.05). Only the 14-day-old pups of dams receiving either mace dose showed significantly elevated (P < 0.001) levels of hepatic glutathione peroxidase.

A comparative study of biochemical changes induced by inhalation of aerosols of o-Chloroacetophenone & Dibenz (b,f)-1,4-oxazepine in rats.

The biochemical changes in blood samples of rats at different intervals after O-Chloroacetophenone (CN) and Dibenz (b,f)-1,4 oxazepine (CR) were studied. After a single subacute (1/10 LC50) exposure, both the compounds induced hyperglycaemia which was abolished within 24 h. The level of plasma urea was unaltered. CR exposed animals did not show any significant changes in plasma GOT, acid and alkaline phosphatase activities at different intervals. However, in CN exposed animals, a significant elevation of the activities of GOT, GPT, acid and alkaline phosphatase was observed at different intervals. All the parameters became normal within seven days after the exposure. Inhalation of CN aerosols can thus lead to tissue damaging effects in rats.

Examination of mouse exorbital lacrimal gland after exposure to 2-chloroacetophenone vapor.

The compound 2-chloroacetophenone has been used to stimulate tear secretion from human subjects, yet the morphological response of the lacrimal gland to this agent has not been experimentally determined. This study used light and electron microscopic techniques to examine the effect of this agent with time on the morphology of the mouse exorbital lacrimal gland. In brief, exposure to 2-chloroacetophenone vapor causes rapid exocytosis of acinar cell granules as well as vacuolation of the secretory and ductal epithelial cells. Concomitantly, intralobular ductal epithelial cells which are normally cuboidal in shape, enlarge in size and release electron-dense granules into the duct lumen. Within 15 min after exposure, acinar cells show a preponderance of cells containing only pale granules and the ductal epithelium returns to a more cuboidal shape. By 60 min after exposure, the gland is not readily distinguishable from the unexposed, control gland.

Effects of methyl mercuric chloride and sulfhydryl inhibitors on phospholipid synthetic activity of lymphocytes.

The effect of methyl mercuric chloride on the activity of phospholipid synthesis of rat lymph node lymphocytes was compared with that of sulfhydryl inhibitors. Measurement of the radioactivity of [14C]oleic acid and [14C]acetate incorporated into lecithin of cells during short-term incubation showed that all the inhibitors tested similarly reduced the incorporation. However, methyl mercuric chloride (MMC) was the strongest inhibitor, being effective at 4 microM and causing more than 80% decrease at 20 microM. Inhibition by the sulfhydryl inhibitors, at less than 40 microM, ranked as follows: N-ethylmaleimide greater than alpha-chloroacetophenone greater than hydroquinone greater than iodoacetamide. MMC also obstructed the enhancement by phytohemagglutinin of [14C]oleic acid incorporation into lecithin. MMC was effective at 2 microM, while the other agents had little or no effect at this concentration. Further investigation suggested that inhibition of phospholipid synthesis did not depend on reduced incorporation of oleic acid into the cellular membrane but on decreased turnover of the fatty acid into phospholipids after the incorporation. The viability of lymphocytes incubated with the agents was measured by trypan blue dye-exclusion test. More than 90% of the cells treated with MMC at a concentration as low as 20-40 microM died, but the SH inhibitors, including NEM which greatly inhibited the phospholipid synthesis, produced few cell deaths at these concentrations. These observations show that the SH inhibitors affect enzymes in phospholipid synthesis, whereas MMC not only inhibits the enzymes but kills cells.

[Proton translocation in membranes of submitochondrial particles]. / Translokatsiia protona v membranakh submitokhondrial'nykh chastits.

Effect of an electrophilous inhibitor, chlorophenacyl, on energy-dependent functions of submitochondrial particles is studied. Chlorophenacyl at concentrations up to 1 mM is found practically not to affect the generation of membrane potential under NADH and succinate oxidation and ATP hydrolysis and to be a strong inhibitor of oxidative phosphorylation and reverse electron transport. The mechanism of the inhibition of energy-dependent functions of submitochondrial particles with chlorophenacyl is different from that of electron transport inhibitor, energy transport inhibitors and classical uncoupling agents--protonophors. The data obtained are suggested to be due to the existence of two ways of proton translocation in submitochondrial particle membrane, phosphorylating and non-phosphorylating, the effect of chlorophenacyl being directed on phosphorylating way only.

Studies on cell differentiation: inducing capacity of sulfhydryl-containing amino acids on post-nodal pieces of chick blastoderms.

The inducing capacity of sulfhydryl (SH)-containing amino acids (cysteine and glutathione) on post-nodal pieces (PNPs) of stage 4 chick blastoderms was investigated. PNPs were treated for different lengths of time with chick Ringer's solution (control group) or chick Ringer's solution containing cysteine or glutathione, followed by culturing for 2-10 days on Spratt-Haas agar medium or on the chorioallantoic membrane of 8-day chick embryos. Control PNPs rarely showed differentiation, but those treated with the amino acids for six hours or longer developed structures such as neural tissue, notochord, somite mesoderm, and nephric tubules. The pulsatile tissue was only seen in the PNPs cultured for four days or longer. Two-four hours of treatment was too short to provoke induction in a statistically significant number of PNPs. The highest frequency of induction was noted in those pretreated with glutathione (8 mug/ml) for eight hours, followed by culturing for four days. The magnitude of the inducing capacity and toxicity of the amino acids were concentration dependent: a deleterious effect was observed at 14 mug/ml; the highest frequency of induction occurred at 8 mug/ml, but the frequency decreased as the concentration decreased; at 2 mug/ml all PNPs remained viable, but only a few (9-14%) showed differentiation. The inducing capacity of the amino acids was counteracted by equimolar concentrations of rho-chloromercuribenzoic acid or omega-chloroacetophenone. The effects of glutathione (8 mug/ml) differed from those of Hensen's node grafts in that the former caused sublethal cytolysis and inhibited H-3-uridine uptake in competent ectodermal cells during the first 18 hours of cultivation.