INPs, which are chemically synthesized compounds belonging to a class of acylated hydrazones of salicylaldehydes, can inhibit the growth of Chlamydiaceae. Evidence has been presented that in Yersinia and Chlamydia INPs may affect the type III secretion (T3S) system. In the present study 25 INPs were screened for antichlamydial activity at a concentration of 50 M, and 14 were able to completely inhibit the growth of Chlamydia trachomatis serovar D in McCoy and HeLa 229 cells. The antichlamydial activities of two of these INPs, INPs 0341 and 0400, were further characterized due to their low cytotoxicity. These compounds were found to inhibit C. trachomatis in a dose-dependent manner; were not toxic to elementary bodies; were cidal at a concentration of >20 M; inhibited all Chlamydiaceae tested; and could inhibit the development of C. trachomatis as determined by the yield of progeny when they were added up to 24 h postinfection. INP 0341 was able to affect the expression of several T3S genes. Compared to the expression in control cultures, lcrH-1, copB, and incA, all middle-to late-expressed T3S genes, were not expressed in the INP 0341-treated cultures 24 to 36 h postinfection. Iron, supplied as ferrous sulfate, as ferric chloride, or as holo-transferrin, was able to negate the antichlamydial properties of the INPs. In contrast, apo-transferrin and other divalent metal ions tested were not able to reverse the inhibitory effect of the INPs. In conclusion, the potent antichlamydial activity of INPs is directly or indirectly linked with iron, and this inhibition of Chlamydia has an effect on the T3S system of this intracellular pathogen.The type III secretion (T3S) system is known to be a potent virulence mechanism shared by several pathogenic bacteria, including the Chlamydiaceae (10). All T3S systems share common structural components, while their effector proteins and methods of gene regulation vary widely. Targeting and inactivating common T3S components has been proposed as a strategy to fight infections caused by pathogens that require a T3S system for virulence (13). In an attempt to identify such compounds, Kauppi et al. (13) used a chemical genetics approach to screen a large number of synthetic compounds for the ability to inhibit Yersinia T3S gene expression. They identified compounds with the general structure of an acylated hydrazone of salicylaldehydes that were able to inhibit the pathogenic Yersinia T3S system, neutralizing the virulence while not affecting the growth of the organism (13, 17).We have previously reported that INP 0400 was able to inhibit the growth of Chlamydophila pneumoniae (27a). We reported that this compound inhibited C. pneumoniae development in a dose-dependent manner, was not cytotoxic, was not directly toxic to elementary bodies (EBs), and was effective at inhibiting the growth of Chlamydia trachomatis and Chlamydia muridarum. The appearance of inclusions at lower concentrations (Ͻ20 M) of INP 0400 resembled the appearance of inclusions seen in persistent infections resulting...

Scope and limitations of the base-free oxidative Heck reaction with arylboronic acids have been explored. Under our conditions, the dmphen-palladium(II)-catalyzed arylation proceeded with air or p-benzoquinone as reoxidants of palladium(0). We found that ambient temperature and mild aerobic conditions allow for the use of substrates sensitive to palladium(II)-catalyzed oxidation. Oxidative Heck couplings, employing different arylboronic acids, were smoothly and regioselectively conducted with both electron-rich and electron-poor olefins, providing high yields even with disubstituted butyl methacrylate, sensitive acrolein, and a vinylboronate ester. Controlled microwave processing was used to reduce reaction times from hours to minutes both in small scale and in 50 mmol scale batch processes.

Electrospray ionization mass spectrometry (ESI-MS) and subsequent MS/MS analyses were used to directly detect palladium-containing cationic reaction intermediates in a ligand controlled palladium(II)-catalyzed oxidative Heck arylation. All potential intermediates were observed as dmphen-ligated palladium(II) species, suggesting that the dmphen bidentate ligand is attached to the metal center during the entire catalytic cycle. The study supports previous mechanistic propositions and provides new information regarding the composition of aryl-containing Pd(II) complexes in an ongoing oxidative Heck reaction. In addition, sodium acetate was found to be a useful base alternative to previously used tertiary amines.