Synthesis and evaluation of diphyllin ß-hydroxyl amino derivatives as novel V-ATPase inhibitors.

Natural diphyllin glycosides were identified as potent vacuolar H+ -ATPase (V-ATPase) inhibitors. A series of diphyllin ß-hydroxyl amino derivatives were designed and synthesized as novel diphyllin derivatives. Most of these derivatives displayed potent cytotoxicity against six cancer cell lines with IC50 values in the submicromolar to nanomolar concentration range. Compounds 2b, 2c, 2l, 2m, and 2n showed similar V-ATPase inhibitory potency to Bafilomycin A1. Compound 2l exhibited potent activity of modulation of lysosomal pH and cytoplasmic pH.

Chemoenzymatic synthesis of enantiomerically enriched diprophylline and xanthinol nicotinate.

A concise chemoenzymatic route toward enantiomerically enriched active pharmaceutical ingredients (API) - diprophylline and xanthinol nicotinate - is reported for the first time. The decisive step is an enantioselective lipase-mediated methanolysis of racemic chlorohydrin-synthon acetate, namely 1-chloro-3-(1,3-dimethyl-2,6-dioxo-1,2,3,6-tetrahydro-7H-purin-7-yl)propan-2-yl acetate, performed under kinetically-controlled conditions on a preparative 500 mg-scale. The best results in terms of reaction enantioselectivity (E = 14) were obtained for the enantiomers resolution performed with lipase type B from Candida antarctica immobilized on acrylic resin (CAL-B, Novozym 435) suspended in homophasic acetonitrile-methanol mixture. The elaborated biocatalytic system furnished the key chlorohydrin intermediate (in 71% ee and 38% yield), which was then smoothly converted into enantioenriched active agents: (R)-(-)-diprophylline (57% ee) and (S)-(+)-xanthinol nicotinate (65% ee). To support the assignment of absolute configurations of EKR-products as well as to confirm the stereochemical outcome of the remaining reaction steps, docking studies toward the prediction of enantiomers binding selectivity in CAL-B active site as well as the respective chemical correlations with enantiomerically enriched analytical standards obtained from commercially available (R)-(-)-epichlorohydrin, were applied. In addition, single-crystal X-ray diffraction (XRD) analyses were performed for the synthesized optically active APIs furnishing by this manner a first crystal structures of nicotinic acid salt of xanthinol.

Preparation and characterization of potential prodrugs of dyphylline.

Four diesters and four monoesters of dyphylline were synthesized as prodrugs proposed to prolong the duration of action of dyphylline. They were characterized by IR, 1H NMR, HPLC, and MS. Appropriate solvent-programming conditions for the HPLC separation of dyphylline and the newly synthesized mono- and diesters were developed. It was confirmed by low-temperature 1H NMR at approximately -40 degrees C that all four monoesters were located on the primary hydroxy position. Attempts to produce the secondary monoesters yielded the primary monoesters during purification. Monoesters were shown by HPLC and MS to migrate between the primary and secondary hydroxy groups in aqueous solution.