Complementary techniques were applied for the investigation of the chiral recognition and enantiomeric resolution of lenalidomide using various cyclodextrins and polysaccharides as chiral selectors. The high-performance liquid chromatography enantioseparation of the anticancer drug was achieved using polysaccharide-type chiral stationary phases in polar organic mode. Elution order and absolute configuration were elucidated by combined circular dichroism spectroscopy and time-dependent density functional theory calculations after the isolation of pure enantiomers. Chiral selector dependent and mobile-phase dependent reversal of the enantiomer elution order was observed, and the nonracemic nature of the lenalidomide sample was also demonstrated. Eight anionic cyclodextrins were screened for their ability to discriminate between the uncharged enantiomers by using capillary electrophoresis. Only two derivatives presented chiral interactions, these cases being interpreted in terms of apparent stability constants and complex mobilities. The best results were delivered by sulfobutylether-β-cyclodextrin, where quasi-equal stability constants were recorded and the enantiodiscrimination process was mainly driven by different mobilities of the transient diastereomeric complexes. The optimized high-performance liquid chromatography (Chiralcel OJ column, pure ethanol with 0.6 mL/min flow rate, 40°C) and capillary electrophoresis methods (30 mM sulfobutylether-β-cyclodextrin, 30 mM phosphate pH 6.5, 12 kV applied voltage, 10°C) were validated for the determination of 0.1% (R)-lenalidomide as a chiral impurity, which could be important if a racemic switch is achieved.

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On the basis of the knowledge that the proline-rich hot spot PPPRPP region of P( 151)PSNPPPRPP( 160), an oligopeptide derived from the cytosolic portion of p22 phox (p22), binds to the single functional bis-SH3 domain of the regulatory protein p47 phox (p47), we designed a mimetic of the tripeptide PPP based on NMR and X-ray crystallographic data for the p22(151−161) peptide PPSNPPPRPPA with a peptide construct. Incorporation of the synthetic pseudo-triproline mimetic Pro-Pro-Cyp in a molecule derived from molecular modeling studies led to only a 7-fold diminution in activity in a surface plasmon resonance assay relative to the same molecule containing the natural Pro-Pro-Pro tripeptide. The alternative sequence corresponding to a Pro-Cyp-Pro insertion was inactive. This is a first example of the use of a triproline mimetic to interfere with the formation of the p47−p22 complex, which is critical for the activation of NOX, leading to the production of reactive oxygen species as superoxide anions.

A sensitive and validated liquid chromatography with mass spectrometry method was developed for the enantioseparation of the racemic mixture of pomalidomide, a novel, second-generation immunomodulatory drug, using β-cyclodextrin-bonded stationary phases. Four cyclodextrin columns (β-, hydroxypropyl-β-, carboxymethyl-β-, and sulfobutyl-β-cyclodextrin) were screened and the effects of eluent composition, flow rate, temperature, and organic modifier on enantioseparation were studied. Optimized parameters, offering baseline separation (resolution = 2.70 ± 0.02) were the following: β-cyclodextrin stationary phase, thermostatted at 15°C, and mobile phase consisting of methanol/0.1% acetic acid 10:90 v/v, delivered with 0.8 mL/min flow rate. For the optimized parameter at multiple reaction monitoring mode 274.1-201.0 transition with 20 eV collision energy and 100 V fragmentor voltage the limit of detection and limit of quantitation were 0.75 and 2.00 ng/mL, respectively. Since enantiopure standards were not available, elution order was determined upon comparison of the circular dichroism signals of the separated pomalidomide enantiomers with that of enantiopure thalidomide. The mechanisms underlying the chiral discrimination between the enantiomers were also investigated. Pomalidomide-β-cyclodextrin inclusion complex was characterized using nuclear magnetic resonance spectroscopy and molecular modeling. The thermodynamic aspects of chiral separation were also studied.

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It was previously reported that 6β-aminomorphinan derivatives show high affinity for opiate receptors. Novel 6β-heteroarylamidomorphinanes were designed based on the MOR selective antagonist NAP. The 6β-aminomorphinanes were prepared with stereoselective Mitsunobu reaction and subsequently acylated with nicotinic acid and isonicotinic acid chloride hydrochlorides. The receptor binding and efficacy were determined in vitro and the analgesic activity was studied in vivo. The in vitro studies revealed moderate selectivity for MOR. At least two compounds in this series exhibited long-acting analgesic response when administered subcutaneously and intracerebroventricularly. When the substances were given intracerebroventricularly to mice, they showed analgesic potency comparable to morphine.

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