Characterization of a novel degradation product of 2,2'-dithiobis[N-isoleucylbenzamide], an inhibitor of HIV nucleocapsid protein zinc fingers.
Zinc finger motifs have been found to be important in a variety of protein structures including transcription factors and viral nucleocapsid proteins. Recently, it was demonstrated that various aromatic disulfides effectively remove the metal ion from the zinc finger, resulting in an alteration of tertiary structure in this region of the protein, thereby inhibiting transcription. Among these compounds, 2,2'-dithiobis[N-isoleucylbenzamide] exhibits activity against human immunodeficiency virus (HIV)-type 1 in vitro and has been selected for preclinical development as an anti-HIV agent. Analysis of this agent by reversed-phase high-performance liquid chromatography (HPLC) indicated a significant quantity of two additional compounds. Identifying the parent disulfide was accomplished by scanning eluting peaks with positive ion thermospray ionization (TSP) mass spectrometry (MS). Solution-induced disproportionation of the disulfide into its sulfhydryl monomer was demonstrated by treating the drug with dithiothreitol (DTT) prior to HPLC analysis. TSP-MS analysis of the remaining chromatographic peak suggested a molecular weight of 265, which, with 1H-nuclear magnetic resonance (NMR) data of the isolated material, allowed us to elucidate the chemical structure as N-isoleucyl-benzisothiazolone. Contact with stainless steel, such as that employed in an HPLC system, was found to accelerate degradation of the parent disulfide to the benzisothiazolone.
Laboratory of Drug Discovery, Research, and Development, Developmental Therapeutics Program, Division of Cancer Treatment and Diagnosis, National Cancer Institute, Frederick, MD 21701, USA.
L R
LR
Phillips
L
L
Malspeis
E K
EK
Tubbs
J G
JG
Supko
D019380:Anti-HIV Agents / Q000032:analysis* / Q000494:pharmacology; D001549:Benzamides / Q000032:analysis* / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D007532:Isoleucine / Q000031:analogs & derivatives / Q000032:analysis* / Q000494:pharmacology; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D013056:Spectrophotometry, Ultraviolet; D016335:Zinc Fingers
C416806:2,2'-dithio(N-isoleucylbenzamide); D019380:Anti-HIV Agents; D001549:Benzamides; D007532:Isoleucine