ENZYMATIC PRODUCTION OF OPTICALLY PURE (2'R-CIS)-2'-DEOXY-3'-THIACYTIDINE (3TC, LAMIVUDINE) - A POTENT ANTI-HIV AGENT

Although equipotent in terms of antiviral activity, the two enantiomer s of 2'-deoxy-3'-thiacytidine (BCH 189) differ markedly in their cytot oxicity.1-4 (2'R-cis)-2'-deoxy-3'-thiacytidine (3TC) is substantially less toxic than its optical antipode, and is undergoing development fo r the therapy of HIV infection. Cytidine deaminase from Escherichia co li is shown here to deaminate 2'-deoxy-3'-thiacytidine enantioselectiv ely to leave 3TC essentially optically pure. This reaction has been us ed to develop a process for production of 3TC in multikilogram amounts . The production of cytidine deaminase was enhanced by strain improvem ent, fermentation development, and finally by cloning and overexpressi on of the gene. The enzyme was immobilized on Eupergit-C, which allowe d it to be reused many times. The biotransformation conditions were op timized so that the best use could be made of the catalyst. A robust s caleable product isolation process was developed to yield the crystall ine product. Overall, yields through the resolution process of 76% wer e obtained. All aspects of this process are capable of substantial fur ther scaleup with only minor modifications.