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Efficient Biocatalytic Synthesis of Dihalogenated Purine Nucleoside Analogues Applying Thermodynamic Calculations

Authors
  • Yehia, Heba1, 2
  • Westarp, Sarah1, 3
  • Röhrs, Viola4
  • Kaspar, Felix1, 3
  • Giessmann, Robert T.1
  • Klare, Hendrik F.T.
  • Paulick, Katharina1
  • Neubauer, Peter1
  • Kurreck, Jens4
  • Wagner, Anke1, 3
  • 1 (P.N.)
  • 2 Chemistry of Natural and Microbial Products Department, Pharmaceutical and Drug Industries Research Division, National Research Centre, Dokki, 12622 Cairo, Egypt
  • 3 BioNukleo GmbH, Ackerstr. 76, 13355 Berlin, Germany
  • 4 (J.K.)
Type
Published Article
Journal
Molecules
Publisher
MDPI AG
Publication Date
Feb 19, 2020
Volume
25
Issue
4
Identifiers
DOI: 10.3390/molecules25040934
PMID: 32093094
PMCID: PMC7070685
Source
PubMed Central
Keywords
License
Green

Abstract

The enzymatic synthesis of nucleoside analogues has been shown to be a sustainable and efficient alternative to chemical synthesis routes. In this study, dihalogenated nucleoside analogues were produced by thermostable nucleoside phosphorylases in transglycosylation reactions using uridine or thymidine as sugar donors. Prior to the enzymatic process, ideal maximum product yields were calculated after the determination of equilibrium constants through monitoring the equilibrium conversion in analytical-scale reactions. Equilibrium constants for dihalogenated nucleosides were comparable to known purine nucleosides, ranging between 0.071 and 0.081. To achieve 90% product yield in the enzymatic process, an approximately five-fold excess of sugar donor was needed. Nucleoside analogues were purified by semi-preparative HPLC, and yields of purified product were approximately 50% for all target compounds. To evaluate the impact of halogen atoms in positions 2 and 6 on the antiproliferative activity in leukemic cell lines, the cytotoxic potential of dihalogenated nucleoside analogues was studied in the leukemic cell line HL-60. Interestingly, the inhibition of HL-60 cells with dihalogenated nucleoside analogues was substantially lower than with monohalogenated cladribine, which is known to show high antiproliferative activity. Taken together, we demonstrate that thermodynamic calculations and small-scale experiments can be used to produce nucleoside analogues with high yields and purity on larger scales. The procedure can be used for the generation of new libraries of nucleoside analogues for screening experiments or to replace the chemical synthesis routes of marketed nucleoside drugs by enzymatic processes.

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