Synthesis of 5-ethynyl-2′-deoxyuridine-5′-boranomono phosphate as a potential thymidylate synthase inhibitor

Article abstract of DOI:10.1081/NCN-200060046

The 5-ethynyl-2′-deoxyuridine nucleoside and the 5′- boranomonophosphate nucleotide were synthesized as analogs of 5-fluoro-2′-deoxyuridine monophosphate (5-FdUMP), a widely used mechanism-based inhibitor of thymidylate synthase. Synthesis was carried out

Full text of DOI:10.1081/NCN-200060046

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SYNTHESIS OF 5-ETHYNYL-2′-DEOXYURIDINE-5′-
BORANOMONO PHOSPHATE AS A POTENTIAL
THYMIDYLATE SYNTHASE INHIBITOR
Shoeb I. Khan a , Mikhail I. Dobrikov a & Barbara Ramsay Shaw a
a Department of Chemistry, P. M. Gross Chemical Laboratory, Duke University, Durham, North
Carolina, USA
Published online: 15 Nov 2011.
To cite this article: Shoeb I. Khan , Mikhail I. Dobrikov & Barbara Ramsay Shaw (2005) SYNTHESIS OF 5-ETHYNYL-2′-
DEOXYURIDINE-5′-BORANOMONO PHOSPHATE AS A POTENTIAL THYMIDYLATE SYNTHASE INHIBITOR, Nucleosides, Nucleotides
and Nucleic Acids, 24:5-7, 1047-1049, DOI: 10.1081/NCN-200060046
To link to this article: http://dx.doi.org/10.1081/NCN-200060046
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Nucleosides, Nucleotides, and Nucleic Acids, 24 (5–7):1047–1049, (2005)
Copyright D Taylor & Francis, Inc.
ISSN: 1525-7770 print/ 1532-2335 online
DOI: 10.1081/NCN-200060046
SYNTHESIS OF 5-ETHYNYL-2’-DEOXYURIDINE-5’-BORANOMONO
PHOSPHATE AS A POTENTIAL THYMIDYLATE SYNTHASE INHIBITOR
5
Shoeb I. Khan, Mikhail I. Dobrikov, and Barbara Ramsay Shaw
Department of
Chemistry, P. M. Gross Chemical Laboratory, Duke University, Durham, North Carolina, USA
5
The 5-ethynyl-2’-deoxyuridine nucleoside and the 5’-boranomonophosphate nucleotide were
synthesized as analogs of 5-fluoro-2’-deoxyuridine monophosphate (5-FdUMP), a widely used
mechanism-based inhibitor of thymidylate synthase. Synthesis was carried out from protected 5-iodo-2’-
deoxyuridine and trimethylsilylacetylene by Sonogashira palladium-catalyzed cross coupling reaction
followed by selective phosphorylation and finally boronation.
Keywords Oligonucleotides, Thymidylate Synthase, Boranomonophosphate
INTRODUCTION
Nucleoside analogs with modifications in the base, sugar, or phosphate residue
can be used as probes for substrate properties, kinetic pathways, and transition
states of enzymes involved in nucleic acid metabolism. 5-Fluorouracil (FU) and its
derivative, 5-fluoro-2’-deoxyuridine monophosphate (5-FdUMP),[1] are strong
inhibitors of thymidylate synthase (TS), an enzyme responsible for de novo
synthesis of DNA thymidine and a major enzyme target for cancer therapeutics.
Intrinsic and acquired drug resistance is a limitation of cancer chemotherapy using
5-fluoropyrimidines, which reduce the response rates. It is thus worthwhile to
synthesize new analogs and their prodrugs combining the modifications at the
phosphate center and C5 pyrimidine position. The substitution of a non-bridging
oxygen in a phosphate ester group by a borane (BH3) results in a class of phosphate
modified nucleotides, e.g., boranomonophosphate.[2] Because the BH3 group is
isoelectronic with oxygen and isosteric with a methyl group, it shares a number of
chemical and biochemical properties such as good aqueous solubility, increased
lipophilicity, and phosphatase resistance. Boranophosphates of nucleosides and
Received 15 October 2004, accepted 19 January 2005.
This research was supported by NIH grant 2RO1-GM57693 to B.R.S.
Address correspondence to Barbara Ramsay Shaw, Department of Chemistry, P. M. Gross Chemical
Laboratory, Duke University, Durham, NC 27708, USA; E-mail: brshaw@chem.duke.edu
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S. I. Khan, M. I. Dobrikov, and B. R. Shaw
SCHEME 1 Synthesis of 5-ethynyl-3’-O-TBDPS-2’-deoxyuridine.
oligonucleotides have recently attracted attention due to their possible therapeutic
applications as anticancer and antiviral[3,4] prodrugs and antisense agents.[5] The
aim is to design and synthesize nucleotide prodrugs combining the best nucleoside
inhibitors with the advantages of a boranophosphate modification. We hypothesize
that nucleoside boranomonophosphates should resist further phosphorylation by
nucleoside monophosphate kinase and dephosphorylation by alkaline phosphatase,
and thus will increase the effective concentration of the active (monophosphate)
drug in the cell.
5-Ethynyl-3’-O-TBDPS-2’-deoxyuridine nucleoside 4 was synthesized following
Scheme 1 in 75% yield. Commercially available 5-iodo-2’-deoxyuridine was
selectively protected at the 5’- and 3’-hydroxyls by trityl 1 and t-butyldiphenylsilyl 2,
respectively. The palladium mediated cross-coupling reaction was carried out using
the Sonogashira method to obtain fully protected nucleoside, which after
purification, was treated with sodium methoxide to deprotect the trimethylsilyl
group to obtain 3 in 77% yield.
The nucleoside 3 was treated with formic acid and ether to remove the 5’-O-
trityl to obtain the 3’-TBDPS protected nucleoside 4 in 75% yield. Finally,
nucleoside 4 was phosphorylated by biscyanoethyldiisopropylphosphoramidite
using tetrazole in CH3CN as an activator followed by boronation by borane-
dimethylsulfide complex, as shown in Scheme 2 to obtain compound 5 in 71%
yield. The 3’-silyl protecting group was removed by TEA:3HF to obtain nucleotide
SCHEME 2 Synthesis of 5-ethynyl-2’-deoxyuridine-5’-boranomonophosphate.

S. I. Khan, M. I. Dobrikov, and B. R. Shaw
1049
6 in 67% yield. Compound 6 on treatment with NH4OH/MeOH solution gave
5-ethynyl-2’-deoxyuridine-5’-borano monophosphate 7 in good yield.
In conclusion, of several approaches (not shown) tried to obtain 7, the
approaches described in Schemes 1 and 2 were found to be optimal for synthesis of
the 5-ethynyl-2’-deoxyuridine-5’-boranomonophosphate. Biscyanoethyldiisopropyl-
phosphoramidite was the best phosphorylating reagent to obtain the final
boranomonophosphate nucleoside 7.
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1. Kalman, T.I.; Nie, Z. Rational design of selective antimetabolites of DNA-thymine biosynthesis: 5-
propynylpyrimidine nucleoside derivatives. Nucleosides Nucleotides Nucleic Acids 2000, 19, 357–369.
2. Shaw, B.R.; Dobrikov, M.; Wang, X.; Wan, J.; He, K.; Lin, J.; Li, P.; Rait, V.; Sergueeva, Z.A.; Sergueev, D.
Reading, writing, and modulating genetic information with boranophosphate mimics of nucleotides, DNA, and
RNA. Ann. N. Y. Acad. Sci. 2003, 1002, 12–28.
3. Dobrikov, M.; Sergueeva, Z.A.; Shaw, B.R. Incorporation of (a-P-borano)-2’,3’-dideoxycytidine-5-triphosphate
into DNA by drug-resistant MMLV reverse transcriptase and Taq DNA polymerase. Nucleosides Nucleotides
Nucleic Acids 2003, 22, 1651–1655.
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dideoxynucleotide resistance by K65R human immunodeficiency virus reverse transcriptase using an
a-boranophosphate nucleoside analogue. J. Biol. Chem. 2001, 276, 48466–48472.
5. Wang, X.; Dobrikov, M.; Sergueeva, D.; Shaw, B.R. RNase H activation by stereoregular boranophosphate
oligonucleotide. Nucleosides Nucleotides Nucleic Acids 2003, 22, 1151–1153.