A novel PNA-monomer for recognition of thymine in triple-helix structures

Article abstract of DOI:10.1081/NCN-120022958

To expand the triplex recognition repertoire of Nucleic Acids, novel nucleobases that recognize thymine in a T-A base pair are still required. A novel conformationally constrained PNA-monomer (II) capable of binding T in a triplex motif was designed and s

Full text of DOI:10.1081/NCN-120022958

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A Novel PNA-Monomer for Recognition of Thymine in
Triple-Helix Structures
Anne Goldbech Olsen a c , Otto Dahl a & Peter E. Nielsen b
a Department of Chemistry , University of Copenhagen , Copenhagen Ø, Denmark
b Department of Medical Biochemistry and Genetics , The Panum Institute, University of
Copenhagen , Copenhagen N, Denmark
c Lab. II, Department of Chemistry , University of Copenhagen , Universitetsparken 5,
DK-2100, Copenhagen, Denmark
Published online: 31 Aug 2006.
To cite this article: Anne Goldbech Olsen , Otto Dahl & Peter E. Nielsen (2003) A Novel PNA-Monomer for Recognition
of Thymine in Triple-Helix Structures, Nucleosides, Nucleotides and Nucleic Acids, 22:5-8, 1331-1333, DOI: 10.1081/
NCN-120022958
To link to this article: http://dx.doi.org/10.1081/NCN-120022958
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NUCLEOSIDES, NUCLEOTIDES & NUCLEIC ACIDS
Vol. 22, Nos. 5–8, pp. 1331–1333, 2003
A Novel PNA-Monomer for Recognition of Thymine in
Triple-Helix Structures
1
2
Anne Goldbech Olsen,1, Otto Dahl, and Peter E. Nielsen
*
1Department of Chemistry, University of Copenhagen, Copenhagen Ø, Denmark
2Department of Medical Biochemistry and Genetics, The Panum Institute,
University of Copenhagen, Copenhagen N, Denmark
ABSTRACT
To expand the triplex recognition repertoire of Nucleic Acids, novel nucleobases
that recognize thymine in a T-A base pair are still required. A novel conforma-
tionally constrained PNA-monomer (II) capable of binding T in a triplex motif
was designed and synthesized in 7 steps starting from commercially available
dimethyl 2-oxoglutarate.
Triplex targeting of double-stranded DNA is essentially limited to homopurine
stretches. Therefore novel nucleobases that recognise C(-G) and especially T (-A) are
required. The known E-base PNA-monomer (I)[1] synthesized some years ago is able
to recognise T when incorporated into the Hoogsteen strand of a bis-PNA. But bind-
ing affinity is low which might be due to excessive flexibility and=or steric clash with
the 5-methyl group of thymine. E-base PNAs bind somewhat stronger to uracil than
thymine containing targets.[1] The new analogue (II) contains a double bond, thereby
introducing restricted flexibility in the linker. The hope is to preorganise the mono-
*Correspondence: Anne Goldbech Olsen, Lab. II, Department of Chemistry, University of
Copenhagen, Universitetsparken 5, DK-2100 Copenhagen, Denmark; Fax: þ45 35 52 02
12; E-mail: ago@kiku.edu.
1331
DOI: 10.1081/NCN-120022958
Copyright # 2003 by Marcel Dekker, Inc.
1525-7770 (Print); 1532-2335 (Online)
www.dekker.com

1332
Olsen, Dahl, and Nielsen
Figure 1. Structures of the known E-base PNA-monomer[1] (I) and the new analogue (II).
mer in a conformation more favourable for binding to thymine. The binding motif is
the same for both monomers. The NH hydrogen functions as a donor and binds the
4-oxo group of thymine.
RESULTS AND DISCUSSION
In order to prepare the conformationally constrained PNA-monomer (8) com-
mercially available dimethyl 2-oxoglutarate (1) was chosen as starting material.
The first step was a ring closure to the known pyridazinone (2)[2] using hydrazine
and acid catalysis in methanol. To improve hydrophobicity compound 2 was reacted
with p-methoxybenzyl chloride and sodium hydride in DMF to obtain the PMB pro-
tected product (3). The ester functionality of 3 was subsequently reduced to the pri-
mary alcohol (4) using sodium borohydride in a refluxing mixture of THF and
methanol.[3] Some cleavage of the ester was also observed due to the alkaline condi-
tions, but to a much lesser extent than when pure hydroxylic solvent was used. Treat-
ment of compound 4 with activated manganese(IV) oxide in refluxing toluene
afforded a tandem reaction where the primary alcohol was first oxidised to the alde-
hyde and next an oxidative aromatisation took place to give compound 5. The reduc-
tion-oxidation strategy to obtain the aldehyde was chosen since an earlier attempt
with direct Dibal-H reduction of the ester was unsuccessful. The key step was a
Wittig reaction between aldehyde (5) and 2-carboxyethyltriphenylphosphonium bro-
mide.[4] The reaction took place in a 1:1 mixture of THF and DMSO using sodium
hydride as base and afforded only the desired E-isomer (6) as expected.[5] Compound
6 was then condensed with methyl N-(2-Boc-aminoethyl)glycinate[6] using DCC
and DHbtOH as coupling reagents. Basic hydrolysis of the resulting ester (7) gave
the monomer 8. Deprotection of p-methoxybenzyl is expected to take place under
the acidic conditions required for cleavage of the other protecting groups when
incorporated in PNA.

Novel PNA-Monomer
1333
Scheme 1. Reagents, conditions and yields: a) NH2NH2, ACOH, MeOH, reflux (92%); b) p-
methoxybenzylchloride, NaH, DMF, 0ꢀC- > RT (84%); c) NaBH4, THF, MeOH, reflux
(58%); d) activated MnO2, toluene, reflux (30%); e) [P(Ph)3(CH2)2COOH]þBrꢁ, NaH,
THF=DMSO, 0ꢀC- > RT (42%); f) methyl N-(2-Boc-aminoethyl)glycinate, DhbtOH, DCC,
DMF, 0ꢀC- > RT (50%); g) 2M NaOH, MeOH, 0ꢀC (46%).
CONCLUSION
The synthesis of a novel PNA-monomer has been accomplished. The triplex
recognition properties of this E-base analogue is currently under investigations.
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57, 1948–1953.
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