A new protecting group for the exocyclic amino groups of nucleosides

Article abstract of DOI:10.1016/j.tetlet.2005.01.028

A new protecting group has been developed for the exocyclic amino groups of nucleosides that occur in DNA. 3-Phenyl-[{N-(2-trimethylsilyl-ethoxycarbonyl)- 2-amino}]-propanoic acid used as the protective agent.

Full text of DOI:10.1016/j.tetlet.2005.01.028

Tetrahedron
Letters
Tetrahedron Letters 46 (2005) 2163–2165
A new protecting group for the exocyclic amino
groups of nucleosides
Chamakura V. N. S. Varaprasad*, and Francis Johnson
Departments of Chemistry and Pharmacological Sciences, State University of New York at Stony Brook,
Stony Brook, NY 11794-3400, USA
Received 18 November 2004;revised 26 December 2004;accepted 7 January 2005
Abstract—A new protecting group has been developed for the exocyclic amino groups of nucleosides that occur in DNA. 3-Phenyl-
[{N-(2-trimethylsilyl-ethoxycarbonyl)-2-amino}]-propanoic acid used as the protective agent.
ꢀ 2005 Elsevier Ltd. All rights reserved.
Currently modified synthetic DNA, required for DNA
damage and mutagenic studies1 is obtained essentially
in one of two ways, either by a presynthetic or a post-
synthetic strategy. Nevertheless, at present, most modi-
fied DNA is prepared by the former method in which
the modified base is pre-synthesized in a protected form2
and then introduced into the oligomeric chain either by
solution-based methods or by an automated resin-based
procedure.3 In a few instances postsynthetic strategy4
was used successfully whereas more rarely a combina-
tion of both of these strategies has been employed.
The latter approach, for example, was adopted to in-
corporate both the base-labile acrolein adduct of deoxy-
guanosine5a and the abasic site5b into DNA.
The deprotected base because of its greater reactivity
than the other protected bases will react with a suitable
electrophilic reagent to give the desired modified nucle-
oside residue. Unfortunately few amino protective
groups can be removed selectively while the DNA is still
attached to resin. The amino protective groups that are
currently in use in the deoxynucleoside series are base la-
bile and are uniformly removed by treatment either with
ammonium hydroxide at 55 ꢁC for 16 h or methylamine7
at 23 ꢁC. This allows no discrimination amongst the
amino groups available for site-specific modification.
3-(2-Aminoaryl)-propionamides are a class of com-
pounds, which disintegrate in a facile manner by internal
cyclization to the corresponding free amino residue and
a lactam. Because of their spontaneous internal cycliza-
tion8 the 2-amino group in 3-(2-aminoaryl)-propion-
amides is always generated from the corresponding
3-(2-nitroaryl)-propionamides. The nitro group serves
as a masked amino group until itÕs reduced to the corre-
sponding amino group either under neutral,6 acidic,9a
basic,9b reductive hydrogenation9c or bioreductive9d
conditions. For the first time, however, we were able
to devise a method to introduce the above 2-amino
In our initial studies towards the synthesis of modified
DNA we developed a protecting group, 1 (Fig. 1), which
would be resistant to the normal basic conditions used
in the deprotection of synthetic DNA.6 Although this
strategy was highly successful in cases where the oligo-
mer contained only two dG residues, its incomplete
resistance to alkali left it wanting in examples where
more than two dG residues were present. An alternative
strategy that might be used to obtain a site-specifically
modified DNA by a postsynthetic strategy is to
deprotect site-specifically an exocyclic amino group in
a nucleoside while the DNA is still attached to resin.
NO2
O
NHR
Keywords: Deoxy nucleosides;Protecting groups.
*
NO2
Corresponding author. Tel.: +1 714 545 0100;fax: +1 714 641
7233;e-mail: vchamakura@valeant.com
1
Present address: Valeant Pharmaceuticals International, 3300 Hyland
Avenue, Costa Mesa, CA 92626, USA.
Figure 1.
0040-4039/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2005.01.028

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C. V. N. S. Varaprasad, F. Johnson / Tetrahedron Letters 46 (2005) 2163–2165
group in a carbamate form, in situ, capitalizing on Cur-
tius rearrangement of the corresponding benzoyl azide
in presence of the desired alcohol.
B-NHCOR
B-NH2
HO
HO
7, Transient protection
O
O
conditions
Herein we describe the synthesis of 3-phenyl-[{N-(2-
trimethylsilyl-ethoxycarbonyl)-2-amino}]-propanoic acid
(PTAP, 2), which has several salient characteristics,
namely, (i) it is readily prepared in four steps in overall
high yield (Scheme 1);(ii) using this agent the three
deoxynucleosides dC, dA and dG were derivatized un-
der transient protection conditions in a facile manner
and (iii) it is deprotected by anhydrous terabutlyammo-
nium fluoride (TBAF) in a cascade of spontaneous steps
that culminate in the free nucleoside and co-generation
of lactam 14 (Scheme 2). Only one other example of
the use of this type of protective group has been re-
ported to protect specifically the exocyclic amino groups
of DNA nucleosides, that of by Van Boom and co-
workers10 in which the 2-(t-butylphenylsilyloxymethyl)-
benzoyl group was used. This was subsequently
removed by fluoride ion. To our knowledge this kind of
strategy using protected 3-(2-aminoaryl)-propionamides
has never been applied in the protection of exocyclic
amino groups of DNA nucleosides.
OH
OH
Anhy. TBAF, DMSO,
50oC, 1-2h
8: B = C
9: B = A
11: B = C
12: B = A
13: B = G
10: B = G
R =
O
N
N
H
14
O
H
TMS
O
Scheme 2.
The acid chloride derivative 7 of the acid 2 was prepared
by means of SOCl2 (1 equiv, 23 ꢁC, 2 h) in the presence
of NEt3 (1 equiv) in dichloromethane. The acid chloride
was then used to derivatise12 the nucleosides 8, 9 and 10
as the corresponding N-acyl derivatives13 11, 12 and 13
(yields: 60–65%) by following the standard transient
protection method.14 Interestingly, unlike in the other
reported methods10 we didnÕt observe any di-substituted
derivatives with dA and dG.
The desired acid 2 was prepared from the readily avail-
able and inexpensive 2-carboxybenzaldehyde, 3. This
was treated with commercially available (carbethoxy-
methylene)-triphenylphosphorane first at 0 ꢁC and
thereafter with stirring at 23 ꢁC for 4 h to obtain the
geometrically isomeric mixture of the ethyl esters 4.
The crucial carbamate 5 was obtained in a one-pot Cur-
tius rearrangement11 from 4 by treatment with diphenyl-
phosphoryl azide (DPPA), NEt3 and 2-trimethylsilyl-
ethanol in dioxane at 80–90 ꢁC for 5 h, in 89% yield after
silica gel chromatography. Catalytic hydrogenation (H2,
10% Pd/C, 50 psi) of the product mixture 5 then gave the
corresponding saturated ester 6, which on mild alkaline
hydrolysis with LiOH (10 equiv, DME:H2O:: 1:2) at
23 ꢁC for 5 h afforded the target acid 2 in 90% yield.
The deprotection15 of nucleosides 11, 12 and 13 was car-
ried out by treatment with anhydrous TBAF16 (3 equiv)
in DMSO for 2 h at 50 ꢁC. This cleanly regenerated the
corresponding nucleosides 8, 9 and 10 in excellent yields
accompanied by the lactam 14. The cyclization of 2-
aminoarylamide released upon deprotection of amino
group was instantaneous and it didnÕt need any assis-
tance from the methyl groups alpha to the carbonyl
group as suggested in earlier literature.9c The appear-
ance of the lactam 14 can be measured spectrophoto-
metrically to ascertain the extent of deprotection.6
Further work is in progress to utilize 2 as a protec-
tive agent in the synthesis of site-specifically modified
DNA.
OH
O
O
H
In conclusion we have developed a new protective group
for the exocyclic amino groups of deoxy nucleosides. We
believe that this protective group will find general use
for the protection of the amino functionality wherein
the conventional methods are ineffective.
a
OEt
H
O
O
HO
3
4
b
Acknowledgements
O
O
H
c
X
We thank both NIEHS (Grant ES 04068) and NCI
(Grant CA17395) for their support of this work.
OEt
H
NH
O
NH
O
Si(CH3)3
Si(CH3)3
O
O
6: X = OEt
2: X = OH
7: X = Cl
Supplementary data
d
5
e
The supplementary data including 1H NMR and EI-MS
or FAB-MS data for compounds 6, 11, 12, 13 is avail-
able online with the paper in Science Direct. Supplemen-
tary data associated with this article can be found, in the
online version, at doi:10.1016/j.tetlet.2005.01.028.
Scheme 1. Reagents and conditions: (a) Ph3P@CHCO2Et (1.0 equiv),
THF, 4 h, 23 ꢁC;(b) DPPA, NEt 3, dioxane, 85 ꢁC, 5 h;(c) H 2 (50 psi),
10% Pd/C (w/w), EtOH, 23 ꢁC, 4 h;(d) LiOH (10.0 equiv), DME–H 2O
(1:1), 23 ꢁC, 5 h;(e) SOCl 2, (1.0 equiv), NEt3, (1.5 equiv), 23 ꢁC, 2 h.

C. V. N. S. Varaprasad, F. Johnson / Tetrahedron Letters 46 (2005) 2163–2165
2165
10. Dreef-Tromp, C. M.;Hoogerhout, P.;Van der Marel, G.
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