2,3-Dicyclohexylsuccinimide as a directing/protecting group for the regioselective glycosylation or alkylation of purines

Article abstract of DOI:10.1039/c3cc37265k

Here we describe the synthesis and application of a novel 2,3-dicyclohexylsuccinimide (Cy₂SI) protecting group towards regioselective purine glycosylation and alkylation reactions. This bulky protecting group promotes high regioselectivity during the glycosylation (as well as diastereoselectivity) or alkylation of purines using Hoffer's chlorosugar or tert-butyl bromoacetate, respectively. Cy₂SI offers the additional synthetic advantage that other base-labile protecting groups, such as toluoyl esters, can be selectively removed in its presence without affecting the imide. The Royal Society of Chemistry 2013.

Full text of DOI:10.1039/c3cc37265k

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2,3-Dicyclohexylsuccinimide as a directing/protecting
group for the regioselective glycosylation or alkylation
of purines†
Cite this: Chem. Commun., 2013,
49, 2936
Received 4th October 2012,
Accepted 25th February 2013
Ayan Pal, Kerry J. Salandria, Joseph W. Arico, Mark K. Schlegel* and
Larry W. McLaughlin*
DOI: 10.1039/c3cc37265k
www.rsc.org/chemcomm
Here we describe the synthesis and application of
a novel site. One disadvantage of these reagents is that the purine
2,3-dicyclohexylsuccinimide (Cy2SI) protecting group towards substrates must be prepared from their corresponding oxopur-
regioselective purine glycosylation and alkylation reactions. This ines or chloropurines, thereby creating additional synthetic
bulky protecting group promotes high regioselectivity during the steps. Moreover, studies in our group have demonstrated that
glycosylation (as well as diastereoselectivity) or alkylation of displacement of the imidazole/triazole derivative of 3-deazapurine
purines using Hoffer’s chlorosugar or tert-butyl bromoacetate, nucleosides was difficult.8
respectively. Cy2SI offers the additional synthetic advantage that
We have previously described the use of 2,2,3,3-tetramethyl-
other base-labile protecting groups, such as toluoyl esters, can be succinimide (M4SI) protected aminopurines during nucleoside
selectively removed in its presence without affecting the imide.
synthesis.8 This imide was quite effective in functioning both as
a protecting group for the exocyclic C6-amino group, as well as
Convergent approaches to synthesize nucleoside derivatives a directing group to favor regioselective glycosylation at the N9
require a glycosylation event to couple the sugar and heterocycle position of purines. Although we anticipated the corresponding
components.1 Typically, purine 20-deoxynucleosides are prepared phthalimide (NPhth) would function similarly, the crystal
via the reaction of a 6-chloropurine heterocycle with 2-deoxy-3,5-di- structure of adenyl-N6-tetramethylsuccinimide suggested that
O-(p-toluoyl)-a-D-erythro-pentofuranosyl chloride (Hoffer’s chloro- it is not the imide carbonyl that blocks access to the N7 of the
sugar)2,3 employing the sodium salt method.4 In this method, the purine substrate, but rather the methyl groups present in
anion of the purine base is generated at the tautomerically M4SI.8 The phthalimide, lacking such methyl groups, was much
equivalent N9 and N7 positions, which commonly results in the less effective at directing the products towards the desired N9
presence of both N9 and N7 glycosylation products in the final regioisomer, especially when using modified purine acceptors.
product mixture. The use of 6-chloropurine as an acceptor results
With this in mind, we wanted to determine whether increasing
in a regioselectivity that favors the desired b-N9 product by an the steric bulk above the purine N7 would further optimize the
approximate factor of five.1 This ratio, however, is reversed in the directing ability of substituted succinimide derivatives. Although a
case of 3-substituted purines such as 6-chloro-3-deaza-3-methyl- variety of protecting groups could be envisioned, we chose the
purine in which the formation of N7 products is favored.5 bicyclohexyl derivative 3 owing to the readily available starting
An additional problem is that many purine acceptors typically material. We could easily convert 1,1-azobis(cyclohexanecarbo-
generate notable quantities of the a-nucleoside products, further nitrile) (1) to the corresponding bicyclohexyl-1,10-dinitrile (2)
reducing yields and complicating purifications.1
(Scheme 1) in 89% yield under refluxing conditions in n-heptane.
Previous work by Robins and colleagues has described the use
of purines bearing a 2-alkylimidazole6,7 or triazole7 in the 6-position
of the purine heterocycle to control regio- and diastereoselectivity
during glycosylation reactions. It was proposed that the 2-alkyl
substituent covers the area over the N7 position of the purine
heterocycle, sterically blocking access of the glycosyl donor to that
Boston College, Department of Chemistry, Merkert Chemistry Center, 2609 Beacon
Street, Chestnut Hill, MA 02467, USA. E-mail: larry.mclaughlin@bc.edu,
mark.schlegel@bc.edu; Fax: +001 617 552 2705; Tel: +001 617 552 3622
† Electronic supplementary information (ESI) available: Synthetic procedures,
analytical data, oligonucleotide synthesis. CCDC 904677. For crystallographic
data in CIF or other electronic format see DOI: 10.1039/c3cc37265k
Scheme 1 Preparation of 2,3-dicyclohexylsuccinic anhydride and the subse-
quent protection of adenine or 3-deaza-3-methyladenine.
c
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Table 1 Regioselectivity and isolated yields for glycosylation and alkylation reactions
increased as the amount of steric bulk was increased in the
cyclo-imide protecting group; the use of Cy2SI significantly
improved the isolated yield of the desired b-N9 product. Clearly,
one big advantage of using Cy2SI protection during glycosylation
was that the resulting product mixture was more homogeneous,
and its purification therefore much simpler, than a product
mixture which potentially contained a complex blend of four
compounds (N9/N7 regioisomers, a/b anomers).
Encouraged by those results, we wanted to investigate the
influence of Cy2SI protection on glycosylation reactions of purine-
like substrates in which the N3 in the purine heterocycle is replaced
with a C–CH3 (substrate set ‘‘B’’). It had been previously shown that
the introduction of a methyl group at the C3 position had a
dramatic effect on both product ratios and isolated yields.8 While
the b-N9 : b-N7 ratio was found to favor the undesired b-N7
regioisomer for the Ndmf, 6-chloro, and NPhth protected deriva-
tives (Entries 6–8), the use of the M4SI-protected derivative (Entry 9)
provided a regioselectivity (6.4 : 1) which favored the desired b-N9
regioisomer. Interestingly, the Cy2SI-protected derivative performed
substantially better with a regioselectivity of 20 : 1 during this
reaction (Entry 10). Moreover, the isolated yield of the desired
product was significantly higher with the Cy2SI derivative when
compared to the use of M4SI (85% vs. 59%).
Glycosylation reactions
% Yield
Alkylation reactions
% Yield
Purine
substrate
Entry b-N9 : N7b b9 (other)c Entry N9 : N7b N9 (other)c
A
R = Ndmf a
R = Cl
1
0.8 : 1
4.7 : 1
18e (48)
57 (13)
36 (0)
14
0.7 : 1 30 (44)
2d
R = NPhth
R = M4SI
R = Cy2SI
R = Ndmf
R = Cl
3d >99 : 1
4d >99 : 1
71 (6)
5
>99 : 1
0.7 : 1
0.6 : 1
0.9 : 1
6.4 : 1
20 : 1
0.7 : 1
7 : 1
94 (3)
15
16
15 : 1 78 (5)
B
6
6e (65)
39 (61)
15 (47)
59 (27)
85 (12)
15e (35)
57 (21)
82 (10)
0.5 : 1
0 (44)
7d
8d
9d
R = NPhth
R = M4SI
R = Cy2SI
R = Ndmf
R = M4SI
R = Cy2SI
10
17
18
12 : 1 75 (6)
0.8 : 1 0 (52)
C
11
12
13
Acceptors containing more bulk attached to the C3 position
(formerly N3 of adenine) of the substrate would appear to offer a
more significant challenge under these glycosylation conditions
by also sterically blocking the N9 position of the heterocycle
(substrate set ‘‘C’’). However, this group did not appear to have
much of an overall effect. While the reactions with both M4SI
16 : 1
19
10 : 1 71 (8)
a
b
For synthesis, see ref. 10. Determined using 1H NMR integration on
c
d
e
the crude products. Isolated yields. See ref. 8. Determined using
1H NMR integration on an isolated mixture of b-N9 and b-N7 products.
The dinitrile 2 was then subjected to strongly acidic conditions9 to and Cy2SI protection (Entries 12–13) showed a similar trend to
form the corresponding cyclic anhydride 3 in 67% yield. that of the 3-deaza-3-methyladenine series (Substrate set ‘‘B,’’
Introduction of this Cy2SI protecting group to a series of Entries 9–10), those using Cy2SI protection showed a major
6-amino purines was accomplished in a single step by simply improvement in both regioselectivity (16: 1 vs. 7 : 1) and isolated
refluxing the purine, for example 4, cyclic anhydride 3, and yield (82% vs. 57%). Furthermore, it was also noteworthy that the
DBU in pyridine to afford compound 6 in 89% yield. The crystal purine heterocycles protected using Cy2SI produced significantly
structure of compound 7 illustrates that the two cyclohexyl lower amounts of the undesired a-isomers across all three
groups position themselves directly above the N7 of the purine substrate sets during these glycosylation reactions.
base; specifically a methylene group from each of the cyclohexyl
To expand the utility of Cy2SI as a protecting/directing
moieties hinders access to the purine N7 (see ESI†). We envisioned group, we also chose to study the alkylation reactions of this
that this would make the use of Cy2SI more advantageous towards same set of purine substrates (Table 1). Studies have shown that
regioselective purine reactions than M4SI since the cyclohexyl alkylation reactions of purines also suffer from the production of
groups flank both sides at the top of the purine nucleobase rather N9 : N7 regioisomeric mixtures.10,11 The ability to perform regio-
than just one in the case of M4SI.8
selective alkylation of purines would be advantageous for the
To determine how effective Cy2SI was in promoting regio- production of N9 regioisomers in research fields that require them,
selective glycosylations using Hoffer’s chlorosugar, a series of such as peptide nucleic acids (PNA).12 A simple alkylation of
reactions was performed in acetonitrile using the sodium salt adenine, protected using Ndmf, with tert-butyl bromoacetate
method (Table 1). Glycosylation of adenine protected as the resulted in a 0.7 : 1 ratio of the N9 : N7 products (Entry 14). The
N-dimethylformamidine10 (Ndmf) resulted in a complex mixture use of Cy2SI as a protecting group resulted in a dramatic reversal of
of b-N9 : b-N7 isomers in a ratio of 0.8 : 1, as well as a significant this ratio, with the desired N9 regioisomer being produced in a
amount of the unwanted a isomers (Entry 1). Glycosylation using 15-fold excess and isolated in 78% yield (Entry 15). Alkylation
6-chloropurine (which can be converted to adenine) resulted in a reactions of purine-like substrates with modifications at the
slightly higher b-N9 : b-N7 ratio of 4.7 : 1, but still contained a fair 3-position of the heterocycle also followed the same trend observed
amount of a isomeric products (Entry 2). When the exocyclic in the corresponding glycosylation reactions; that is, Cy2SI protec-
amine of adenine was protected using phthalimide (NPhth), tion resulted in a 12 : 1 and 10 : 1 N9 : N7 ratio for substrates
M4SI, or Cy2SI cyclo-imide protecting groups, the desired b-N9 B and C, respectively (Entries 17 and 19). This was in stark contrast
products were formed in a greater than 99: 1 ratio with respect to to the corresponding reactions using Ndmf protection in which the
the b-N7 products (Entries 3–5). However, the isolated yields N7 regioisomer was formed in excess (Entries 16 and 18). It should
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Cy2SI protecting group from the purified oligos with [NH4OH], 7 M
NH3 in MeOH, or AMA (1 : 1 mixture of [NH4OH] and 40% aq.
methylamine) at 55 1C was unfortunately met with little success.
In summary, we have developed a new directing/protecting
group that provided an effective means to access 6-aminopurine-
20-deoxynucleosides with a high regioselectivity for the desired
b-N9 product over the b-N7 product. This Cy2SI protecting group
also provided this advantage to alkylation reactions in which the
N9 product was formed in a significant excess over the undesired
N7 product. Structural analysis using X-ray crystallography con-
firmed that portions of both cyclohexyl groups are positioned to
sterically block access to the N7 nitrogen, thereby promoting
reaction at the N9 position instead. This advantage became
especially apparent in the synthesis of valuable C3 substituted
purines in which the desired products could be easily isolated in
high yields.14 Since it has been previously shown that running
purine glycosylation reactions in less polar solvents can reduce
the number of a-nucleosides produced during the reaction,15
further development of directing/protecting groups is ongoing in
our laboratory.
Scheme 2 Selective removal of toluoyl and Cy2SI protecting groups.
We gratefully acknowledge funding from the NSF (MCB
0958515) for this research. We would also like to thank
Dr Han Yueh and Ms Alena Carlson for their generous donation
of synthetic intermediates.
Scheme 3 Synthesis of phosphoramidite 14.
also be noted that although both the N9 and N7 regioisomers
were detected in the crude 1H NMR spectra for the corres-
ponding heterocycles containing Ndmf protection (Entries 16
and 18), only the N7 product could be isolated via column
chromatography.
Notes and references
1 L. J. Wilson, M. W. Hager, Y. A. El-Katan and D. C. Liotta, Synthesis,
1995, 1465–1479.
2 M. Hoffer, Chem. Ber., 1960, 93, 2777–2781.
3 V. Rolland, M. Kotera and J. Lhomme, Synth. Commun., 1997, 27,
3505–3511.
4 Z. Kazimierczuk, H. B. Cottam, G. R. Revankar and R. K. Robins,
J. Am. Chem. Soc., 1984, 106, 6379–6382.
5 R. J. Irani and J. Santalucia, Nucleosides Nucleotides Nucleic Acids,
2002, 21, 737–751.
6 M. Zhong, I. Nowak, J. F. Cannon and M. J. Robins, J. Org. Chem.,
2006, 71, 4216–4221.
7 M. Zhong, I. Nowak and M. J. Robins, J. Org. Chem., 2006, 71,
7773–7779.
8 J. W. Arico, A. K. Calhoun, K. J. Salandria and L. W. McLaughlin,
Org. Lett., 2010, 12, 120–122.
9 C. G. Overberg, H. Biletch, P. T. Huang and H. M. Blatter, J. Org.
Chem., 1955, 20, 1717–1720.
The hydrolytic stability of the Cy2SI protecting group also
presented a synthetic advantage over other protecting groups.
After glycosylation, the toluoyl esters of the newly formed
nucleoside, for example 8, could be selectively removed using
7 M ammonia in methanol at ambient temperature to produce
compound 9 in 92% yield without affecting the Cy2SI group
(Scheme 2). Further reaction of compound 9 in concentrated
aqueous ammonium hydroxide at 55 1C completely removed
the imide protecting group and produced the free nucleoside
10 in 84% yield.
Encouraged by those results, we decided to test the feasi-
bility of using Cy2SI as a protecting group during DNA oligo- 10 K. Okumura, T. Oine, Y. Yamada, M. Tomie, T. Adachi, T. Nagura,
M. Kawazu, T. Mizoguchi and I. Inoue, J. Org. Chem., 1971, 36,
1573–1579.
11 (a) T. Fujii and T. Itaya, Heterocycles, 1998, 48, 1673–1724;
nucleotide synthesis. The synthesis of phosphoramidite 14
proceeded using well established procedures starting from
compound 11 (Scheme 3). After removal of the toluoyl esters
in 11 using 7 M NH3 in MeOH, the free 50-OH in 12 was
selectively protected using 4,40-dimethoxytrityl chloride in
(b) E. Enkvist, G. Raidaru, A. Uri, R. Patel, C. Redick, J. L. Boyer,
J. Subbi and I. Tammiste, Nucleosides Nucleotides Nucleic Acids, 2006,
25, 141–157; (c) Q. Zhang, G. Cheng, Y.-Z. Huang, G.-R. Qu, H.-Y. Niu
and H.-M. Guo, Tetrahedron, 2012, 68, 7822–7826.
pyridine to afford compound 13 in 84% yield. Compound 13 12 (a) P. E. Nielsen, Acc. Chem. Res., 1999, 32, 624–630; (b) P. E. Nielsen,
Chem. Biodiversity, 2010, 7, 786–804; (c) E. Uhlmann, A. Peyman,
G. Breipohl and D. W. Will, Angew. Chem., Int. Ed., 1998, 37,
2796–2823.
was then transformed into phosphoramidite 14 using 2-cyanoethyl
N,N,N0,N0-tetraisopropylphosphordiamidite and tetrazole in DCM
in 63% yield. With this phosphoramidite in hand, we were able 13 (a) M. K. Schlegel and E. Meggers, J. Org. Chem., 2009, 74,
4615–4618; (b) M. K. Schlegel, J. Hu¨tter, M. Eriksson, B. Lepenies
and P. H. Seeberger, ChemBioChem, 2011, 12, 2791–2800.
14 K. J. Salandria, J. W. Arico, A. K. Calhoun and L. W. McLaughlin,
to successfully synthesize two 15-mer oligonucleotides which
incorporated one or two modified nucleotides containing the Cy2SI
protecting group. However, after cleavage/deprotection and purifi-
cation over a C18 Sep-Pak column13 we observed exclusively the
oligonucleotide product which still contained full Cy2SI protection
in both oligos (see ESI†). Further attempts at the removal of the
J. Am. Chem. Soc., 2011, 133, 1766–1768.
15 (a) G. Ritzmann, R. S. Klein, D. H. Hollenberg and J. J. Fox,
Carbohydr. Res., 1975, 39, 227–236; (b) H. G. Howell,
P. R. Brodfuehrer, S. P. Brundlage, D. A. Benigni and C. Sapino,
J. Org. Chem., 1988, 53, 85–88.
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