Convenient synthesis and cyclization of dimeric abasic PNA

Article abstract of DOI:10.1016/j.tetlet.2012.10.032

The synthesis of an abasic PNA-based dimer block has been achieved. An alkyl chain stapling the two base sites conformationally restricts the PNA backbone, and serves as an example of preorganization by direct base site linkage. Other possible examples of

Full text of DOI:10.1016/j.tetlet.2012.10.032

Tetrahedron Letters 53 (2012) 6943–6945
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Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
Convenient synthesis and cyclization of dimeric abasic PNA
⇑
Daniel C. Cooper, J. William Suggs
Department of Chemistry, Box H, Brown University, Providence, RI 02912, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
The synthesis of an abasic PNA-based dimer block has been achieved. An alkyl chain stapling the two base
sites conformationally restricts the PNA backbone, and serves as an example of preorganization by direct
base site linkage. Other possible examples of this strategy, such as enforced stacking of adjacent intra-
strand bases to give base pair step analogs, are put forward.
Received 22 August 2012
Revised 5 October 2012
Accepted 8 October 2012
Available online 22 October 2012
Ó 2012 Elsevier Ltd. All rights reserved.
Keywords:
Peptide nucleic acid
Base pair step
Stacking
DNA analog
DNA analogs have been reported with almost every imaginable
structural substitution.1,2 Different base pairs, backbones, and su-
gar types are well known and in a few cases have led to marketable
drugs.3 What is retained is some mechanism to limit the conforma-
tional flexibility. One feature of natural double-helical deoxyribo-
nucleic acids that leads to a predictable but variable structure is
intrastrand base stacking.4,5 In DNA, some steps, such as 50-TpdA-
30, are highly flexible, while 50-dApdA-30 steps adopt the canonical
B-DNA conformation.6 Different preferences for the step parame-
ters twist, roll, tilt, rise, slide, and shift lead to the differences in lo-
cal helical parameters seen in structural studies and to differences
in flexibility.7 While a great deal of research has focused on nucleic
acid base pair analogs2,8–10 we are unaware of studies directed to-
ward the design and synthesis of intrastrand base pair stacking
analogs.11,12 In our opinion, what is crucial for a base pair step is
that it forms a pseudo-ring, preorganizing the backbone in a helical
conformation with flexibility in at least some step parameters to
accommodate adjacent sequences. In native DNA, hydrophobic
and dispersion forces act like a pseudoatom (of the type one can
use in molecular modeling) in the structure to keep adjacent bases
stacked. There are other conceivable alternatives to these forces.
Examples of molecules that illustrate the idea of a base pair step
stacking analog are 1–3, Figure 1. Since the distance between cyclo-
pentadienyl rings in ferrocene is 3.32 Å while the distance between
bases in B-DNA is 3.4 Å (a similarity first pointed out by Inouye and
co-workers13,14), ferrocenes make attractive base pair step analogs,
either for deoxyribose-phosphodiester backbones or peptide nu-
cleic acid backbones, with the iron atom acting as the pseudoatom.
Figure 1. DNA/Fc conjugate 1, PNA/Fc conjugate 2, and PNA/alkyl 3.
O
O
H
N
H
N
PG
R
N
H
N
H
O
4
Base
O
Base
O
PG
N
N
R
N
N
O
H
H
5
Figure 2. PNA scaffolding 4 and desired linked bases 5 PG = protecting group.
However, direct attachment of ferrocene to a deoxyribose C10
leads to acid instability and epimerization at the anomeric center
mediated by ferrocene’s strong stabilization of adjacent carboca-
tions (unpublished results, J. W. Suggs). Alternatively, a peptide
⇑
Corresponding author. Tel.: +1 401 863 3436.
E-mail address: j_suggs@brown.edu (J.W. Suggs).
0040-4039/$ - see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tetlet.2012.10.032

6944
D. C. Cooper, J. W. Suggs / Tetrahedron Letters 53 (2012) 6943–6945
O
O
O
shown in Figure 2 (idealized as 5). Note in this initial realization
H
N
H
N
the linkage functions only to constrain the conformation of one
chain by creating a ring, not to interact with bases in the adjacent
strand. In this way it mimics the nucleic acid universal bases.19
A Boc/Z protecting group strategy was chosen for 4 due to its lit-
erature precedence, chemical orthogonality, and applicability to
PNA automated synthesis.20 We therefore chose 6 as the backbone
target, which has the literature precedent, 7.21
BocHN
BocHN
N
H
O
6
O
Cbz
N
Cbz
N
N
H
OH
In our experience, the published procedure for 721 Figure 3 was
difficult to scale, required careful pH control, and was low yielding.
We have discovered conditions that provide multigram quantities
and are scalable and economical. The key Boc/Z monomer 9 was
made from hydrochloride salt 822 in 88% (Scheme 1).
7
Figure 3. Desired backbone material 6 and the literature precursor 7.
With bulk 9 in hand, Boc deprotection using AcCl/EtOH23 di-
rectly afforded hydrochloride 10 while hydrolysis afforded free
acid 11. Conveniently, both monomeric compounds were obtained
as white solid powders (Scheme 2).
nucleic acid backbone can be used as in 2, which solves any epi-
merization problems and introduces no new stereogenic centers.
The simplest ferrocene base pair step analog would contain a Fc-
CH2CONR2 group. Since ferrocenes with FcCH2COX groups undergo
autoxidation at CH2,15–17 compounds with this kind of step would
be air sensitive. Thus, the most direct realization of a nonreactive
group capable of preorganizing a nucleic acid chain into a helical
conformation would be an alkyl chain as depicted in 3. Such a
strategy has been used to preorganize peptide backbones into an
As a major goal was operational ease, we wished to avoid any
use of polar aprotic solvents typically employed when attempting
peptide coupling directly with amine salts. Carbonyl diimidazole,
used industrially for peptide couplings24 smoothly delivered the
desired dimer 12 in good yield. The simplicity of the reaction is
notable: solid hydrochloride salt was added in a single portion to
a solution of activated 11 and allowed to stir overnight. Catalytic
a
-helical conformation (stapled peptides).18 The generalized PNA
(peptide nucleic acid) architecture for a base pair step analog is
O
O
TEA (2 equiv), BzCl
Toluene
H
N
Cbz
x HCl
N
BocHN
O
BocHN
O
9
88%
8
Scheme 1. Preparation of fully protected monomer 9.
AcCl
O
Cbz
N
HCl
9
H2N
O
EtOH
10 92%
CDI
O
O
Cbz
N
Cbz
N
BocHN
N
H
O
CH2Cl2
O
NaOH
12 69%
Cbz
N
9
BocHN
OH
3:1 MeOH:H2O
11 72%
Scheme 2. Route to fully protected dimer 12.
O
Cl
O
O
O
O
O
O
O
H
H
N
N
N
N
BocHN
N
BocHN
N
O
THF, Cs2CO3
H
H
6
85%
13 76%
O
1. Grubbs II, CH2Cl2
2. Pd/C, H2, MeOH
N
BocHN
N
O
H
13
N
O
O
O
14 55%
Scheme 3. Preparation of desired alkyl-ring macrocycle 14.

D. C. Cooper, J. W. Suggs / Tetrahedron Letters 53 (2012) 6943–6945
6945
Figure 4. Calculated lowest energy conformers of 14 (
D
E kcal/mol 0.00, 2.77, and 3.69 for 15, 16, and 17, respectively).
hydrogenation of 12 afforded deprotected 6 in 85% yield, repre-
senting our desired scaffolding 4.
References and notes
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4. Hunter, C. A. J. Mol. Biol. 1993, 230, 1025–1054.
To illustrate linking, we reacted 6 with 4-pentenoic acid chlo-
ride in THF/CsCO3 smoothly affording diacylated 13 in 76% yield.
Treatment of 13 with Grubbs’ II catalyst followed by catalytic
hydrogenation afforded macrocycle 14 in 55% over two steps. HPLC
provided an analytical sample of 14 (single peak, 97% purity by UV
trace integration) (Scheme 3).
5. Gardiner, E. J.; Hunter, C. A.; Packer, M. J.; Palmer, D. S.; Willett, P. J. Mol. Biol.
2003, 332, 1025–1035.
6. Svozil, D.; Hobza, P.; Sponer, J. J. Phys. Chem. B 2010, 114, 1191–1203.
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As was expected due to restricted rotation about tertiary amide
bonds,25 the NMR spectra of 14 were obtained as a complex mix-
ture of conformers. A computational investigation of the ring sys-
tem was performed by running geometry optimizations on all
possible cis/trans amide conformers at the B3LYP/6-31G⁄⁄ level of
theory (details in Supplementary data). The 2D COSY spectrum
clearly revealed that at least three conformers were present in sig-
nificant amounts, and the computational results indeed indicated
that four of the possible eight conformers are within only 5 kcal/
mol. The 3 calculated lowest energy conformations of 14 are
shown in Figure 4.
In summary an expedient synthesis of scaffolding 6 and a sim-
ple example of a ‘universal’ base-pair step analog 14 have been
achieved. It is notable that every reaction toward preparing 14 is
either catalytic or uses exceedingly common and inexpensive re-
agents. Every step toward backbone 6 is high yielding and opera-
tionally simple. This allows easy access to research quantities of
an abasic PNA dimer.
24. Montalbetti, C. A. G. N.; Falque, V. Tetrahedron 2005, 61, 10827–10852.
25. Richards, S. A.; Hollerton, J. C. Essential Practical NMR for Organic Chemistry;
John Wiley and Sons, 2011.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.tetlet.2012.10.
032.