Synthesis of a novel tetrafluoropyridine-containing amino acid and tripeptide

Article abstract of DOI:10.1016/j.tetlet.2013.06.124

The synthesis of a novel fluoropyridine-containing amino acid from pentafluoropyridine by a nucleophilic substitution process is reported. The orthogonal protecting groups can be manipulated and the fluoropyridine amino acid incorporated into a tripeptide

Full text of DOI:10.1016/j.tetlet.2013.06.124

Accepted Manuscript
Synthesis of a novel tetrafluoropyridine-containing amino acid and tripeptide
Alex S. Hudson, Alex Hoose, Christopher R. Coxon, Graham Sandford, Steven
L. Cobb
PII:
S0040-4039(13)01109-X
DOI:
http://dx.doi.org/10.1016/j.tetlet.2013.06.124
TETL 43177
Reference:
To appear in:
Tetrahedron Letters
Received Date:
Revised Date:
Accepted Date:
22 March 2013
12 June 2013
26 June 2013
Please cite this article as: Hudson, A.S., Hoose, A., Coxon, C.R., Sandford, G., Cobb, S.L., Synthesis of a novel
tetrafluoropyridine-containing amino acid and tripeptide, Tetrahedron Letters (2013), doi: http://dx.doi.org/10.1016/
j.tetlet.2013.06.124
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Graphical Abstract
Synthesis of a novel tetrafluoropyridine-
containing amino acid and tripeptide
Leave this area blank for abstract info.
Alex S. Hudson, Alex Hoose, Christopher R. Coxon, Graham Sandford, Steven L. Cobb∗
O
O
O
N
H
N
OBn
O
H2N
BocHN
N
H
OH
H2N
O
O
OH
OH
O
N
F
F
F
F
F
F
F
F

1
Tetrahedron Letters
journal homepage: www.elsevier.com
Synthesis of a novel tetrafluoropyridine-containing amino acid and tripeptide
Alex S. Hudson, Alex Hoose, Christopher R. Coxon, Graham Sandford, Steven L. Cobb∗
Department of Chemistry, Durham University, Science Laboratories, South Road, Durham DH1 3LE, U.K.
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
The synthesis of a novel fluoropyridine-containing amino acid from pentafluoropyridine by a
nucleophilic substitution process is reported. The orthogonal protecting groups can be
manipulated and the fluoropyridine amino acid incorporated into a tripeptide.
2012 Elsevier Ltd. All rights reserved.
Available online
Keywords:
Novel amino acid
pentafluoropyridine
peptides
A significant hurdle in the potential clinical application of
peptide-based drugs is their inherent in vivo susceptibility
towards enzymatic degradation.1 Several different strategies have
been developed in an effort to overcome this problem, with one
of the most commonly used methods involving cyclisation of
amino acid side chains to prepare cyclic peptides.2,3 The resulting
cyclic peptides have been shown, in many cases, to have both
enhanced stability and biological properties.4 Backbone
cyclisation can be achieved using proteinogenic amino acids, for
example, by forming an amide bond between lysine and glutamic
acid residues.5 Non-proteinogenic amino acids, such allylglycine
(1) and propargylglycine (2), which can be used to form cyclic
peptides via ring-closing metathesis or click chemistry
respectively, can also be exploited.6,7 The applications of 1 and 2
offer several advantages over the formation of backbone amide
bridges derived from naturally occurring amino acids. In
particular, the reactive functionality present in 1 and 2, that
permits the formation of cyclic peptides is orthogonal to standard
peptide coupling conditions and thus there is no need to utilize a
protecting group strategy. In designing new non-proteinogenic
amino acids that can be utilized to prepare backbone cyclized
peptides, a suitably reactive functional group must be present
within the side chains. The functional group that is selected must
be stable to typical protection/coupling/deprotection reaction
conditions common in peptide chemistry and offer functionality
that has a different reactivity profile, ‘orthogonal reactivity’, to
these processes. This would enable, for example, cyclisation to
be carried out at any stage of a multi-step peptide coupling
sequence on demand, leaving the growing peptide backbone
structurally intact.
O
O
H2N
H2N
OH
OH
2
1
Figure 1. Structures of non-proteinogenic amino acids commonly used to
prepare cyclic peptides.
Perfluoroheteroaromatic systems are very reactive towards
nucleophilic attack due to the presence of several highly-electron
withdrawing fluorine atoms attached to the heterocyclic ring and
many processes involving reactions of, for example,
pentafluoropyridine, with a wide range of nucleophiles have been
developed.8,9 Reactions of oxygen- and nitrogen-centered
nucleophiles with pentafluoropyridine (3) occur, in general,
regioselectively at the 4-positon, para to the ring nitrogen, to
give 4-substituted derivatives which, in turn, may react
sequentially with further nucleophilic species to give, in general,
2,4- and 2,4,6-polysubstituted systems, regioselectively (Scheme
———
∗ Corresponding author. Tel.: +44-191-334-2086; fax: +44-191-384-4737; e-mail: s.l.cobb@durham.ac.uk (S.L. Cobb)

2
Tetrahedron
1). This reactivity profile has been utilised recently for the
used.16 Prolonged reaction times (>72 h) at room temperature,
synthesis of a wide range of macrocycles,10,11 fused ring
systems,12 polyfunctional heterocycles13,14 and glycosyl donors15
derived from the pentafluoropyridine scaffold in convenient, high
yielding, regioselective processes.
and the use of heating did not improve the recovered yield of 5.
With 5 in hand we next sought to confirm that the Boc and
methyl ester protecting groups could be removed selectively, and
that 5 would tolerate standard peptide coupling conditions. To
this end, 5 was treated with TFA/ CH2Cl2 (1:1, rt, 4 h) and the
resulting TFA salt obtained was immediately coupled to Boc-
Ala-OH via PyBOP-mediated amide bond formation to give the
di-peptide 6 in an excellent yield (78%) (Scheme 3). The fact that
6 was isolated in a good yield confirmed that the fluoropyridyl
group was stable to both the acidic conditions required for
removal of the Boc group and also the PyBop mediated-peptide
coupling conditions used. Furthermore, both H and 13C NMR
1
spectra gave no indication of the presence of diastereoisomers
indicating that the formation of
5 does not affect the
stereochemistry at the alpha-carbon of the amino acid substrate.
Attempts to remove the methyl ester protecting group from 6
using standard reaction conditions (e.g. aqueous base / MeOH, rt)
proved to be problematic and several degradation products were
observed.
Scheme 1. Regioselective sequential SNAr reactions of pentafluoropyridine
Given this reactivity profile (Scheme 1), we were eager to
investigate if we could access a fluoropyridine-containing amino
acid that could be incorporated into linear peptide chains,
because, in principle, subsequent reaction with nucleophilic sites
on adjacent amino acid side chains could be utilized to prepare
novel cyclic peptides. As part of a wider developing research
programme in fluorinated peptide systems, we now report
reactions between pentafluoropyridine and the amino acid serine,
and the subsequent use of the tetrafluorinated residue for the
synthesis of a representative tripeptide as the first stage in the
synthesis of cyclic peptide derivatives.
Scheme 3. Synthesis of dipeptide 6. Reagents and conditions: (a) TFA,
CH2Cl2, rt, 4 h (b) Boc-Ala-OH, PyBop, NMM, CH2Cl2, rt, 18 h.
Given the instability of 6 to basic hydrolysis of the methyl
ester, an alternative protecting group strategy was investigated. In
order to retain protecting group orthogonality, we chose to
investigate the suitability of the benzyl ester protecting group.
Boc-Ser-OBn (8), which can be prepared in excellent yield (71%)
from commercially available Boc-Ser-OH (7), was reacted with
pentafluoropyridine utilizing the reaction conditions previously
developed for the preparation of 5. After purification by column
chromatography, amide 917 was obtained in moderate yield
(55%) (Scheme 4). Again 19F NMR analysis of 9 confirmed that
the nucleophilic substitution had occurred, as expected, at the 4-
position, para to the ring nitrogen [δ -89.84 (2F, m, F-2), -158.22
(2F, m, F-3)].
Our aim was to prepare the desired building block with
orthogonal protecting groups so that further synthetic
manipulations could be carried out selectively on either the amine
or acid functionality. Thus, initially, we chose to investigate Boc-
Ser-OMe (4) as a nucleophile as it can easily be prepared on a
large scale. The reaction of 4 with pentafluoropyridine (2.0 eq.)
in the presence of potassium carbonate (1.0 eq.) afforded 5 in
moderate 30% yield (Scheme 2).
O
BocHN
OMe
O
(a)
BocHN
O
OMe
F
F
F
F
30%
OH
O
O
N
4
BocHN
BocHN
(a)
OH
OH
OBn
5
Scheme 2. Synthesis of fluoropyridine amino acid 5. Reagents and
conditions: (a) pentafluoropyridine, K2CO3, MeCN, rt, 20 h.
71%
OH
(b)
7
8
1
Despite the low yield, the reaction proceeded cleanly and H
NMR analysis of the crude reaction mixture showed that only
starting material 4 and product 5 were present, and these could be
easily separated by column chromatography. 19F NMR analysis
of 5, in comparison to literature data,13 confirmed that
nucleophilic substitution had occurred as expected at the 4-
positon, para to the ring nitrogen. Further, investigation of the
reaction conditions showed that increased yields of 45-55% could
be obtained if an excess of pentafluorpyridine (4 equiv.) was
55%
O
O
BocHN
OBn
F
F
F
F
N
9

3
Supplementary Material
Scheme 4. Synthesis of fluoropyridine amino acid 9. Reagents and
conditions: (a) BnBr, NaHCO3, H2O, EtOAc, rt, 16 h, (b)
pentafluoropyridine, K2CO3, MeCN, rt, 20 h.
Supplementary data associated with compounds 5, 9, and 12 can be found,
in the online version.
With 9 in hand we sought to extend the peptide chain in both
directions and prepare a model tripeptide. Firstly, removal of the
benzyl ester was achieved via Pd/carbon-catalyzed hydrogenation
in an excellent yield (86%). PyBop-mediated coupling of the
resultant free acid 10 to NH2-Ala-OBn afforded the dipeptide 11
in excellent yield (77%). Finally, removal of the Boc protecting
group under acidic conditions and subsequent coupling to Boc-
Ala-OH afforded the tripeptide 12,18 demonstrating that the
tetrafluoropyridine side-group was able to tolerate a typical
peptide coupling reaction protocol (Scheme 5).
References and notes
1.
2.
Frokjaer, S.; Otzen, D.D. Nat. Rev. Drug Discov. 2005, 4, 298-306
(a) Jiang, S.; Li, Z.; Ding, K.; Roller, P.P. Curr. Org, Chem. 2008, 12,
1502–1542 (b) White, C.J.; Yudin, A.K. Nat. Chem. 2011, 3, 509-524
(c) Davies, J.S. J. Peptide Sci. 2003, 9, 471-501
3.
Ling, R.; Yoshida, M.; Mariano, P. S. J. Org. Chem. 1996, 61, 4439–
4449.
4.
5.
6.
Kessler, H. Angew. Chem. Int. Ed. 1982, 21, 512–523.
Shao, Y.; Lu, W. Y.; Kent, S. B. H. Tetrahedron Lett., 1998, 39, 3911
(a) Miller, S.J.; Blackwell, H.E.; Grubbs, R. H. J. Am. Chem. Soc.
1996, 118, 9606-9614 (b) Kim, Y.W.; Kutchukian, P.S.: Verdine, G.L.
Org. Lett. 2010, 12, 3046-2049 (c) Kim, Y.-W.; Grossmann, T. N.;
Verdine, G. L. Nat. Protoc. 2011, 6, 761-771.
O
7.
(a) Kawamoto, S.A.; Coleska, A.: Ran, X.; Yi, H.; Yang, C.-Y.; Wang,
S. J. Med. Chem. 2012, 55, 1137-1146 (b) Cantel, S.: Isaad Ale, C.;
Scrima, M.; Levy, J.J.; DiMarchi, R.D.; Rovero, P.; Halperin, J. A.;
D’Ursi, A.M.; Chorev, M. J. Org. Chem. 2008, 73, 5663-5674.
Brooke, G. M. J. Fluorine Chem. 1997, 86, 1-76.
O
BocHN
BocHN
OBn
OR
N
H
(b)
O
O
O
8.
9.
F
F
F
77%
F
F
F
F
Chambers, R. D. Fluorine in Organic Chemistry; Blackwell: Oxford,
2004.
N
F
10. Sandford, G. Chem. Eur. J. 2003, 9, 1464-1469.
N
11. Ranjbar-Karimi, R.; Sandford, G.; Yufit, D. S.; Howard, J. A. K. J.
9
R = Bn
11
Fluorine Chem. 2008, 129, 307-313.
(a)
86%
10 R = H
12. Cartwright, M. R.; Convery, L.; Kraynck, T.; Sandford, G.; Yufit, D.
S.; Howard, J. A. K.; Christopher, J. A.; Miller, D. D. Tetrahedron
2010, 66, 519-52
(c), (d)
90%
13. Chambers, R. D.; Hoskin, P. R.; Sandford, G.; Yufit, D. S.; Howard, J.
A. K. J. Chem. Soc., Perkin Trans. 1 2001, 2788-2795.
14. Christopher, J. A.; Brophy, L.; Lynn, S. M.; Miller, D. D.; Sloan, L.
A.; Sandford, G. J. Fluorine Chem. 2008, 129, 447–454.
15. Hargreaves, C. A.; Sandford, G.; Davis, B. G. Polyfluoropyridyl
glycosyl donors, in Fluorinated Synthons; Soloshonok, V. A. Ed.;
ACS Symposium Series 949, ACS: Washington D.C., 2007; pp. 323-
336.
16. Preparation of 5: Pentafluoropyridine (3) (0.7 mL, 6.2 mmol) was
added via a syringe to a stirred solution of BocNH-Ser-OMe (0.34 g,
1.5 mmol) and K2CO3 (0.20 g, 1.5 mmol) in MeCN (30 mL). The
mixture was stirred at rt for 20 h. Filtration followed by removal of the
solvent under reduced pressure yielded the crude product, which was
O
H
N
OBn
BocHN
N
H
O
O
O
F
F
F
N
F
12
purified by column chromatography (hexane-EtOAc, 9:1) to give 5 as a
23.9
Scheme 5. Synthesis of a fluoropyridine-containing tripeptide 12. Reagents
and conditions: (a) H2, Pd/carbon, rt (b) NH2-Ala-OBn, PyBop, NMM,
CH2Cl2, rt, 18 h (c) TFA, CH2Cl2, rt, 4 h (d) Boc-Ala-OH, PyBop, NMM,
CH2Cl2, rt, 18 h.
white powder (0.27 g, 48%). Rf = 0.5 (hexane – EtOAc, 2:1); [α]D
=
+26.90 (c 1.0, CHCl3); m.p. 40-42 °C; IR νmax/cm-1 1644, 1678, 1730,
2992, 3362; 1H NMR (400 MHz, CDCl3) δ 1.44 (9H, s, C(CH3)3), 3.81
(3H, s, OCH3), 4.68 (1H, m, α-CHCH2OArF), 4.80 (2H, m, β-
CH2OArF), 5.48 (1H, d, J 7.4 Hz, BocNH); 13C NMR (176 MHz,
CDCl3) δ 28.2 (3C, s, C(CH3)3), 53.1, 54.0, 74.4 (1C, s, CH2), 81.5,
1
1
In conclusion, this work describes the synthesis of two
135.0 (2C, dm, JCF 257 Hz, C-3), 144.1 (2C, dm, JCF 243 Hz, C-2),
146.8 (1C, m, C-4), 154.8, 169.3; 19F NMR δ (376 MHz, CDCl3) -
89.81 (2F, m, F-2), -158.23 (2F, m, F-3); m/z (ESI+) 391.09 ([M +
Na]+, 100%); HRMS (ESI+) C14H16N2O5F4Na+ ([M + Na]+); requires
391.0893; found 391.0893.
orthogonally protected amino acids that contain
a
tetrafluoropyridine functionality on the side chain (5 and 9). The
tetrafluoropyridine group was found to be stable to manipulations
of both the Boc and Bn protecting groups, but the attempted
removal of a methyl ester resulted in degradation of the starting
material. Incorporation of the tetrafluoropyridine amino acid 9
17. Preparation of 9: To a stirred solution of BocNH-Ser-OBn (8) (1.20 g,
4.3 mmol) and K2CO3 (0.60 g, 4.3 mmol) in MeCN (40 mL) was added
pentafluoropyridine (3) (1.9 mL, 17.2 mmol). The mixture was stirred
at rt for 20 h before the solution was filtered and the solvent removed
under reduced pressure. Purification by column chromatography
(hexane-EtOAc, 9:1) gave 9 (0.95 g, 55%); as a white powder. Rf =
into
a test tripeptide demonstrated the stability of the
tetrafluoropyridine amino acids to standard peptide coupling
reaction conditions. Efforts are now underway to investigate the
potential applications of 9 in the synthesis of backbone-cyclized
peptides.
29.8
0.23 (hexane – EtOAc, 9:1); m.p. 50-51 °C; [α]D = +7.22 (c 1.0,
CHCl3); IR νmax/cm-1 1681, 1728, 2984, 3356; 1H NMR (400 MHz,
CDCl3) δ 1.44 (9H, s, C(CH3)3), 4.71 (1H, m, α-CHCH2OArF), 4.81
(2H, m, β-CH2OArF), 5.23 (2H, dd, J 32.7, 12.0 Hz, CH2Ph), 5.56 (1H,
d, J 7.8 Hz, BocNH), 7.31 (5H, m, Ph); 13C NMR (176 MHz, CDCl3) δ
27.2 (3C, s, C(CH3)3), 53.1, 66.9, 73.4, 79.7, 127.0, 127.9, 132.08 –
135.5 (2C, dm, 2JCF 257 Hz, C-3), 133.8, 143.0 (1C, dm, JCF 242.4 Hz,
C-2), 145.6 (1C, m, C-1), 154.1, 167.7; 19F NMR (376 MHz, CDCl3) δ
-89.84 (2F, m, F-2), -158.22 (2F, m, F-3); m/z (ESI+) 467.8 ([M + Na]+,
Acknowledgments
This work was supported financially by the University of Durham (A. S. H)
and the Nuffield Foundation (A. H. summer studentship).
100%); HRMS (ESI+) C20H20N2O5F4Na+ ([M
467.1206; found 467.1194.
+
Na]+); requires
18. Preparation of 12 from dipeptide 11: Boc Deprotection (i): TFA (xs, 4
mL) was carefully syringed into a solution of BocNH-SerArF-Ala-
OBn (11) (0.12 g, 0.23 mmol) in CH2Cl2 (4 mL) and left to stir for 4 h.

4
Tetrahedron
Et2O was added and subsequently evaporated (4 x 30 mL) to remove
excess TFA. Peptide Coupling (ii): The deprotected dipeptide was
dissolved in CH2Cl2 (10 mL) and stirred at room temperature. To this
solution BocNH-Ala-OH (0.044 g, 0.23 mmol) and PyBOP (0.12 g,
0.23 mmol) were added. NMM (0.08 mL, 0.69 mmol) was slowly
added and the mixture was left to stir for 18 h at rt. The solvent was
evaporated and the crude material was purified by column
chromatography (hexane-EtOAc, 7:3). The product BocNH-SerArF-
Ala-OBn (11) was isolated (0.13 g, 90%); as a white powder. Rf = 0.61
(hexane-EtOAc, 1:1); IR νmax/cm-1 1477, 1509, 1643, 2954, 2998,
3304; 1H NMR (600 MHz, CDCl3) δ 1.37 (3H, d, J 7.1 Hz, CH3), 1.40
(9H, s, C(CH3)3), 1.42 (3H, d, J 7.2 Hz, CH3), 4.15 (1H, m, α-CH),
4.54 - 4.58 (2H, m, α-CH, β-CHOArF), 4.89 (1H, m, α-CHCH2OArF),
4.93 (1H, d, J = 6.5 Hz, NHBoc), 5.02 – 5.07 (1H, m, β-CHOArF),
5.07 – 5.17 (2H, m, CH2Ph), 7.22 (1H, d, J = 7.5 Hz, NH), 7.27 – 7.33
(5H, m, Ph); 13C NMR (151 MHz, CDCl3) δ 17.7, 28.1, 29.7, 48.7,
1
50.9, 52.5 , 67.1, 73.5, 80.8, 128.0, 128.3, 128.5, 134.8 (2C, dm, JCF
257 Hz, C-3), 135.2, 142.9 – 145.0 (2C, dm, JCF 242.4 Hz, C-2), 146.5
(1C, m, C-1), 155.9, 167.6, 171.9, 172.9; 19F NMR (376 MHz, CDCl3)
δ -89.84 (2F, m, F-2), -158.27 (2F, m, F-3); m/z (ESI+) 609.3 ([M +
Na]+, 100%); HRMS (ESI+) C26H30N4O7F4Na+ ([M + Na]+); requires
609.1948; found 609.1948;