(2R)- and (2S)-3-fluoroalanine and their N-methyl derivatives: Synthesis and incorporation in peptide scaffolds

Article abstract of DOI:10.1021/ol062434q

(Diagram presented) A convergent synthetic methodology has been developed to access both (2S)- and (2R)-3-fluoroalanine and their corresponding N-methyl analogues, in optically pure form, through a common oxazolidinone intermediate that can be obtained fr

Full text of DOI:10.1021/ol062434q

ORGANIC
LETTERS
2006
Vol. 8, No. 25
5849-5852
(2R)- and (2S)-3-Fluoroalanine and Their
N-Methyl Derivatives: Synthesis and
Incorporation in Peptide Scaffolds
Hamid R. Hoveyda* and Jean-Franc¸ois Pinault
Tranzyme Pharma Inc., Institut de pharmacologie de Sherbrooke, 3001,
12e AVenue Nord, Sherbrooke, Que´bec J1H 5N4, Canada
hhoVeyda@tranzyme.com
Received October 3, 2006
ABSTRACT
A convergent synthetic methodology has been developed to access both (2S)- and (2R)-3-fluoroalanine and their corresponding N-methyl
analogues, in optically pure form, through a common oxazolidinone intermediate that can be obtained from - or -serine. In addition, a
procedure for incorporation of these unnatural amino acids in peptide scaffolds is also disclosed herein that minimizes the occurrence of
-elimination during amide bond formation.
L
D
â
Fluorine substitution as a means of modifying the physico-
chemical properties of a biologically active molecule is a
strategy that finds great utility in medicinal chemistry and
in biomedical applications.1
In the context of a medicinal chemistry program, we
required access to both enantiomers of N-methyl-3-fluoro-
alanine.2 To the best of our knowledge, no reports for the
chiral synthesis of N-alkyl derivatives of 3-fluoroalanines
have appeared in the literature.3 Therefore, we attempted to
adapt some of the known procedures for the synthesis of
3-fluoroalanine, or related derivatives, and subsequently
apply the known strategies to prepare the N-methyl deriva-
tives as well.4 As these efforts proved futile, we devised a
convergent strategy, using oxazolidinone intermediates, to
gain access to both N-methylated and primary analogues of
3-fluoroalanines in enantiopure form. A summary of these
efforts is provided herein, as well as optimal amide bond
formation conditions for efficient incorporation of such
amino acids in peptide scaffolds.
(1) For a selection of review articles on this topic, see: (a) Welch, J. T.
Tetrahedron 1987, 43, 3123-3197. (b) Imperiali, B. AdV. Biotechnol.
Processes 1988, 10, 97-129. (c) Welch, J. T.; Eswarakrishnan, S. Fluorine
in Bioorganic Chemistry; John Wiley & Sons: New York, 1991. (d) Filler,
R.; Kobayashi, Y.; Yagupolskii, L. M. Organofluorine Compounds in
Medicinal Chemistry and Biomedical Applications; Elsevier: New York,
1993. (e) O’Hagan, D.; Rzepa, H. S. Chem. Commun. 1997, 645-652. (f)
Park, K. B.; Kitteringham, N. R.; O’Neill, P. M. Annu. ReV. Pharmacol.
Toxicol. 2001, 41, 443-470. (g) Ismail, F. M. D. J. Fluorine Chem. 2002,
118, 27-33. (h) Bo¨hm, H.-S.; Banner, D.; Bendels, S.; Kansy, M.; Kuhn,
B.; Mu¨ller, K.; Obst-Sander, U.; Stahl, M. ChemBioChem 2004, 5, 637-
643. (i) Thayer, A. M. C&EN 2006, June 5, 15-24.
(2) For a few recent review articles on the synthesis of fluorinated amino
acids, see: (a) Sutherland, A.; Willis, C. L. Nat. Prod. Rep. 2000, 17, 621-
631. (b) Qiu, X.-L.; Meng, W.-D.; Qing, F.-L. Tetrahedron 2004, 60, 6711-
6745. For a review on â-fluoro-R-amino acids, see: Sewald, N.; Burger,
K. In Fluorine Containing Amino Acids; Kukhar, V. P., Soloshonok, V.
A., Eds.; John Wiley & Sons: New York, 1995; Chapter 4, pp 139-220.
Fluorodehydroxylation in the side chain of serine can be
effected through reaction with sulfur tetrafluoride in liquid
hydrogen fluoride at low temperature with little racemiza-
tion.5 However, given the extremely corrosive nature of the
(3) For a report on the racemic synthesis of N-alkylated fluoroamino
acids, in very low yields, see ref 6a.
(4) For recent reviews of synthetic methodologies for the preparation of
N-methyl-R-amino acids, see: (a) Sagan, S.; Karoyan, P.; Lequin, O.;
Chassaing, G.; Lavielle, S. Curr. Med. Chem. 2004, 11, 2799-2822. (b)
Aurelio, L.; Brownlee, R. T. C.; Hughes, A. B. Chem. ReV. 2004, 104,
5823-5846 and references cited therein. For a review on the use of
N-methyl-substituted amino acids in macrocyclic peptidomimetics, see: (c)
Fairlie, D.; Abbenante, G.; March, D. R. Curr. Med. Chem. 1995, 2, 654-
686.
10.1021/ol062434q CCC: $33.50
© 2006 American Chemical Society
Published on Web 11/07/2006

reagents employed in this approach, we looked for more user-
friendly synthetic procedures. Several other literature reports
exist on the synthesis of 3-fluoroalanines or related deriva-
tives (excluding N-alkylated derivatives). Of these reports,2
the majority include the use of nucleophilic fluorinating
agents,6 although other approaches such as use of electro-
philic fluorination7 and enzymatic chiral resolution8 are also
precedented. One of the commonly used nucleophilic fluo-
rination reagents for the fluorodehydroxylation reaction is
DAST.9,10a Nonetheless, it is well appreciated that reagents
such as DAST can also cause â-elimination to result in
formation of dehydroalanine derivatives as byproducts.11a,12
The work of Pansare and Vederas is notable in this regard,
as they were able to successfully use DAST to prepare
several â-fluoro-R-amino acids, albeit by employing 4,5-
diphenyl-4-oxazolin-2-one as the amine protective group (cf.
1, Table 1).12 However, with substrates such as 1, we were
Table 1. Screening of Fluorodehydroxylation Reaction
Conditions on Substrate 1
HPLC yields, %
entry
1
reagents and conditions
1
2
3
DAST (1.1 equiv), CH2Cl2,
-78 °C f rt, 7 h
6
55
20a
(5) (a) Kollonitsch, J.; Marburg, S.; Perkins, L. M. J. Org. Chem. 1979,
44, 771-777. (b) Reider, P. J.; Conn, R. S. E.; Davis, P.; Grenda, V. J.;
Zambito, A. J.; Grabowski, E. J. J. Org. Chem. 1987, 52, 3326-3334.
(6) For a few specific references, in addition to those cited in ref 2, see:
(a) Cohen, A.; Bergmann, E. D. Tetrahedron 1966, 22, 3545-3547.
(Racemic synthesis of N,N-dimethyl 3-fluoroalanine in overall ca. 11% yield,
using 1,1,2-trifluoro-2-chloroethyl-diethylamine as fluorinating reagent.) (b)
Gershon, H.; McNeil, M. W.; Bergmann, E. D. J. Med. Chem. 1973, 16,
1407-1409. (Racemic synthesis of 3-fluoroalanines, using liquid HF, in
poor yield.) (c) Groth, U.; Scho¨llkopf, U. Synthesis 1983, 673-675. (DAST
was used in this asymmetric synthesis for fluorodehydroxylation. Fluoro-
alanine, however, cannot be obtained by this route.) (d) Yang, D.; Kuang,
L.-R.; Cherif, A.; Tansey, W.; Li, C.; Lin, W. J.; Liu, C.-W.; Kim, E.;
Wallace, S. J. Drug Targeting 1993, 1, 259-267. ([18F]Fluoroalanine
prepared in 0.5-1.% yield by side chain displacement in Boc-Ser(OTs)-
OMe with K18F.)
(7) In addition to those cited in ref 2, for a few specific references see:
(a) Davis, F. A.; Sriajan, V.; Titus, D. A. J. Org. Chem. 1999, 64, 6931-
6934. (Electrophilic fluorination with N-fluorobenzenesulfonimide, followed
by an interesting but lengthy asymmetric synthesis with chiral sulfinimines.
Fluoroalanine was not provided in the examples synthesized, presumably
because fluorine alone is a poor stereodirecting group, as stated by the
authors.) (b) Gerus, I. I.; Kolomeitsev, A. A.; Kolycheva, M. I.; Kukhar,
V. P. J. Fluorine Chem. 2000, 105, 31-33. (Racemic synthesis of
3-fluoroalanine; no characterization data offered.)
2
3
Deoxo-Fluor (1.3 equiv), CH2Cl2,
-78 °C f rt, 7 h
nC4H9SO2F (2 equiv), iPr2NEt
(4.5 equiv), Et3N(HF)3 (1.5 equiv),
MeCN, 45 °C, 24 h
52
0
44
0
0
100
a In addition, 18% of an intractable byproduct was detected.
unable to extend their approach (entry 1) to form 3-fluoro-
alanines despite attempting different reaction conditions (e.g.,
entries 2 and 3).13 In all cases, â-elimination byproduct 2 or
a derivative, i.e., 3, prevailed as depicted below; the desired
product 4 was not detected under any of the experimental
conditions attempted.
Other potential approaches were investigated in hopes of
finding a synthetic route that obviates â-elimination during
the nucleophilic fluorodehydroxylation of serine (cf. Schemes
1 and 2). One such effort consisted of employing the
(8) (a) Schmitt, L.; Boniface, J. J.; Davis, M. M.; McConnel, H. M. M.
J. Mol. Biol. 1999, 286, 207-218. (b) Gonc¸alves, L. P. B.; Antunes, O. A.
C.; Pinto, G. F.; Oestericher, E. G. J. Fluorine Chem. 2003, 124, 219-
227. (c) Gonc¸alves, L. P. B.; Antunes, O. A. C.; Oestericher, E. G. Org.
Process. Res. DeV. 2006, 10, 673-677.
(9) Abbreviations used herein: DAST, (diethylamino)sulfur trifluoride;
Deoxo-Fluor, bis(2-methoxyethyl)aminosulfur trifluoride; MorphoCDI,
N-cyclohexyl-N′-(2-morpholinoethyl)carbodiimide methyl-p-toluenesulfonate;
DPPA, diphenylphosphoryl azide; NMM, N-methylmorpholine; IBCF,
isobutyl chloroformate; EDCI, 1-ethyl-3-(3′-dimethylaminopropyl)carbo-
dimide hydrochloride; HOAt, 7-aza-1-hydroxybenzotriazole; Ms, methane-
sulfonyl; TBSCl, tert-butyldimethylsilyl chloride; TBS, tert-butyldimeth-
ylsilyl; TBAF, tert-butylammonium fluoride; PTSA, p-toluenesulfonic acid.
(10) (a) Middleton, W. J. J. Org. Chem. 1975, 40, 574-578. (b) Deoxo-
Fluor is a more thermally stable variant of DAST; see: Lal, G. S.; Pez, G.
P.; Pesaresi, R. J.; Prozonic, F. M.; Cheng, H. J. Org. Chem. 1999, 64,
7048-7054. (c) For a recent publication on new methodologies for direct
conversion of alcohols to fluorides see: Yin, J.; Zarkowsky, D. S.; Thomas,
D. W.; Zhao, M. M.; Huffman, M. A. Org. Lett. 2004, 6, 1465-1468. (Cf.
Conditions used in entry 3, Table 1.)
(11) (a) Somekh, L.; Shanzer, A. J. Org. Chem. 1983, 48, 907-908.
The authors used DAST/pyridine to effect stereospecific synthesis of R,â-
dehydroamino acids. (b) Use of fluorotrimethylsilanes to effect SN2
fluorodehydroxylation of serine side chain resulted in a cyclodehydration
reaction instead, leading to the formation of corresponding oxazoline, see:
Choi, D.; Stables, J. P.; Kohn, H. J. Med. Chem. 1996, 39, 1907-1916.
(12) Pansare, S. V.; Vederas, J. C. J. Org. Chem. 1987, 52, 4804-4810.
The following points are noteworthy. (i) The use of 4,5-diphenyl-4-oxazolin-
2-one as the amine protective group was reported as a means of suppressing
the intramolecular attack of the carbamate oxygen (e.g., with Boc or Cbz)
on the activated â-carbon. (ii) All the successful examples cited by the
authors bear an alkyl substituent at â-carbon, unlike the case with 1. (iii)
Elimination byproducts were attenuated but not abrogated; nevertheless,
apart from cases in which the â-carbon substituent was extremely bulky
(e.g., iPr group), the fluorodehydroxylation product predominated.
Scheme 1. Attempted Use of Weinreb Amide Derivatives of
Serine To Effect Fluorodehydroxylationa
a Reagents and conditions: (a) DAST, CH2Cl2; (b) Deoxo-Fluor,
CH2Cl2; (c) nC4H9SO2F, iPr2NEt, Et3N(HF)3; (d) TBAF, THF.
(With substrate 5a, conditions a-c and with 5b conditions a and
c-d were attempted.)
corresponding Weinreb amide of serine, 5a,b (Scheme 1),
for the fluorination chemistrysa strategy that has been
effective in circumventing â-elimination under Mitsunobu
conditions.14 Another attempt consisted of testing the utility
(13) We did not have much success with direct fluorination of protected
serine derivatives, using DAST or similar reagents, either.
(14) See for example: Panda, G.; Rao, N. V. Synlett 2004, 714-716.
5850
Org. Lett., Vol. 8, No. 25, 2006

of reaction conditions were then explored for the conversion
of the silyl-protected oxazolidinones (Scheme 3, 8a,b) to their
corresponding fluoro congeners (Scheme 3, 9a,b). Initially,
we carried out the fluorodehydroxylation reaction after
having isolated the free alcohol by treatment of the silylated
oxazolidinones (8a, 8b) with HF-pyridine. Upon additional
optimization studies, we observed that desilylation and
fluorodehydroxylation steps can be conducted in a one-pot
manner through treatment of the silylated intermediate with
HF-pyridine (8 equiv) and Deoxo-Fluor (2 equiv).10b,20 The
fluorinated oxazolidinones (9a, 9b) were then reduced by
using the triethylsilane/TFA conditions reported by Fre-
idinger17 and co-workers, although rather sluggishly, requir-
ing 2 days for its completion.21 During this prolonged
acidolytic reductive ring-opening, the Cbz protective group
was partially cleaved. As we needed an acid labile protective
group, such as Boc, in the final product, we simply proceeded
to the hydrogenolytic deprotection of the remaining Cbz,
followed by protection of the secondary amine by the Boc
protective group to obtain both antipodes of N-methyl-3-
fluoroalanine (Scheme 3, 10a,b) in g92% ee (chiral HPLC)
and with an overall yield of g50%.
Scheme 2. Attempted Use of the â-Lactone Derived from
Serine to Nucleophilic Fluorination Reactiona
a Reagents and conditions: (a) TBAF, THF; (b) HF-pyridine;
(c)16 Et3N(HF)3 (4 equiv), 110 °C, 18 h.
of the â-lactone derived from serine, 7 (Scheme 2), under
nucleophilic fluorination conditions.15 None of these efforts
proved fruitful: either no reaction was observed, or an
intractable mixture, which occasionally contained traces of
the desired product, was obtained (cf. Schemes 1 and 2).
The utility of oxazolidinone derivatives of amino acids as
synthetic intermediates both for the synthesis of N-alkylated
amino acids17 and as a protective group strategy is well
established.18 We thus decided to investigate the potential
utility of such oxazolidinone derivatives as a common
intermediate for accessing both 3-fluoroalanine and its
N-methyl derivatives in enantiopure form.
As depicted in Scheme 4, treatment of intermediates 9a
or 9b with HCl (2 N) in dioxane18c furnished the deprotection
As the oxazolidinone 8 showed little propensity toward
â-elimination during fluorodehydroxylation, we were able
to successfully employ this approach for the synthesis of the
title compounds (Scheme 3). Starting with Cbz-Ser-OH or
Scheme 4. Synthesis of Cbz-(2R)-3-fluoroalanine and
Cbz-(2S)-3-fluoroalanine from 9a and 9b
Scheme 3. Synthesis of Boc-N-Me-(2R)-3-fluoroalanine and
Boc-N-Me-(2S)-3-fluoroalanine
of the oxazolidinone, thus providing both (2R)- and (2S)-3-
fluoroalanine in good yields and high enantiomeric excess.
Our efforts in using alternative conditions18a,b,d,f for cleavage
of the oxazolidinone met with little or no success.
With both enantiomers of N-Me-fluoroalanine in hand, we
proceeded to the subsequent step in the synthesis of our target
(18) See for example: (a) Itoh, M. Chem. Pharm. Bull. 1969, 17, 1679-
1686. (b) Blaser, D.; Seebach, D. Liebigs Ann. Chem. 1991, 1067-1078.
(c) We modified the conditions reported in this reference to avoid
deprotection of the Cbz protective group (cf. Scheme 4): Natalini, B.;
Mattoli, L.; Pellicciari, R.; Carpenedo, R.; Chiarugi, A.; Moroni, F. Bioorg.
Med. Chem. Lett. 1995, 5, 1451-1454. (d) Thompson, M. J.; Mekhalfia,
A.; Hornby, D. P.; Blackburn, G. M. J. Org. Chem. 1999, 64, 7467-7473.
(e) Allevi, P.; Cribiu`, R.; Anastasia, M. Tetrahedron Lett. 2004, 45, 5841-
5843. (f) Allevi, P.; Anastasia, M. Tetrahedron Lett. 2003, 44, 7663-7665.
(19) (a) Aurelio, L.; Brownlee, R. T. C.; Hughes, A. B.; Sleebs, B. E.
Aust. J. Chem. 2000, 53, 425-433. (b) Luo, Y.; Evindar, G.; Fishlock, D.;
Lajoie, G. Tetrahedron Lett. 2001, 42, 3807-3809. (c) Aurelio, L.; Box, J.
S.; Brownlee, R. T. C.; Hughes, A. B.; Sleebs, B. E. J. Org. Chem. 2003,
68, 2652-2667 and references cited therein.
Cbz-D-Ser-OH, the TBS9 protection of the side chain alcohol,
followed by the formation of the oxazolidinone with
paraformaldehyde, was effected through standard conditions
to obtain intermediates 8a and 8b, respectively.19 A number
(15) Both 3-chloroalanine and 3-bromoalanine, but not 3-fluoroalanine,
were reported to be accessible in this manner by Vederas and co-workers;
see: (a) Arnold, L. D.; Kalantar, T. H.; Vederas, J. C. J. Am. Chem. Soc.
1985, 107, 7105-7109. (b) Arnold, L. D.; May, R. G.; Vederas, J. C. J.
Am. Chem. Soc. 1988, 110, 2237-2241.
(16) These conditions are similar to those reported for ring opening of
epoxides to form fluorohydrins; see: Muehlbacher, M.; Poulter, C. D. J.
Org. Chem. 1988, 53, 1026-1030.
(17) Freidinger, R. M.; Hinkle, J. S.; Perlow, D. S.; Arison, B. H. J.
Org. Chem. 1983, 48, 77-81 Also see references cited in footnote 4.
(20) We obtained better results when using Deoxo-Fluor as compared
to alternative fluorinating reagents such as DAST or morpholinosulfur
trifluoride.
(21) Zhang, S.; Govender, T.; Norstro¨m, T.; Arvidsson, P. I. J. Org.
Chem. 2005, 70, 6918-6920. New and improved conditions are reported
in this publication. Despite multiple attempts, use of the conditions reported
by Zhang et al. with oxazolidinones such as 9a,b did not faciliate the
reaction.
Org. Lett., Vol. 8, No. 25, 2006
5851

conditions, the amide bond formation proceeded without
detectable epimerization (cf. Table 2). Once incorporated in
the peptide scaffold, the fluoroalanine moiety proved stable
toward further â-elimination.24,25
Table 2. Screening of the Amide Bond Formation Conditions
for the Synthesis of N-Methylfluoroalanine Peptides9
In summary, we have discovered a convergent route for
the synthesis of both enantiomers of 3-fluoroalanine and their
respective N-Me analogues through the use of oxazolidinones
8a,b (cf. Scheme 3) as synthetic intermediates. Moreover,
we report optimized amide coupling conditions that allow
effective incorporation of such amino acids in peptide
scaffolds in good yields and with minimal â-elimination and
no detectable epimerization. In view of the preponderance
of fluorine-containing pharmaceuticals, including fluoropep-
tides, the advances discussed herein should prove helpful in
the context of future endeavors in this area.
yields, %a
entry
1
reagents and conditions
12b 14
MorphoCDI, Et3N (1.5 equiv),
MeCN (0.05 mM), 16 h, rt
14
0
2
3
4
5
DPPA, NMM (1.5 equiv),
DMF (0.2 mM), 16 h, 0 °C f rt
IBCF, NMM (1.5 equiv),
THF (0.2 mM), 16 h, -20 °C f rt
EDCI, HOAt, Et3N (1.5 equiv),
THF:CH2Cl2 (1:1, 0.2 mM), 5 h, 0 °C f rt
EDCI, HOAt, NMM (1.5 equiv),
THF: CH2Cl2 (1:1, 0.2 mM), 5 h, 0 °C f rt
3
5
Supporting Information Available: Experimental pro-
cedures and full characterization of compounds 1, 8a,b, 9a,b,
10a,b, 11a,b, 12a,b, and 13a,b. This material is available
free of charge via the Internet at http://pubs.acs.org.
15
62b
87b
0
10
10
OL062434Q
(22) Mitra, A. K.; Ostashevsky, I.; Brewer, C. F. Int. J. Peptide Protein
Res. 1983, 22, 495-501. The reaction times reported in their optimized
conditions are fairly long (48 h at rt), attributed to a “deactivating effect”
by fluorine.
a Refers to partially purified HPLC yields (UV) except where indicated
otherwise. b Isolated and purified yield.
(23) Carpino, L. A.; Ionescu, D.; El-Faham, A. J. Org. Chem. 1996, 61,
2460-2465.
(24) Several observations are noteworthy in this regard. (i) No â-elimina-
tion side product was detected upon treatment of 13b with neat Et3N (>800
equiv) for 16 h at room temperature (HPLC). (ii) In ref 22, the authors
prepare Cbz-3-fluoroalanine in 82% yield in a procedure that involves
treatment for 2.5 h with NaOH (2N). (iii) In the work described by Patchett
et al. (see below), 21 equiv of LDA was required to eliminate HF in
[3-fluoro-D-Ala8]cyclosporin A. (Cf.: Patchett, A. A.; Taub, D.; Hensens,
O. D.; Goegelman, R. T.; Yang, L.; Dumont, F.; Peterson, L.; Sigal, N. H.
J. Antibiot. 1992, 45, 94-102.) Collectively these observations tend to
indicate that the tendency of 3-fluoroalanine to undergo â-elimination is a
liability mainly at the activated ester stage, but the fluoropeptide itself is
stable to this side reaction.
structures, i.e., formation of the N-Me-fluoroalanine-contain-
ing peptides. The susceptibility of fluoroalanine toward
â-elimination during the amide bond formation is docu-
mented in the literature.22 However, the optimal 3-fluoro-
alanine amide bond coupling conditions reported were not
very successful in the case of formation of 12b (entry 1,
Table 2). After several experiments, satisfactory conditions
for the formation of 12b were obtained through the use of
EDCI/HOAt coupling reagents together with the milder base
NMM (pKa ) 7.38,23 entry 5, Table 2). Under these
(25) For a recent report on the synthesis of 3-fluorodehydroalanines
see: Zhou, H.; van der Donk, W. A. Org. Lett. 2001, 3, 593-596.
5852
Org. Lett., Vol. 8, No. 25, 2006