Chemoselective protection of solid-phase compatible Fmoc-phosphinic building blocks

Article abstract of DOI:10.1021/jo061535z

(Chemical Equation Presented) An efficient four-step synthetic strategy able to fully discriminate hydroxyphosphinyl and carboxylic groups of Fmoc-phosphinic building blocks and related analogues has been developed. The proposed method applies selective a

Full text of DOI:10.1021/jo061535z

strategy to phosphinic peptides requires the use of an orthogonal
protection scheme, compatible with Fmoc solid-phase chemistry.
Although two alternative synthetic strategies provide such
blocks,6 both require careful planning in terms of protection
and deprotection steps of the numerous functional groups of
the pseudodipeptidic unit. The chemoselective deprotection of
either the hydroxyphosphinyl by TFA,7 TMSBr,8 LiI,9 LiBr,10
and phosphodiesterase I11 or the carboxylate group by Carlsberg
subtilisin,12 pig liver esterase,13 or (Bu3Sn)2O14 of phosphinic
peptide blocks has been achieved for methyl and ethyl esters,
which are not compatible with the Fmoc solid-phase peptide
synthesis (SPPS) protocol.
Chemoselective Protection of Solid-Phase
Compatible Fmoc-Phosphinic Building Blocks
Magdalini Nasopoulou,† Magdalini Matziari,†
Vincent Dive,‡ and Athanasios Yiotakis*,†
UniVersity of Athens, Department of Chemistry, Laboratory of
Organic Chemistry, Panepistimiopolis Zografou 15771, Athens,
Greece, and CEA, De´partement d’Inge´nierie et d’ Etudes des
Prote´ines, 91191 Gif/YVette Cedex, France
yiotakis@chem.uoa.gr
We report herein a new chemoselective strategy, able to
discriminate between the two acidic groups of the phosphinic
units, namely, the hydroxyphosphinyl and the carboxylic acid
functionalities. Although phosphinic building blocks of type 1
(Scheme 1) which are unprotected at the hydroxyphosphinyl
group have been successfully used in SPPS,2,15,16 the use of
protected blocks of type 5 remains the method of choice for
the development of phosphinic peptide libraries devoid of
byproducts and incomplete reactions.
ReceiVed July 24, 2006
Fmoc-phosphinic building blocks of type 1 have been recently
reported to be easily synthesized by a three-component con-
densation reaction.16 Any attempt to discriminate between the
two acidic functionalities, using a variety of protecting groups,
(e.g., tert-butyl, benzyl, diphenylmethyl, trityl, etc.) has failed.17
In practice, a phenacyl (Pac) protecting group offered the
expedient method for the efficient discrimination between the
two acidic functionalities. The simultaneous protection of both
hydroxyphosphinyl and carboxylic functional groups by Pac was
easily achieved using PacBr in the presence of Et3N, in very
good yields.
An efficient four-step synthetic strategy able to fully
discriminate hydroxyphosphinyl and carboxylic groups of
Fmoc-phosphinic building blocks and related analogues has
been developed. The proposed method applies selective
acidic removal of the phenacyl (Pac) group from the
hydroxyphosphinyl functionality and protection by the 1-ada-
mantyl (Ad) group. Reductive removal of the Pac group from
the carboxylic functionality leads to Fmoc-protected phos-
phinic pseudodipeptidic units suitable for phosphinic peptide
and library development using solid-phase peptide synthesis
(SPPS).
Completely chemoselective deprotection of the hydroxyphos-
phinyl functional group of building blocks 2 (Scheme 1) was
accomplished under acidic conditions (TFA/CH2Cl2), where the
carboxylate Pac-ester remains perfectly intact. Hydroxyphos-
The intensive use of combinatorial chemistry in the discovery
of novel biologically active compounds has provided large
amounts of information concerning structure-activity relation-
ships. Solid-phase synthesis has been the major tool of com-
binatorial chemistry with apparent advantages over solution-
phase chemistry.1
Synthesis of phosphinic peptides has been successfully
accomplished on solid phase, using either parallel2 or combi-
natorial strategies.3 Such approaches have led to the rapid access
and identification of not only highly potent but also selective
inhibitors of various zinc metalloproteases, such as angiotensin-
converting enzyme (ACE)4 and matrix metalloproteinases
(MMPs),5 among others.
(4) (a) Dive, V.; Cotton, J.; Yiotakis, A.; Michaud, A.; Vassiliou, S.;
Jiracek, J.; Vazeaux, G.; Chauvet, M.-T.; Cuniasse, P.; Corvol, P. Proc.
Natl. Acad. Sci. U.S.A. 1999, 96, 4330. (b) Georgiadis, D.; Beau, F.; Czarny,
B.; Cotton, J.; Yiotakis, A.; Dive, V. Circ. Res. 2003, 93, 148.
(5) Devel, L.; Rogakos, V.; David, A.; Makaritis, A.; Beau, F.; Cuniasse,
F.; Yiotakis, A.; Dive, V. J. Biol. Chem. 2006, 281, 11152.
(6) (a) Yiotakis, A.; Vassiliou, S.; Jiracek, J.; Dive, V. J. Org. Chem.
1996, 61, 6601. (b) Georgiadis, D.; Matziari, M.; Yiotakis, A. Tetrahedron
2001, 57, 3471.
(7) Reiter, L. A.; Jones, B. P. J. Org. Chem. 1997, 62, 2808.
(8) Allen, M. C.; Fuhrer, W.; Tuck, B.; Wade, R.; Wood, J. M. J. Med.
Chem. 1989, 32, 1652.
(9) Parsons, W. H.; Patchett, A. A.; Bull, H. G.; Schoen, W. R.; Taub,
D.; Davidson, J.; Combs, P. L.; Springer, J. P.; Gadebusch, H.; Weissberger,
B.; Valiant, M. E.; Mellin, T. N.; Busch, R. D. J. Med. Chem. 1988, 31,
1772.
(10) Cristau, H. J.; Herve, A.; Virieux, D. Tetrahedron 2004, 60, 877.
(11) Natchev, I. A. Phosphorus Sulfur 1988, 37, 133.
(12) Morgan, B. P.; Scholtz, J. M.; Ballinger, M. D.; Zipkin, I. D.;
Bartlett, P. A. J. Am. Chem. Soc. 1991, 113, 297.
The most popular strategy to synthesize natural peptides on
solid support applies the Fmoc protocol. Application of this
(13) Ross, F. C.; Botting, N. P.; Leeson, P. D. Biorg. Med. Chem. Lett.
1996, 22, 2643.
(14) Miller, D. J.; Hammond, S. M.; Anderluzzi, D.; Bugg, T. D. H. J.
Chem. Soc., Perkin Trans. 1 1998, 131.
(15) (a) Makaritis, A.; Georgiadis, D.; Dive, V.; Yiotakis, A. Chem.-
Eur. J. 2003, 9, 2079. (b) Matziari, M.; Georgiadis, D.; Dive, V.; Yiotakis,
A. Org. Lett. 2001, 3, 659.
* To whom correspondence should be addressed. Tel/Fax: 0030-210-7274498.
† University of Athens.
‡ CEA.
(1) Fassina, G.; Miertus, S. In Combinatorial Chemistry and Technolo-
gies. Methods and Applications; Marcel Dekker Ltd: New York, 2005.
(2) Matziari, M.; Beau, F.; Cuniasse, P.; Dive, V.; Yiotakis, A. J. Med.
Chem. 2004, 47, 325.
(3) Jiracek, J.; Yiotakis, A.; Vincent, B.; Checler, F.; Dive, V. J. Biol.
Chem. 1996, 271, 19606.
(16) Matziari, M.; Yiotakis, A. Org. Lett. 2005, 7, 4049.
(17) These results will be published elsewhere.
10.1021/jo061535z CCC: $33.50 © 2006 American Chemical Society
Published on Web 11/15/2006
J. Org. Chem. 2006, 71, 9525-9527
9525

TABLE 1. Yields and Side Chains of Compounds 2-5
yields of
yields of
yields of
yields of
entry
R1
R2
PhCH2-
2 (%)
3 (%)
4 (%)
5 (%)
a
b
c
d
e
f
H-
70
78
72
80
82
81
81
82
62
82
65
75
78
60
71
70
61
90
60
85
74
84
73
62
89
81
85
83
91
87
80
86
CH3-
PhCH2-
PhCH2-
PhCH2-
PhCH2-
PhCH2-
(CH3)2CHCH2-
H
(CH3)2CH-
CH3OOCCH2CH2-
Ph-
PhCH2OCH2-
CH3-
(CH3)2CHCH2-
g
h
SCHEME 1. Chemoselective Protection of Fmoc
Pseudodipeptidic Building Blocks 5
SCHEME 2. Phosphinic Ester Acidic Cleavage via
â-Carbamido-Induced Assistance
was synthesized19 and subjected to the same acidic conditions
as compounds 2 (Scheme 1). In this case, Pac-cleavage did not
take place from the hydroxyphosphinyl or from the carboxylic
functional groups.
On the contrary, acidic treatment of compound 7 (Scheme
2) resulted in the selective Pac-phosphinyl deprotection, leading
to compound 8 (Scheme 2). These results are consistent with
pertinent reports of the literature, where assisted acidic cleavage
of methyl â-carboxyphosphinates,20 or â-carboxamides,7 was
observed. According to those results, the cleavage of methyl
phosphinate is promoted by the â-carboxamide or the â-car-
boxylic groups due to the formation of a five-membered reactive
intermediate. In the present case, a similar mechanistic pathway
is proposed, involving an R-fluorenylmethyl carbamide-induced
cleavage via a five-membered cyclic intermediate formation
(Scheme 3). Alternatively, electrophilic activation of the PdO
group has also been proposed to proceed through hydrogen
bonding with the carbamate N-H.21 However, compound 6
cannot form such a reactive intermediate, and therefore, the Pac-
cleavage is not favorable in this case.
In conclusion, we have described here an efficient method
for the chemoselective protection of the hydroxyphosphinyl
group of Fmoc-phosphinopeptidic building blocks, suitable for
SPPS and combinatorial library development. To the best of
our knowledge, the proposed strategy constitutes the first
successful method to the vexing problem of discriminating
between the two acidic functionalities of the Fmoc-phosphinic
units. The â-carbamido-induced selective Pac-phosphinic ester
cleavage can thus be applied to any Fmoc-protected R-amino-
phosphinic analogues. Such blocks are of great importance in
the development of potent and selective Zn-metalloprotease
phosphinic inhibitors.
phinyl reprotection by the 1-Ad group6 (3) and final removal
of the Pac-protecting group from the carboxylic functionality
of 4 by metal reduction18 afforded the target compounds 5 in
good yields (Table 1). As it has been previously shown,
phosphinic blocks of type 1 are formed as two pairs of
enantiomers, namely, four diastereoisomers, in nearly the same
amount, which can be easily separated by RP-HPLC, once
incorporated into a peptide sequence.16 Although protection of
the hydroxyphosphinyl group, as present in compounds 5,
creates a new stereogenic center, removal of the Ad protecting
group simultaneously with peptide cleavage from the resin
reconstitutes the number and ratio of the aforementioned
diastereoisomers.
To examine the applicability of the proposed method to
nonpeptidic phosphinic compounds, compound 6 (Scheme 2)
(19) For experimental details, see the Supporting Information file.
(20) Georgiadis, D.; Dive, V.; Yiotakis, A. J. Org. Chem. 2001, 66, 6604.
(21) Smith, A. B.; Ducry, L.; Corbett, M.; Hirschmann, R. Org. Lett.
2000, 2, 3887.
(18) Kokinaki, S.; Leondiadis, L.; Ferderigos, N. Org. Lett. 2005, 7, 1723.
9526 J. Org. Chem., Vol. 71, No. 25, 2006

1
SCHEME 3. Proposed Mechanism for the Acid-Catalyzed
Cleavage of the Phosphinic Ester via â-Carbamido-Induced
Assistance
) 4:1) 0.30. HPLC tR 48.42, 48.69. H NMR (200 MHz, CDCl3)
δ 1.27-1.43 (m, 3H), 1.94-2.06 (m, 1H), 2.24-2.36 (m, 1H),
2.89-3.00 (m, 1H), 3.22-3.28 (m, 2H), 4.08-4.35 (m, 4H), 5.18-
5.36 (m, 2H), 5.77-5.84 (m, 1H), 7.18-7.83 (m, 18H). 13C NMR
(50 MHz, CDCl3) δ 13.9, 14.1, 26.3, 26.8, 28.2, 28.6, 29.6, 39.2,
40.7, 44.2, 44.7, 46.4, 47.0, 66.2, 67.2, 119.9, 125.1, 126.7, 127.0,
127.7, 127.8, 128.5, 128.7, 129.1, 133.9, 137.6, 141.2, 143.7, 143.8,
155.9, 173.4, 192.4. 31P NMR (81 MHz, CDCl3) δ 53.94, 54.42.
ESMS m/z calcd for C35H35NO7P (M + H)+ 612.2, found 612.3.
General Method for Phosphinate Adamantyl Protection for
Compounds of Type 4. Phosphinic ester of type 3 (1 mmol) and
1-AdBr (1.2 mmol) are dissolved in CHCl3 (20 mL), and the
reaction mixture is refluxed. To this refluxing mixture, silver oxide
(1.2 mmol) is added in five equal portions over 50 min. After the
solution is refluxed overnight, the solvent is removed in vacuo and
the residue is diluted in Et2O and filtered through a pad of celite.
Evaporation of the solvent and purification by column chromatog-
raphy, using P.E./AcOEt ) 1:2 as the eluent system, afford the
pure products as white solids, after trituration with petroleum ether
40-60 °C.
3-{(Adamantan-1-yloxy)-[1-(9H-fluoren-9-ylmethoxycarbony-
lamino)-ethyl]-phosphinoyl}-2-benzyl-propionic Acid 2-Oxo-2-
phenyl-ethyl Ester (4b). White solid. Yield 90%, 671 mg. TLC
1
Rf (CHCl3/MeOH )95:5) 0.62. H NMR (200 MHz, CDCl3) δ
1.25-1.42 (m, 3H), 1.56-1.58 (m, 7H), 1.91-2.08 (m, 9H), 2.27-
2.55 (m, 1H), 2.81-2.90 (m, 1H), 3.22-3.30 (m, 2H), 4.06-4.23
(m, 1H), 4.31-4.37 (m, 3H), 5.15-5.94 (m, 3H), 7.18-7.86 (m,
18H). 13C NMR (50 MHz, CDCl3) δ 14.6, 14.8, 30.3, 30.8, 31.1,
31.2, 35.6, 39.7, 39.9, 41.7, 42.1, 44.1, 44.2, 45.0, 47.1, 66.1, 66.3,
67.2, 83.6, 119.9, 125.2, 125.5, 126.8, 127.0, 127.7, 128.5, 128.6,
128.8, 129.3, 133.8, 134.1, 138.0, 138.1, 141.2, 143.8, 143.9, 155.9,
173.8, 192.1. 31P NMR (81 MHz, CDCl3) δ 47.81, 48.56. ESMS
m/z calcd for C45H49NO7P (M + H)+ 746.3, found 746.2.
General Method for Selective Deprotection of Carboxylate
Esters for Compounds of Type 5. To a solution of phosphinic
diester of type 4 (1 mmol) in MeOH/DMF (4:1) (17 mL) were
added AcOH (35 mmol) and Mg turnings (17 mmol). After stirring
for 4 h at rt, the reaction mixture was filtered. The filtrate is
concentrated in vacuo, and the residue is diluted with DCM, washed
with HCl (1 N), and brine and dried over Na2SO4. Evaporation of
the solvent and purification by column chromatography, using
CHCl3/MeOH ) 9.8:0.2 as the eluent system, afford the pure
products as white solids, after trituration with petroleum ether 40-
60 °C.
3-{(Adamantan-1-yloxy)-[1-(9H-fluoren-9-ylmethoxycarbony-
lamino)-ethyl]-phosphinoyl}-2-benzyl-propionic Acid (5b). White
solid. Yield 81%, 508 mg. TLC Rf (CHCl3/MeOH ) 95:5) 0.31.
1H NMR (200 MHz, CDCl3) δ 1.25-2.40 (m, 20H), 2.42-2.79
(m, 1H), 2.98-3.41 (m, 2H), 3.85-4.42 (m, 4H), 6.62-6.57 (m,
1H), 7.17-7.89 (m, 13H). 13C NMR (50 MHz, CDCl3) δ 13.6,
14.1, 14.9, 29.3, 29.7, 30.8, 31.1, 35.2, 35.4, 40.2, 40.4, 41.8, 42.6,
44.0, 44.1, 44.2, 44.3, 47.0, 67.5, 83.1, 84.6, 119.8, 125.4, 125.7,
126.0, 126.6, 127.1, 127.6, 128.5, 129.0, 129.4, 138.0, 138.5, 140.0,
141.2, 143.7, 144.0, 144.3, 156.4, 157.1, 176.1, 176.4. 31P NMR
(81 MHz, CDCl3) δ 48.99, 52.09. ESMS m/z calcd for C37H43-
NO6P (M + H)+ 628.3, found 628.1.
Experimental Section
General Method for Diphenacyl Protection for Compounds
of Type 2. Phosphinic acid of type 1 (1 mmol) is dissolved in
AcOEt (1 mL) then cooled at 0 °C, and PacBr (4 mmol) is added,
followed by Et3N (4 mmol). After stirring at rt overnight, the
reaction mixture is diluted with AcOEt, washed with HCl (1 N),
and brine and dried over Na2SO4. Evaporation of the solvent and
purification by column chromatography, using CHCl3/MeOH ) 9.8:
0.2 as the eluent system, afford the pure products as white solids,
after trituration with petroleum ether 40-60 °C.
2-Benzyl-3-[[1-(9H-fluoren-9-ylmethoxycarbonylamino)-ethyl]-
(2-oxo-2-phenyl-ethoxy)-phosphinoyl]-propionic Acid 2-Oxo-2-
phenyl-ethyl Ester (2b). White solid. Yield 78%, 569 mg. TLC
1
Rf (CHCl3/MeOH ) 95:5) 0.63. H NMR (200 MHz, CDCl3) δ
1.24-1.46 (m, 3H), 2.06-2.19 (m, 2H), 2.28-2.60 (m, 1H), 2.88-
3.07 (m, 1H), 3.27-3.40 (m, 2H), 4.09-4.41 (m, 3H), 5.14-5.60
(m, 4H), 5.71-6.34 (m, 1H), 7.12-7.90 (m, 23H). 13C NMR (50
MHz, CDCl3) δ 14.6, 14.8, 27.6, 39.3, 39.5, 40.8, 40.9, 45.2, 47.0,
66.1, 66.3, 67.2, 119.9, 125.1, 126.7, 127.0, 127.7, 128.5, 128.8,
128.9, 129.1, 129.2, 133.8, 133.9, 134.2, 137.6, 141.1, 143.7, 155.8,
173.6, 191.7. 31P NMR (81 MHz, CDCl3) δ 55.28, 55.45, 56.14.
ESMS m/z calcd for C43H41NO8P (M + H)+ 730.3, found 730.3.
General Method for Selective Cleavage of Phosphinate
Phenacyl Ester for Compounds of Type 3. A phosphinic diester
of type 2 (1 mmol) is dissolved in CH2Cl2 (1.6 mL), and TFA (6.4
mL) is added. The reaction mixture is allowed to stir at 50 °C
overnight. Evaporation of the solvent and purification by column
chromatography, using CHCl3/MeOH/AcOH ) 7:0.2:0.2 as the
eluent system, afford the pure products as white solids, after
trituration with petroleum ether 40-60 °C.
Acknowledgment. This work was supported in part by the
European Commission (FP5RDT, QLK3-CT02-02136 and
FP6RDT, LSHC-CT-2003-503297), by funds from the Labora-
tory of Organic Chemistry, and by the Special Account for
Research Grants of National Athens University (NKUA).
Supporting Information Available: Detailed experimental
procedures, spectroscopic and analytical data for all compounds,
copies of 1H, 13C, and 31P NMR spectra for compounds 3a-h, 5a-
h, 2b, 3b, and 4b, and HPLC chromatograms for compounds 3a-
h. This material is available free of charge via the Internet at
http://pubs.acs.org.
2-Benzyl-3-{[1-(9H-fluoren-9-ylmethoxycarbonylamino)-ethyl]-
hydroxy-phosphinoyl}-propionic Acid 2-Oxo-2-phenyl-ethyl Es-
ter (3b). White solid. Yield 82%, 502 mg. TLC Rf (CHCl3/MeOH
JO061535Z
J. Org. Chem, Vol. 71, No. 25, 2006 9527