Natural product derived inhibitors of lipoprotein associated phospholipase a2, synthesis and activity of analogues of sb-253514

Article abstract of DOI:10.1016/S0960-894X(00)00395-4

The synthesis of analogues of SB-253514, a novel natural product derived inhibitor of lipoprotein associated phospholipase A₂ (Lp-PLA₂), is described together with their ability to inhibit Lp-PLA₂. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0960-894X(00)00395-4

Bioorganic & Medicinal Chemistry Letters 10 (2000) 2015±2017
Natural Product Derived Inhibitors of Lipoprotein Associated
Phospholipase A2, Synthesis and Activity of Analogues of
SB-253514
Ivan L. Pinto,* Helen F. Boyd and Deirdre M. B. Hickey
SmithKline Beecham Pharmaceuticals, New Frontiers Science Park, Third Avenue, Harlow, Essex, CM19 5AW, UK
Received 4 May 2000; revised 3 July 2000; accepted 4 July 2000
AbstractÐThe synthesis of analogues of SB-253514, a novel natural product derived inhibitor of lipoprotein associated phospho-
lipase A2 (Lp-PLA2), is described together with their ability to inhibit Lp-PLA2. # 2000 Elsevier Science Ltd. All rights reserved.
Lipoprotein associated phospholipase A2 (Lp-PLA2),
also known as platelet-activating factor acetylhydro-
lyase, is a novel serine phospholipase that is responsible
for hydrolysing PAF and oxidatively modi®ed phospha-
tidylcholine.1 The hydrolysis products, lysophospha-
tidylcholine and oxidised free fatty acids, have been
shown to be pro-in¯ammatory causing macrophage
proliferation and endolithial dysfunction.2 4 It would be
expected therefore that inhibitors of Lp-PLA2 would
have a bene®cial role in the treatment of in¯ammatory
diseases such as atherosclerosis.5 As a result of a natural
product screening exercise for inhibitors of Lp-PLA2,
SB-253514 was identi®ed as one of a novel family of
cyclic enol-carbamates isolated from Pseudomonas
¯uorescens which were found to be potent inhibitors of
the enzyme.6 The novel structure and potency of this
natural product provided an attractive starting point for
a programme directed at developing inhibitors of Lp-
PLA2 for the treatment of atherosclerosis.
It has already been shown that the sugar moiety, an a-
rhamnose, did not contribute to the in vitro activity of
SB-253514 against the isolated Lp-PLA2 enzyme,6 and
it was speculated that the enol-carbamate moiety rather
than the imide function was acting as an acylating agent
to inactivate the lipase. As such we targetted the amide
analogue 1 as a suitable analogue for investigating the
scope of the enol-carbamate bicycle as a novel serine
lipase inhibitor.
Bicyclic enol-carbamates of this nature have not been
exempli®ed in the literature to our knowledge and ret-
rosynthetic analysis suggested that proline would be a
appropriate starting material for the synthesis. Since the
stereochemistry of the ring junction in the natural pro-
duct SB-253514 was not established at the outset of this
work we chose to use the natural (S) enantiomer of
proline (Scheme 1). Borane reduction of (S)-Boc-proline
followed by oxidation with pyridium chlorochromate
provided the chiral aldehyde 4, which was treated with
the anion of diazomethane to yield the acetylene 5 in
moderate yield.7 Cyclisation to the enol-carbamate was
e€ected by treatment with iodine and 1-[3-(dimethyl-
amino)propyl]-3-ethylcarbodiimide hydrochloride (DEC)
*Corresponding author. Fax: +44-1279-627841; e-mail: ivan_pinto-
1@sbphrd.com
0960-894X/00/$ - see front matter # 2000 Elsevier Science Ltd. All rights reserved.
PII: S0960-894X(00)00395-4

2016
I. L. Pinto et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2015±2017
Scheme 1.
in the presence of silver tetra¯uoroborate to yield a 9:1
E:Z mixture of vinyl iodides.8
be potent inhibitors of Lp-PLA2 (Table 1). A similar
time dependancy was observed for the natural product
inhibitor SB-253514 (Table 1).3 Potency for the amide
series 9 improved with increasing chain length and
lipophilicity, with the C-12 and C-16 chains 9a and 9c
demonstrating IC50s of 170 and 25 nm respectively. The
inhibitory activity of 9c was about 2-fold more potent that
the natural product SB-253514. However lipophilicity
does not appear to be the sole determinant of potency
since the alkynyl derivatives 10a±c were devoid of activity.
It may be possible that the amidic carbonyl, which is con-
jugated into the enol ether, activates the carbamate
towards the enzyme. It is unlikely that the inhibition of
Lp-PLA2 by 9 is simply a function of a highly reactive
acylating system as 7 was devoid of any activity. It
therefore appears that not only the activating capacity
of the amidic carbonyl but also a long fatty chain is
required for potency against the lipase. Stability studies
were also carried out on 9a at pH 7 (aqueous media)
which demonstrated a half-life of 62 h, suggesting this
series is not overtly hydrolytically unstable.
Conversion of the E-vinyl iodide 6 to the amide 9 was
achieved by ®rst carbonylation of 6 with carbon mon-
oxide in the presence of 2-trimethylsilylethanol and
palladium(0) followed by deprotection with ¯uoride
a€ording the acid 8 (Scheme 2).9 The acid 8 was a key
intermediate in the preparation of amide analogues such
as 9 through coupling with suitable amines using DEC,
while the iodide could also be converted to alkynyl
derivatives such as 10 in good yield (Scheme 2).9
The enol-carbamates were assayed against Lp-PLA2
using 1-decanoyl-2-(4-nitrophenylglutaryl)phosphatidyl-
choline as a substrate.5 In the case of the amide ana-
logues 9, a time dependant inhibition of the enzyme was
observed, such that without pre-incubation the IC50
varied from 2 to >10 mM. However a 10 minute pre-
incubation was sucient to observe maximum inhibition
of the enzyme, and the amide analogues 9a±c proved to
Scheme 2.

I. L. Pinto et al. / Bioorg. Med. Chem. Lett. 10 (2000) 2015±2017
2017
Table 1.
R
IC50 nM (10 min pre-incubation)
R1
IC50 nM (10 min pre-incubation)
9a=C12H25
9b=C14H29
9c=C16H31 (S isomer)
9d=C16H31 (R isomer)
170
28
25
12
10a=C4H9OH
10b=C14H29
Inactive
Inactive
Inactive
51
10c=C4H9O(CˆO)C7H15
SB-253514
References and Notes
All the above compounds were prepared from natural
(S)-proline, however the absolute stereochemistry of
SB-253514 was unknown at the inception of this pro-
gramme of work. We therefore prepared the (R) isomer
9d of the C16 compound 9c to determine the in¯uence of
the ring stereochemistry on potency. The (R) isomer 9d
was only two fold more potent than the (S) isomer 9c
and it suggests that inhibition of the enzyme is not cri-
tically dependant on the ring junction stereochemistry.
It was subsequently found that the stereochemistry of
the natural product ring junction was in fact (R).6 Inhi-
bitory tests of 9c against a range of serine proteases
including elastase, trypsin and chymotrypsin as well as
the metallo-protease thermolysin demonstrated no inhi-
bition of these enzymes, demonstrating this novel series
of serine lipase inhibitor is speci®c to Lp-PLA2.
1. Tew, D. G.; Southan, C.; Rice, S. Q.; Lawrence, M. P.; Li,
H.; Boyd, H. F.; Moores, K.; Gloger, I. S.; MacPhee, C. H.
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Biochem. J. 1999, 338, 479.
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In conclusion, a novel and potent series of lipase inhi-
bitor which is selective for Lp-PLA2 has been disclosed.
The inhibition of Lp-PLA2 could have therapeutic
potential for the treatment of atherosclerosis.
7. Miwa, K.; Aoyama, T.; Shioiri, T. Synlett 1994, 107.
8. Tam, T. F.; Thomas, E.; Krantz, A. Tetrahedron Lett.
1987, 28, 1127.
Acknowledgements
We would like to thank Simon Readshaw for NMR deter-
minations and David Tolson for mass spectrum studies.
9. Heck, R. F. Palladium Reagents in Organic Synthesis,
Academic, 1985.