Synthesis of photoactivatable acyclic analogues of the lobatamides

Article abstract of DOI:10.1021/jo0477751

(Chemical Equation Presented) The lobatamides and related salicylate enamide natural products are potent mammalian V-ATPase inhibitors. To probe details of binding of the lobatamides to mammalian V-ATPase, three photoactivatable analogues bearing benzophe

Full text of DOI:10.1021/jo0477751

Synthesis of Photoactivatable Acyclic Analogues of the
Lobatamides
Ruichao Shen,† Takao Inoue,‡ Michael Forgac,‡ and John A. Porco, Jr.*,†
Department of Chemistry and Center for Chemical Methodology and Library Development (CMLD-BU),
Boston University, 590 Commonwealth Avenue, Boston, Massachusetts 02215, and Tufts University
School of Medicine, Department of Physiology, Boston, Massachusetts 02111
porco@chem.bu.edu
Received December 20, 2004
The lobatamides and related salicylate enamide natural products are potent mammalian V-ATPase
inhibitors. To probe details of binding of the lobatamides to mammalian V-ATPase, three
photoactivatable analogues bearing benzophenone photoaffinity labels have been prepared. The
analogues were designed on the basis of a simplified acyclic analogue 2. Late-stage installation of
the enamide side chain and tandem deallylation/amidation were employed in synthetic routes to
these derivatives. Simplified analogue 2 showed strong inhibition against bovine clathrin-coated
vesicle V-ATPase (10 nM). Analogues 3-5 were also evaluated for inhibition of bovine V-ATPase
in order to select a suitable candidate for future photoaffinity labeling studies.
Introduction
c′′, d, and Ac45.5 Unlike other potent V-ATPase inhibitors
such as the bafilomycins and concanamycins which
inhibit both yeast and mammalian V-ATPases and have
been shown to bind to subunit c of the V0 domain,6,7 the
salicylate enamide natural products selectively inhibit
mammalian V-ATPases.3 Although extensive synthetic
studies have been performed on these natural products,8
studies regarding determination of the binding site of
these compounds on V-ATPase are limited at this stage.
A recent publication by Xie and co-workers disclosed that
the salicylate enamide natural product salicylihalamide
A binds to the V0 domain of bovine V-ATPase.9 However,
the particular subunit of the V0 domain for binding
remains to be determined. In addition, recent studies by
Huss et al.6 indicated that salicylihalamide A does not
compete with concanamycin A for binding to subunit c
of the V0 domain.
The salicylate enamide natural products, including the
lobatamides1 and salicylihalamides,2 are potent antitu-
mor compounds and mammalian vacuolar type proton
ATPase (V-ATPase) inhibitors.3 V-ATPases are essential
proton-translocating pumps of eukaryotic cells and play
important roles in many processes including receptor-
mediated endocytosis, acid secretion, bone degradation,
and control of cytoplasmic pH.4 The V-ATPases are
composed of two distinct domains: the cytoplasmic V1
domain responsible for binding and hydrolysis of ATP
and a membrane-embedded V0 domain responsible for
proton translocation across the membrane. For mam-
malian V-ATPase, the V0 domain contains subunits a, c,
† Boston University.
‡ Tufts University School of Medicine.
(1) McKee, T. C.; Galinis, D. L.; Pannell, L. K.; Cardellina, J. H., II;
Laasko, J.; Ireland, C. M.; Murray, L.; Capon, R. J.; Boyd, M. R. J.
Org. Chem. 1998, 63, 7805.
(5) For a recent structural study on V-ATPase, see: Wilkens, S.;
Inoue, T.; Forgac, M. J. Biol. Chem. 2004, 279, 41942.
(6) Huss, M.; Ingenhorst, G.; Ko¨nig, S.; Gassel, M.; Dro¨se, S.; Zeeck,
A.; Altendorf, K.; Wieczorek, H. J. Biol. Chem. 2002, 277, 40544.
(7) Bowman, E. J.; Graham, L. A.; Stevens, T. H.; Bowman, B. J. J.
Biol. Chem. 2004, 279, 33131.
(8) For a review on the chemistry and biology of salicylate enamide
natural products, see: Yet, L. Chem. Rev. 2003, 103, 4283.
(9) Xie, X.-S.; Padron, D.; Liao, X.; Wang, J.; Roth, M. G.; De
Brabander, J. K. J. Biol. Chem. 2004, 279, 19755.
(2) (a) Erickson, K. L.; Beutler, J. A.; Cardellina, J. H., II; Boyd, M.
R. J. Org. Chem. 1997, 62, 8188 (b) Erickson, K. L.; Beutler, J. A.;
Cardellina, J. H., II; Boyd, M. R. J. Org. Chem. 2001, 66, 1532.
(3) Boyd, M. R.; Farina, C.; Belfiore, P.; Gagliardi, S.; Kim, J. W.;
Hayakawa, Y.; Beutler, J. A.; McKee, T. C.; Bowman, B. J.; Bowman,
E. J. J. Pharmacol. Exp. Ther. 2001, 297, 114.
(4) For a comprehensive review on the V-ATPases, see: Nishi, T.;
Forgac, M. Nature Rev. Mol. Cell Biol. 2002, 3, 94.
10.1021/jo0477751 CCC: $30.25 © 2005 American Chemical Society
Published on Web 04/02/2005
3686
J. Org. Chem. 2005, 70, 3686-3692

Photoactivatable Acyclic Analogues of Lobatamides
Since photoaffinity reagents derived from natural
products and drugs have become powerful tools in explor-
ing the binding sites of target proteins,10 we have
initiated studies to prepare photoactivatable analogues
of the lobatamides in order to identify their binding
subunits on V-ATPase. In our previous synthesis of
lobatamide C and simplified analogues,11 acyclic analogue
1 was uncovered as a potent inhibitor against bovine
chromaffin granule membrane V-ATPase (IC50 ) 60 nM).
We thus selected three photoactivatable analogues 3-5
as synthetic targets. These three probe reagents were
designed on the basis of simplified analogue 2 in which
the base-sensitive â-hydroxy ester moiety of analogue 1
was altered to a 2-hydroxyethyl unit. This modification
provided an opportunity to install the enamide side chain
at a late stage using our methodology for Cu(I)-catalyzed
amidation of vinyl iodides.12 In addition, the primary
alcohol could also be utilized as a site of attachment for
photoactive groups. The benzophenone moiety was ini-
tially employed as a photoactive group due to its easy
availability, mild conditions required for photoactivation
(350-360 nm), and propensity for selective C-H abstrac-
tion.13 To obtain information on the influence of the site
of probe attachment with regard to the V-ATPase inhibi-
tion of these analogues, the benzophenone moiety was
positioned in different locations of analogue 2, i.e., the
enamide side chain, primary alcohol, and the salicylate
ring.14 After evaluation of nonradiolabeled probes for
mammalian V-ATPase inhibition, radiolabeled versions
incorporating tritium may be prepared for use in photo-
affinity studies.15 Herein, we report the synthesis of
lobatamide probe derivatives 2-5 and preliminary evalu-
ation of these compounds as bovine V-ATPase inhibitors.
Results and Discussion
Retrosynthesis of analogue 3 (Figure 1) led to N-
allyloxycarbonyl (alloc)-protected salicylate enamide 6
and N-succinimidyl p-benzoyldihydrocinnamate 7, the
latter developed by Prestwich and co-workers.15a The
N-alloc group was chosen due to its stability to basic
conditions, and the mild deprotection conditions16 com-
patible with the acid-sensitive enamide fragment.17 Cleav-
age of the enamide bond of protected derivative 6 affords
amide 8 and vinyl iodide 9, the latter a substrate for
CuTC-catalyzed amidation.11c,12 In contrast to our studies
toward the total synthesis of lobatamide C11a,c in which
the enamide moiety was installed at relatively early
stage, construction of the salicylate bond before the
enamide synthesis would provide divergent access to
analogues 2-4 from vinyl iodide 9. Further disconnection
(10) (a) Fleming, S. A. Tetrahedron, 1995, 46, 12479. (b) Dorma´n,
G. Top. Curr. Chem. 2000, 211, 169. (c) Hatanaka, Y.; Sadakane, Y.
Curr. Top. Med. Chem. 2002, 2, 271.
(11) (a) Shen, R.; Lin, C.-T.; Porco, J. A., Jr. J. Am. Chem. Soc. 2002,
124, 5650. (b) Shen, R.; Lin, C.-T.; Bowman, E. J.; Bowman, B. J.; Porco,
J. A., Jr. Org. Lett. 2002, 4, 3103. (c) Shen, R.; Lin, C.-T.; Bowman, E.
J.; Bowman, B. J.; Porco, J. A., Jr. J. Am. Chem. Soc. 2003, 125, 7889.
For syntheses of salicylihalamide A and apicularen A analogues, see:
(d) Wu, Y.; Liao, X.; Wang, R.; Xie, X-s.; De Brabander, J. K. J. Am.
Chem. Soc. 2002, 124, 3245. (e) Smith, A. B., III.; Zheng, J. Tetrahedron
2002, 58, 6455. (f) Lebreton, S.; Xie, X.-S.; Ferguson, D.; De Brabander,
J. K. Tetrahedron 2004, 60, 9635. (g) Nicolaou, K. C.; Kim, D. W.; Baati,
R.; O’Brate, A.; Giannakakou, P. Chem. Eur. J. 2003, 9, 6177.
(12) Shen, R.; Porco, J. A., Jr. Org. Lett. 2000, 2, 1333. For additional
reports concerning Cu-catalyzed synthesis of enamides, see: (b) Lam,
P. Y. S.; Vincent, G.; Bonne, D.; Clark, C. G. Tetrahedron Lett. 2003,
44, 4927. (c) Jiang, L.; Job, G. E.; Klapars, A.; Buchwald, S. L. Org.
Lett. 2003, 5, 3667. (d) Pan, X.; Cai, Q.; Ma, D. Org. Lett. 2004, 6,
1809.
(15) (a) Olszewski, J. D.; Dorman, G.; Elliott, J. T.; Hong, Y.; Ahern,
D. G.; Prestwich, G. D. Bioconjugate Chem. 1995, 6, 395. For photo-
affinity studies employing a tritium-labeled benzophenone moiety,
see: (b) Ojima, I.; Duclos, O.; Dorma´n, G.; Simonot, B.; Prestwich, G.
D. J. Med. Chem. 1995, 38, 8, 3891. (c) Andrus, M. B.; Turner, T. M.;
Sauna, Z. E.; Ambudkar, S. V. Bioorg. Med. Chem. Lett. 2000, 10, 2275.
(16) Guibe, F. Tetrahedron 1998, 54, 2967.
(13) Dorma´n, G.; Prestwich, G. D. Biochemistry 1994, 33, 5661.
(14) For representative studies on the synthesis of natural product
analogues by attachment of photoaffinity probes at diverse locations,
see: (a) Wang, P.; Blank, D. H.; Spencer, T. A. J. Org. Chem. 2004,
69, 2693. (b) Radeke, H. S.; Digits, C. A.; Bruner, S. D.; Snapper, M.
L. J. Org. Chem. 1997, 62, 2823.
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A.; Nowlan, V. J.; Tidwell, T. T. J. Am. Chem. Soc. 1979, 101, 974.
J. Org. Chem, Vol. 70, No. 9, 2005 3687

Shen et al.
FIGURE 1. Retrosynthetic analysis of photoactivatable analogue 3.
SCHEME 1. Synthesis of Vinyl Iodide 9
of 9 employs base-mediated ring-opening18 of benzo[1,3]-
dioxin-4-one 10 with alcohol 11.
trimethylsilylacetylene and subsequent protection of the
secondary alcohol 13 with p-methoxybenzyl trichloro-
acetimidate using camphorsulfonic acid as catalyst21
provided compound 14, which was further converted to
vinyl iodide 11 by desilylation, hydrozirconation/iodina-
tion, and deprotection of the p-methoxybenzyl ether.
Benzo[1,3]dioxin-4-one 10 was obtained by Stille coupling
of aryltriflate 1522 and tributylprenylstannane using
Synthesis of Analogue 3. Synthesis of alcohol 11
commenced with chiral epoxide 12,19 which was prepared
in good yield and high enantiopurity (>99% ee) from
3-buten-1-ol by silyl protection, epoxidation, and hydro-
lytic kinetic resolution20 (Scheme 1). BF3‚Et2O-mediated
epoxide opening with the lithium acetylide derived from
(20) Schaus, S. E.; Brandes, B. D.; Larrow, J. F.; Tokunaga, M.;
Hansen, K. B.; Gould, A. E.; Furrow, M. E.; Jacobsen, E. N. J. Am.
Chem. Soc. 2002, 124, 1307.
(18) (a) Bhattacharjee, A.; De Brabander, J. K. Tetrahedron Lett.
2000, 41, 8069. (b) Wang, X.; Porco, J. A., Jr. J. Am. Chem. Soc. 2003,
125, 6040.
(19) Cf. Romeril, S. P.; Lee, V.; Baldwin, J. E.; Claridge, T. D. W.;
Odell, B. Tetrahedron Lett. 2003, 44, 7757.
(21) Walkup, R. D.; Kane, R. R.; Boatman, P. D., Jr.; Cunningham,
R. T. Tetrahedron Lett. 1990, 31, 7587.
(22) Fu¨rstner, A.; Konetzki, I. Tetrahedron 1996, 52, 15071.
3688 J. Org. Chem., Vol. 70, No. 9, 2005

Photoactivatable Acyclic Analogues of Lobatamides
SCHEME 2. Synthesis of Dienoic Acid Amide 8
SCHEME 3. Enamide Coupling of 8 and 9
Engler’s protocol.23 NaHMDS-mediated ring-opening18 of
10 with secondary alcohol 11 furnished vinyl iodide 9
(91%).
3). Aldehyde 20 is likely derived from hydrolysis of vinyl
ether intermediate 9a which may be formed by inter-
molecular Cu(I)-mediated Ullman coupling of 9.27 After
considerable evaluation of different bases (K2CO3, Rb2-
CO3), supporting ligands (1,10-phenanthroline),11a and
solvents (THF)12c in the coupling reaction, no satisfactory
results were obtained. Protection of the phenol-OH with
either a triisopropylsilyl (TIPS) ether or 2-nitrobenzyl
group28 did not further improve the yield of the desired
salicylate enamide 6.
However, we were delighted to find that when the
phenol-OH of compound 9 was protected as an O-allyl
ether (Scheme 4), amidation of vinyl iodide 21 using
CuTC and N,N′-dimethylethylenediamine12c,29 proceeded
cleanly to afford enamide 22 in 51% yield. Although
O-allyl ethers and N-alloc group react differently under
Pd(0)-catalyzed deallylation conditions, it was anticipated
that, by choice of the appropriate allyl scavenger, both
the N-alloc and the O-allyl groups could be removed in a
one-pot operation.30 Application of Pd(PPh3)2Cl2/HSn-
Bu3,31 Pd(OAc)2/HSiMe2tBu/Et3N,32 and Pd(PPh3)4/di-
medone33 to enamides 6 and 22 either caused severe
decomposition or led to recovery of starting material.
The synthesis of dienoic acid amide 8 was initiated
from allylic alcohol 16, prepared from ethyl sorbate by a
bromination/hydrolysis sequence following the protocols
of Durrant24 and Closa25 (Scheme 2). Mitsunobu coupling
of 16 and diallyl imidodicarbonate 1726 smoothly provided
the desired bis(N-alloc)-protected allylic amine 18, which
was carefully hydrolyzed (aq LiOH, t-BuOH/MeOH) to
dienoic acid 19. In this transformation, one of the N-alloc
groups was simultaneously removed. Compound 8 was
prepared using mixed anhydride formation of dienoic acid
19 followed by quenching with aqueous ammonium
hydroxide.
Initial attempts at Cu(I)-catalyzed amidation of vinyl
iodide 9 with dienoic acid amide 8 were found to be
problematic due to likely interference by the free phenol-
OH. We found that aldehyde 20 was consistently the
major product of attempted amidation reactions with only
10% of the desired enamide product 6 obtained (Scheme
(23) Engler, T. A.; Reddy, J. P.; Combrink, K. D.; Velde, D. V. J.
Org. Chem. 1990, 55, 1248.
(24) Durrant, G.; Green, R. H.; Lambeth, P. F.; Lester, M. G.; Taylor,
N. R. J. Chem. Soc., Perkin Trans. 1 1983, 2211.
(25) Closa, M.; De March, P.; Figueredo, M.; Font, J.; Soria, A.
Tetrahedron 1997, 53, 16803.
(26) (a) Arnould, J. C.; Landier, F.; Pasquet, M. J. Tetrahedron Lett.
1992, 33, 7133. For a Mitsunobu reaction using 17, see (b) Kanno, O.;
Kawamoto, I. Tetrahedron 2000, 56, 5639.
(27) For a recent review of Cu-mediated Ullman condensation, see:
Ley, S. V.; Thomas, A. W. Angew. Chem., Int. Ed. 2003, 42, 5400.
(28) Amit, B.; Hazum, E.; Fridkin, M.; Patchornik, A. Int. J. Pept.
Protein Res. 1977, 9, 91.
(29) Klapars, A.; Huang, X.; Buchwald, S. L. J. Am. Chem. Soc. 2002,
124, 7421.
J. Org. Chem, Vol. 70, No. 9, 2005 3689

Shen et al.
SCHEME 4. Synthesis of Analogue 3
SCHEME 5. Synthesis of Analogues 2 and 4
Considering the labile amine intermediate 23, a tandem
deallylation/amidation employing polymer-supported al-
lyl scavengers would be ideal for the transformation to
the final target.30a,b Using Pd(PPh3)4 as catalyst, polymer-
supported N-hydroxyphthalimide,34 1-hydroxybenzotri-
azole-6-sulfonamidomethyl polystyrene (PS-HOBt),35 and
macroporous triethylammonium methylpolystyrene boro-
hydride (MP-BH4)36 were evaluated for in situ generation
of free amine 23 and tandem amide formation with
activated ester 7. Both PS-N-hydroxyphthalimide and
PS-HOBt were not effective in this process, while PS-
HOBt was found to remove the N-alloc group without
removal of the O-allyl ether. However, treatment of
enamide 22 with MP-BH4 (7.0 equiv), Pd(PPh3)4 (0.2
equiv) and activated ester 7 (2.0 equiv) in THF smoothly
provided the desired benzophenone-attached amide 24,
which was rapidly transformed to analogue 3 by de-
silylation with HF-pyridine/pyridine (30%, two steps)
(Scheme 4).
Syntheses of Analogues 2 and 4. CuTC/N,N′-di-
methylethylenediamine-catalyzed amidation of O-allyl-
protected salicylate 21 with butenamide 2511c afforded
enamide 26 (Scheme 5), an intermediate required for
preparation of analogues 2 and 4. While unprotected
salicylate 9 was used instead of 21, enamide 27 was
obtained in only 16% yield. Removal of the O-allyl group
of 26 using Pd(PPh3)4/MP-BH4, followed by deprotection
of the TIPS ether using HF-pyridine/pyridine, provided
analogue 2 (51%). Since the acid-sensitive enamide side
chain was not stable to other esterification conditions,11c
analogue 4 was obtained employing a Mitsunobu protocol
from analogue 2 and 3-(4-benzoylphenyl)propionic acid
28.15a,37
Synthesis of Analogue 5. Synthesis of analogue 5
was performed on the basis of a similar strategy em-
ployed for analogue 3. Since the benzophenone fragment
was attached to the aromatic ring of analogue 5, prepa-
ration of substituted benzo[1,3]dioxin-4-one 35 was re-
quired (Scheme 6). Therefore, Mitsunobu coupling of
commercial available compound 29 and monosilylated
ethylene glycol 3038 smoothly afforded aryl ether 31,39
which was subsequently transformed to aryl triflate 32.
(30) For simultaneous reductive deprotection of both O-allyl and
N-alloc groups using Pd(PPh3)4/LiBH4, followed by reaction with
activated esters to prepare amides, see: (a) Beugelmans, R.; Neuville,
L.; Bios-Choussy, M.; Chastanet, J.; Zhu, J. Tetrahedron Lett. 1995,
36, 3129. (b) Bios-Choussy, M.; Beugelmans, R.; Bouillon, J.-P.; Zhu,
J. Tetrahedron Lett. 1995, 36, 4781. For simultaneous deprotection of
O-allyl and N-alloc groups using Pd(PPh3)4/morpholine, see: (c) Pelish,
H. E.; Westwood, N. J.; Feng, Y.; Kirchhausen, T.; Shair, M. D. J. Am.
Chem. Soc. 2001, 123, 6740.
(31) Dangles, O.; Guibe, F.; Balavoine, G. J. Org. Chem. 1987, 52,
4984.
(32) Sakaitani, M.; Ohfune, Y. J. Org. Chem. 1990, 55, 870.
(33) Kunz, H.; Unverzagt, C. Angew. Chem., Int. Ed. Engl. 1984,
23, 436.
(34) Han, C.; Shen, R.; Su, S.; Porco, J. A., Jr. Org. Lett. 2004, 6,
27.
(35) For further information on PS-HOBT, see:
www.argotech.com.
http://
(36) For alloc deprotection using a polymer-supported Pd(0) catalyst
and a resin-bound borohydride, see: Bhattacharyya, S.; Vo, Lanchi.;
Gooding, O. W.; Labadie, J. W. Abstracts of Papers. 227th ACS
National Meeting; Anaheim, CA, March 2004; American Chemical
Society: Washington, DC, 2004; ORGN-101. For information on
MP-BH4, see: http://www.argotech.com/products/lead/resins/solutions/
mp_borohydride.html.
(37) De Kort, M.; Luijendijk, J.; Van der Marel, G. A.; Van Boom,
J. H. Eur. J. Org. Chem. 2000, 3085.
(38) McDougal, P. G.; Rico, J. G.; Oh, Y.-I.; Condon, B. D. J. Org.
Chem. 1986, 51, 3388.
(39) Cf. Takahashi, S.; Kamisuki, S.; Mizushina, Y.; Sakaguchi, K.;
Sugawara, F.; Nakata, T. Tetrahedron Lett. 2003, 44, 1875.
3690 J. Org. Chem., Vol. 70, No. 9, 2005

Photoactivatable Acyclic Analogues of Lobatamides
SCHEME 6. Synthesis of Compound 35
SCHEME 7. Synthesis of Analogue 5
TABLE 1. V-ATPase Inhibition of Analogues 2-5a
Stille coupling of tributylprenyl stannane and aryltriflate
32 afforded prenyl benzo[1,3]dioxin-4-one 33 in 93%
yield.23 Since Pd(0) was used in the Stille reaction, it was
necessary to install the N-alloc-protected amine at a later
stage. Desilylation of 33 with TBAF, followed by Mit-
sunobu coupling of alcohol 34 and diallyl imidodicarbon-
ate 17,26 furnished the desired product 35a, which was
converted to compound 35 by in situ removal one of the
N-alloc groups using aqueous LiOH (84%).
compd
IC50
lobatamide C
2 nM
10 nM
4 µM
100 nM
>10 µM
2
3
4
5
a Inhibition of bovine clathrin-coated vesicle V-ATPase. See the
Supporting Information for experimental details.
Treatment of compound 35a and alcohol 11 under basic
conditions (1.1 equiv of NaHMDS) provided 35 as the
major product instead of the desired desired salicylate
ester indicating the necessity for removal of the ad-
ditional N-alloc group of 35a prior to transesterification
of the benzo[1,3]dioxin-4-one moiety. Accordingly, treat-
ment of benzo[1,3]dioxin-4-one 35 and alcohol 11 with
2.0 equiv NaHMDS afforded vinyl iodide 36 (Scheme 7).
Protection of the phenol of 36 with allyl bromide followed
by Cu(I)-catalyzed enamide formation furnished the
desired enamide 37 in 48% yield (two steps). The tandem
deallylation/amidation process was performed similarly
as reported for analogue 3. Final deprotection of the TIPS
ether on the primary alcohol using HF-pyridine/pyridine
provided analogue 5 in good yield (67%, two steps).
V-ATPase Inhibition of Analogues 2-5. Analogues
2-5 were next evaluated for inhibition of bovine clathrin-
coated vesicle V-ATPase (Table 1). Simplified analogue
2 showed high potency for V-ATPase inhibition (10 nM)
in line with the potency of the natural product lobatamide
C (2 nM). Analogue 3 was found to be much less active
than the parent compound 2 (4 µM). Analogue 4 bearing
the benzophenone probe as an ester linkage was also
determined to be a weaker V-ATPase inhibitor than
compound 2 (100 nM). However, analogue 5 exhibited
negligible V-ATPase inhibition activity compared to other
analogues, which indicated that the 4-alkoxy subsititu-
tion of the salicylate to afford a resorcylate ring was not
well tolerated in the V-ATPase binding site. Analogue 4
effectively retained useful levels of V-ATPase inhibition
J. Org. Chem, Vol. 70, No. 9, 2005 3691

Shen et al.
which bodes well for further use of this probe reagent
and related derivatives as photoaffinity reagents.
all analogues tested and afforded the most potent simpli-
fied acyclic lobatamide analogue11c identified to date.
Photoactivatable analogue 4 retained much of the inhibi-
tion activity of parent analogue 2 and was superior to
analogues 3 and 5. Further studies employing analogue
4 and related compounds in photoaffinity labeling of
mammalian V-ATPase are currently in progress and will
be reported in due course.
Conclusion
In an effort to explore the binding site of lobatamides
on mammalian V-ATPase, photoactivatable analogues
3-5 based on the simplified lobatamide analogue 2 were
prepared. In initial studies, a benzophenone moiety was
employed as a photoactive group and attached to different
positions of parent analogue 2. The synthesis of ana-
logues included late stage installation of enamides from
amidation of vinyl iodides bearing allyl ether-protected
salicylate esters. An efficient tandem deallylation/ami-
dation process was used for final deprotection and
photoaffinity probe attachment. The synthetic route also
permits installation of other photoactive groups in addi-
tion to the benzophenone moiety or other reporting/
modifying groups (e.g., biotin) as required for particular
applications. Finally, analogues 2-5 have been tested for
inhibition of bovine clathrin-coated vesicle V-ATPase.
Compound 2 showed the highest potency of inhibition of
Acknowledgment. We thank the National Insti-
tutes of Health (GM 62842 to J.A.P., Jr., GM 34478 to
M.F.) for financial support of this study and Bristol-
Myers Squibb for an Unrestricted Grant in Synthetic
Organic Chemistry (J.A.P, Jr.). T.I. was supported by a
Postdoctoral Fellowship from the Northeast Affiliate of
the American Heart Association.
Supporting Information Available: Experimental pro-
cedures and characterization data for new compounds. This
material is available free of charge via the Internet at
http://pubs.acs.org.
JO0477751
3692 J. Org. Chem., Vol. 70, No. 9, 2005