Dihydroxyphenylisoindoline amides as orally bioavailable inhibitors of the heat shock protein 90 (Hsp90) molecular chaperone

Article abstract of DOI:10.1021/jm901209q

The discovery and optimization of potency and metabolic stability of a novel class of dihyroxyphenyl-isoindoline amides as Hsp90 inhibitors are presented. Optimization of a screening hit using structure-based design and modification of log D and chemical

Full text of DOI:10.1021/jm901209q

J. Med. Chem. 2010, 53, 499–503 499
DOI: 10.1021/jm901209q
Dihydroxyphenylisoindoline Amides as Orally Bioavailable Inhibitors of the Heat Shock
Protein 90 (Hsp90) Molecular Chaperone†
Pei-Pei Kung,* Buwen Huang, Gang Zhang, Joe Zhongxiang Zhou, Jeff Wang, Jennifer A. Digits, Judith Skaptason,
Shinji Yamazaki, David Neul, Michael Zientek, Jeff Elleraas, Pramod Mehta, Min-Jean Yin, Michael J. Hickey,
Ketan S. Gajiwala, Caroline Rodgers, Jay F. Davies II, and Michael R. Gehring
Pfizer Global Research and Development, La Jolla Laboratories, 10770 Science Center Drive, San Diego, California 92121
Received June 16, 2009
The discovery and optimization of potency and metabolic stability of a novel class of dihyroxyphenyl-
isoindoline amides as Hsp90 inhibitors are presented. Optimization of a screening hit using structure-
based design and modification of log D and chemical structural features led to the identification of a
class of orally bioavailable non-quinone-containing Hsp90 inhibitors. This class is exemplified by 14
and 15, which possess improved cell potency and pharmacokinetic profiles compared with the original
screening hit.
Introduction
throughput screening. They are currently in phase I/II, phase
II, and phase I trials, respectively, and their preclinical
pharmacokinetic and efficacy results have been reported in
the literature.7-9
Molecular chaperones are protein machines that are res-
ponsible for the correct folding, stabilization, and function of
other proteins in the cell.1 Among all the molecular chaper-
ones, a 90 kDa heat shock protein (Hsp90a) has became an
exciting target for cancer therapy.2 Overexpression of Hsp90
in solid and hematologic tumors also suggests a role for the
chaperone in oncogenesis.3 The known Hsp90 antibiotics
include the N-terminus ATP-competitive binders (such as
geldanamycin (GA) and radicicol4) and the C-terminus
ATP-site binder, such as novobiocin and its derivatives.5
Compound 1a (GA) and its 17-substituted semisynthetic
analogues (17-allylaminogeldanamycin (17-AAG), 17-di-
methylaminogeldanamycin (17-DMAG), and 17-propylami-
nogeldanamycin (17-PGA), Figure 1) were reported in the
literature over the past decade. 1a and 1b (17-AAG) display
efficacy in preclinical tumor models but suffer from poor
solubility,6a limited bioavailaility,6b hepatotoxicity,6c and ex-
tensive metabolism by polymorphic enzymes.6d These liabil-
ities complicate further pharmaceutical development of these
agents. The undesirable chemical features associated with 1a
and its analogues (quinone moiety, relatively high molecular
weight) have led to siginificant efforts to identify novel non-
quinone-containing small molecule inhibitors of Hsp90.
NVP-AUY922,7 BIIB021,8 and SNX54229 (Figure 2) are
recent examples of such inhibitors that were identified by high
Result and Discussion
Our Hsp90 research efforts led to the initial discovery of
dihydroxylphenylpyrrolidine 2a (Table 1) which displayed
reasonable competitive binding properties (IC50 of 20 nM)
and an IC50 of 1 μM as assessed by the degradation of Hsp90
client protein, Akt, in H1299 cells.10 This compound was
stable in a human liver microsome assay (HLM)11 but was
unstable in a human hepatocyte metabolism assessment
(hHep). The instability in the hHep assay of 2a was identified
by mass difference from the parent compound and is due to
phase II conjugation of the phenolic groups present in the
molecule.12 Similar in vitro and in vivo instability was ob-
served for a related resorcinol-containing Hsp90 inhibitor
previously described in the literature.7 Therefore, the main
priorities of the lead optimization effort associated with this
dihydroxyphenyl class of inhibitor were to improve Hsp90
inhibitory potency and compound stability in the hHep assay.
Our first laboratory objectives were to improve the cellular
inhibitory potency of 2a and to explore the correlation
between predicted lipophilicity (clogD) of related compounds
and their clearance13 in the hHep assay. Compound 3
(Table 1) reduced lipophilicity by 2 log units relative to 2a
†Coordinates of Hsp90 complexes with 2a, 2b, and 15 have been
deposited in the Protein Data Bank under accession codes 3K97, 3K99,
and 3K98.
*To whom correspondence should be addressed. Phone: 858-526-
4867. Fax: 877-481-1781. E-mail: peipei.kung@pfizer.com.
a Abbreviations: Hsp90, 90 kDa heat shock protein; 17-AAG, 17-
allylamino-17-demethoxygeldanamycin; 17-DMAG, 17-dimethylami-
no-17-demethoxygeldanamycin; 17-PGA, 17-propylamino-17-demeth-
oxygeldanamycin; HTS, high throughput screening; EtOAc, ethyl acet-
ate; NaOAc-HOAc, sodium acetate and acetic acid; DMSO-d6, hex-
adeuteriodimethyl sulfoxide; DCM, dichloromethane; MeOH,
methanol; THF, tetrahydrofuran; DMF, N,N-dimethylformamide;
LC/MS, liquid chromatogrpahy-mass spectrametry.
Figure 1. Structures of quinone ansamycins (1a-d).
r
2009 American Chemical Society
Published on Web 11/12/2009
pubs.acs.org/jmc

500 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Kung et al.
Figure 2. Structures of known Hsp90 inhibitors.
Table 1. Relationship between Lipophilicity and in Vitro Human
Hepatocyte Clearance
Figure 4. Schematic diagram showing the key interactions of 2b
with Hsp90 enzyme. Water molecules in red represent structurally
conserved water molecules.
Figure 5. Overlap of the cocrystal structures of 2a and 2b with
human Hsp90 enzyme.
Table 2. SAR of 5-Chloro-2,4-dihydroxyphenylisoindoline-1-carboxy-
amide
a Enzymatic Ki was calculated from enzymatic IC50 using Cheng-
Prusoff equation.
cellb
IC50
CLhHep
(μL/min)/
a
Ki
compd chirality
R
(nM) (μM) clogD million cells
7
R/S
R/S
R/S
R/S
R/S
R/S
R/S
R
N(CH3)2
N(CH2)4
20
10
7
1.5
0.6
1.5
0.3
1.7
1.5
0.7
0.6
0.7
1.6
1.4
0.9
0.4
1.2
2.2
1.6
0.9
0.6
2.4
2
8
9
NH(CH2)2CH3
NHCH2CH3
NHCH3
9
10
11
12
13
14
15
1c
4
4.7
<2
8.5
25
11
7
40
5
NHCH2CHdCH2
NHCH2Ph
N(CH2)4
20
4
R
NHCH2CH3
<1 0.3
250 0.1
Figure 3. Hsp90 isoindoline lead structures.
<3
a Enzyme Ki. b H1299 cell.
by removing the 2-methylphenyl moiety (from 2.9 to 0.8) but
did not reduce the corresponding clearance in the hHep assay.
Compounds 4, 5, and 6 were designed to introduce an amide
moiety to occupy a hydrophobic Hsp90 pocket (formed by
Gly135, Val136, and Tyr139) that was described in a previous
publication.10 These inhibitors displayed reduced lypophilicity
relative to 2a. Surprisingly, 6, which has a similar clogD
compared with 3, improved the in vitro clearance from 56 to
9 (μL/min)/million cells. Compounds 4, 5, and 6 (Table 1) all
displayed reduced in vitro clearance compared with 2a and 3

Brief Article
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 501
Scheme 1. General Synthesis of the 2-(5-Chloro-2,4-dihydroxybenzoyl)-2,3-dihydro-1H-isoindole-1-carboxylic Amidesa
a Reagents and conditions: (a) 4-methylmorpholine, N-(3-dimethylaminopropyl)-N0-ethylcarbodiimide hydrochloride, 1-hydroxybenzotriazole,
DMF, 23 °C, 12 h, 64%; (b) NaOH(aq) (2 M), MeOH, 23 °C, 12 h, 94%; (c) various amines (both commercially available or synthetic amines),
4-methylmorpholine, N-(3-dimethylaminopropyl)-N0-ethylcarbodiimide hydrochloride (EDC), 1-hydroxybenzotriazole, DMF, 23 °C, 12 h, 64.2%;
(d) HCl (in dioxane, 4 M), DCM, 23 °C, 12 h, 82%.
Table 3. Rat PK Profiles of Two Compounds
14
15
In Vitro
In Vivo
hHep CL ER
rat Hep CL ER
0.2
0.2
0.3
0.2
Cl ((L/h)/kg)
AUC0-inf (μg h/mL)
1.4
2.9
1.4
1.3
3
(po) (10 mg/kg)
AUC0-inf (μg h/mL)
(iv) (2.5 mg/kg)
2.1
1.7
3
V
dss (L/kg)
8.1
7.9
35
3.3
5.2
20
T1/2 (h)
F (%)a
a Foral = (doseiv ꢀ AUCpo)/(dosepo ꢀ AUCiv).
Figure 6. Crystal structure of 15 with human Hsp90 enzyme.
Table 4. Mouse Plama Levels of Compound 15 through Different
Administration Routes
and possessed clogD values from -0.3 to 1.4. We therefore
speculate that 4-6 contain a unique structural feature that
reduces the glucuronidation of the associated resorcinol phe-
nolic moieties.
adm route
dose (mg/kg)
iv
20
ip
100
po
100
8.3
810
T1/2 (h)
free plasma
5.2
286
To facilitate potency improvements of 4-6, we sought to
gain additional interactions with the Hsp90 enzyme. Another
previously discovered HTS hit, 2b (Figure 3), was used to
assess the possibility of improving the inhibitors’ potency
throughgainingdifferent interactions withthe Hsp90 enzyme.
A chlorine atom was introduced next to the para-hydroxy
group to confirm a SAR similar to that observed for the
previous dihydroxyphenylpyrrolidine series.10,14 Compound
2c (Figure 3) displayed a competitive binding IC50 of 20 nM, a
cellular IC50 of 1 μM, and high clearance in the hHep assay (93
(μL/min)/million cells). These results provided evidence that
the two classes of inhibitors (pyrrolidine and isoindoline)
shared the same SAR derived from occupying the small
Hsp90 hydrophobic pocket formed by Leu107, Val150, and
16000
410
concn at 1 h (nM)
free plasma
concn at 4 h (nM)
76
330
Several other amides were designed and synthesized to
further elucidate the relationship between lipophilicity and
clearance in the hHep assayfor this classofcompounds. These
2,3-dihydro-1H-isoindole-1-carboxylic acid amides (7-13,
Table 2), which have clogD between 0.4 and 1.6, displayed
reasonable clearance values in the hHep assay. Compound 13
has a higher clogD value (2.2) than the other amides and
displayed higher clearance (>10 (μL/min)/million cells). This
result demonstrated that the 2,3-dihydro-1H-isoindole-1-car-
boxylic acid amides, which have a clogD of 0.4-2.2, could
reduce the binding of the resorcinol-containing compounds to
the uridine glucuronosyltransferase (UGT) enzymes.
Dihydroxylphenylpyrrolidine 2a was prepared following
the route previously described.10 Compounds 3-6 were pre-
pared following the same amide coupling procedure using
commercially available D-proline methyl ester hydrochloride.
Compounds 7-13 were prepared using key intermediates 1615
and 1710 as shown in Scheme 1.
˚
Phe138 (Figure 4). We also obtained a 2.1 A cocrystal
structure of 2b bound to Hsp90. In the overlap of the cocrystal
structures of 2a (PDB code 3K97) and 2b (PDB code 3K99),
Lys58 was displaced by the phenyl portion of the isoindoline
moiety present in 2b to form a new hydrophobic pocket
formed by Lys58, Leu56, and Ala55 (Figure 5). In addition,
three other Hsp90 residues that surrounded the isoindoline
moiety (Ile110, Thr109, and Gly108) were pushed away by the
phenyl group of 2a (Figure 5). Compounds 7 and 8 were then
designed based on 4 and 5 to occupy this space. Compounds 7
and 8 improved the binding affinity 20-fold relative to 4 and 5,
respectively, and exhibited cell potencies of 0.6-1.5 μM.
The active enantiomers of the two most potent compounds
14 and 15 were isolated from their racemic counterparts 8 and

502 Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1
Kung et al.
10, respectively, and further tested in rat PK experiments.16
A
concentrated to get a brown oil. This oil residue (20) was used in
the next step without further purification.
cocrystal structure of 15 and Hsp90 was obtained (Figure 6,
PDB code 3K98) and confirmed that the carboxylamide
moiety occupied the same space as the 2-methylphenyl moiety
exists in 2a. Both 14 and 15 displayed a good correlation
between in vitro and in vivo hepatic clearances in rat along
with acceptable oral bioavailabilities (Table 3). We also
examined 15 in a mouse PK experiment using different
adminstration routes (Table 4). Both ip and po routes af-
forded compound plasma levels above the in vitro cellular
IC50 for up to 4 h. In comparison, iv administration of 1c to
mice (6 mg/kg) afforded a T1/2 of 1.8 h.
Hydrogen chloride (42 mL, 4 M in dioxane, 167 mmol) was
added to a solution of 20 in DCM (20 mL). MeOH was added,
and the mixture was stirred at room temperature for 12 h. The
solvents were evaporated. Water (50 mL) was added, and the pH
of the resulting solution was adjusted to 7 by the careful addition
of saturated NaHCO3(aq) to basicify the aqueous solution.
EtOAc (2 ꢀ 100 mL) was added to extract the aqueous solution.
The combined organic layer was concentrated to get a brown oil.
DCM (20 mL) was added and the pink precipitate was collected
to afford the desired 10 (5.2 g, 86% yield). 1H NMR (400 MHz,
DMSO-d6) δ 1.01 (t, J = 7.2 Hz, 3 H), 2.82-2.91 (m, 1 H),
3.08-3.22 (m, 1 H), 4.62-4.90 (m, 2 H), 5.55 (s, 1 H), 7.22-7.35
(m, 4 H), 7.40 (d, J = 7.3 Hz, 1 H), 8.07 (s, 1 H), 10.06 (s, 1 H),
10.42 (s, 1 H). Anal. Calcd for C18H17N2O4Cl: C, 59.92; H, 4.75;
N, 7.76. Found: C, 59.93; H, 4.84; N, 7.62.
On the basis of these experiments, we identified the dihyr-
oxybenzoyl 2,3-dihydro-1H-isoindole-1-carboxylic acid ami-
des as bioavailable small molecule inhibitors of the Hsp90
target with the potential to display therapeutic effects.17
(R)-2-(5-Chloro-2,4-dihydroxybenzoyl)-2,3-dihydro-1H-isoind-
ole-1-carboxylic Acid Ethylamide (15). Two enantiomers of 10
(5 g) were separated by supercritical fluid chromatography (SFC)
using Chiralpak AD-H SFC column (21.2 mm ꢀ 250 mm)
and eluted with 25% MeOH in carbon dioxide at 120 bar (flow
rate at 60 mL/min). Two peaks that have retention times of 4.89
and 6.05 min were separated. The peak with the retention time of
4.89 min was identifed to be 15 (the active enantiomer) and to be
the “R” enantiomer based on the crystal structure (Figure 6).
Compound 15 displayed chiral purity of >95% ee.
Conclusion
In summary, we have identified a series of dihydroxyphe-
nylisoindoline amide compounds as potent, metabolically
stable, and orally bioavailable non-quinone Hsp90 inhibitors.
The process of lead optimization utilized structure-based drug
design techniques, and the exploration between compound
lipophilicity and phase II conjugation metabolism is also
discussed. Additional results from our Hsp90 program will
be reported in the near future.
Acknowledgment. We thank Dr. Ping Kang for his meta-
bolite identification work and Dr. Martin Wythes for his
helpful discussions.
Experimental Section
Supporting Information Available: Experimental details and
spectral data for all compounds other than 18, 19, 10, and 15.
This material is available free of charge via the Internet at http://
pubs.acs.org.
2-(5-Chloro-2,4-bis-methoxymethoxybenzoyl)-2,3-dihydro-1H-
isoindole-1-carboxylic Acid Methyl Ester (18). 4-Methylmorpho-
line (39 mL, 351 mmol), 1,(3-dimethylaminopropyl)-3-ethylcar-
bodiimide hydrochloride (EDC) (9 g, 47 mmol), and 1-hydro-
xybenzotrizole (HOBt) (6.3 g, 47 mmol) were added to a solution
of 16 (5 g, 20 mmol) and 17 (6.5 g, 23.4 mmol) in DMF (100 mL).
The mixture was stirred at room temperature for 12 h. Water
(100 mL) was added to the mixture, and EtOAc (2 ꢀ 300 mL)
was added to extract the aqueous solution. The combined organic
layer was dried, filtered, and concentrated to get a brown oil.
The crude product was purified by silica gel chroma-
tography (eluting with 30% EtOAc in hexanes) to afford
compound 18 (6.4 g, 60% yield) as a pale-yellow solid. 1H
NMR (400 MHz, DMSO-d6) δ 3.69 (s, 3 H), 4.71-4.89 (m, 2
H), 5.67-5.76 (m, 1 H), 6.61 (s, 1 H), 7.16 (s, 1 H), 7.24-7.53 (m,
4 H), 10.34 (s, 1 H), 10.51 (s, 1 H). Anal. Calcd for
References
(1) Jolly, C.; Morimoto, R. I. Role of the heat shock response and
molecular chaperones in oncogenesisand cell death. J. Natl. Cancer
Inst. 2000, 92, 1564–1572.
(2) Isaacs, J. S.; Xu, W.; Neckers, L. Heat shock protein 90 as a
molecular target for cancer therapeutics. Cancer Cell 2003, 3, 213–
217.
(3) Ferrarini, M.; Heltai, S.; Zocchi, M. R.; Rugarli, C. Unusual
expression and localization of heat-shock proteins in human tumor
cells. Intl. J. Cancer 1992, 51, 613–619.
(4) Soga, S.; Shiotsu, Y.; Akinaga, S.; Sharma, S. V. Development of
radicicol analogues. Curr. Cancer Drug Targets 2003, 3, 359–369.
(5) Burlison, J. A.; Avila, C.; Vielhauer, G.; Lubbers, D. J.; Holzbeierlein,
J.; Blagg, B. S. J. Development of novobiocin analogues that manifest
anti-proliferative activity against several cancer cell lines. J. Org.
Chem. 2008, 73, 2130–2137.
(6) (a) Xiong, M. P.; Yanez, J. A.; Kwon, G. S.; Davies, N. M.; Forrest,
M. L. A cremophor-free formulation for tanespimycin (17-AAG)
using PEG-b-PDLLA micelles: characterization and pharmacoki-
netics in rats. J. Pharm. Sci. 2009, 98, 1577–1586. (b) Egorin, M. J.;
Zuhowski, E. G.; Rosen, D. M.; Sentz, D. L.; Covey, J. M.; Eiseman,
J. L. Plasma pharmcokinetics and tissue distribution, and metabolism of
17-(allylamino)-17-demethoxy-geldanamycin (NSC 330507) in
CD2F1 mice. Cancer Chemother. Pharmacol. 2001, 47, 291–302.
(c) Solit, D. B.; Ivy, S. P.; Kopil, C.; Sikorski, R.; Morris, M. J.; Slovin,
S. F.; Kelly, W. K.; DeLaCruz, A.; Curley, T.; Heller, G.; Larson, S.;
Schwartz, L.; Egorin, M. J.; Rosen, N.; Scher, H. I. Phase 1 trial of 17-
AAG in patients with advanced cancer. Clin. Cancer Res. 2007, 13,
1775–1782. (d) Egorin, M. J.; Rosen, D. M.; Wolff, J. H.; Callery, P. S.;
Musser, S. M.; Eiseman, J. L. Metabolism of 17-(allylamino)-17-
demethoxy-geldanamycin (NSC 330507) by murine and human hepatic
preparations. Cancer Res. 1998, 58, 2385–2396.
C17H14NO5Cl 0.25H2O: C, 57.97; H, 4.15; N, 3.98. Found: C,
58.06; H, 4.17; N, 4.00.
3
2-(5-Chloro-2,4-bis-methoxymethoxybenzoyl)-2,3-dihydro-1H-
isoindole-1-carboxylic Acid (19). Lithium hydroxide (73.4 mL,
4 M, 294 mmol) was added to a solution of 18 (6.4 g, 15 mmol) in
dioxane (30 mL). The mixture was heated at 60 °C for 3 h and
monitored by LC/MS. The solvent was evaporated and acidified
with NaOAc-HOAc buffer. EtOAc (2 ꢀ 200 mL) was added to
extract the aqueous solution. The combined organic layer was
dried, filtered, and concentrated to give 19 as a light-brown foam
(6.3 g, quant. yield). Compound 19 was used in the next step
without further purification.
2-(5-Chloro-2,4-dihydroxybenzoyl)-2,3-dihydro-1H-isoindole-
1-carboxylic Acid Ethylamide (10). 4-Methylmorpholine (9.2
mL, 83. 4 mmol), dimethylaminopropyl-3-ethylcarbodiimide
hydrochloride (6.4 g, 33.4 mmol), and 1-hydroxybenzotriazole
(4.5 g, 33.4 mmol) were added to a solution of 19 (6.3 g, 17
mmol) and ethylamine (25 mL, 2 M in THF) in DMF (100 mL).
The mixture was stirred at room temperature for 12 h and
monitored by LC/MS. Water (50 mL) was added to the mixture,
and EtOAc (2 ꢀ 300 mL) was added to extract the aqueous
solution. The combined organic layer was dried, filtered, and
(7) Brough, P. A.; Aherne, W.; Barril, X.; Borgognoni, J.; Boxall, K.;
Cansfield, J. E.; Cheung, K.-M. J.; Collins, I.; Davies, N. G. M.;
Drysdale, M.; Dymock, B.; Eccles, S. A.; Finch, H.; Fink, A.;
Hayes, A.; Howes, R.; Hubbard, R. E.; James, K.; Jordan, A. M.;
Lockie, A.; Martins, V.; Massey, A.; Matthews, T. P.; McDonald,
E.; Northfield, C. J.; Pearl, L. H.; Prodromou, C.; Ray, S.;

Brief Article
Raynaud, F. I.; Roughley, S. D.; Sharp, S. Y.; Surgenor, A.;
Journal of Medicinal Chemistry, 2010, Vol. 53, No. 1 503
(12) Smith, N. F.; Hayes, A.; James, K.; Nutley, B. P.; McDonald, E.;
Henley, A.; Dymock, B.; Drysdale, M. J.; Raynaud, F. I.;
Workman, P. Preclinical pharmacokinetics and metabolism of a
novel diaryl pyrazole resorcinol series of heat shock protein 90
inhibitors. Mol. Cancer Ther. 2006, 5, 1628–1637.
Walmsley, D. L.; Webb, P.; Wood, M.; Workman, P.; Wright, L.
4,5-Diarylisoxazole Hsp90 Chaperone inhibitors: potential thera-
peutic agents for the treatment of cancer. J. Med. Chem. 2008, 51,
196–218.
(8) Kasibhatla, S. R.; Hong, K.; Biamonte, M. A.; Busch, D. J.;
Karjian, P. L.; Sensintaffar, J. L.; Kamal, A.; Lough, R. E.;
Brekken, J.; Lundgren, K.; Grecko, R.; Timony, G. A.; Ran, Y.;
Mansfield, R.; Fritz, L. C.; Ulm, E.; Burrows, F. J.; Boehm, M. F.
Rationally designed high-affinity 2-amino-6-halopurine heat
shock protein 90 inhibitors that exhibit potent antitumor activity.
J. Med. Chem. 2007, 50, 2767–2778.
(9) Chandarlapaty, S.; Sawai, A.; Ye, Q.; Scott, A.; Silinski, M.;
Huang, K.; Fadden, P.; Partdrige, J.; Hall, S.; Steed, P.; Norton,
L.; Rosen, N.; Solit, D. B. Clin. SNX2112, a synthetic heat
shock protein 90 inhibitor, has potent antitumor activity
against HER kinase-dependent cancers. Cancer Res. 2008, 14,
240–248.
(13) Reddy, A.; Heimbach, T.; Freiwald, S.; Smith, D.; Winters, R.;
Michael, S.; Surendran, N.; Cai, H. Validation of a semi-auto-
mated human hepatocyte assay for the determination and predic-
tion of intrinsic clearance in discovery. J. Pharm. Biomed. Anal.
2005, 37, 319–326.
(14) Example ofa chloride-containingradicicol: Roe, S. M.; Prodromou,
C.; O’Brien, R.; Ladbury, J. E.; Piper, P. W.; Pearl, L. H. Structural
basis for inhibition of the Hsp90 moleular chaperone by the anti-
tumor antibiotics radicicol and geldanamycin. J. Med. Chem. 1999,
42, 260–266.
(15) Cignarella, G.; Cerri, R.; Grella, G.; Sanna, P. Researches on
isoindoles. VIII. Synthesis and reactivity of 1-carbomethoxyiso-
indoles. Gazz. Chim. Ital. 1976, 106, 65–75.
(10) Kung, P.-P.; Funk, L.; Meng, J.; Collins, M.; Zhou, J. Z.; Johnson,
M. C.; Ekker, A.; Wang, J.; Mehta, P.; Yin, M.-J.; Rodgers, C.;
Davies, J. F.; Bayman, E.; Smeal, T.; Maegley, K. A.; Gehring, M.
R. Dihydroxypheyl amides as inhibitors of the Hsp90 molecular
chaperone. Bioorg. Med. Chem. Lett. 2008, 18, 6273–6278.
(11) Giuliano, C.; Jairaj, M.; Zafiu, C. M.; Laufer, R. Direct determina-
tion of unbound intrinsic drug clearance in the microsomal stability
assay. Drug Metab. Dispos. 2005, 33, 1319–1324.
(16) The “S” enantiomers corresponding to 8 and 10 had Ki of 2000 and
400 nM, respectively, in the biochemical assay.
(17) Mehta, P.; Kung, P.-P.; Yamazaki, S.; Nguyen, L.; Gehring, M. R.;
Los, G.; Yin, M.-J.
A Novel Class of Specific Hsp90
Small Molecule Inhibitors Demonstrate in Vitro and in Vivo
Anti-Tumor Activity in Human Melanoma Cells. Presented
at the America Association for Cancer Research (AACR), April
2009.