Design and Synthesis of Tetrazole- And Pyridine-Containing Itraconazole Analogs as Potent Angiogenesis Inhibitors

Article abstract of DOI:10.1021/acsmedchemlett.9b00438

Itraconazole, a widely used antifungal drug, was found to possess antiangiogenic activity and is currently undergoing multiple clinical trials for the treatment of different types of cancer. However, it suffers from extremely low solubility and strong interactions with many drugs through inhibition of CYP3A4, limiting its potential as a new antiangiogenic and anticancer drug. To address these issues, a series of analogs in which the phenyl group is replaced with pyridine or fluorine-substituted benzene was synthesized. Among them the pyridine- and tetrazole-containing compound 24 has significantly improved solubility and reduced CYP3A4 inhibition compared to itraconazole. Similar to itraconazole, compound 24 inhibited the AMPK/mTOR signaling axis and the glycosylation of VEGFR2. It also induced cholesterol accumulation in the endolysosome and demonstrated binding to the sterol-sensing domain of NPC1 in a simulation study. These results suggested that compound 24 may serve as an attractive candidate for the development of a new generation of antiangiogenic drug.

Full text of DOI:10.1021/acsmedchemlett.9b00438

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Letter
Design and Synthesis of Tetrazole- and Pyridine-containing
Itraconazole Analogs as Potent Angiogenetic Inhibitors
Yingjun Li, Kalyan Kumar Pasunooti, Hanjing Peng, Ruo-Jing Li, Wei
Q. Shi, Wukun Liu, Zhiqiang Cheng, Sarah A. Head, and Jun O. Liu
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.9b00438 • Publication Date (Web): 08 Apr 2020
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Design and Synthesis of Tetrazole- and Pyridine-containing Itracon-
azole Analogs as Potent Angiogenetic Inhibitors
Yingjun Li,† Kalyan Kumar Pasunooti,†,⊥ Hanjing Peng,† Ruo-Jing Li,†,§ Wei Q. Shi,†,†† Wukun Liu,†,‡‡
Zhiqiang Cheng,† Sarah A. Head,†,§§ Jun O. Liu*,†,‡
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†Department of Pharmacology and Molecular Sciences, Johns Hopkins School of Medicine, Baltimore, Maryland 21205,
United States
‡Department of Oncology, Johns Hopkins School of Medicine, Baltimore, Maryland 21205, United States
KEYWORDS: angiogenesis inhibitor, itraconazole, CYP3A4, NPC1.
ABSTRACT: Itraconazole, a widely used antifungal drug, was found to possess anti-angiogenic activity and is currently un-
dergoing multiple clinical trials for the treatment of different types of cancer. However, it suffers from extremely low solubility
and strong interactions with many drugs through inhibition of CYP3A4, limiting its potential as a new anti-angiogenic and
anticancer drug. To address these issues, a series of analogs in which the phenyl group is replaced with pyridine or fluorine-
substituted benzene was synthesized. Among them the pyridine- and tetrazole-containing compound 24 has significantly
improved solubility and reduced CYP3A4 inhibition compared to itraconazole. Similar to itraconazole, compound 24 inhibited
the AMPK/mTOR signaling axis and the glycosylation of VEGFR2. It also induced cholesterol accumulation in the endolyso-
some and demonstrated binding to the sterol-sensing domain of NPC1 in a simulation study. These results suggested that
compound 24 may serve as an attractive candidate for the development of a new generation of anti-angiogenic drug.
Itraconazole (1, Figure 1) is a triazole-containing anti-
fungal drug that has been found to possess anti-angiogenic
activity.1 The anti-angiogenic and anticancer activities of
itraconazole have been demonstrated in animal models and
it has shown efficacy in multiple phase II clinical trials for
the treatment of lung cancer, prostate cancer and basal cell
carcinoma.2-4 The underlying mechanism of angiogenesis
inhibition has been extensively investigated.5 Itraconazole
was found to bind two distinct molecular targets, Niemann-
Pick Type C 1 (NPC1) and Voltage-Dependent Anion Chan-
nel 1 (VDAC1) in primary human umbilical vein endothelial
cells (HUVECs).6,7 NPC1, a membrane protein localized to
the late endosomes and lysosomes (collectively called endo-
lysosomes), mediates cholesterol trafficking from the endo-
lysosome to other cellular compartments.8 The inhibition of
NPC1 by itraconazole, or the structurally related drug
posaconazole, leads to the accumulation of cholesterol in
the endolysosome9,10. VDAC1, a channel on the mitochon-
drial outer membrane, serves as a key gateway for a multi-
tude of important cellular metabolites including adenosine
diphosphate (ADP) and adenosine triphosphate (ATP) to
enter and exit the mitochondria.11 Inhibition of VDAC1 by
itraconazole increases the AMP/ATP ratio, thereby activat-
ing the AMP-activated protein kinase (AMPK) signaling
pathway in endothelial cells. Dual inhibition of NPC1 and
VDAC1 culminates in a synergistic inhibition of mechanistic
target of rapamycin (mTOR), blocking the proliferation of
endothelial cells and angiogenesis.9 In addition, itracona-
zole has also been shown to inhibit the glycosylation and
trafficking of vascular endothelial growth factor receptor 2
(VEGFR2), which is also required for pathological angiogen-
esis.12
Despite its potential as a novel anti-angiogenic drug,
itraconazole has three major limitations. First, the inhibi-
tion of Cytochrome P450 3A4 (CYP3A4) prevents combina-
tion therapy of itraconazole with the majority of other anti-
cancer drugs.13,14 Second, the high lipophilicity of itracona-
zole (cLogP= 5.3) results in its accumulation in adipose
Figure 1. Structures of itraconazole (1), IT-C (2) and com-
pound 3.
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tissues, with concentrations in skin and fat tissues to be 19-
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inhibition and hepatotoxicity,14,19 the hydrophobicity and
accompanying low water solubility remains to be resolved.
The hydrophobicity of itraconazole can be attributed in
large part to the two phenyl groups in the “core” region of
the molecule (W1-piperizin-1-yl-W2, Figure 1). The phe-
nyl-piperizin-1-yl-phenyl core portion has a symmetrical
and rigid configuration. It was reported that symmetrical
and rigid compounds have high crystal packing energies,
and therefore have low solubility in water as well as in or-
ganic solvents.22,23 It has been shown previously that re-
placement of the W2 phenyl with a pyridyl group can signif-
icantly increase the solubility and bioavailability of itracon-
azole analogs.24 We took a similar approach by using pyri-
dine to replace phenyl group or adding a fluorine substitu-
tion on the phenyl ring. The synthesis and anti-angiogenic
activity characterization of the novel analogs are reported
herein.
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fold and 17-fold greater than that in the plasma, respec-
tively.15 Third, itraconazole is relatively insoluble in water
and has limited oral bioavailability. Although a clinically-
used suspension formulation is able to improve its bioavail-
ability, itraconazole has a highly variable absorption and
plasma concentrations from individual to individual.16,17
The high lipophilicity and the low solubility of itraconazole,
though tolerable for the treatment of fungal infections, pose
major problems in the treatment of cancer.
9
In our attempts to improve the potency and reduce the
toxicity of itraconazole, we conducted systematic structure
and activity relationship (SAR) studies on the two ends of
itraconazole. Compound IT-C (2) with cis-(2S,4R) stereo-
chemistry in the dioxolane ring, wherein the isobutyl
sidechain with a 2’S configuration, showed the strongest
anti-angiogenic activity among the 8 stereoisomers of itra-
conazole and significantly reduced hepatotoxicity.18,19 Com-
pound 3 with tetrazole in place of the 1,2,4-triazole in the
head position had increased activity and significantly re-
duced CYP3A4 inhibition while a methyl substitution in the
same position resulted in the loss of anti-angiogenic activ-
ity.20 Moreover, the sec-butyl tail (or similar alkyl group)
was required for angiogenesis inhibition.21
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The syntheses of novel itraconazole analogs were ac-
complished using modified synthetic routes reported previ-
ously (Scheme 1).14 Nucleophilic displacement of the bro-
mine in 4 with 1,2,4-triazole, imidazole or tetrazole, fol-
lowed by ketalization ring-closure reaction with enantio-
merically pure glyceryl tosylate 6 under strong acid condi-
tions, provided 1,3-dioxolane intermediate as a cis- and
trans- mixture in a ratio of 3:1. The cis-2S,4R diastereomers
7a-c were separated with column chromatograph. For
Although modifications at the “head” and “tail” sections
of itraconazole have effectively reduced the CYP3A4
Scheme 1: Synthesis of Compounds 4, 5, 8-12 and 15-26a
aReagents and conditions: (a) azoles, K2CO3, MeCN, 25oC, 16 h, 40-70%; (b) 6, TfOH, toluene, rt, 60 h, 55%; (c) Pd2(dba)3,
o
BINAP, NaOtBu, toluene, 80 C, 16 h, 80%; (d) 10% Pd/C, N2H4.H2O, EtOH, reflux, 3.5h, 93%; (e) Phenyl chloroformate, pyri-
dine, rt, 16h, 82%; (f) i.NH2NH2.H2O, 1,4-dioxane, reflux, 3h; ii. formamidine acetate, 1-propanol, reflux, 3h, 66%; (g) 2-bro-
mobutane, K2CO3, DMSO, 80 oC, 16 h, 82%; (h) 48% aqueous HBr, reflux, 76%; (i) 7a-c, NaH, DMF, 80 oC, 16 h, 60-75%.
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analogs in which the W1 group was a pyridyl, Buchwald-
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itra c o n a z o le
16
Hartwig cross coupling of bromides with 8a-b was used to
generate nitrobenzene 9a-e.25 Then the nitro group was
reduced to an amine with 10% Pd/C and hydrazine mono-
hydrate in ethanol. The triazolone ring was built in three
steps by reacting the aniline intermediates with phenyl
chloroformate, hydrazine monohydrate, and formamidine
acetate, successively.20 N-alkylation of triazolone 12a-e
with 2-bromobutane under basic conditions afforded the
intermediates 13a-e. The methyl protecting group in 13a-
e was unmasked with 48% aqueous hydrobromic acid
(HBr). The resulting phenol 14a-e was coupled with tosyl-
ate 7a-c to give the desired compounds 15-26. Com-
pounds 27-30 were synthesized under similar conditions
(Supplementary Scheme 1). All desired compounds were
evaluated as racemic mixtures (2-S-butanyl group and 2-
R-butanyl group) for their biological activities.
1 0 0
5 0
0
21
24
2 5
26
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lo g (M )
Figure 2. Dose-response curves of CYP3A4 enzyme inhibi-
tion by itraconazole (1) and analogs 5, 12, 24, 25 and 26.
their 1,2,4-triazole counterparts, which was unfavorable
for drug distribution. Moreover, the 1,2,4-triazole- and im-
idazole-containing compounds displayed inhibition of
CYP3A4, albeit weaker than itraconazole. For example, the
IC50 of compounds 16 and 21 for CYP3A4 inhibition were
3.4 µM and 2.6 µM, respectively. (Figure 2).
The inhibitory effects of the new analogs on HUVEC
proliferation were determined using a [3H]-thymidine in-
corporation assay.26 The cLogP values, used as an indicator
of lipophilicity, were calculated using ALOGPS2.1 soft-
ware. Aqueous solubility is a key physiochemical property
relevant for oral absorption. We determined the solubility
of each analog in 0.001 N HCl (pH = 3), which is similar to
that in the acidic human gastric fluid.22,27 The IC50 values
for inhibition of HUVEC proliferation, cLogP and solubility
of all analogs are shown in Table 1. Itraconazole was
slightly soluble in 0.001N HCl (10.1 ng/mL). In order to
disrupt the symmetricity of the phenyl-piperizin-1-yl-phe-
nyl core (W1- piperizin-1-yl-W2), we used pyridyl moiety
to replace one of the two phenyl rings in the W1 or W2
position or 3`-fluorophenyl to replace W2. Both pyridine
and fluoro-benzene are reasonable benzene isosteres and
were not expected to cause significant steric perturbations
of the W1-W2 portion of the molecules. Initially, com-
pounds 15-19, 27 and 28 with 1,2,4-triazole in the R1 po-
sition were synthesized and characterized. Pyridyl substi-
tution at either the W1 (15, 16) or W2 (27, 28) position
and 3`-fluoro-phenyl at W2 (17-19) all resulted in in-
creases in solubility by 5-84 folds. The pyridin-2-yl and 3`-
fluorophenyl combination (19) resulted in the best solu-
bility in this series of analogs (847.7 ng/mL). Pyridine
(=N), benzene (=CH), and fluorine-substituted benzene
(=CF) are similar in van der Waals radius but quite differ-
ent in lipophilicity.28 The lipophilicity of the three groups
follows the order: pyridyl < benzyl < fluoro-benzyl.29 As ex-
pected, the analogs containing a pyridine (15, 16, 27, 28)
exhibited decreases in cLogP, while the fluoro-phenyl ana-
log (17) had an increased clogP. In the HUVEC prolifera-
tion assay, compounds 28, 17, and 19 showed similar ac-
tivity to that of itraconazole, while the other modifications
led to decreases in potency.
Our previous SAR study showed that 1H-tetrazole-1-yl
in the R1 position significantly reduced CYP3A4 inhibition
and increased anti-angiogenic potency.14 Thus, three new
analogs with either pyridin-2-yl, 3`-fluorophenyl or the
combination in the core region and tetrazole in the R1 po-
sition were synthesized. As expected, the resulting te-
trazole-containing analogs had reduced CYP3A4 inhibition
and the IC50 values of 24 and 25 are greater than 90 μM
(Figure 2). To our delight, the calculated logP values of the
three new analogs were further decreased and the clogP of
24 was reduced to 4.36, falling into the range of orally ac-
tive drugs according to Lipinski`s rule of five.30 Among the
three tetrazole-containing analogs, compound 24 also had
the highest potency for HUVEC inhibition (IC50 of 77 nM)
and highest solubility. Compared to itraconazole, the solu-
bility of 24 was increased by over 90-fold.
Tetrazole compounds 24 and 26 were selected for en-
dothelial cell tube formation assay to further assess their
anti-angiogenic potential, with compound 3 as a positive
Next, six analogs with imidazole in the R1 position were
synthesized and characterized. Except for compound 20
(pyridin-3-yl) and 30 (pyridin-2’-yl), the other four ana-
logs (21, 29, 22, 23) exhibited increased activities for in-
hibition of HUVEC proliferation. In general, replacement of
the triazole with an imidazole led to a significant increase
in solubility ranging from 6 μg/mL to 125 μg/mL, much
greater than itraconazole. However, the imidazole com-
pounds were more lipophilic (higher cLogP value) than
Figure 3. Inhibition of HUVEC tube formation. HUVECs
were seeded on Matrigel-coated plates and treated with
DMSO or 3μM of 3, 24 and 26 for 24h. (A) Cells were stained
with Calcein-AM and vascular networks were imaged using
fluorescence microscopy. (B) Quantified total tube lengths
from the fluorescence images. Data represent mean ± SD of
three independent experiments.
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Table 1: Inhibition of HUVEC Proliferation, Aqueous Solubility and cLogP of Itraconazole Analogs
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2
3
4
5
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7
8
9
HUVEC
Solubility in
cLogPb 0.001N HClc
(ng/mL)
R1
W1
W2
inhibitiona
IC50 (nM)
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15
16
17
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19
20
21
22
23
24
25
26
27
28
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1
170 ± 13.1
5.35
10.1
3
73 ± 17.0
4.92
5.00
321.4
116.1
15
686.8 ± 78.1
16
27
28
379.8 ± 50.8
221.9 ± 23.1
187.8 ± 17.5
4.99
5.02
5.04
113.6
53.9
496.5
17
18
19
128.4 ± 59.7
446.5 ± 38.3
173.1 ± 27.6
5.75
5.12
5.11
29.1
255.4
847.7
20
21
29
30
468.5 ± 42.2
69.1 ± 7.0
5.48
5.47
5.49
6483.0
2938.9
7988.0
41338.6
98.5 ± 36.3
636.0 ± 177.2 5.52
22
70.1 ± 15.4
6.16
17150.0
23
24
25
68.0 ± 15.5
77.1 ± 11.9
153.4 ± 32.7
5.61
4.36
4.96
124904.5
951.3
52.8
26
108.5 ± 8.6
4.40
707.6
aIC50 in HUVEC was evaluated using [3H]-thymidine incorporation assay. Values represent the mean ± SD in three independent
experiments carried out in triplicate; b cLogP was the calculated partition coefficient between n-octanol and water log(coc-
tanol/cwater) using ALOGPS2.1 software; cThe thermodynamic solubility in 0.001N HCl was measured using HPLC.
control. In this assay, HUVEC assembled into three-dimen-
sional networks and formed tubular structures in Mat-
rigel-coated wells, recapitulating many key aspects of new
blood vessel formation in vivo. Compound 24 inhibited
70% of HUVEC tube formation at 3 μM, as judged by total
tube length, further demonstrating its anti-angiogenetic
potency (Figure 3).
NPC1 plays an essential role in cholesterol export from
the endolysosome.31 Previously, we and others reported
itraconazole and its structurally related drug posaconazole
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Next, we determined the effects of 24 on AMPK/mTOR
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activity and VEGFR2 glycosylation. Similar to itraconazole,
24 activated AMPK as judged by the phosphorylation of its
substrate acetyl CoA carboxylase (ACC) in a dose-depend-
ent manner (Figure 5A). AMPK activation and NPC1 inhi-
bition were demonstrated to lead to synergistic mTOR in-
hibition by itraconazole.33 Indeed, 24 potently inhibited the
phosphorylation of the mTOR substrate p70 S6 Kinase
(S6K) in a dose-dependent manner.
We have previously shown that itraconazole inhibits
VEGFR2 glycosylation and surface expression.12 We ob-
served two VEGFR2 bands by Western blotting, represent-
ing differentially glycosylated forms of the receptor, with
the higher molecular weight band being more predominant
in untreated HUVEC. Treatment with 24 caused a mobility
shift to the lower molecular weight, hypoglycosylated band.
The high molecular weight species of VEGFR2 disappeared
upon treatment with 24 at 0.5 μΜ or higher concentrations
(Figure 5A). We next determined whether 24 and other
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Figure 4. Inhibition of NPC1. (A): HUVECs were treated
with 0.2 μM itraconazole, 3, 24, 25, 26 or DMSO for 24 h.
Intracellular cholesterol was stained with filipin and fluo-
rescent images were captured using LSM710 confocal mi-
croscope with 25x objective. Predicted binding mode of
itraconazole (magenta) and 24 (green) with NPC1 (PDB
code: 5I31) by AutoDock Vina software (B). The predicted
interaction of compound 24 (green) with SSD (C).
target NPC1 and induce NPC phenotype in endothelial
cells.9,10 Using filipin staining, we observed that compound
24 caused a massive build-up of cholesterol in the peri-
nuclear structure, in the same manner as itraconazole and
the other tetrazole-containing analogs 3, 25 and 26 (Fig-
ure 4A).7 These results suggested that the pyridinyl and
fluorophenyl analogs retained NPC1 inhibitory activity of
itraconazole. To further assess the potential binding of 24
to NPC1 protein, we performed docking of compound 24
and itraconazole into the pocket within the sterol-sensing
domain (SSD) using AutoDock Vina software as we have
previous shown that itraconazole binds to the SSD domain
of NPC1.7,32 As shown in Figure 4B, itraconazole and 24
were predicted to bind to NPC1 in a similar manner. The
linear pyridinyl-piperizin-1-yl-phenyl core of 24 nestled
into the hydrophobic channel created by the transmem-
brane helices (Figure 4C). The isobutyl tail faced the open
entrance of this channel. The tetrazole moiety pointed to
the closed end of the pocket and interacted with the hy-
droxyl group of Tyr1225 via hydrogen bonding. These re-
sults offered a plausible explanation of how the benzene bi-
oisostere replacement could modify the physiochemical
properties of itraconazole without compromising NPC1 in-
hibition.
Figure 5. Inhibition of VEGFR2 and AMPK/mTOR in
HUVEC. (A) HUVECs were treated with 0.05μM, 0.25μM,
0.5μM, 1μM or 2μM 24 or DMSO for 24 h. VEGFR2, p-ACC,
total-ACC, p-S6K, total S6K and β-actin proteins were ana-
lyzed by western blot. (B) HUVECs were treated with DMSO,
2μM compound 24 or itraconazole (1). The cells were
stained with VEGFR2 (green), GM130 (red) antibody and
DAPI (blue). Images were captured using LSM 700 confocal
microscopy.
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††Department of Chemistry, Ball State University, Muncie, Indi-
analogs affected cellular localization of VEGFR2 using im-
munofluorescence staining (Figure 5B). In 24 and itracon-
azole-treated cells, VEGFR2 accumulated in the perinuclear
region and colocalized with the Golgi marker, GM130. In
contrast, in the untreated cells, VEGFR2 uniformly distrib-
uted in small puncta throughout the cytoplasm. As a critical
component of lipid rafts, cholesterol is pivotal to intracellu-
lar transport and cell signaling. In our previous study, we
demonstrated that the hypoglycosylation of VEGFR2 was
rescued by supplementation with exogenous cholesterol. It
is therefore possible that VEGFR2 relocalization and inhibi-
tion by itraconazole and its analogs may be mediated
through inhibition of NPC1. Taken together, these results
indicated that 24, like itraconazole, inhibits endothelial cell
growth and angiogenesis by concurrent inhibition of NPC1
and VDAC1, leading to activation of AMPK, and inhibition of
mTOR and VEGFR2 signaling.
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ana 47306, United States
‡‡Nanjing University of Chinese Medicine,Jiangsu, Nan-
jing210023, China,
§§Centre for Genomic Regulation (CRG), The Barcelona Institute
of Science and Technology, Barcelona 08003, Spain
Author Contributions
The manuscript was written through contributions of all au-
thors. All authors have given approval to the final version of the
manuscript.
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Funding Sources
This work was supported by the National Cancer Institutes
(Grant R01CA184103) and the Flight Attendant Medical Re-
search Institute.
Notes
Anti-angiogenic therapy has been clinically validated
for the treatment of a number of diseases including cancer,
autoimmune disorders, retinopathy, obesity, macular de-
generation and others.34 Itraconazole has great potential as
a newly identified angiogenesis inhibitor and is under in-
vestigation in multiple clinical trials. However, its wider use
as an anti-angiogenic agent in general, and its use in combi-
nation with other drugs for treating cancer in particular, has
been limited by its inhibition of CYP450 and unfavorable
physicochemical properties. To improve its solubility and
decrease its lipophilicity, we used pyridyl or fluorophenyl
groups to replace the phenyl group in the core region of itra-
conazole. Among the newly synthesized analogs, 24 with 2-
pyridyl in W1 and 1H-tetrazol-1-yl in R1 position exhibited
improved anti-angiogenic activity, solubility and hydro-
philicity with negligible effects on CYP3A4. The anti-angio-
genic activity of 24 was further validated using a tube for-
mation assay. Moreover, 24 bears all the hallmarks of itra-
conazole activity in endothelial cells, including activation of
AMPK and inhibition of mTOR, induction of cholesterol ac-
cumulation in the endolysosome and binding to NPC1, and
inhibition of VEGFR2 glycosylation, suggesting that the
structural changes required to improve its pharmacological
properties did not alter its mechanism of action. This work
paves the way for 24 to undergo further preclinical studies
as a novel anti-angiogenic and anticancer drug candidate.
The authors declare no competing financial interest.
ABBREVIATIONS
HUVEC, human umbilical vein endothelial cell; NPC1, Niemann-
Pick disease, type C1; VDAC1, voltage-dependent anion channel
1; AMPK, AMP-activated protein kinase; ATP, adenosine tri-
phosphate; ADP, adenosine diphosphate; mTORC, mammalian
target of rapamycin complex; CYP3A4, cytochrome P450 3A4;
SAR, structure-activity relationship; VEGFR, vascular endothe-
lial growth factor receptor; IC50, half-maximal inhibitory con-
centration; DMSO, dimethyl sulfoxide; DMF, dimethylforma-
mide; HBr, hydrobromic acid; THF, tetrahydrofuran; DCM, di-
chloromethane; TfOH, trifluoromethanesulfonic acid; NaH, so-
dium hydride; SSD, sterol-sensing domain; ACC, acetyl CoA car-
boxylase; S6K, p70 S6 kinase; , chemical shifts; MeCN, acetoni-
trile;
Pd2(dba)3,
tris(dibenzylideneacetone)dipalladium;
EtOAc, ethyl acetate; NaOtBu, sodium t-butoxide; BINAP, 2,2′-
bis(diphenyl phosphino)-1,1′-binaphthalene.
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ASSOCIATED CONTENT
Supporting Information
The Supporting Information is available free of charge on the
ACS Publications website.
Synthetic procedures, assay protocols, concentration standard
curve of 24, IC50 of CYP3A4 inhibition, western blot of itracon-
azole and analytical data (PDF)
AUTHOR INFORMATION
Corresponding Author
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Y.; Colombini, M.; Liu, J. O. Antifungal drug itraconazole targets
*Phone, 410-955-4619; fax, 410-955-4520; e-mail, jo-
liu@jhu.edu
Present Addresses
⊥
ProSAMBio, Hyderabad,Telangana500049, India
§FDA, Silver Spring, Maryland 20993, United States
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