Dual inhibitors of inosine monophosphate dehydrogenase and histone deacetylases for cancer treatment

Article abstract of DOI:10.1021/jm070864w

Mycophenolic acid (MPA), an inhibitor of IMP-dehydrogenase (IMPDH), is used worldwide in transplantation. Recently, numerous studies showed its importance in cancer treatment. Consequently, MPA entered clinical trials in advanced multiple myeloma patients

Full text of DOI:10.1021/jm070864w

J. Med. Chem. 2007, 50, 6685–6691
6685
Dual Inhibitors of Inosine Monophosphate Dehydrogenase and Histone Deacetylases for Cancer
Treatment
Liqiang Chen,† Daniel Wilson,† Hiremagalur N. Jayaram,‡ and Krzysztof W. Pankiewicz*,†
Center for Drug Design, UniVersity of Minnesota, Minneapolis, Minnesota 55455, and Department of Biochemistry and Molecular Biology, Indiana
UniVersity School of Medicine and Richard Roudebush Veterans Affairs Medical Center, 1481 West Tenth Street, Indianapolis, Indiana 46202
ReceiVed July 17, 2007
Mycophenolic acid (MPA), an inhibitor of IMP-dehydrogenase (IMPDH), is used worldwide in transplanta-
tion. Recently, numerous studies showed its importance in cancer treatment. Consequently, MPA entered
clinical trials in advanced multiple myeloma patients. Suberoylanilide hydroxamic acid (SAHA), a potent
differentiation agent acting through inhibition of histone deacetylases (HDACs), was recently approved for
treatment of cutaneous T cell lymphoma. We report herein the synthesis of dual inhibitors of IMPDH and
HDACs. We found that mycophenolic hydroxamic acid (9, MAHA) inhibits both IMPDH (Ki ) 30 nM)
and HDAC (IC50 ) 5.0 µM). A modification of SAHA with groups known to interact with IMPDH afforded
a SAHA analogue 14, which inhibits IMPDH (Ki ) 1.7 µM) and HDAC (IC50 ) 0.06 µM). Both MAHA
(IC50 ) 4.8 µM) and SAHA analogue 14 (IC50 ) 7.7 µM) were more potent than parent compounds as
antiproliferation agents. They were also significantly more potent as differentiation inducers.
Introduction
Chronic myelogenous leukemia (CMLa) is a malignant cancer
of the bone marrow that causes rapid growth of blood forming
cells (myeloid precursors) in the bone marrow, peripheral blood,
and body tissues. The hallmark of CML is the Philadelphia
chromosome translocation, which fuses the bcr and c-abl genes,
resulting in expression of a constitutively active tyrosine kinase.
There is often little hope for patients with CML upon the onset
of blast crisis (BC). Currently there are only two agents that
are in clinical use for CML-BC, 4-[(4-methylpiperazin-1-yl)
methyl]-N-[4-methyl-3-[(4-pyridin-3-ylpyrimidin-2-yl)amino]
phenyl]benzamide (imatinib, tyrosine kinase inhibitor) and
tiazofurin (TR, inosine monophosphate dehydrogenase, IMPDH
inhibitor). TR is a prodrug that requires metabolic activation
into its active form, tiazofurin adenine dinucleotide (TAD,
Figure 1
Figure 1), a mimic of nicotinamide adenine dinucleotide (NAD).
Patients develop resistance to both agents after several treatment
(plant), induces apoptosis in leukemic stem cells but not in
cycles, and therefore, this condition is exceptionally difficult
normal stem cells. Recently it was found6 that induction of
to repress. While imatinib is a phenomenal success, the drug
differentiation severely limited the cancer stem cell’s ability to
does not affect a small population of leukemic stem cells (less
form new tumors.1,7,8 The mechanisms of apoptosis and
than 1% of all tumor cells)1–4 and resistance develops.
differentiation have recently been studied extensively in CML
As the cells emerge from the progenitor stem cell, they are
cells, and these processes are now well characterized when
destroyed by imatinib, which needs to be taken every day for a
triggered by Bcr-Abl inhibitors.9 TR proved to be the most
lifetime. Little is known about the differences between normal
effective inducer of erythroid differentiation in CML leukemic
and cancer stem cells that would allow for designing drugs that
(K562) cells among numerous compounds studied.10,11 It
specifically target the malignant cancer stem cells. However,
showed good anticancer activity in both acute myelogenous
cancer stem cells, especially leukemic stem cells, can be
leukemia (AML) and CML patients and regardless of its general
selectively disarmed by induction of apoptosis and/or dif-
toxicity was approved as an orphan drug for treatment of patients
ferentiation.1 Parthenolide,5 a natural product from feverfew
in CML blast crisis.11
Inhibitors of IMPDH, in general, show a significant ability
to trigger differentiation and/or apoptosis.12–14 Mycophenolic
* To whom correspondence should be addressed. Phone: 612-625-7968.
Fax: 612-625-8252. E-mail: panki001@umn.edu.
acid (MPA, Figure 2), one of the most potent inhibitors of
human IMPDH (Ki ) 10 nM), binds at the NAD binding domain
of the enzyme and is used in clinic as an immunosuppressant.
However, its potential anticancer activities are of current interest.
Thus, MPA assisted differentiation of human prostate cancer
PC-3 cells and DU145 cells has been reported by Huberman et
al.15–17 and it was found to block tumor-induced angiogenesis
in ViVo.18
†
University of Minnesota.
University School of Medicine and Richard Roudebush Veterans Affairs
‡
Medical Center.
aAbbreviations: MPA, mycophenolic acid; IMPDH, inosine monophos-
phate dehydrogenase; SAHA, suberoylanilide hydroxamic acid; MAHA,
mycophenolic hydroxamic acid; HDACs, histone deacetylases; CML,
chronic myelogenous leukemia; BC, blast crisis; TR, tiazofurin; TAD,
tiazofurin adenine dinucleotide; MAD, mycophenolic adenine dinucleotide;
NAD, nicotinamide adenine dinucleotide.
10.1021/jm070864w CCC: $37.00
2007 American Chemical Society
Published on Web 11/27/2007

6686 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 26
Chen et al.
Figure 2
In CML, synergy between MPA and imatinib was reported
in Bcr-Abl-expressing cell lines.13 The authors suggested that
these results “lay the foundation for clinical trials in which
IMPDH inhibitors are added to imanitib in patients who have
suboptimal molecular responses to single agent therapy”.
Meanwhile, because of significant apoptotic properties, MPA
entered phase I clinical trial in advanced multiple myeloma
patients.19,20
Our group has reported novel IMPDH inhibitors, mycophe-
nolic adenine dinucleotides (MAD analogues), that mimic NAD
cofactor. Among them, C2-MAD (Figure 2) was found to be
more potent than MPA in both antileukemic activity and the
ability to induce differentiation.21 At concentrations as low as
1.5 µM, it converted 65% of CML leukemic cells (K562 cells)
into mature, nonproliferating cells able to produce hemoglobin.21
C2-MAD showed superior anticancer activity and lower toxicity
than TR in mouse model of CML.22
All the above inhibitors (e.g., TR, MPA, and MAD analogues)
mimic NAD cofactor (or part of it) and inhibit NAD-dependent
IMPDH, a key enzyme in de novo synthesis of purine nucle-
otides. They induce differentiation due to depletion of guanine
nucleotides. Such G-restriction affects DNA and RNA metabo-
lism as well as the activity of GTP-binding proteins involved
in transduction pathways.23 In recent years IMPDH has emerged
as a major therapeutic target24 and its mechanism of action has
been recently reviewed.25
Figure 3
Histone deacetylases (HDACs), enzymes that catalyze the
removal of acetyl groups from lysine residues of histones
represent new class of anticancer targets.26 HDACs inhibitors
alter gene transcription and exert antitumor effects through
growth arrest, apoptosis, differentiation, and inhibition of tumor
angiogenesis. Thus, differentiation triggered by HDAC inhibitors
is based on a fundamentally different mechanism27–30 than that
caused by inhibition of IMPDH.
Figure 4
MPA’s structure consists of an aromatic moiety and a linker;
however, it does not contain a zinc binding group. Thus, we
replaced the carboxylic group of MPA with a hydroxamic acid
moiety and synthesized hydroxamic acid analogue 9 (MAHA,
Figure 4). We also modified SAHA by addition of groups known
to interact with IMPDH and prepared SAHA analogue 14. Both
compounds 9 and 14 were found to act as novel dual inhibitors
of IMPDH and HDAC. In this paper we focused our efforts on
synthesis and evaluation of dual IMPDH/HDAC inhibitors
against CML; however, a similar anticancer activity of these
novel inhibitors is expected (and was confirmed) for other
cancers cell lines where SAHA proved to be highly effective.
To date, four classes of human HDACs with 18 members
have been identified. The class I, II, and IV HDACs require
zinc for catalytic deacetylation, while class III HDACs consists
of seven sirtuins31–33 that are NAD-dependent and do not share
homologies with the zinc-dependent histone deacetylases.
Chemically, the zinc-dependent HDACs inhibitors can be
divided into several classes such as short-chain fatty acids (e.g.,
compound 1), hydroxamic acids 2–6, and benzamides 7 and 8
(Figure 3). Three structural regions can be distinguished in these
molecules; a cap (usually aromatic) region, a linker, and a metal
binding group. For the metal binding group, hydroxamic acids
usually exhibit higher potency than the corresponding benzamides.
The most representative member of HDACs inhibitors is
suberoylanilide hydroxamic acid (SAHA), which was recently
approved for treatment of cutaneous T cell lymphoma.34 SAHA
was studied in combination with imatinib35,36 and was found
to promote apoptosis in imatinib-resistant leukemic cells (Bcr/
Abl positive). Recently, numerous studies showed synergism
not only between imatinib and SAHA but also for combinations
of new generation anti-CML drugs with SAHA.9,37,38
Results and Discussion
1. Inhibition of IMPDH, HDAC, and Proliferation of
K562 Cell. A conversion of the carboxylic group of MPA into
hydroxamic acid moiety afforded MAHA (9) which, like other
HADCs inhibitors, contain an aromatic cap (5-methoxy-4-
methylphtalan-1-one), a linker (3-methylpent-2-enyl group), and
a zinc binding group (hydroxamic acid). MAHA did not lose
inhibitory activity against human IMPDH (Ki ) 30 nM, type 2
isoform, Table 1) and was found to have a similar potency

Dual Inhibitors for Cancer Treatment
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 26 6687
Table 1. Inhibition of IMPDH, HDAC, and Proliferation of K562 Cells
by SAHA, MPA, TR, and Novel Compoundsa
IMPDH Ki (µM)
HDAC IC50
nuclear extract
(µM)
% of differ.
K562 cells
(µM)
K562 IC50
(µM)
inhibitor type I type II
SAHA
MPA
MAHA (9) 0.07
10
11
12
13
14
NA
0.04
NA
0.01
0.03
NA
1.10
0.54
0.65
1.7
0.41
NA
5.0
14.0
10.4
20.1
>100
0.06
NA
0.75 47.4 (at 0.50)
7.7
4.8
46.4 (at 0.25)
75.0 (at 0.25)
NA
42.3
10.8
44.7
60
ND
ND
ND
ND
0.40
2.07
2.83
5.0
0.29 87.8 (at 0.005)
TR (TAD) 0.1
0.1
3.0b 80.0 (at 5.30)b
Figure 5
a NA, not active. ND, not determined. b These data were reported
previously by us.21
TR (TAD) when compared against compounds 11–13. Clearly,
more compounds with equally adjusted activity against IMPDH/
HDACs are needed for a better understanding of their antileu-
kemic activity.
against IMPDH as MPA (Ki ) 10 nM, type 2). It also inhibited
a nuclear extract of HDACs enzymes (IC50 ) 5.0 µM) as well
as proliferation of K562 cells at a similar concentration (IC50
) 4.8 µM). MAHA’s antiproliferative activity was slightly
higher than that of MPA (IC50 ) 7.7 µM).
Finally, we became interested in a chemical modification of
SAHA that would result in inhibition of IMPDH without losing
SAHA’s ability to inhibit HDAC enzymes. In a series of IMPDH
inhibitors developed by Vertex and Bristol-Myers-Squibb such
as (S)-tetrahydrofuran-3-yl 3-(3-(3-methoxy-4-(oxazol-5-yl)phe-
nyl)ureido)benzylcarbamate (15, merimepodib, Figure 5) and
related inhibitors 16 and 17, the phenyloxazole moiety with an
ortho methoxy group was introduced and found to bind at the
nicotinamide subdomain of NAD-binding pocket of IMPDH.40–44
Thus, we modified SAHA by addition of oxazole group and
methoxy group in ortho configuration, resulting in SAHA
derivative 14.
We found that compound 14 is more potent than SAHA as
inhibitor of HDAC enzymes (IC50 ) 0.06 µΜ) and proliferation
of K562 cells (IC50 ) 0.29 µM). It showed inhibition of IMPDH
at low micromolar level. Again, compound 14 is 7-fold more
potent than SAHA as an HDAC inhibitor; however, in spite of
having both components for the activity, it is only 2.6-fold more
potent than SAHA in the antiproliferative assay.
We expected that our double IMPDH/HDAC inhibitors would
show an improved ability to differentiate leukemic cells such
as K562 cells. Indeed, MAHA at concentration of 0.25 µM
converted significantly larger number of K562 cells (75%) than
SAHA (47%) at 2-fold higher concentration (0.5 µM). Interest-
ingly, our modified SAHA analogue 14 was found to be one of
the most potent (if not the most potent) K562 differentiation
inducers (88% at 5 nM concentration) ever reported. In fact,
compound 14 is a much more powerful differentiation inducer
(3 orders of magnitude) than TR, a clinically used nucleoside
with well established differentiation ability. Since compound
14 is much more active in cells than against isolated target
enzymes, its differentiation activity could be due to additional
interaction with cellular targets other than IMPDH and HDACs.
It was recently found that differentiation severely limits cancer
stem cells’ ability to form new tumors.6 Thus, we are currently
in the process of preparation of the second generation of dual
IMPDH/HDAC inhibitors for their evaluation against cancer
stem cells.
2. Chemical Synthesis. Chemical synthesis of MAHA (9)
and its analogues 10–13 (Figure 4) requires preparation of the
corresponding carboxylic acids and their subsequent conversion
to the protected hydroxamates. Triphenylmethyl (trityl) was
chosen as a hydroxylamine protecting group, and its facile
removal under acidic conditions would afford the desired
hydroxamic acids. As depicted in Scheme 1, carboxylic acids.
19–21 were prepared from mycophenolic aicd (MPA).
Methylation of the phenolic hydroxyl group in mycophenolic
At enzymatic level MAHA showed more potent inhibition
of IMPDH than HDACs. Thus, the potential effect of HDACs
inhibition on proliferation and differentiation of K562 cells may
be negligible. In order to estimate the importance of HDACs
inhibition by MAHA, we synthesized MAHA analogue 10 in
which the 7-OH group is replaced by an OMe group. It is well-
known that the 7-phenol group is crucial for MPA inhibitory
activity. All attempts to protect this group or to replace it with
groups such as -F, -NH2, -CN resulted in inactive deriva-
tives.39 Indeed, 7-OMe analogue of MPA did not show any
activity against human isoforms of IMPDH. However, replace-
ment of the carboxylic group of the 7-OMe-MPA analogue with
hydroxamic acid afforded 7-OMe-MAHA (10), which was only
3-fold less active against HDACs nuclear extract than MAHA
and weakly inhibited K562 cell proliferation with IC50 ) 42.3
µM. These results indicate that antileukemic activity of MAHA
in K562 cells is related to its ability to inhibit IMPDH rather
HDAC enzymes. Nevertheless, the inhibitory activity of com-
pound 10 against K562 cells growth is only 14-fold less potent
than tiazofurin. It is likely that this effect is related to HDACs
inhibition.
In order to find more about structure–activity relationship
(SAR) of these novel inhibitors, we synthesized compounds
11–13, in which the linker of MAHA was extended by a one
carbon (11), the double bond of the linker was reduced (12),
and the hydroxamic group of MAHA was replaced by the ortho
aminobenzamide group (13; see structures of HDACs inhibitors,
Figure 2). None of these modifications produced a better
inhibitor than MAHA. Interestingly, compound 11 is 1–2 orders
of magnitude less potent an inhibitor of IMPDH [Ki ) 0.4 µM
(type 1), Ki ) 1.1 µM (type 2)] than MAHA or MPA and only
twice less potent (IC50 ) 10.4 µM) than MAHA (IC50 ) 5.0
µM) as HDACs inhibitor. It was found to inhibit proliferation
of K562 cells (IC50 ) 10.8 µM) at a concentration similar to
that of MPA (IC50 ) 7.7 µM) and MAHA (IC50 ) 4.8 µM).
These results suggest that at least for compound 11 the two
components, IMPDH and HDACs inhibition, contribute to the
antileukemic activity. On the other hand, the above structure–
activity relationship appears to correlate again with the potency
against IMPDH rather than HDACs inhibitory activity. Com-
pounds 12 and 13 show similar inhibitory activity against
IMPDH and similar antiproliferative potency against K562 cells
despite the HDAC component for compound 12, which is at
least 5-fold higher than that of 13. The same trend is valid for

6688 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 26
Chen et al.
Scheme 1a
a Reaction conditions: (a) cat. H2SO4, MeOH; (b) MeI, Cs2CO3, CH3CN, reflux; (c) NaOH, THF/H2O and then H+; (d) (COCl)2, cat. DMF, dichloroethane;
(e) TMSCHN2, CH3CN, 0 °C to room temp; (f) AgOBz, THF/H2O; (g) H2, Pd/C, MeOH.
Scheme 2a
a Reaction conditions: (a) TrONH2, EDC, HOBt, CH2Cl2; (b) TFA, Et3SiH, CH2Cl2.
Scheme 3a
Scheme 4a
a Reaction conditions: (a) 28, EDC, HOBt, CH2Cl2; (b) NH2OH·H2O,
MeOH.
the product was readily precipitated from aqueous solution, this
method is amenable to scale up of MAHA.
a Reaction conditions: (a) (COCl)2, cat. DMF, CH3CN and then
Benzamide derivative 13 was readily prepared by coupling
of MPA and 1,2-phenylenediamine (26), which was mediated
by benzotriazol-1-yloxytris(dimethylamino)phosphonium hexaflu-
orophosphate (BOP). The preparation of SAHA analogue 14 is
depicted in Scheme 4. A coupling of known 3-methoxy-4-(5-
oxazolyl)phenylamine (27)48 and suberic acid monomethyl ester
(28) afforded methyl ester 29, which was converted into
hydroxamic acid 14 under treatment with hydroxylamine
aqueous solution.
NH2OH·H2O; (b) 26, BOP, Et3N, DMF.
acid methyl ester followed by hydrolysis of the resulting methyl
ester 18 provided acid 19.45 Arndt-Eistert homologation46 of
MPA furnished acid 20, which contains a side chain one carbon
longer than that of MPA. Saturated acid 2147 was readily
obtained by hydrogenation of MPA.
With these carboxylic acids available, coupling with trityl
protected hydroxylamine gave hydroxamates 22–25 (Scheme
2). As expected, the removal of trityl protecting was ac-
complished under mildly acidic conditions, and the desired
hydroxamic acids 9-12 were obtained.
Even though the method procedures described above pro-
duced hydroxamic acids smoothly, a more efficient one-pot
procedure was used for the preparation of MAHA (9). It
involved in situ formation of acyl chloride and subsequent
treatment of hydroxylamine aqueous solution (Scheme 3). Since
In summary, we demonstrated that chemical modification of
clinically used drugs such as MPA (IMPDH inhibitor) and
SAHA (HDACs inhibitor) resulted in new molecules that show
inhibitory activity of both IMPDH and HDAC enzymes. New
inhibitors, MAHA and SAHA analogue 14, were found to be
more potent antiproliferative agents than parent drugs. Because
of a dual mechanism of action, our novel inhibitors may be of
therapeutic interest in cancer treatment. Since the inhibitory
activity of our best compounds, MAHA and 14, is not yet

Dual Inhibitors for Cancer Treatment
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 26 6689
1
24 as a pale solid (69.3 mg, 27%), mp 132.8–134.3 °C. H NMR
perfectly balanced, we are working now on other SAHA-MPA
(CDCl3, 600 MHz) δ 7.70 (s, 1H), 7.66 (s, 1H), 7.52–7.24 (m,
15H), 5.19 (s, 2H), 5.10 (brs, 1H), 3.73 (s, 3H), 3.35 (d, J ) 6.6
Hz, 2H), 2.14 (s, 3H), 1.86–1.66 (m, 6H), 1.53 (brs, 2H), 1.33 (brs,
1H). HRMS calcd for C37H37NO6Na 614.2513 (M + Na)+, found
614.2513.
hybrid combinations and evaluation of their biological potential.
Experimental Section
General Methods. All commercial reagents (Sigma-Aldrich,
Acros) were used as provided unless otherwise indicated. An
anhydrous solvent dispensing system (J. C. Meyer) using two
packed columns of neutral alumina was used for drying THF, Et2O,
and CH2Cl2, while two packed columns of molecular sieves were
used to dry DMF. Solvents were dispensed under argon. Flash
chromatography was performed with Ultra Pure silica gel (Silicycle)
with the indicated solvent system. Analytical HPLC was performed
on a Varian Microsorb column (C18, 5 µm, 4.6 mm × 250 mm)
with a flow rate of 0.5 mL/min with an isocratic or linear gradient
of water with 0.1% (v/v) TFA and MeOH with 0.1% (v/v) TFA.
Melting points were determined on a Mel-Temp apparatus and were
uncorrected. 1H nuclear magnetic resonance spectra were recorded
on a Varian 300 or 600 MHz with Me4Si, DDS, or signals from
residual solvent as the internal standard. Chemical shifts are reported
in ppm, and signals are described as s (singlet), d (doublet), t
(triplet), q (quartet), m (multiplet), brs (broad singlet), and dd
(double–doublet). Values given for coupling constants are first-
order. High-resolution mass spectra were recorded on an Agilent
TOF II TOF/MS instrument equipped with either an ESI or APCI
interface.
Hydroxamate 25. In a similar manner carboxylic acid 2147 (294
mg, 0.91 mmol) was coupled with O-tritylhydroxylamine to give
25 as a white solid (315 mg, 60%), mp 141.2–142.8 °C. 1H NMR
(CDCl3, 600 MHz) δ 7.52–7.26 (m, 15H), 5.19 (s, 2H), 3.75 (s,
3H), 2.64–2.48 (m, 2H), 2.14 (s, 3H), 1.67–1.10 (m, 7H), 0.90-
.072 (m, 3H). HRMS calcd for C36H37NO6Na 602.2513 (M + Na)+,
found 602.2518.
Hydroxamic Acids 9–12. Hydroxamic Acid 1 (MAHA)
from Hydroxamate 22. To a solution of protected hydroxamate
22 (103 mg, 0.18 mmol) in anhydrous CH2Cl2 (3 mL) was added
TFA (0.15 mL). The resulting yellow solution was treated with
Et3SiH till it was colorless. After being stirred for an additional 10
min, the reaction mixture was concentrated and the residue was
triturated with hot hexanes (2 × 10 mL). The syruplike residue
was dissolved in CH2Cl2 (0.3 mL), and hexanes (10 mL) were
added. After the organic solvents were discarded, the syrup formed
was collected and dried under high vacuum to give hydroxamic
acid 9 as a white solid (52.7 mg, 87%).
Hydroxamic Acid 9 (MAHA) Directly from Mycophenolic
Acid. To a suspension of mycophenolic acid (552 mg, 1.72 mmol)
in anhydrous CH3CN were added oxalyl chloride (0.18 mL, 2.10
mmol) and then five drops of DMF. The reaction mixture turned
clear, was stirred at room temperature for 1.5 h, and then cooled
to 0 °C. A solution of 50% aqueous hydroxylamine (0.51 mL, 8.3
mmol) was added slowly, and the mixture was stirred at 0 °C for
an additional 2 h. After addition of water (5 mL), the reaction
mixture was warmed to room temperature and CH3CN was removed
in vacuo. A syrup that formed on the wall of the reaction flask was
washed with water (5 mL) and then dissolved in CH3CN (10 mL)
and water (5 mL). A syrup was obtained after the removal of
CH3CN, and it was redissolved in CH2Cl2 (100 mL) and water (50
mL). After CH2Cl2 and some of the water were removed in vacuo,
the solid formed was filtered, washed with water, and dried under
high vacuum to give MAHA as a white solid (357 mg, 62%), mp
136.0–136.9 °C. 1H NMR (CDCl3, 300 MHz) δ 5.36–5.08 (m, 3H),
3.75 (s, 3H), 3.74 (d, J ) 6.3 Hz, 2H), 2.29 (brs, 4H), 2.13 (s,
3H), 1.77 (s, 3H). HRMS calcd for C17H22NO6 336.1447 (M +
H)+, found 336.1451.
Hydroxamic Acid 10. In a similar manner protected hydrox-
amate 23 (164 mg, 0.28 mmol) was treated with TFA to give 10
as a pale solid (50.6 mg, 52%), mp 142.0–144.0 °C. 1H NMR
(CD3OD, 300 MHz) δ 5.26–5.14 (m, 3H), 3.98 (s, 3H), 3.78 (s,
3H), 3.40 (d, J ) 6.9 Hz, 2H), 2.27 (t, J ) 7.2 Hz, 2H), 2.22–2.12
(m, 5H), 1.82 (s, 3H). HRMS calcd for C18H24NO6 350.1603 (M
+ H)+, found 350.1607.
Hydroxamic Acid 11. In a similar manner protected hydrox-
amate 24 (65.2 mg, 0.11 mmol) was treated with TFA to give 11
as a pale syrup (3.1 mg, 8%). 1H NMR (CDCl3, 600 MHz) δ
5.24–5.18 (m, 3H), 3.80 (s, 3H), 3.40 (d, J ) 6.6 Hz, 2H), 2.16 (s,
3H), 2.06 (t, J ) 6.6 Hz, 2H), 2.01 (t, J ) 6.9 Hz, 2H), 1.82–1.72
(m, 5H). HRMS calcd for C18H23NO6Na 372.1423 (M + Na)+,
found 372.1432.
Chemical Syntheses. Carboxylic Acid 20. To a solution of
mycophenolic acid (811 mg, 2.53 mmol) in anhydrous CH2Cl2 (25
mL) were added oxalyl chloride (0.44 mL, 5.12 mmol) and three
drops of DMF. The solution was stirred at room temperature for
2 h and concentrated. The residue was dissolved in anhydrous
CH3CN (25 mL) at 0 °C, and a solution of TMSCHN2 (2.0 M in
Et2O, 5.1 mL, 10.2 mmol) was added. After being stirred at 0 °C
for 30 min, the mixture was allowed to warm to room temperature
and stirred for 48 h. After concentration, the resulting residue was
dissolved in THF (12 mL) and H2O (12 mL), and silver benzoate
(120 mg, 0.52 mmol) was added. The mixture was stirred at room
temperature overnight, diluted with EtOAc (150 mL), and subse-
quently washed with 0.5 N HCl (30 mL), H2O (30 mL), and brine
(2 × 60 mL). The organic layer was dried over Na2SO4 and filtered.
The filtrate was concentrated and the residue was chromatographed
on a silica gel column (0%–4% MeOH/CH2Cl2) to give carboxylic
1
acid 20 as a pale solid (673 mg, 80%), mp 124.2–126.0 °C. H
NMR (CDCl3, 600 MHz) δ 5.24–5.18 (m, 3H), 3.77 (s, 3H), 3.39
(d, J ) 6.6 Hz, 2H), 2.28 (t, J ) 7.5 Hz, 2H), 2.15 (s, 3H), 2.02
(t, J ) 7.5 Hz, 2H), 1.78 (s, 3H), 1.73 (t, J ) 7.5 Hz, 2H). HRMS
calcd for C18H23O6 335.1489 (M + H)+, found 335.1489.
Protected Hydroxamates 22–25. Hydroxamate 22. A solution
of mycophenolic acid (107 mg, 0.33 mmol), O-tritylhydroxylamine
(102 mg, 0.37 mmol), 1-ethyl-3-(3-dimethylaminopropyl)carbodi-
imide hydrochloride (EDC) (104 mg, 0.53 mmol), and 1-hydroxy-
benzotriazole (HOBt) (51 mg, 0.38 mmol) in anhydrous CH2Cl2
(5 mL) was stirred at room temperature for 20 h. After concentra-
tion, the residue was diluted with EtOAc (20 mL) and washed with
water (3 × 10 mL) and brine (20 mL). The organic layer was dried
over Na2SO4 and filtered. The filtrate was concentrated and the
residue was purified by flash column chromatography (20–40%
EtOAc/hexanes) to give protected hydroxamate 22 as a white solid
(118 mg, 62%), mp 187.0–188.5 °C. 1H NMR (CDCl3, 300 MHz)
δ 7.37–7.24 (m, 15H), 5.34–5.16 (m, 3H), 3.80 (s, 3H), 3.42 (d, J
) 6.0 Hz, 2H), 2.34 (brs, 2H), 2.17 (s, 3H), 1.82 (brs, 2H), 1.27
(s, 3H). HRMS calcd for C36H35NO6Na 600.2362 (M + Na)+, found
600.2363.
Hydroxamic Acid 12. In a similar manner protected hydrox-
amate 25 (275 mg, 0.47 mmol) was treated with TFA to give 12
as a pale syrup (19.1 mg, 12%). H NMR (CD3OD, 600 MHz) δ
1
5.23 (s, 2H), 3.78 (s, 3H), 2.73–2.61 (m, 2H), 2.18–2.04 (m, 5H),
1.78–1.68 (m, 1H), 1.60–1.44 (m, 3H), 1.43–1.34 (m, 1H), 0.99
(d, J ) 6.0 Hz, 3H). HRMS calcd for C17H24NO6 338.1598 (M +
H)+, found 338.1598.
Benzamide 13. A solution of mycophenolic acid (217 mg, 0.68
mmol), BOP (457 mg, 1.03 mmol), Et3N (0.43 mL, 3.08), and 1,2-
phenylenediamine (26, 184 mg, 1.70 mmol) in dry DMF (5 mL)
was stirred at room temperature for 18 h. The reaction mixture
was diluted with toluene (60 mL) and washed with water (2 × 10
mL). The organic layer was concentrated and the residue was
Hydroxamate 23. In a similar manner carboxylic acid 1945 (248
mg, 0.74 mmol) was coupled with O-tritylhydroxylamine to give
23 as a white solid (190 mg, 43%), mp 169.3–170.5 °C. 1H NMR
(CDCl3, 300 MHz) δ 7.48–7.25 (m, 15H), 5.12 (s, 2H), 4.98 (brs,
1H), 3.99 (s, 3H), 3.72 (s, 3H), 3.32 (d, J ) 6.6 Hz, 2H), 2.15 (s,
3H), 1.90 (brs, 2H), 1.70–1.50 (m, 5H). HRMS calcd for
C37H37NO6Na 614.2518 (M + Na)+, found 614.2515.
Hydroxamate 24. In a similar manner carboxylic acid 20 (145
mg, 0.43 mmol) was coupled with O-tritylhydroxylamine to give

6690 Journal of Medicinal Chemistry, 2007, Vol. 50, No. 26
Chen et al.
purified on a Chromatotron (2 mm, 0–4% MeOH/CH2Cl2) to give
benzamide 13 as a tan solid (51.4 mg, 18%), mp 187.6–189.4 °C.
1H NMR (CD3OD, 300 MHz) δ 6.97 (dd, J ) 7.2, 1.5 Hz, 1H),
6.88 (dd, J ) 8.0, 1.0 Hz, 1H), 6.78 (dd, J ) 7.8, 1.5 Hz, 1H),
6.63–6.55 (m, 1H), 5.34 (td, J ) 7.0, 1.2 Hz, 1H), 5.21 (s, 2H),
3.74 (s, 3H), 3.41 (d, J ) 6.9 Hz, 2H), 2.54–2.45 (m, 2H), 2.42–2.34
(m, 2H), 2.11 (s, 3H), 1.88 (s, 3H). HRMS calcd for C23H27N2O5
411.1919 (M + H)+, found 411.1931.
Differentiation of K562 Cells. Logarithmically growing K562
cells (1.0 × 103 cells/0.1 mL) were plated in triplicate into 96-
well plates in RPMI 1640 medium containing 10% heat-inactivated
fetal bovine serum and antibiotics and incubated at 37 °C in an
atmosphere of air and 5% CO2. Twenty-four hours later, compounds
were added and further incubated for 5 days. For examination of
the effect of compounds on cytotoxicity, 20 µL of MTS reagent
was added and further incubated for 3 h and the absorbance was
read at 490 nm. For examination of the effect on cellular
differentiation, 25 µL of 5-day incubated cell suspension was taken
and mixed with 25 µL of differentiation reagent (0.2% benzidine
in acetic acid and 0.15% H2O2) and about 500 cells were counted
to determine the percent of differentiated cells.
Monomethyl Ester 29. A solution of suberic acid monomethyl
ester (28, 411 mg, 2.18 mmol), EDC (650 mg, 3.28 mmol), HOBt
(295 mg, 2.18 mmol), and aniline 27 (458 mg, 2.41 mmol) in
anhydrous CH2Cl2 (15 mL) was stirred at room temperature for
48 h. After concentration, the residue was diluted with EtOAc (100
mL) and washed with 0.1 N HCl (50 mL), water (50 mL), saturated
NaHCO3 (50 mL), and brine (50 mL). The organic layer was dried
over Na2SO4 and filtered. The filtrate was concentrated and the
residue was purified by silica gel column chromatography (0–4%
MeOH/CH2Cl2) to give amide 29 as a yellow solid (571 mg, 73%),
Acknowledgment. This research was supported by the
Center for Drug Design (CDD) in the Academic Health Center
of the University of Minnesota. We thank Dr. Courtney Aldrich
of CDD for his help in establishing enzymatic methods at the
CDD. We also thank Dr. Liz Hedstrom for providing constructs
of IMPDH1 and IMPDH2.
1
mp 115.3–116.6 °C. H NMR (CDCl3, 600 MHz) δ 7.88 (s, 1H),
7.71 (brs, 2H), 7.67 (d, J ) 8.4 Hz, 1H), 7.49 (s, 1H), 6.91 (dd, J
) 8.4, 1.8 Hz, 1H), 3.96 (s, 3H), 3.67 (s, 3H), 2.38 (t, J ) 7.5 Hz,
2H), 2.31 (t, J ) 7.5 Hz, 2H), 1.75 (q, J ) 7.2 Hz, 2H), 1.65 (q,
J ) 7.2 Hz, 2H), 1.44–1.30 (m, 4H). HRMS calcd for C19H25N2O5
361.1757 (M + H)+, found 361.1787.
Supporting Information Available: HPLC profiles of com-
pounds 9–14. This material is available free of charge via the
Internet at http://pubs.acs.org.
Hydroxamic Acid 14. To a solution of monomethyl ester 29
(253 mg, 0.70 mmol) in MeOH (6.6 mL) was added 50% aqueous
hydroxylamine solution (2.2 mL), and additional MeOH (2.0 mL)
was then added to clear the mixture. The reaction mixture was
stirred at room temperature for 4 days, during which time
hydroxylamine solution/MeOH was added to drive the reaction to
completion. The solid formed was filtered, washed with MeOH/
water, air-dried, and dried under high vacuum to give hydroxamic
References
(1) Everts, S. Getting to the root of cancer. Chem. Eng. News 2007, 85,
28–29.
(2) Kucia, M.; Ratajczak, M. Z. Stem cells as a two edged swordsfrom
regeneration to tumor formation. J. Physiol. Pharmacol. 2006, 57 (7),
5–16.
(3) Passegue, E. Hematopoietic stem cells, leukemic stem cells and chronic
myelogenous leukemia. Cell Cycle 2005, 4, 266–268.
(4) Bonnet, D. Normal and leukaemic stem cells. Br. J. Hamaetol. 2005,
130, 469–479.
(5) Jordan, C. T. The leukemic stem cell. Best Pract. Res. Clin. Haematol.
2007, 20, 13–8.
(6) Piccirillo, S. G.; Reynolds, B. A.; Zanetti, N.; Lamorte, G.; Binda,
E.; Broggi, G.; Brem, H.; Olivi, A.; Dimeco, F.; Vescovi, A. L. Bone
morphogenetic proteins inhibit the tumorigenic potential of human
brain tumour-initiating cells. Nature 2006, 444, 761–765.
(7) Deshpande, A. J.; Cusan, M.; Rawat, V. P.; Reuter, H.; Krause, A.;
Pott, C.; Quintanilla-Martinez, L.; Kakadia, P.; Kuchenbauer, F.;
Ahmed, F.; Delabesse, E.; Hahn, M.; Lichter, P.; Kneba, M.;
Hiddemann, W.; Macintyre, E.; Mecucci, C.; Ludwig, W. D.;
Humphries, R. K.; Bohlander, S. K.; Feuring-Buske, M.; Buske, C.
Acute myeloid leukemia is propagated by a leukemic stem cell with
lymphoid characteristics in a mouse model of CALM/AF10-positive
leukemia. Cancer Cell 2006, 10, 363–374.
1
acid 14 as a pale solid (167 mg, 66%), mp 186.5–187.2 °C. H
NMR (DMSO, 600 MHz) δ 10.31 (s, 1H), 10.04 (s, 1H), 8.64 (s,
1H), 8.35 (s, 1H), 7.60 (d, J ) 8.4 Hz, 1H), 7.55 (s, 1H), 7.42 (s,
1H), 7.27 (d, J ) 8.4 Hz, 1H), 3.89 (s, 3H), 2.30 (t, J ) 7.2 Hz,
2H), 1.93 (t, J ) 7.2 Hz, 2H), 1.62–1.54 (m, 2H), 1.52–1.44 (m,
2H), 1.34–1.20 (m, 4H). HRMS calcd for C18H24N3O5 362.1710
(M + H)+, found 362.1725.
Biological Evaluation. Enzyme Assays. IMPDH inhibition
assays were performed as previously described.49 Briefly, assays
were set up in duplicate using two different concentrations of
IMPDH type 1 (87 and 155 nM) and type 2 (33 and 66 nM) and
varying concentrations of inhibitor. IMPDH and inhibitors were
added to 100 µL of reaction buffer (50 mM Tris, pH 8.0, 100 mM
KCL, 1 mM DTT, 100 µM IMP, 100 µM NAD) at 25 °C and mixed
gently, and the production of NADH was monitored by following
changes in absorbance at 340 nm on a Molecular Devices M5e
multimode plate reader. Steady-state velocities were used to
determine IC50 and KiApp values by fitting the velocities vs inhibitor
concentration to the sigmoidal concentration–response curve (vari-
able slope) and the Morrison equation respectively using GraphPad
Prism.50
(8) Yang, Z. J.; Wechsler-Reya, R. J. Hit ’em where they live: targeting
the cancer stem cell niche. Cancer Cell 2007, 11, 3–5.
(9) Jacquel, A.; Colosetti, P.; Grosso, S.; Belhacene, N.; Puissant, A.;
Marchetti, S.; Breittmayer, J. P.; Auberger, P. Apoptosis and erythroid
differentiation triggered by Bcr-Abl inhibitors in CML cell lines are
fully distinguishable processes that exhibit different sensitivity to
caspase inhibition. Oncogene 2007, 26, 2445–2458.
HDAC inhibition assays were performed using the HDAC
Activity/Inhibitor Screening Assay Kit (Cayman Chemical) per the
manufacturer’s instructions. Inhibitors were suspended in either
DMSO or methanol. End point readings were used to determine
IC50 values by fitting the fractional fluorescence vs inhibitor
concentration to the sigmoidal concentration–response curve (vari-
able slope) using GraphPad Prism. Curves were corrected for
autofluorescence by making a standard concentration curve, sub-
stituting compound for solvent using the background well conditions.
Inhibition of Proliferation of K562 Cells. About 2000 cells/
well of logarithmically growing human myelogenous leukemia
K562 cells were plated into 96-well plates and incubated at 37 °C
for 24 h. Compounds at final concentrations up to 100 µM were
added in duplicate wells in 0.15% DMSO (final concentration),
mixed, and incubated for 72 h. At the end of the incubation period,
20 µL of MTS reagent was added, mixed, and further incubated
for 3 h and then absorbance read at 490 nm in a plate reader. Control
cells exhibited three doublings (doubling time was 24 h).
(10) Olah, E.; Csokay, B.; Prajda, N.; Kote-Jarai, Z.; Yeh, Y. A.; Weber,
G. Molecular mechanisms in the antiproliferative action of Taxol and
tiazofurin. Anticancer Res. 1996, 16, 2469–2477.
(11) Grifantini, M. Tiazofurine ICN Pharmaceuticals. Curr. Opin. InVest.
Drugs 2000, 1, 257–262.
(12) Inai, K.; Tsutani, H.; Yamauchi, T.; Nakamura, T.; Ueda, T. Dif-
ferentiation and reduction of intracellular GTP levels in HL-60 and
U937 cells upon treatment with IMP dehydrogenase inhibitors. AdV.
Exp. Med. Biol. 1998, 431, 549–553.
(13) Gu, J. J.; Santiago, L.; Mitchell, B. S. Synergy between imatinib and
mycophenolic acid in inducing apoptosis in cell lines expressing Bcr-
Abl. Blood 2005, 105, 3270–3277.
(14) Inai, K.; Tsutani, H.; Yamauchi, T.; Fukushima, T.; Iwasaki, H.;
Imamura, S.; Wano, Y.; Nemoto, Y.; Naiki, H.; Ueda, T. Differentia-
tion induction in non-lymphocytic leukemia cells upon treatment with
mycophenolate mofetil. Leuk. Res. 2000, 24, 761–768.
(15) Floryk, D.; Tollaksen, S. L.; Giometti, C. S.; Huberman, E. Dif-
ferentiation of human prostate cancer PC-3 cells induced by inhibitors
of inosine 5′-monophosphate dehydrogenase. Cancer Res. 2004, 64,
9049–9056.

Dual Inhibitors for Cancer Treatment
Journal of Medicinal Chemistry, 2007, Vol. 50, No. 26 6691
(16) Floryk, D.; Huberman, E. Differentiation of androgen-independent
prostate cancer PC-3 cells is associated with increased nuclear factor-
kappaB activity. Cancer Res. 2005, 65, 11588–11596.
(17) Floryk, D.; Huberman, E. Mycophenolic acid-induced replication arrest,
differentiation markers and cell death of androgen-independent prostate
cancer cells DU145. Cancer Lett. 2006, 231, 20–29.
(18) Chong, C. R.; Qian, D. Z.; Pan, F.; Wei, Y.; Pili, R.; Sullivan, D. J.,
Jr.; Liu, J. O. Identification of type 1 inosine monophosphate
dehydrogenase as an antiangiogenic drug target. J. Med. Chem. 2006,
49, 2677–2680.
(19) Takebe, N.; Cheng, X.; Wu, S.; Bauer, K.; Goloubeva, O. G.; Fenton,
R. G.; Heyman, M.; Rapoport, A. P.; Badros, A.; Shaughnessy, J.;
Ross, D.; Meisenberg, B.; Tricot, G. Phase I clinical trial of the inosine
monophosphate dehydrogenase inhibitor mycophenolate mofetil (cellcept)
in advanced multiple myeloma patients. Clin. Cancer Res. 2004, 10,
8301–8308.
Abl-positive human acute leukemia cells. Blood 2003, 101, 3236–
3239.
(36) Yu, C. R.; Rahmani, M.; Almenara, J.; Subler, M.; Krystal, G.; Conrad,
D.; Varticovski, L.; Dent, P.; Grant, S. Histone deacetylase inhibitors
promote STI571-mediated apoptosis in STI571-sensitive and -resistant
Bcr/Abl(+) human myeloid leukemia cells. Cancer Res. 2003, 63,
2118–2126.
(37) Rahmani, M.; Reese, E.; Dai, Y.; Bauer, C.; Kramer, L. B.; Huang,
M.; Jove, R.; Dent, P.; Grant, S. Cotreatment with suberanoylanilide
hydroxamic acid and 17-allylamino 17-demethoxygeldanamycin syn-
ergistically induces apoptosis in Bcr-Abl+ cells sensitive and resistant
to STI571 (imatinib mesylate) in association with down-regulation of
Bcr-Abl, abrogation of signal transducer and activator of transcription
5 activity, and Bax conformational change. Mol. Pharmacol. 2005,
67, 1166–1176.
(38) Fiskus, W.; Pranpat, M.; Balasis, M.; Bali, P.; Estrella, V.; Kuma-
raswamy, S.; Rao, R.; Rocha, K.; Herger, B.; Lee, F.; Richon, V.;
Bhalla, K. Cotreatment with vorinostat (suberoylanilide hydroxamic
acid) enhances activity of dasatinib (BMS-354825) against imatinib
mesylate-sensitive or imatinib mesylate-resistant chronic myelogenous
leukemia cells. Clin. Cancer Res. 2006, 12, 5869–5878.
(20) Takebe, N.; Cheng, X.; Fandy, T. E.; Srivastava, R. K.; Wu, S.;
Shankar, S.; Bauer, K.; Shaughnessy, J.; Tricot, G. IMP dehydrogenase
inhibitor mycophenolate mofetil induces caspase-dependent apoptosis
and cell cycle inhibition in multiple myeloma cells. Mol. Cancer Ther.
2006, 5, 457–466.
(21) Pankiewicz, K. W.; Lesiak-Watanabe, K. B.; Watanabe, K. A.;
Patterson, S. E.; Jayaram, H. N.; Yalowitz, J. A.; Miller, M. D.;
Seidman, M.; Majumdar, A.; Prehna, G.; Goldstein, B. M. Novel
mycophenolic adenine bis(phosphonate) analogues as potential dif-
ferentiation agents against human leukemia. J. Med. Chem. 2002, 45,
703–712.
(39) Franklin, T. J.; Jacobs, V.; Jones, G.; Ple, P. Human collorectal
carcinoma cells in vitro as means to assess the metabolism of analogues
of mycophenolic acid. Drug Metab. Dispos. 1997, 25, 367–374.
(40) Sintchak, M. D.; Nimmesgern, E. The structure of inosine 5′-
monophosphate dehydrogenase and the design of novel inhibitors.
Immunopharmacology 2000, 47, 163–184.
(22) Patterson, S. E.; Black, P. L.; Clark, J. L.; Risal, D.; Goldstein, B. M.;
Jayaram, H. N.; Schinazi, R. F.; Pankiewicz, K. W. The Mechanism
of Action and Antileukemic Activity of Bis(phosphonate) Analogue
of Mycophenolic Adenine Dinucleotide (C2-MAD). An Alternative
for Tiazofurin? In DeVelopments in Nucleic Acids; Schinazi, R. F. L. D.,
Ed.; IHL Press: Arlingtion, MA, 2004.
(23) Yalowitz, J. A.; Jayaram, H. N. Molecular targets of guanine
nucleotides in differentiation, proliferation and apoptosis. Anticancer
Res. 2000, 20, 2329–2338.
(24) Pankiewicz, K. W., Goldstein, B. M., Eds. Inosine Monophosphate
Dehydrogenase: A Major Therapeutic Target; American Chemical
Society: Washinghton, DC, 2003; Vol. 839.
(25) Hedstrom, L.; Gan, L. IMP dehydrogenase: structural schizophrenia
and an unusual base. Curr. Opin. Chem. Biol. 2006, 10, 520–525.
(26) Mei, S.; Ho, A. D.; Mahlknecht, U. Role of histone deacetylase
inhibitors in the treatment of cancer (review). Int. J. Oncol. 2004, 25,
1509–1519.
(27) Kim, D. H.; Kim, M.; Kwon, H. J. Histone deacetylase in carcino-
genesis and its inhibitors as anti-cancer agents. J. Biochem. Mol. Biol.
2003, 36, 110–119.
(28) Grunstein, M. Histone acetylation in chromatin structure and transcrip-
tion. Nature 1997, 389, 349–352.
(29) Mahlknecht, U.; Hoelzer, D. Histone acetylation modifiers in the
pathogenesis of malignant disease. Mol. Med. 2000, 6, 623–644.
(30) Kouraklis, G.; Theocharis, S. Histone deacetylase inhibitors: a novel
target of anticancer therapy (review). Oncol. Rep. 2006, 15, 489–494.
(31) Mai, A.; Massa, S.; Lavu, S.; Pezzi, R.; Simeoni, S.; Ragno, R.;
Mariotti, F. R.; Chiani, F.; Camilloni, G.; Sinclair, D. A. Design,
synthesis, and biological evaluation of sirtinol analogues as class III
histone/protein deacetylase (sirtuin) inhibitors. J. Med. Chem. 2005,
48, 7789–7795.
(41) Jain, J.; Almquist, S. J.; Shlyakhter, D.; Harding, M. W. VX-497: a
novel, selective IMPDH inhibitor and immunosuppressive agent.
J. Pharm. Sci. 2001, 90, 625–637.
(42) Jain, J.; Almquist, S. J.; Heiser, A. D.; Shlyakhter, D.; Leon, E.;
Memmott, C.; Moody, C. S.; Nimmesgern, E.; Decker, C. Charac-
terization of pharmacological efficacy of VX-148, a new, potent
immunosuppressive inosine 5′-monophosphate dehydrogenase inhibi-
tor. J. Pharmacol. Exp. Ther. 2002, 302, 1272–1277.
(43) Ishitsuka, K.; Hideshima, T.; Hamasaki, M.; Raje, N.; Kumar, S.;
Podar, K.; Le Gouill, S.; Shiraishi, N.; Yasui, H.; Roccaro, A. M.;
Tai, Y. Z.; Chauhan, D.; Fram, R.; Tamura, K.; Jain, J.; Anderson,
K. C. Novel inosine monophosphate dehydrogenase inhibitor VX-
944 induces apoptosis in multiple myeloma cells primarily via caspase-
independent AIF/Endo G pathway. Oncogene 2005, 24, 5888–5896.
(44) Dhar, T. G.; Guo, J.; Shen, Z.; Pitts, W. J.; Gu, H. H.; Chen, B. C.;
Zhao, R.; Bednarz, M. S.; Iwanowicz, E. J. A modified approach to
2-(N-aryl)-1,3-oxazoles: application to the synthesis of the IMPDH
inhibitor BMS-337197 and analogues. Org. Lett. 2002, 4, 2091–2093.
(45) Lee, Y. M.; Fujiwara, Y.; Ujita, K.; Nagatomo, M.; Ohta, H.; Shimizu,
I. Syntheses of mycophenolic acid and its analogs by palladium
methodology. Bull. Chem. Soc. Jpn. 2001, 74, 1437–1443.
(46) Aoyama, T.; Shioiri, T. New methods and reagents in organic-
synthesis. 8. Trimethylsilyldiazomethane, a new, stable, and safe
reagent for the classical Arndt-Eistert synthesis. Tetrahedron Lett.
1980, 21, 4461–4462.
(47) Jones, D. F.; Mills, S. D. Preparation and antitumor properties of
analogs and derivatives of mycophenolic acid. J. Med. Chem. 1971,
14, 305–311.
(48) Watterson, S. H.; Liu, C.; Dhar, T. G.; Gu, H. H.; Pitts, W. J.; Barrish,
J. C.; Fleener, C. A.; Rouleau, K.; Sherbina, N. Z.; Hollenbaugh, D. L.;
Iwanowicz, E. J. Novel amide-based inhibitors of inosine 5′-mono-
phosphate dehydrogenase. Bioorg. Med. Chem. Lett. 2002, 12, 2879–
2882.
(49) Umejiego, N. N.; Li, C.; Riera, T.; Hedstrom, L.; Striepen, B.
Cryptosporidium parvum IMP dehydrogenase: identification of func-
tional, structural, and dynamic properties that can be exploited for
drug design. J. Biol. Chem. 2004, 279, 40320–40327.
(50) Morrison, J. F. Kinetics of the reversible inhibition of enzyme-catalysed
reactions by tight-binding inhibitors. Biochim. Biophys. Acta 1969,
185, 269–286.
(32) Yang, T.; Sauve, A. A. NAD metabolism and sirtuins: metabolic
regulation of protein deacetylation in stress and toxicity. Am. Assoc.
Pharm. Sci. J. 2006, 8, E632–E643.
(33) Sauve, A. A.; Moir, R. D.; Schramm, V. L.; Willis, I. M. Chemical
activation of Sir2-dependent silencing by relief of nicotinamide
inhibition. Mol. Cell 2005, 17, 595–601.
(34) Marks, P. A. Discovery and development of SAHA as an anticancer
agent. Oncogene 2007, 26, 1351–1356.
(35) Nimmanapalli, R.; Fuino, L.; Stobaugh, C.; Richon, V.; Bhalla, K.
Cotreatment with the histone deacetylase inhibitor suberoylanilide
hydroxamic acid (SAHA) enhances imatinib-induced apoptosis of Bcr-
JM070864W