H2O2/Peroxynitrite-Activated Hydroxamic Acid HDAC Inhibitor Prodrugs Show Antileukemic Activities against AML Cells

Article abstract of DOI:10.1021/acsmedchemlett.8b00057

Occurrence of acute myeloid leukemia (AML) results in abundant endogenous reactive oxygen species (ROS)/reactive nitrogen species (RNS) in AML cells and in disease-relevant microenvironments. Histone deacetylase inhibitor (HDACi) prodrug approach was designed accordingly by masking the hydroxamic acid zinc binding group with hydrogen peroxide (H₂O₂)/peroxynitrite (PNT)-sensitive, self-immolative aryl boronic acid moiety. Model prodrugs 5-82 and 5-23 were activated in AML cells to release cytotoxic HDACis, evidenced by inducing acetylation markers and reducing viability of AML cells. Intracellular activation and antileukemic activities of prodrug were increased or decreased by ROS/PNT inducers and scavengers, respectively. Prodrugs 5-82 and 5-23 also enhanced the potency of chemotherapy drug cytarabine, supporting the potentials of this prodrug class in combinatorial treatment.

Full text of DOI:10.1021/acsmedchemlett.8b00057

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Letter
H2O2/peroxynitrite-activated hydroxamic acid HDAC inhibitor
prodrugs show antileukemic activities against AML cells
Yi Liao, Liping Xu, Siyu Ou, Holly Edwards, Daniel Luedtke, Yubin Ge, and Zhihui Qin
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.8b00057 • Publication Date (Web): 13 Jun 2018
Downloaded from http://pubs.acs.org on June 14, 2018
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H2O2/peroxynitrite-activated hydroxamic acid HDAC inhibitor pro-
drugs show antileukemic activities against AML cells
Yi Liao,† Liping Xu,† Siyu Ou,† Holly Edwards,§ Daniel Luedtke,§, # Yubin Ge,§ and Zhihui Qin†,*
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† Department of Pharmaceutical Sciences, Eugene Applebaum College of Pharmacy and Health Sciences, Wayne State Uniꢀ
versity, Detroit, Michigan 48201, United States.
§ Department of Oncology and the Molecular Therapeutics Program of the Karmanos Cancer Institute, Wayne State Univerꢀ
sity School of Medicine, Detroit, Michigan 48201, United States.
# Cancer Biology Graduate Program, Wayne State University School of Medicine.
KEYWORDS: HDAC inhibitor, HDAC inhibitor prodrugs, acute myeloid leukemia, hydrogen peroxide, peroxynitrite, reac-
tive oxygen species
ABSTRACT: Occurrence of acute myeloid leukemia (AML) results in abundant endogenous reactive oxygen species
(ROS)/reactive nitrogen species (RNS) in AML cells and in diseaseꢀrelevant microenvironments. Histone deacetylase inhibitor
(HDACi) prodrug approach was designed accordingly by masking the hydroxamic acid zinc binding group with hydrogen peroxide
(H2O2)/peroxynitrite (PNT)ꢀsensitive, selfꢀimmolative aryl boronic acid moiety. Model prodrugs 5-82 and 5-23 were activated in
AML cells to release cytotoxic HDACis, evidenced by inducing acetylation markers and reducing viability of AML cells. Intracelꢀ
lular activation and antileukemic activities of prodrug were increased or decreased by ROS/PNT inducers and scavengers, respecꢀ
tively. 5-82 and 5-23 also enhanced the potency of chemotherapy drug cytarabine, supporting the potentials of this prodrug class in
combinatorial treatment.
Acute myeloid leukemia (AML) remains a challenging disꢀ
ease, overall 5ꢀyear survival is less than 5% in patients over
the age of 65.1 Histone deacetylase inhibitors (HDACis) target
aberrant epigenetic modifications of cancer and have been
studied as an attractive “targeted” treatment for AML.2 Altꢀ
hough promising in preclinical models, HDACis as monotherꢀ
apy only resulted in modest or poor clinical outcomes in AML
trials.3 Multiple factors, such as undesired cardiovascular and
vorinostat), a FDAꢀapproved drug to treat cutaneous T cell
lymphoma14 and 1-58, a piperlongumine (PL)ꢀHDACi hybrid
molecule with potent antiꢀAML activity,15 were converted to
prodrugs 5-82 and 5-23, respectively (Fig. 1a, b). We used
boronic acid instead of boronic acid ester as masking group to
retain water solubility, and at the same time, to increase lipoꢀ
philicity of the parental hydroxamic acid HDACis. For inꢀ
stance, cLogP of 5-82 is 3.26, compared with that of SAHA at
0.99 (calculated by using ChemDraw 12.0), predicting better
tissue/tumor penetration. Hydroxamic acid is liable to O-
glucoronidation16 and hydrolysis, derivatizations with boronic
acid or other “promoiety” could potentially increase metabolic
stability of hydroxamic acidꢀbased HDACis.7
5
gastrointestinal toxicity,4, fast elimination and poor tissue
penetration caused by metabolically labile and highly polar
hydroxamic acid zinc binding group (ZBG),6, 7 hindered the
use of HDACi in AML and other cancer indications. Several
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prodrug approaches, e.g., using carbamate6, and quinone7ꢀ
based moieties to protect hydroxamic acid have been proposed
to partially address these shortcomings.
Different from the reported approaches, we designed novel
HDACi prodrugs by considering chemical properties of the
“promoiety” and the biological characteristics of AML. The
prodrugs were constructed by masking the hydroxamic acid
ZBG with hydrogen peroxide (H2O2)/peroxynitrite (PNT)ꢀ
sensitive, selfꢀimmolative aryl boronic acid promoiety (Fig.
1a). Aromatic boronates/boronic acids specifically react with
H2O2 (a ROS)9 or PNT (a RNS)10 under physiological condiꢀ
tions and have been used to design anticancer prodrugs and
ROS probes.11 Similar to arylboronateꢀbased nitrogen musꢀ
tard12 and aminoferrocene13 prodrugs, boronic acid HDACi
prodrugs could form phenol intermediates during activation
and subsequently release free hydroxamic acid and a shortꢀ
lived, chemically reactive quinone methide (QM) through 1,6ꢀ
elimination (Fig. 1a). In this study, two HDACis, SAHA (i.e.
Figure 1. (a) The design of arylboronic acidꢀmasked hydroxamic
acid prodrug and the mechanism of activation, exemplified using
SAHA and 5-82. (b) Chemical structures of HDACi 1-58, prodrug
5-23, and compound 12.
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Scheme 1. Synthesis of prodrugs 5-23, 5-82 and the 5-82 analogue 12.
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Reagents and conditions: (a) NꢀHydroxyphthalimide, PPh3, DEAD, THF, 82%; (b) Methylhydrazine, THF, 0°C, 77%; (c) Aniline,
BOP, DIPEA, DCM, 70%; (d) LiOH, THF/H2O (1:1), 80%; (e) 1. HBTU, DIPEA, THF; 2. NaIO4, NH4OAc, acetone/H2O (1:1);
57% for 5-23, 60% for 5-82; 75% for 12; (f) (BOC)2O, Et3N, 95%; (g) Bis(pinacolato)diboron, AcOK, Pd(dppf)Cl2, 1,4ꢀdioxane,
80°C, 80%; (h) trifluoroacetic acid, DCM, 91%.
Besides chemical properties, the design of H2O2/PNTꢀ
activated HDACi prodrugs is also reasoned by abundant enꢀ
dogenous ROS/RNS in AML cells and in diseaseꢀrelevant
microenvironments. Due to malfunctioned redox homeostasis
pathways, such as alteration of mitochondrial electron transfer
chain, FLT3ꢀITDꢀcaused upregulation of NADPH oxidase and
the elevated xanthine oxidoreductase activity,17 AML blasts
display higher levels of ROS compared to normal
leukocytes.18ꢀ20 The occurrence of AML also abnormally upꢀ
regulates and activates NOX4 (an superoxideꢀgenerating enꢀ
zyme) and nitric oxide synthase 3 (i.e. eNOS) in bone marrow
(BM) endothelial cells and results in overproduction of nitric
oxide (NO) and ROS.21 This event could eventually produce
higher levels of PNT from the reaction of NO and superoxide
(Fig. S1 a). Peak B eventually disappeared, and the disapꢀ
pearance was accompanied by the increased peak area of
SAHA. At a given time point (e.g., at 30 min, Fig. 2a), the
formation of phenol intermediate and SAHA were increased
along with the elevated H2O2 concentrations (Fig. 2b). 5-82
(100 ꢁM) was completely converted to SAHA by equivalent
(100 ꢁM) or excessive (500 ꢁM) H2O2, whereas only half of
the 5-82 was consumed by 50 ꢁM of H2O2 as determined by
comparisons of the peak area of SAHA with the standard
sample (Fig 2b and Fig. S1 b).
.ꢀ
(O2 ) in AML BM microenvironment. PNT reacts with boꢀ
ronic acid up to a millionꢀfold faster than H2O210, 22 and could
be another activator of boronic acidꢀbased HDACi prodrugs.
Compared with healthy tissue, the elevated H2O2 and PNT
may activate prodrug and release higher concentrations of
cytotoxic HDACi in AML cells and in AMLꢀrelevant,
ROS/RNSꢀenriched microenvironments.
Starting from pinacol ester 1, hydroxylamine 3 was made
by deprotecting Mitsunobu reaction product 2 with methylhyꢀ
drazine. Coupling of 3 with carboxylic acids 415 and 7,23 folꢀ
lowed by the treatment of NaIO4/NH4OAc, gave prodrugs 5-
23 and 5-82 respectively (Scheme 1). To study the potential
biological effects of aryl boronic acid, we prepared a close
structural analogue of 5-82, i.e. compound 12 (Fig. 1b) that
does not convert to HDACi due to replacing the oxygen of
ZBG with a methylene group. 12 was made by condensing 7
with amine 11 which was synthesized from 8 via sequential
amine protection, borylation and acidic deprotection (Scheme
1).
Figure 2. (a) Prodrug 5-82 was activated by H2O2. 5-82 (100 µM)
was incubated with H2O2 in PBS (pH 7.4) at 37°C for 30 min.
Aliquots were analyzed by using HPLC. (b) 5-82 (100 µM) timeꢀ
dependently released SAHA in the presence of H2O2 in 4h in PBS
(pH 7.4) at 37°C. (c) Prodrug 5-82 was activated by PNT. 5-82
(100 µM) was incubated with indicated reagents in PBS (pH 7.4)
at room temperature for 15 min. Aliquots were analyzed by using
HPLC. Benzophenone was used as internal standard (IS). X, xanꢀ
thine; XO, xanthine oxidase; Cat., catalase; PAPA/NO, PAPAꢀ
NONOate.
By using SAHA prodrug 5-82 as a model compound, we
firstly investigated if the prodrug can be activated by H2O2 or
PNT in cellꢀfree incubations. 5-82 was incubated with H2O2
in PBS buffer (pH 7.4) at 37°C. As revealed by HPLC analyꢀ
sis (Fig. 2a), it reacted with H2O2 to form two new peaks: the
retention time of A was identical to that of SAHA, and peak B
was assigned as the phenol intermediate (Fig. 1a) as the measꢀ
ured m/z value matched F.W. of the proposed structure in a
high resolution mass spectrometryꢀbased LCꢀMS analysis
5-82 was also activated by PNT (Fig. 2c). PNT was generꢀ
ated by a mixture of xanthine (X, 200 mM), xanthine oxidase
(XO, 10 mU/mL) and NO donor PAPAꢀNONOate (100 ꢁM).10
.ꢀ
X and XO produce fluxes of superoxide anion (O2 ) which
further reacts with NO to form PNT. Even at room temperaꢀ
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ture, 5-82 was completely converted to the phenol intermediꢀ
QM is formed along with HDACi release (Fig. 1a), and is
antiproliferative to cancer cells.25 4ꢀ(chloromethyl)phenyl
acetate (4ꢀCPA) is a QM precursor:26 the acetate is hydrolyzed
within cells by esterases, and the resulted 4ꢀ
(chloromethyl)phenol simultaneously converts to QM (Fig. S4
a). 4ꢀCPA decreased viability of MV4ꢀ11 cells with IC50 value
(1.13 ꢁM, Fig. S4 b) higher than those of SAHA (0.50 µM)
and 1-58 (0.23 µM). Because the inferior potency vs. SAHA
and the quick trapping of QM by water and GSH, combinaꢀ
tions of 4ꢀCPA (up to 500 nM) and SAHA didn’t outperform
SAHA alone when used in 1:1 ratio to mimic prodrug setting
(Fig. S4 c). However, in treatments where the mole ratio of 4ꢀ
CPA were in excess of SAHA/1-58 and/or concentrations of
4ꢀCPA were increased beyond the cellular buffering capacity
(Fig S4 d, e), 4ꢀCPA indeed enhanced the potency of HDACis,
suggesting that although the QM may not be critical for the
activities of 5-82 or 5-23 in current work, it could become a
positive contributor to the overall cytotoxicity especially when
the released HDACis are less cytotoxic than QM.
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ate by X/XO/PAPAꢀNONOate within 15 min. In sharp conꢀ
trast, PAPAꢀNONOate itself barely induced any change of 5-
82, emphasizing the specific reaction between boronic acid
and PNT. X/XO alone partially consumed 5-82 to generate the
phenol, and this conversion was abrogated by the addition of
catalase (100 U/mL), indicating that 5-82 was specifically
.ꢀ
oxidized by H2O2, a disproportionate product of O2 yielded
from the X/XO system. The addition of catalase also reduced
consumption of 5-82 in the X/XO/PAPAꢀNONOate system,
suggesting that both H2O2 and PNT were generated and were
responsible for the oxidation of aryl boronic acid in this incuꢀ
bation. Since catalase removes H2O2, 5-82 was mainly conꢀ
sumed by PNT in the X/XO/PAPAꢀNONOate/catalase system
to generate the phenol intermediate that gradually released
SAHA (Fig. S2). Taken together, we conclude that 5-82 can
rapidly react with either H2O2 or PTN and subsequently reꢀ
leases SAHA in PBS buffer via a phenol intermediate.
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We also measured cellular HDAC inhibition markers, the
acetylated histone H4 and αꢀtubulin, to further confirm intraꢀ
cellular activation of prodrugs. Both 5-82 and 5-23 concentraꢀ
tionꢀdependently increased acetylation of histone H4 and αꢀ
tubulin within 6 h in MV4ꢀ11 (Fig. 3b) and U937 cells (Fig.
S5). Compared to the same standard SAHA, densitometry
analysis revealed that 1-58 prodrug 5-23 caused more promiꢀ
nent global acetylation than SAHA prodrug 5-82 in MV4ꢀ11
(Fig. 3b) and U937 cells (Fig. S5). To understand this observaꢀ
tion, we measured intrinsic HDAC inhibition activity of parenꢀ
tal HDACis using enzymatic assay against nuclear HDACs
extracted from Hela cells. Interestingly, 1 ꢁM 1-58 and SAHA
treatment resulted in 32% and 65% inhibition, respectively
(Fig. S6), demonstrating that 1-58 is weaker than SAHA at
nuclear HDAC inhibition. Therefore, compared to 5-82, the
increased H4 acetylation induced by 5-23 was likely attributed
to the higher levels of intracellular free HDACi (but not higher
intrinsic HDACi activity), potentially caused by: 1) increased
intracellular prodrug concentration since 5-23 is more lipoꢀ
philic than 5-82 (clogP 4.73 vs. 3.26); and 2) the presence of
ROSꢀinducing piperlongumine functionality27 may enhance
oxidative stress in AML cells, which facilitates prodrug actiꢀ
vation. 5-23 was also more potent than 5-82 in reducing cell
viability, reflected by the IC50s of the two prodrugs: 0.70 ꢁM
vs. 3.07 ꢁM in MV4ꢀ11 cells and 1.61 ꢁM vs. 6.13 ꢁM in
U937 cells (Table S1). At least two factors, the higher intraꢀ
cellular free HDACi concentration and the stronger antiprolifꢀ
erative activity of the released HDACi (1-58 vs. SAHA),15
may have contributed to the greater in vitro activities of 5-23
than 5-82.
Figure 3. (a) Prodrugs 5-82 and 5-23 reduced viability of both
U937 and MV4ꢀ11 AML cells. Viability were measured using
MTT assay. Bars represent mean ± SD, n=8. ** indicates P <
0.01, *** indicates P<0.001. (b) Prodrugs 5-82 and 5-23 concenꢀ
trationꢀdependently increased acetylation of histone H4 (AcꢀH4)
and αꢀtubulin (Tub) in MV4ꢀ11 cells determined by Western
Blotting. MV4ꢀ11 cells were treated with 5-82 or 5-23 at indicatꢀ
ed concentrations for 6 h. The fold changes for the AcꢀH4 and
AcꢀαꢀTub were obtained using densitometry measurements, norꢀ
malized to GAPDH, and then compared with SAHA treatment.
One representative blot from three experiments is shown.
We next tested antileukemic activities of prodrugs 5-82 and
5-23 in AML cells. Both prodrugs concentrationꢀdependently
reduced the viability of U937 and MV4ꢀ11 cells, with IC50s in
sub µM to low µM range (Fig. 3a; Table S1). MV4ꢀ11 is more
sensitive to prodrug treatments than U937 cells, consistent
with their sensitivity to parental HDACis SAHA and 1-58
(Table S1).15 Since boronic acid is a structural component of
several approved (e.g., bortezomib) or clinically tested
drugs,24 we used 12 (Fig. 1b, Scheme 1), an analogue of 5-82,
to assess biological effects of aryl boronic acid of the proꢀ
drugs. 12 didn't affect the viability of MV4ꢀ11 cells at the
concentrations up to 10 ꢁM (Fig. S3 a), suggesting that aryl
boronic acid does not contribute to 5-82ꢀinduced growth inhiꢀ
bition. We also compared prodrug 5-23 and the methyl ester of
4 (i.e. 2715 in our previous research, Fig. S3 b) in U937 cells.
Although sharing the same PL hybrid drug chemical scaffold,
27 was significantly less effective than 5-23 in MTT assay
(Fig. S3 b), providing another evidence supporting the imꢀ
portance of HDACi release for the antileukemic activities of
prodrugs.
We further evaluated selectivity of prodrugs using three
human peripheral blood mononuclear cell (PBMNC) samples
obtained from healthy donors (two for MTT and one for
Western blot assays). Compared to vehicle control, prodrugs
5-82 and 5-23 induced little to no acetylation of histone H4
and αꢀtubulin in PBMNCs at 5 and 10 µM (Fig. S7 a), which
is in contrast to the substantial increase of HDAC inhibition
markers in AML cells (Fig. 3b, Fig. S5). In MTT assays, the
IC50 of 5-82 was greater than 10 µM in both PBMNC samples,
and the IC50 of 5-23 was 7.6 µM in one sample and was greatꢀ
er than 10 µM in another sample (Fig. S7 b). The higher IC50
values (Table S1) and lower induction of HDAC inhibition
markers in PBMNC samples vs. AML cells support the selecꢀ
tivity of 5-82 and 5-23 to AML cells over normal cells.
HDACi SAHA also showed selectivity, however, SAHA was
more potent against normal cells than the prodrugs (Table S1,
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Fig.7 b). Besides cancer cells, the cancer microenvironment
in an AML clinical trial.29 To explore the feasibility of HDACi
prodrugs in combinatorial therapy, MV4ꢀ11 cells were treated
with 5-82 or 5-23 in combination with cytarabine. Both proꢀ
drugs significantly enhanced the potency of cytarabine (0.05ꢀ
0.5 µM) (Fig. 5a), and the inhibitory effects of drug combinaꢀ
tion were accompanied by the substantially increased PARPꢀ1
and caspaseꢀ3 cleavage compared to the individual drug
treatments (Fig. 5b).
may also generate higher levels of ROS/RNS21 that promote
the release of HDACis. Selectivity of this HDACi prodrug
class can be further tested in in vitro and in vivo models conꢀ
taining microenvironmental components.
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Figure 5. Prodrugs 5-82 (2.5 µM) and 5-23 (0.5 µM) enhanced
the antileukemic effects of cytarabine in MV4ꢀ11 cells. (a) MV4ꢀ
11 cells were treated with drugs at indicated concentrations for 72
h, cell viability was measured using MTT assay. Bars represent
mean±SD, n=6. *** indicates P < 0.001. (b) MV4ꢀ11 cells were
treated with prodrugs and/or cytarabine (0.5 µM) for 24 h, expresꢀ
sion of cleaved caspaseꢀ3 and PARPꢀ1 were analysed using Westꢀ
ern Blotting. One representative blot from three experiments is
shown.
Figure 4. Antileukemic activities of prodrug 5-82 were influꢀ
enced by H2O2/PNT inducers and scavengers. (a, b) MV4ꢀ11 cells
were pretreated with GSH (5 mM), GSHEE (5 mM), catalase
(0.125 mg/mL), SOD (0.05 mg/mL) or the combination of LPS
(10 µg/mL) and BSO (2.5 ꢁM) for 2 h, respectively, followed by
5-82 (2.5 ꢁM) treatment for 72 h. Viable cells were measured
using MTT assay. Bars represent mean ± SD, n=6. *** indicates
P < 0.001. (c) MV4ꢀ11 cells were pretreated with catalase (Cat,
0.5 mg/mL), SOD (0.1 mg/mL), LPS (10 µg/mL) or BSO (10
ꢁM) for 2 h, followed by 5-82 (2.5 ꢁM) treatment for 24h. Acetyꢀ
lation of histone H4 and αꢀtubulin were analysed using Western
Blotting. The fold changes for the AcꢀH4 and AcꢀαꢀTub were
obtained by densitometry measurements, normalized to GAPDH,
and then compared to 5-82 treatment. One representative blot
from three experiments is shown.
In summary, we report new HDACi prodrugs designed by
masking the hydroxamic acid ZBG with an aryl boronic acidꢀ
based promoiety. Model prodrugs were activated by H2O2 or
PNT, leading to intracellular release of active HDACi and
reduction of viable AML cells. Supported by enhancing the
potency of cytarbine, this HDACi prodrug class could be
combined with conventional chemotherapy drugs as well as
other “targeted” antiꢀAML agents. Given the broad pharmacoꢀ
logical effects of HDACi30 and the critical roles of ROS31 and
PNT32 in human diseases, H2O2 and PNTꢀactivated HDACi
prodrugs may have great potential in the treatment of cancer
and nonꢀcancer indications.
We then investigated if antileukemic activities of prodrug
could be affected by exogenous redox reagents. Pretreatment
with an antioxidant, such as GSH, GSH ethyl ester (GSHEE)
or catalase, significantly rescued viability of MV4ꢀ11 cells
from 5-82 treatment (Fig. 4a). 5-82 was stable in GSH coꢀ
incubation (Fig. S8), suggesting GSH or GSHEE rescued cells
via scavenging intracellular ROS/RNS instead of their reacꢀ
tions with the electrophilic boron center. On the contrary,
H2O2 and/or PNT inducers, such as superoxide dismutase
(SOD) and a combination of lipopolysaccharide (LPS) and
buthionineꢀ(S,R)ꢀsulfoximine (BSO), notably enhanced inhibiꢀ
tory effect of 5-82 in MV4ꢀ11 cells (Fig. 4b). Besides potenꢀ
tially affecting biological activities of the released HDACis,
redox agents could modulate prodrug activation. Indeed, SOD,
the combination of LPS and BSO increased and catalase deꢀ
creased acetylation of histone H4 and αꢀtubulin (Fig. 4c). Catꢀ
alase suppresses whereas SOD promotes intracellular H2O2
production, consistent with their opposite effects on HDAC
inhibition markers. LPS induces iNOS and BSO depletes
GSH, their combination could boost ROS (e.g., H2O2) and
PNT levels and generate oxidative environment favouring
prodrug activation. Collectively, antileukemic activities of
HDACi prodrugs could be enhanced by exogenous ROS/RNS
inducers whereas being weakened by antioxidants, modulating
prodrug activation could be one of the factors responsible for
the observed cellular effects.
ASSOCIATED CONTENT
Supporting Information
The Supporting Information, include general experimental methꢀ
ods, HPLC analysis of incubation mixtures, detailed synthetic
procedures and compound characterization (1H, 13CꢀNMR and
MS), is available free of charge on the ACS Publications website.
AUTHOR INFORMATION
Corresponding Author
*Phone: 313ꢀ577ꢀ1567. Fax: 313ꢀ577ꢀ2033. Eꢀmail:
zqin@wayne.edu
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENT
This study was supported by a Wayne State University startup
fund (Z.Q.) and pilot grant from Karmanos Cancer Institute Canꢀ
cer Epigenetics Strategic Research Funding (Z.Q. and Y.G.). The
authors would like to thank Dr. Maozi Liu (Agilent, Inc.) for
characterizing phenol intermediate using high resolution mass
spectrometry.
Because of pleiotropic anticancer mechanisms, HDACi is a
versatile component of combinatorial AML treatments.28 For
instance, SAHA was combined with idarubicin and cytarabine
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and association between SAHA glucuronidation activity phenotype
ABBREVIATIONS
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1
2
3
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AML, acute myeloid leukemia; SAHA (vorinostat), suberaniloꢀ
hydroxamic acid; HDACis, histone deacetylase inhibitors; H2O2,
hydrogen peroxide; PNT, peroxynitrite; NOX, NADPH oxidase;
X, xanthine; XO, xanthine oxidase; PAPA/NO, PAPAꢀNONOate;
GSHEE, glutathione ethyl ester; LPS, lipopolysaccharide; BSO,
buthionineꢀ(S,R)ꢀsulfoximine; SOD, superoxide dismutase; PBS,
phosphateꢀbuffered saline.
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White, P.; Marquez, N.; Hills, R. K.; Burnett, A. K.; Tonks, A.;
Darley, R. L. Overproduction of NOXꢀderived ROS in AML
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H2O2/peroxynitrite-activated hydroxamic acid HDAC inhibitor prodrugs show antileukemic ac-
tivities against AML cells
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Yi Liao,† Liping Xu,† Siyu Ou,† Holly Edwards,§ Daniel Luedtke,§, # Yubin Ge,§ and Zhihui Qin†,*†
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