Selective AKR1C3 Inhibitors Potentiate Chemotherapeutic Activity in Multiple Acute Myeloid Leukemia (AML) Cell Lines

Article abstract of DOI:10.1021/acsmedchemlett.6b00163

We report the design, synthesis, and evaluation of potent and selective inhibitors of aldo-keto reductase 1C3 (AKR1C3), an important enzyme in the regulatory pathway controlling proliferation, differentiation, and apoptosis in myeloid cells. Combination treatment with the nontoxic AKR1C3 inhibitors and etoposide or daunorubicin in acute myeloid leukemia cell lines, elicits a potent adjuvant effect, potentiating the cytotoxicity of etoposide by up to 6.25-fold and the cytotoxicity of daunorubicin by >10-fold. The results validate AKR1C3 inhibition as a common adjuvant target across multiple AML subtypes. These compounds in coadministration with chemotherapeutics in clinical use enhance therapeutic index and may avail chemotherapy as a treatment option to the pediatric and geriatric population currently unable to tolerate the side effects of cancer drug regimens.

Full text of DOI:10.1021/acsmedchemlett.6b00163

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Letter
Selective AKR1C3 inhibitors potentiate chemotherapeutic
activity in multiple acute myeloid leukemia (AML) cell lines
Kshitij Verma, Tianzhu Zang, Nehal Gupta, Trevor M. Penning, and Paul C. Trippier
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.6b00163 • Publication Date (Web): 22 Jun 2016
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Selective AKR1C3 inhibitors potentiate chemotherapeutic ac-
tivity in multiple acute myeloid leukemia (AML) cell lines
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Kshitij Vermaa, Tianzhu Zangb, Nehal Guptac, Trevor M. Penningb, and Paul C. Trippiera,d,*
a Department of Pharmaceutical Sciences, Texas Tech University Health Sciences Center, School of Pharmacy, Amarillo,
TX, 79106, United States
b Center of Excellence in Environmental Toxicology, Department of Systems Pharmacology & Translational Therapeutics,
Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104-6160, United States
c Department of Biomedical Sciences, Texas Tech University Health Sciences Center, School of Pharmacy, Amarillo, TX,
79106, United States
d Center for Chemical Biology, Department of Chemistry and Biochemistry, Texas Tech University, Lubbock, TX 79409-
1061, United States
KEYWORDS: AKR1C3 Inhibitor, acute myeloid leukemia, etoposide, daunorubicin, synergism, adjuvant.
ABSTRACT: We report the design, synthesis and evaluation of potent and selective inhibitors of aldoꢀketo reductase 1C3
(AKR1C3), an important enzyme in the regulatory pathway controlling proliferation, differentiation and apoptosis in myeloid cells.
Combination treatment with the nonꢀtoxic AKR1C3 inhibitors and etoposide or daunorubicin in acute myeloid leukemia cell lines,
elicits a potent adjuvant effect, potentiating the cytotoxicity of etoposide by up to 6.25ꢀfold and the cytotoxicity of daunorubicin by
>10ꢀfold. The results validate AKR1C3 inhibition as a common adjuvant target across multiple AML subꢀtypes. These compounds
in coꢀadministration with chemotherapeutics in clinical use enhance therapeutic index and may avail chemotherapy as a treatment
option to the pediatric and geriatric population currently unable to tolerate the side effects of cancer drug regimens.
The aldoꢀketo reductase family 1 member C (AKR1C) enꢀ
zymes, are oxidoreductases which catalyze the NADPHꢀ
dependent reduction of aldehyde and ketone functionalities on
a range of steroids, carbohydrates and prostaglandins.1 The
AKR1C3 enzyme, also known as prostaglandin (PG) F2α synꢀ
thase,2 converts PGD2 to 11βꢀPGF2α, which acts to prevent
myeloid differentiation and facilitates proliferation of tumor
cells by preventing peroxisome proliferatorꢀactivated receptor
(PPAR) γ activation.3, 4 In the absence of AKR1C3, PGD2 can
be converted to the PGJ2 series of prostanoids that activate
PPARγ leading to differentiation and apoptosis.5
promyelocytic leukemia (APL) but its effects are limited to
this one subꢀtype of AML.10 The identification of a therapeutic
agent with activity across all AML subꢀtypes would have subꢀ
stantial clinical implications.
Primary AML cells and the AML cell lines HLꢀ60 (APL, M3
subꢀtype) and KG1a (AML, M0 subꢀtype) predominantly exꢀ
press AKR1C3 with median levels two orders of magnitude
greater than the AKR1C1 isozyme and more than three orders
of magnitude greater than the AKR1C2 isozyme.11ꢀ13 Strong
expression of AKR1C3 is also detected in nonꢀmalignant proꢀ
liferating CD34+ve cells isolated from peripheral blood.13
Overall, this data identifies AKR1C3 as the primary AKR1C
isoform encountered in myeloid progenitors and indicates a
critical role in the regulation of myelopoiesis. Pharmacological
inhibition of AKR1C3 holds the promise of an adjuvant effect,
sensitizing leukemic cells to the cytotoxic action of chemoꢀ
therapeutics delivered synergistically.
Acute myeloid leukemia (AML) is a hematologic neoplasm
characterized by proliferation of poorly differentiated myeloid
progenitor cells.6 The current treatment of AML involves
chemotherapy with cytarabine combined with etoposide and/or
anthracycline chemotherapeutics7. Young age (<1)8 and older
age (>60),7 are widely recognized risk factors for a major
cause of therapeutic failure in AML, the inability to tolerate
high toxicity associated with the most intensive chemotheraꢀ
peutic treatment regimes, resulting in an increased rate of early
death. Adjuvant therapies that increase response to the chemoꢀ
therapeutic agent, yet have no innate toxicity, represent a
promising therapeutic strategy allowing for lower dosing and
hence, less toxicity to nonꢀmalignant cells while preserving
the desired cytotoxic effect in cancerous cells.9 Allꢀtrans retꢀ
inoic acid (ATRA) is an effective differentiation agent in acute
Combination of the weak and nonꢀselective panꢀAKR1C inꢀ
hibitor medroxyprogesterone acetate (AKR1C3 pIC50 = 5.6)
and bezafibrate demonstrated an approximate twoꢀfold potenꢀ
tiation of cytotoxic activity.14 A recent study reported that the
specific AKR1C3 inhibitor 4ꢀMDDT (pIC50 = 6.3) does not
give the adjuvant effect at concentrations up to 50 µM that a
panꢀAKR1C inhibitor does, despite the low expression of othꢀ
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er isozymes in AML cells, casting doubt on the validity of
Scheme 1. Synthesis of para-amide derivative 2.
AKR1C3 as a therapeutic target in AML.15
1
2
3
4
5
6
7
8
O
NH2
The structurally distinct natural product baccharin (1, Figure
1a) demonstrates highly potent inhibitory activity for
AKR1C3 (pIC50 = 7.0). Critically, baccharin exhibits exquisite
selectivity with absolutely no inhibition of the AKR1C1 or
AKR1C2 isoforms.16 Hydrolysis of the ester moiety of 1 proꢀ
vides the known phenol drupanin (1a, Figure S7), which posꢀ
sesses attenuated AKR1C3 inhibitory activity (pIC50 = 4.8)
with only sevenꢀfold selectivity for AKR1C3 over the 1C2
isoform. Molecular modeling of baccharin in the active site of
AKR1C3 predicts formation of a hydrogen bond between the
ester carbonyl and the active site Tyr55 residue.16 Ester bond
hydrolysis is a primary feature of metabolism and the magniꢀ
tude in the reduction of potency for drupanin highlights the
unsuitable pharmacokinetics of baccharin as a drug candidate
or chemical probe.17
NH2
NH2
Br
HN
Ph
Br
i
ii
iii
Br
I
I
CO2tBu
6
CO2tBu
5
7
8
O
O
9
HN
Ph
HN
Ph
iv
v
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t
CO2 Bu
CO2H
9
2
Reagents and conditions: i) Br2, AcOH, 44%; ii) tert-butylacrylate, NEt3, P(Ph)3,
Pd(OAc)2, PhMe, reflux, 64%; iii) 3-phenylpropanoyl chloride, NEt3, DMAP, DCM, 70oC, 10%;
iv) prenylboronic acid pinacol ester, Pd(dppf)Cl2, CsCO3, DMF, 900C, 44%; v) SiO2,
PhMe, reflux, 50%.
We sought to evaluate the suitability of baccharin and rationalꢀ
ly designed hydrolytically more stable derivatives as chemical
probes to evaluate adjuvant effect in AML cell lines. To this
end, we replaced the ester bond of 1 with the hydrolytically
more stable amide bioisostere (2, Scheme 1). Commercially
available 4ꢀiodoaniline (5) was brominated and the desired
metaꢀbromide (6) was obtained as the major product by colꢀ
umn chromatography. Selective Heck reaction with tertꢀ
butylacrylate was achieved by exploiting the greater reactivity
of the iodo substituent over the bromide,18 providing cinnamic
ester (7). Dimethylaminopyridine catalyzed addition of a suitꢀ
ably functionalized acid chloride installed the amide bond (8),
subsequent SuzukiꢀMiyaura reaction19 with prenyl pinacolꢀ
borane provided ester (9) which underwent hydrolysis by exꢀ
posure to silica gel,20 to yield the amide derivative of bacchaꢀ
rin (2). Ring analogues of the hit compound were synthesized
based on the premise that reducing the distance of the hydroꢀ
gen bond between the AKR1C3 active site Tyr55 residue and
the side chain ester carbonyl would result in enhanced potency
coupled with enhanced steric congestion around the ester
providing greater hydrolytic stability. Commercially available
3ꢀhydroxycinnamic acid (10, Scheme 2) was protected as the
methyl ester (11). Subsequent bromination with molecular
bromine provided 4ꢀbromoꢀ3ꢀhydroxycinamic ester (12),
which was readily separable from regioisomers by column
chromatography. Subsequent SuzukiꢀMiyaura coupling with
prenyl pinacolborane and hydrolysis provided cinnamic acid
(14), which underwent addition of a suitably functionalized
acid chloride to yield metaꢀester derivative (3). In a similar
fashion, the metaꢀamide derivative (4, Scheme 3) was acꢀ
cessed utilizing the same synthetic sequence but beginning
with commercial 2ꢀbromoꢀ5ꢀiodoaniline (15).
Scheme 2. Synthesis of meta-ester derivative 3.
CO2R
CO2Me
ii
iii
Br
OH
OH
12
10 R = H
i
11 R = Me
CO2H
CO2R
v
O
OH
13 R = Me
14 R = H
O
iv
3
Reagents and Conditions: i) MeOH, H2SO4, reflux, 93%; ii Br2, AcOH, RT, 35%;
iii) prenylboronic acid pinacol ester, Pd(dppf)Cl2, CsCO3, DMF, 90oC, 27%;
iv) NaOH, H2O, reflux, 99%; v) 3-phenylpropanoyl chloride, DMAP, NEt3, DCM, RT, 82%
Scheme 3. Synthesis of meta-amide derivative 4.
Table 1. AKR1C3 inhibition activity and selectivity of syn-
thesized compounds.
The inhibitory activities of the synthesized baccharin derivaꢀ
tives against AKR1C3 and the highly homologous isozyme
AKR1C2 were determined (Table 1).21 Synthetic baccharin (1)
exhibited potency for AKR1C3 inhibition (pIC50 = 7.0) and
excellent selectivity (510ꢀfold) as expected. The paraꢀamide
Compound AKR1C3
pIC50
AKR1C2
pIC50
Fold selectivity
AKR1C3
1
1a
7.0
4.8
4.3
3.9
510
7
derivative (2) retained some potency and selectivity (pIC50
=
2
3
4
6.4
7.1
4.4
4.6
5.1
5.4
89
6.4 with 89ꢀfold selectivity) for AKR1C3. The meta-ester deꢀ
rivative (3) represents the best combination of potent inhibitor
(pIC50 = 7.1) and selectivity (261ꢀfold for AKR1C3). The me-
taꢀamide derivative (4) is the most potent AKR1C3 inhibitor
of this scaffold class (pIC50 = 7.2) with 109ꢀfold affinity for
AKR1C3.
261
109
0.66
7.2
MPA*32
5.632
*MPA – Medroxyprogesterone Acetate
This selection of compounds enable direct comparison of adꢀ
juvant activity for the four AKR1C3 inhibitors (1ꢀ4) which
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Figure 1. Synergistic activity of AKR1C3 inhibitors with etoposide in HL-60 cells. 72 hr coꢀincubation with etoposide. Values are the
mean ± S.D. (n = 6). The twoꢀtailed tꢀtest analysis was used to compare the statistical difference between control and treatments, ns ꢀ not
significant * p<0.01, ** p<0.05, *** p<0.001, **** p<0.0001.
exhibit potent inhibitory activity and selectivity over AKR1C2
while incorporating hydrolytically labile and stable linkers to a
pharmacophoric moiety.
and S6). In accordance with the literature, 0.1 µM of etoposide
provides no cytotoxic effect in HLꢀ60 cells.26 The IC50 value
of etoposide was calculated as 1.16 µM and 6.70 µM for the
HLꢀ60 and KG1a cell lines respectively (Table 2).
To investigate the general cytotoxicity of baccharin (1) against
a range of cancer cell lines, the compound was submitted to
the National Cancer Institute 60 cancer cell line assay (Figure
S1).22 The results, in agreement with others,15 show
little cytotoxic activity associated with the inhibition of
AKR1C3 at a concentration of 10 µM. Baccharin (1) and the
derivative AKR1C3 inhibitors (2 and 3) demonstrate no toxiciꢀ
ty to the HLꢀ60 human APL cell line at concentrations up to
100 µM (Figure S2). The highly potent metaꢀamide derivative
4 was significantly (p<0.0001) cytotoxic at 50 µM. At the
lower concentrations employed in this study (0.1 – 1 µM) 4
showed up to 8% reduction of cell viability. The toxicity of
AKR1C3 inhibitor 4 may be attributed to its high potency for
AKR1C3 enzyme inhibition that leads to cell death, as has
been reported for similarly potent AKR1C3 inhibitors in prosꢀ
tate cancer cells.23 When the AKR1C3 inhibitors were exposed
to the KG1a AML cell line (AML M0 subꢀtype) which has
much greater expression of AKR1C3 (Figure S3), cytotoxicity
was observed at concentrations above 25 µM for inhibitors 1ꢀ3
indicating direct toxicity to AML cells upon inhibition of
AKR1C3 (Figure S4). Potent inhibitor 4 showed 20% reducꢀ
tion of cell viability at just 1 µM in the KG1a cell line.
Table 2. Adjuvant effect of compounds 1 - 4 to potentiate
the effect of etoposide in human AML cell lines upon co-
treatment at 72 hr.
Compound AKR1C3
pIC50
HL-60
KG1a
*
*
CIa DRIb IC50
CIa DRIb IC50
(ꢀM)
(ꢀM)
1
2
3
4
7.0
6.4
7.1
7.2
N/A
0.44 2.25 0.51 N/A N/A N/A
0.38 2.56 0.45 0.42 2.34 2.88
0.28 3.46 0.33 0.16 6.25 1.08
0.45 2.17 0.53 0.37 2.73 2.47
N/A N/A 1.16 N/A N/A 6.70
Etoposide
*Calculated for etoposide + AKR1C3 inhibitor (ꢀM)
aCombination index
bDose reduction index
N/A; Not Applicable
Combination treatment of HLꢀ60 cells with a range of concenꢀ
trations of etoposide and AKR1C3 inhibitor was performed to
determine effect on cell viability (Figure 1). Etoposide treatꢀ
ment of HLꢀ60 cells alone at 1 µM reduces cell viability by
40%. The natural product hit compound baccharin (1) at a
concentration of 0.1 µM when delivered synergistically with
etoposide (1 µM) provided a significant (p<0.0001) potentiaꢀ
tion of etoposide cytotoxicity providing an overall reduction of
cell viability by 70% (Figure 1a). When amide analogue (2),
the lowest potency AKR1C3 inhibitor of the four lead comꢀ
pounds (pIC50 = 6.4), was delivered synergistically at 0.1 µM
concentration with 1 µM etoposide, a potentiation effect was
observed that provided an overall 57% reduction of cell viabilꢀ
ity (Figure 1b). The ring analogue (3) with greater AKR1C3
The clinically approved drug etoposide, employed as a second
line chemotherapeutic to manage AML24, 25 was chosen as the
cytotoxic agent. The use of etoposide ensures that any potentiꢀ
ation of cytotoxicity would be attributable to AKR1C3 inhibiꢀ
tion rather than prevention of AKR1C3ꢀmediated metabolism
of the chemotherapeutic agent. A doseꢀresponse curve of
etoposide was obtained in HLꢀ60 and KG1a cells (Figure S5
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Figure 2. Synergistic activity of AKR1C3 inhibitors with etoposide in KG1a cells. 72 hr coꢀincubation with etoposide. Values are the
mean ± S.D. (n = 6). The twoꢀtailed tꢀtest analysis was used to compare the statistical difference between control and treatments, ns ꢀ not
significant * p<0.01, ** p<0.05, *** p<0.001, **** p<0.0001.
inhibition potency (pIC50 7.1) at a concentration of 0.1 µM
provided an 80% cell viability reduction in combination with 1
µM etoposide (Figure 1c). The most potent AKR1C3 inhibitor,
amide analogue (4) (pIC50 = 7.2), provided a cell viability
reduction of 53% at 0.1 µM along with 1 µM etoposide. A
significant (p<0.0001) adjuvant effect was observed when 4
was incubated with just 0.5 µM of etoposide. This concentraꢀ
tion of etoposide provides a 20% reduction of cell viability
alone, which was potentiated by the action of 4 at 0.1 µM to
provide a 45% reduction of cell viability (Figure 1d). When
comparison is made at this concentration of etoposide with the
less active inhibitor 2, a reduction of cell viability of only 25%
is observed. These data illustrate that all four AKR1C3 inhibiꢀ
tors (1 ꢀ 4) show a potent adjuvant effect.
This observation can be attributed to the greater inhibition of
AKR1C3 before addition of etoposide.
To quantify the degree of synergism, the results of the coꢀ
treatment and preꢀtreatment experiments were analyzed by
CompuSyn software (Paramus, NJ) based on the median effect
principle or ‘ChouꢀTalalay’ method.27,28 The combination inꢀ
dex (CI) and dose reduction index (DRI) values were calculatꢀ
ed at a constant ratio of etoposide to AKR1C3 inhibitor at
50% cytotoxic effect (Fa = 0.5). In all experiments, CI values
were found to be < 1, indicative of synergism. As outlined in
Table 2, coꢀtreatment of each AKR1C3 inhibitor with etopoꢀ
side in AML cell lines demonstrated a synergistic drug action.
In HLꢀ60 cells up to 3.5ꢀfold reduction in dosing of etoposide
was observed in coꢀtreatment experiments with AKR1C3 inꢀ
hibitors and the dose reduction was increased up to 5.8ꢀfold
upon 24 hr preꢀtreatment with AKR1C3 inhibitor. CI values
exhibited a narrow range from 0.16 (2) to 0.23 (1 and 4), inꢀ
dicative of strong synergism in the preꢀtreatment experiments
in HLꢀ60 cells (Figure 3e). A stronger synergistic action was
observed in KG1a cells, which have much greater AKR1C3
expression, wherein a dose reduction of up to 6.25ꢀfold was
observed upon coꢀtreatment. CI and DRI indices ranged from
0.16 (3) – 0.42 (2) and 2.34 (2) – 6.25 (3) respectively (Table
2).
The adjuvant effect was enhanced in KG1a cells when comꢀ
pounds 2 ꢀ 4 were incubated with etoposide (Figure 2aꢀc). The
coꢀtreatment experiments demonstrated potentiation of etopoꢀ
side toxicity at 1 µM dose from 18% cell viability reduction
(etoposide alone) to 42% when dosed with 0.1 µM of inhibitor
2 (Figure 2a). Ring analogues 3 and 4 at 0.1 µM reduced
KG1a cell viability by 54% and 40% respectively with 1 µM
etoposide (Figure 2b and 2c).
To assess if this adjuvant effect was indeed attributable to
AKR1C3 inhibition we employed the hydrolysis product of 1,
the low potency (pIC50 = 4.8) and nonꢀselective (7ꢀfold selecꢀ
tivity for AKR1C3) inhibitor drupanin (1a) as a control comꢀ
pound. The observed adjuvant effect diminished substantially,
in parallel with AKR1C3 inhibition activity, when 1a was coꢀ
incubated with etoposide in HLꢀ60 cells. Compound 1a
demonstrated simply additive effects, providing no statistically
significant adjuvant effect (p = 0.8512) (Figure S7).
The data presented in Table 2 provides a clear quantification
of synergistic activity of the AKR1C3 inhibitors by computer
simulation. From the calculated DRI values, metaꢀester 3 conꢀ
fers greatest adjuvant activity upon synergistic treatment with
etoposide. The cytotoxicity of etoposide is enhanced by 3.5ꢀ
fold in HLꢀ60 cells and 6.25ꢀfold in KG1a cells, resulting in
new calculated IC50 values (3 and etoposide) of just 0.33 µM
and 1.08 µM in HLꢀ60 and KG1a cells respectively. These
data can perhaps be rationalized to the enhanced stability of
meta-ester 3 to hydrolysis. The hit compound baccharin is
metabolically labile; hydrolysis is known to provide an inacꢀ
tive compound (1a). The metaꢀester 3 experiences greater
steric congestion around the labile ester bond and as such
would be expected to be more resistant to hydrolysis than 1.
Inhibitor 2, by the nature of the amide bond, would be exꢀ
pected to be more stable to hydrolysis than esters 1 and 3 and
To further discern if inhibition of AKR1C3 is indeed responꢀ
sible for sensitizing AML cells to the cytotoxic action of
etoposide, AKR1C3 inhibitors were next incubated in HLꢀ60
cells for 24 hr prior to the addition of etoposide. After a furꢀ
ther 72 hr, cell viability was measured (Figure 3aꢀd). All four
AKR1C3 inhibitors demonstrated a greater potentiation effect
after 24 hr preꢀtreatment in contrast to the coꢀtreatments,
showing a near complete abrogation of cell viability (80 ꢀ 95%
cell viability reduction) when combined with 1 µM etoposide.
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pression was observed. The data are in agreement with prior
1
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reports that show a transient increase in AKR1C3 expression
immediately preceding differentiation.31
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AKR1C3 inhibitors derived from modification of a natural
product have enhanced biological stability and exhibit exꢀ
tremely selective and potent activity. Proofꢀofꢀconcept that
AKR1C3 inhibitors derived from baccharin have a synergistic
effect sensitizing AML cells to the chemotherapeutic effects of
etoposide and daunorubicin is demonstrated. Treatment with
the nonꢀtoxic AKR1C3 inhibitors in two in vitro models of
AML with coꢀadministration of the clinical chemotherapeutics
etoposide or daunorubicin results in a synergistic drug action.
The IC50 value of etoposide was reduced from 1.16 µM to 0.21
µM in HLꢀ60 cells and from 6.70 µM to 1.08 µM in KG1a
cells. Daunorubicin is potentiated by up to 10ꢀfold in both HLꢀ
60 and KG1a cells. This is contrary to recent reports detailing
selective AKR1C3 inhibitors do not perform an adjuvant role
compared to panꢀAKR1C isoform inhibitors.15
Our results suggest that a strategy developing small molecule
AKR1C3 inhibitors may yield powerful adjuvant agents for
the synergistic treatment of leukemia, especially since favoraꢀ
ble toxicity and pharmacokinetic properties can exist within
the baccharin structure The identified highly potent and selecꢀ
tive derivatives represent valuable lead compounds to underꢀ
stand the structural features required for AKR1C3 inhibitory
activity and selectivity, and how these properties interrelate to
further our understanding of the role which AKR1C3 plays to
enable potentiation of chemotherapeutic effect within AML.
Figure 3. Synergistic activity of AKR1C3 inhibitors with
etoposide in HL-60 cells (pre-treatment). a,b,c,d) 24 hour preꢀ
treatment of AKR1C3 inhibitor followed by 72 hr exposure to
etoposide. Values are the Mean ± S.D. (n = 6). The twoꢀtailed tꢀ
test analysis was used to compare statistical difference between
control and treatment ns ꢀ not significant, ** p<0.05, ***
p<0.001, **** p<0.0001. e) Quantification of synergistic activity.
ASSOCIATED CONTENT
Supporting Information
provides the greatest potentiation effect in preꢀtreatment experiꢀ
ments in HLꢀ60 cells, yet is a less potent AKR1C3 inhibitor. Amꢀ
ide 4, while the most potent AKR1C3 inhibitor, possesses innate
cytotoxicity that reduces the DRI index, accounting for the obꢀ
served CI value of 0.45. Comparison of cell viability data (Figure
1) resulting from combination of AKR1C3 inhibitor (0.1 µM) and
1 µM etoposide in HLꢀ60 cells with 72 hr incubation show a lineꢀ
ar relationship between AKR1C3 inhibition potency and reduction
in cell viability. Compound 3 (pIC50 = 7.1) with etoposide proꢀ
vides 80% cell death, compound 1 (pIC50 = 7.0) with etoposide
provides 70% cell death and compound 4 (pIC50 = 6.4) with
etoposide provides 57% cell death. Indeed this is supported by
data from coꢀtreatment experiments in both HLꢀ60 and KG1a
cells; amide 2 provides a lower DRI (2.56 and 2.34 respectively)
than the more potent metaꢀester 3 (3.46 and 6.25 respectively)
relating AKR1C3 inhibition potency to potentiation effect.
The Supporting Information is available free of charge on the
ACS Publications website.
Full experimental details and characterization for all final comꢀ
pounds and descriptions of biological assays (PDF).
AUTHOR INFORMATION
Corresponding Author
*Tel: 1ꢀ806ꢀ414ꢀ9245. Eꢀmail: paul.trippier@ttuhsc.edu
Funding Sources
This work was supported by the NCI and NIEHS (R01ꢀCA90744
and P30ꢀES013508 to T.M.P) from the National Institutes of
Health and by Texas Tech University Health Sciences Center
(P.C.T).
To establish the scope of synergistic activity across other
chemotherapeutics, daunorubicin was employed. Pretreatment
experiments were performed in HLꢀ60 and KG1a cells with
compound 3 followed by daunorubicin incubation for 72 hr. A
dose reduction of approximately 10ꢀfold was observed, reducꢀ
ing IC50 from 42 nM to 4.2 nM and from 1.77 µM to 0.2 µM in
the respective cell lines. (Figure S8).
Notes
The authors declare no competing financial interests.
ACKNOWLEDGMENTS
We thank Dr. J. Lu and Dr. S. Srivastava for valuable advice.
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Selective AKR1C3 inhibitors potentiate chemotherapeutic activity in multiple acute myeloid leukemia
(AML) cell lines
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Kshitij Vermaa, Tianzhu Zangb, Nehal Guptac, Trevor M. Penningb, and Paul C. Trippiera,d,*
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