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Histone Deacetylase Inhibitor Potentiates Anticancer Effect ofDocetaxel via Modulation of Bcl-2 Family Proteins and Tubulin inHormone Refractory Prostate Cancer Cells

Jung Jin Hwang, Yong Sook Kim, Mi Joung Kim, Dong Eun Kim, In Gab Jeong

and Choung-Soo Kim*

From the Institute for Innovative Cancer Research (JJH, YSK, MJK, DEK) and Department of Urology (IGJ), University of Ulsan College of

Medicine, Asan Medical Center (CSK), Seoul, Korea

Purpose: We evaluated the antitumor effects of docetaxel (Sigma®) and histone

deacetylase inhibitors in hormone refractory prostate cancer cells, and analyzed

the mechanism by which combination treatment induced cell death.Materials and Methods: We used LNCaP, DU145 and PC3 cells (ATCC®) toevaluate the in vitro apoptotic effects of histone deacetylase inhibitors and their

combinations with docetaxel as well as the molecular mechanisms. The DU145xenograft model was used to evaluate the in vivo efficacy of PXD101 combined

with docetaxel.Results: Suberoylanilide hydroxamic acid or PXD101 inhibited the growth of

hormone dependent LNCaP cells, and hormone independent DU145 and PC3cells. It increased sub-G1 population and activated caspase-8, 9 and 3, indicating

apoptosis induction. Pretreating DU145 cells with docetaxel followed by histonedeacetylase inhibitors showed significant synergistic cytotoxicity compared with

that of simultaneous co-treatment or reverse sequential treatment. Pretreatment

with docetaxel followed by histone deacetylase inhibitors increased the apoptoticsub-G1 population, caspase activation and tubulin acetylation compared withthat of docetaxel alone. Combination treatment decreased Mcl-1 and Bcl-xl, and

increased t-Bid, Bik and Bim. Combined docetaxel and PXD101 reduced tumorsize with efficacy equivalent to that of a double dose of docetaxel alone in the

DU145 xenograft model.

Conclusions: These preclinical results indicate that the sequential combination

of docetaxel and histone deacetylase inhibitors led to a synergistic increase in the

death of hormone refractory prostate cancer cells via intrinsic and extrinsicapoptotic pathways by modulating Bcl-2 family proteins and tubulin in vitro and

in vivo. Results suggest that this combination may be a new therapeutic modalityin patients with hormone refractory prostate cancer.

Key Words: prostate, prostatic neoplasms, histone deacetylases,docetaxel, apoptosis

Abbreviations

and Acronyms

CI combination index

HDAC histone deacetylase

HDACI HDAC inhibitor

HRPC hormone refractory

prostate cancer

MTS 3-(4,5-dimethylthiazol-2-

yl)-5-(3-carboxymethoxyphenyl)-2-

(4-sulfophenyl)-2H-tetrazolium

SAHA suberoylanilide

hydroxamic acid

v/v volume per volume

Submitted for publication March 23, 2010.

Study received institutional animal care and

use committee approval.

Supported by Grant A062254 from the Korea

Health 21 R & D Project, Ministry of Health, Wel-

fare and Family Affairs, and a grant (2009-450)

from the Asan Institute for Life Sciences, Repub-

lic of Korea.

* Correspondence: Department of Urology,

Asan Medical Center, 388-1 Pungnap 2 dong,

Songpa-gu, Seoul 138-736, Korea (telephone: 82-2-

3010-3734; FAX: 82-2-477-8928; e-mail: cskim@

amc.seoul.kr).

PROSTATE cancer is the most common

nonskin cancer and the second most

common cause of cancer death in men

in the United States with an estimated

186,320 newly diagnosed patients and

27,360 prostate cancer deaths in 2008.1

Although docetaxel based regimens

have palliative and survival bene-

fits, men with metastatic HRPC

have only 16 to 18-month median

survival.2,3 Thus, new therapeutic

modalities aimed to improve man-

0022-5347/10/1846-2557/0 Vol. 184, 2557-2564, December 2010

THE JOURNAL OF UROLOGY® Printed in U.S.A.© 2010 by AMERICAN UROLOGICAL ASSOCIATION EDUCATION AND RESEARCH, INC. DOI:10.1016/j.juro.2010.07.035

www.jurology.com 2557

mailto:[email protected]







agement for advanced HRPC include docetaxel

plus other agents.4

The importance of epigenetic changes in cancer

cells led to the development of agents that modify

histones via acetylation, methylation, phosphoryla-

tion, ubiquitination or adenosine triphosphate ribo-

sylation, or that methylate DNA. Of these modifica-tions the acetylation of lysine residues at the histone

N-terminal tail decreases the affinity of histone for

DNA, resulting in the expression of genes related to

tumor suppression and/or differentiation.5 Thus,

HDACIs show potent activity against many can-

cers.6 Five HDACI classes have been characterized,

including the hydroxamic acids SAHA, PXD101,

LAQ-824, LBH 589 and trichostatin A, the short-

chain fatty acids 4-phenylbutyrate, pivaloyloxy-

methyl butyrate and valproic acid, the cyclic tet-

rapeptides trapoxin, apicidin and depsipeptide (FK

228), and benzamide (MS-275).7–10 Many reportsshow the usefulness of various HDACIs for prostate

cancer.11

Cell death mechanisms of HDACIs include induc-

tion of the cyclin dependent kinase inhibitors p21

and p27, apoptotic Bcl-2 family proteins, death re-

ceptors, death ligands and retinoic acid receptor.8,12

HDACI also acetylates many nonhistone proteins,

including tubulin, heat shock proteins and Ku70,

disrupting protein function and resulting in cell

death.13–15 Although it was reported that SAHA,

valproic acid, MS-275, PXD101 and FR235222 de-

creased prostate specific antigen, and increasedcaspase activation, and p21 and annexin A1 expres-

sion in a prostate cancer cell line,11,16–19 the precise

mechanism of HDACIs must be further elucidated.

The merit of HDACIs is that cytotoxic effects are

specific to cancer cells and not to normal cells or

tissues. HDACIs are well tolerated with a good tox-

icity profile compared to that of other anticancer

agents.7 Many clinical trials have been done or are

under way using HDACIs combined with other che-

motherapy, for example the taxane paclitaxel or

docetaxel combined with the HDACI pivaloyloxy-

methyl butyrate, LBH589, SAHA or PXD101 forsolid tumors, including prostate, breast and lung

cancer.9,20–22 However, the exact mechanism of ac-

tion is not fully understood.

We determined whether HDACIs would enhance

the effects of docetaxel in advanced prostate cancer

cells. We also assessed the mechanism of cell death.

Thus, we evaluated the growth inhibitory effects of

HDACI alone in hormone dependent and indepen-

dent prostate cancer cells. Finally, we determined

the antitumor effect of docetaxel and HDACI in

HRPC DU145 cells in vitro and in vivo, and ana-

lyzed the mechanism by which combination treat-

ment induced cell death.

MATERIALS AND METHODS

Cell Culture and Drug TreatmentWe cultured the human prostate cancer cell lines LNCaP,

DU145 and PC3 in RPMI 1640 medium containing 100

U/ml penicillin, 100 g/ml streptomycin and 10% (v/v)

fetal bovine serum (Invitrogen™). The HDACIs SAHA

and PXD101 were synthesized elsewhere. We treated cellswith SAHA, PXD101 or docetaxel in 5% (v/v) RPMI 1640

medium containing fetal bovine serum. For sequential

combination treatment with HDACI and docetaxel cells

were exposed to the former drug for 24 hours and then to

the next drug for another 48 hours.

Cell Survival and CI AnalysisCells were cultured in 96-well plates and treated with

various drug concentrations for the indicated times. We

measured viability with the CellTiter 96® Aqueous One

Solution Cell proliferation assay (MTS assay). MTS re-

agent was added to each well according to manufacturer

instructions. After 2-hour incubation we determined cell viability by measuring absorbance at 490 nm.

We calculated the CI with CalcuSyn (Biosoft®), which

is based on the Chou and Talalay median effect principle.

The isobologram is a graphic representation of the inter-

action between 2 drugs that is formed by plotting the

individual drug doses required to achieve a single agent

effect on the respective x and y-axes. A line connecting the

2 points is drawn and the concentration of the 2 drugs

used in combination to achieve the same effect is plotted

on the isobologram. Combination data points that fall on

the line represent an additive interaction while points

above and below represent antagonism and synergy, re-

spectively. Similar to the isobologram, CI analysis pro- vides qualitative information on the drug interaction. A

numerical CI value is calculated based on the equation,

CI (D)1/(Dx)1 (D)2/(Dx)2 (D)1(D)2/(Dx)1(Dx)2,

where (D)1 and (D)2 are the doses of drugs 1 and 2 with a

certain percent effect when used in combination, and

(Dx)1 and (Dx)2 are the doses of drugs 1 and 2, respec-

tively, with the same certain percent effect when used

alone. CI greater than 1 indicates antagonism, 1 indicates

an additive effect and less than 1 indicates synergy.

Cell Cycle AnalysisCells were treated with drugs, fixed with 70% (v/v) ethanol

and stained with 60 g/ml propidium iodide (Sigma) con-

taining 10 U/ml ribonuclease A for 30 minutes. We mea-sured the percent of 10,000 cells in the different cell cycle

phases using the FACSCalibur™ flow cytometer built-in

ModFit LT™ 3.0 software.

Western BlotEqual amounts of protein were electrophoresed in sodium

dodecyl sulfate-polyacrylamide gel and transferred to Mil-

lipore polyvinylidene difluoride membranes (Chemicon®).

After blocking with 3% (weight per volume) nonfat dry

milk the membranes were incubated with various primary

antibodies (1:1,000) in blocking solution overnight at 4C.

Appropriate secondary antibodies (1:5,000) conjugated to

horseradish peroxidase (Pierce, Rockford, Illinois) were

incubated for 1 hour at room temperature. Immobilon™

Western ECL solution and Image Station 4000MM

HISTONE DEACETYLASE INHIBITOR POTENTIATES DOCETAXEL IN PROSTATE CANCER2558



(Kodak, Rochester, New York) were used to visualize im-

munoreactive bands. We used antibodies to acetylated

histone 3, histone 3, caspase-3, 8 and 9, Bid, Bik, tubulin,

p21 (Cell Signaling Technology®), actin, acetylated tubu-

lin (Sigma), Bim, Mcl-1 and Bcl-xl (Santa Cruz Biotech-

nology, Santa Cruz, California).

Xenograft Animal ModelFour-week-old male BALB/C nude mice (OrientBio, Seoul,

Korea) were subcutaneously inoculated with 5 106

DU145 cells. Mice bearing tumors with a volume of about

100 mm3 intraperitoneally received docetaxel (10 or 5

mg/kg for 1 day per week at 9:00 a.m.) and PXD101 (30

mg/kg for 5 days per week at 6:00 p.m.) for 3 weeks. Tumor

volume was measured twice weekly and calculated using

the formula, tumor volume in mm3 1/2 (1 x 2

2), where

1 and 2 represent the larger and smaller tumor diame-

ters, respectively.

Statistical Analysis

All data are shown as the mean

SD. Statistical signifi-cance was considered at p0.05 and determined by 1-way

ANOVA.

RESULTS

HDAC Inhibitors

Prostate cancer cell growth inhibition. Incubating

hormone dependent LNCaP prostate cancer cells,

and DU145 and PC3 HRPC cells with 0.04, 0.2, 1, 5,

25 or 100 M SAHA or PXD101 for 48 hours de-creased cell viability in a dose dependent manner

(fig. 1, A). PXD101 was more potent than SAHA in

all preparations. DU145 cells were as sensitive as

LNCaP and more sensitive than PC3 cells (fig. 1, A).

The half maximum inhibitory concentration of

SAHA and PXD101 was 2.80 and 0.75 M in LNCaP

cells, 2.50 and 0.70 M in DU145 cells, and 6.60 and

1.20 M in PC3 cells, respectively. These results

indicate that hormone dependent and independent

prostate cancer cells are sensitive to HDACIs.

To assess the effects of these 2 HDACIs on intra-

cellular HDAC activity we analyzed histone 3 and

tubulin acetylation by Western blot in the 3 cell lines

Figure 1. Cytotoxicity and activity of HDACIs SAHA and PXD101 in LNCaP, DU145 and PC3 cell lines. A, percent viability of 4

preparations of each cell type exposed to 0.04, 0.2, 1, 5, 25 and 100 M SAHA or PXD101 for 48 hours. B , histone 3 acetylation (Ac-H3 )

was increased by HDACIs in cells exposed to 1 or 10 M SAHA or PXD101. h, hours. C , HDACIs increased acetylated tubulin (Ac-tubulin)in cells on 15% gel. Conc , concentration.

HISTONE DEACETYLASE INHIBITOR POTENTIATES DOCETAXEL IN PROSTATE CANCER 2559



exposed to 1 or 10 M SAHA or PXD101 for the

indicated times (fig. 1, B and C). SAHA and PXD101increased histone 3 and tubulin acetylation in a time

and dose dependent manner. The histone 3 or tubu-lin level was not changed by HDACIs.

Effect on cell cycle and HRPC cell apoptotic signaling. To assess the effects of these HDACIs on thecell cycle we analyzed DNA contents by flow cytom-

etry. We observed an increase in the hypodiploidpopulation (sub-G1) of LNCaP, DU145 and PC3 cells

treated with 10 M SAHA or PXD101 for 48 hours(fig. 2, A and B). Exposure of these cells to 5 M

SAHA or PXD101 for 72 hours resulted in activa-tion of caspase-8, an enzyme with a role in the

extrinsic apoptotic pathway (fig. 2, C), and theactivation of caspase-9 and 3, which mediate in-

trinsic apoptosis.

Induction of DU145 cell apoptosis with docetaxel. Since HDACI had an antiproliferative effect

on HRPC cells, we tested the effect of HDACIs ondocetaxel toxicity in DU145 cells, which are sensi-

tive to HDACIs. Although treatment with 1.5 nMdocetaxel, 1.5 M SAHA or 0.5 M PXD101 alone

resulted in modest toxicity (mean SD 69.3%

3.5%, 65.6% 7.7% and 73% 2.7% viability,

respectively), pretreatment with 1.5 nM docetaxelfollowed by 1.5 M SAHA or 0.5 M PXD101 in-

creased toxicity significantly (35.8% 8.0% and35.0% 3.6% viability, respectively, fig. 3, A).

However, administering HDACIs followed by do-cetaxel (HDACIs ¡docetaxel) had no effect on cell

death while simultaneous treatment with HDA-CIs and docetaxel was less effective than docetaxel

followed by HDACIs (fig. 3, B and C).To establish whether the combined effects of do-

cetaxel, and the HDACIs SAHA and PXD101 weresynergistic we exposed DU145 cells to the drugs

while keeping a constant ratio of each drug to theother. Growth inhibition was then measured by

MTS assay (fig. 3, D and G). Cell growth was mark-

edly inhibited by docetaxel and HDACI applied in

combination compared with that of each drug alone. Analysis of the dose effect relationship and isobolo-

grams of these results revealed that sequentialtreatment with a high concentration of docetaxel

and SAHA (docetaxel ¡SAHA) was synergistic (fig.

3, E and F ). The docetaxel and PXD101 combination(docetaxel¡PXD101) similarly showed synergy ex-cept for the highest concentration of the combination

on isobologram (fig. 3, H and I ).To investigate apoptosis induced by combination

treatment we performed flow cytometry of DU145cells sequentially exposed to docetaxel and HDACIs.

Histograms showed that combination treatment in-creased the sub-G1 population compared with cells

exposed to a single agent, that is docetaxel, SAHA or

PXD101 (fig. 3, J and K ).

Combination TreatmentApoptosis inducing mechanisms. Since pan-HDACIsand docetaxel increase in tubulin acetylation, we

examined the accumulation of tubulin acetylation in

DU145 cells using combination treatment (fig. 4, A). Although 1.5 nM docetaxel, 1.5 M SAHA or 0.5 M

PXD101 slightly increased tubulin acetylation, com-bined docetaxel/HDACIs increased it strongly. On

the other hand, adding docetaxel had no effect on thelevel of acetylation on histone 3 due to HDACIs.

Since combination treatment also significantly in-creased activated caspase-8, 9 and 3, we analyzed

the level of proteins that regulate cell survival anddeath (fig. 4, B and C). Combination treatment

decreased levels of the anti-apoptotic Bcl-2 familyproteins (Mcl-1 and Bcl-xl) and increased levels of

pro-apoptotic Bcl-2 family proteins (t-Bid, Bik andBim). This indicates that combined docetaxel and

HDACI treatment (docetaxel¡HDACIs) increasedapoptosis by modulating Bcl-2 family protein ex-

pression. The cell cycle inhibitor p21 protein wasaccumulated by combination treatment, as re-

ported previously.8

Figure 2. Apoptosis induction by SAHA or PXD101 in LNCaP, DU145 and PC3 cells. A, flow cytometry profiles show that HDACI

increased sub-G1 apoptotic population of cells treated with 10 M SAHA or PXD101 for 48 hours. B , sub-G1 population of 3 cell lines

in 3 preparations each. PXD , PXD101. Asterisk indicates p 0.05 vs control. C , Western blots of caspase-8, 9 and 3 in cells treated with5 M SAHA or PXD101 for 72 hours on 15% gel. CTL, control.




Figure 3. Cytotoxicity of combined HDACI and docetaxel treatment in DU145 HRPC cells. A, 6 preparations each of cells were incubated

with 1.5 nM docetaxel (DTX ) for 24 hours and then with or without 1.5 M SAHA, or 0.5 M PXD101 for 48 hours (DTX ¡HDACI ). HDACI

posttreatment significantly potentiated docetaxel induced cell death. B , 6 preparations each of cells were incubated with 1.5 M SAHA

or 0.5 M PXD101 for 24 hours and then with or without 1.5 nM docetaxel for 48 hours (HDACI ¡ DTX ). C , 6 preparations each of cells

were exposed to HDACI, that is 1.5 M SAHA or 0.5 M PXD101, and 1.5 nM docetaxel simultaneously for 48 hours. D , viability curves

in 3 cell preparations each of single agent docetaxel and SAHA, and sequential combinations in cells treated with docetaxel and SAHA

at 1:1,000 ratio. E , CI plot of combined docetaxel and SAHA (D ). F , isobologram shows combined docetaxel and SAHA (D ). G , viability

curves of single agent docetaxel and PXD101, and sequential combinations in 3 preparations each of cells treated with docetaxel and

PXD101 at 1:333 ratio. H , CI plot of the combined docetaxel and PXD101 (G ). I , isobologram shows combined docetaxel and PXD101

(G ). J , flow cytometry profiles reveal that sub-G1 apoptotic population was greater after sequential treatment with 1.5 nM docetaxel

followed by 1.5 M SAHA or 0.5 M PXD101. CTL, control. K , bars represent sub-G1 population of sequentially (DTX ¡ HDACI ) treated

cells in 3 preparations each.




Effect on DU145 xenografts. To confirm the effectsof docetaxel and HDACI on HRPC cells in vivo we

subcutaneously injected nude mice with 5 106

DU145 cells, maintained them until the tumor size

reached 100 mm3 and then injected them intraperi-toneally with docetaxel (10 mg/kg per week) and/or

PXD101 (30 mg/kg per day) for 3 weeks. Combined

treatment reduced tumor volume compared withthat in control mice or mice treated with docetaxel orPXD101 alone but combined treatment had no effect

on body weight (fig. 5, A and B).To determine whether PXD101 decreased the re-

quired dose of docetaxel we treated tumor bearing

mice with 5 mg/kg per week docetaxel and/or 30mg/kg per day PXD101 for 3 weeks and then calcu-

lated tumor size (fig. 5, C). Injecting 5 mg/kg perweek docetaxel plus 30 mg/kg per day PXD101 re-

sulted in a mean tumor volume of 64% 0.2% thatin saline injected controls, similar to the 64% 0.3%

tumor volume in mice treated with 10 mg/kg perweek docetaxel. These results suggest that when

combined, PXD101 may allow a dose reduction of docetaxel, thus decreasing its side effects.

DISCUSSION

The finding that HDAC activity is increased in tu-

mors, including prostate cancer, led to the develop-ment of HDACIs as antitumor agents.11,23 ManyHDACIs are in phase I/II clinical trials of treatment

for hematopoietic cancer and solid tumors6 but todate only SAHA has been approved by the Food and

Drug Administration in the United States as treat-ment for cutaneous T-cell lymphoma only. We eval-

Figure 4. Western blot shows effect of sequential combination treatment with docetaxel followed by HDACI on histone acetylation,

caspase activation and Bcl-2 family protein expression in DU145 cells. A, acetylated histone 3 (Ac-H3 ) and acetylated tubulin(Ac-Tubulin) from cells incubated with 1.5 nM docetaxel for 24 hour and then next with or without 1.5 M SAHA or 0.5 M PXD101 for

48 hours on 15% gel. B , for caspase-8, 9 and 3 from cells treated with sequential combinations on 15% gel. C , Bcl-2 family proteins from

cells treated with sequential combination on 13% gel. Minuses indicate negative. Plus signs indicate positive.

Figure 5. Growth inhibitory effect of combined HDACI and docetaxel (DTX ) in DU145 xenografts in mice. A, tumor volume changes

after 3-week treatment with 10 mg/kg per week docetaxel and/or 30 mg/kg per day PXD101 ( PXD ) for 5 days per week. Asterisk indicates

p 0.05 vs control, PXD101 alone and docetaxel alone. B , total body weight in 3 treatment groups (A). C , tumor volume percent

reduction after treatment with 5 or 10 mg/kg per week docetaxel combined with 30 mg/kg per day PXD101 for 3 weeks. Minusesindicatenegative. Plus signs indicate positive. Asterisk indicates p 0.05.




uated the antitumor effects of HDACIs on hormone

dependent (LNCaP) and hormone independent

(DU145 and PC3) prostate cancer cells. The pan-

HDACIs SAHA and PXD101 inhibited the growth of

hormone dependent and hormone independent cells

in a dose dependent manner (fig. 1, A). Cell cycle

analysis and Western blotting for caspase revealedthat HDACIs induced prostate cancer cell apoptosis

(fig. 2). These results suggest that pan-HDACIs may

be used for hormone dependent and independent

prostate cancer.

Since HDACIs are thought to act by potentiating

the antitumor effects of chemotherapeutic agents

and/or radiation,20,24–26 and they showed cytotoxic-

ity as monotreatment in HRPC cells in our study, we

tested whether SAHA and PXD101 could potentiate

the activity of docetaxel, an agent widely used for

HRPC. Using DU145 cells, the cell line most sensi-

tive to HDACIs of the 3 lines tested, we found thatsequential treatment with docetaxel followed by

HDACI (docetaxel ¡SAHA or PXD101) resulted in

the greatest growth inhibition (fig. 3, A). Although

HDACI cytotoxicity was lower in PC3 than in

DU145 cells, docetaxel¡HDACI combination treat-

ment had a potent antiproliferative effect in PC3

cells (data not shown). Also, median effect analysis

and isobolograms indicated that the docetaxel and

HDACI combination had synergy (fig. 3, D to I ).

These results suggest that sequential treatment

with docetaxel followed by SAHA or PXD101 may be

a useful therapy for HRPC. However, further studyis needed to elucidate the influence of the combina-

tion schedule on the response.

Docetaxel stabilizes polymerized tubulin and in-

creased tubulin acetylation is associated with such

stabilization.27 Also, HDACIs enhance tubulin acet-

ylation by inhibiting HDAC 6, an enzyme that acety-

lates cytosolic nonhistone proteins.7 In accordance

with these reports we found that the docetaxel and

HDACI combination increased the acetylated tubu-

lin level compared with that using either reagent

alone (fig. 4, A). To elucidate other mechanisms of

cell death we analyzed caspase activation and Bcl-2

family protein levels. Western blotting for caspaseshowed that combination treatment activated ex-

trinsic and intrinsic apoptosis pathways (fig. 4, B).Moreover, combination treatment increased the pro-

apoptotic proteins t-Bid, Bik and Bim, and de-

creased the anti-apoptotic proteins Mcl-1 and Bcl-xl(fig. 4, C). Depsipeptide (FK228) potentiated do-cetaxel toxicity in HRPC cells, as shown by p21

expression assays.28,29 We have extended these find-ings by noting that SAHA or PXD101 potentiated

docetaxel toxicity, and using tubulin and Bcl-2 fam-ily proteins as biomarkers in prostate cancer cell

lines.Since all of our in vitro experiments indicated

that combination treatment with docetaxel and

HDACIs has an antitumor effect on HRPC cells byinducing apoptosis, we tested the effects of combin-

ing docetaxel and PXD101 on tumor growth in nudemice injected with DU145 cells. Because it was re-

ported that combining SAH A plus docetaxel ispoorly tolerated in HRPC cases30 and PXD101 is not

a substrate of the multidrug resistance gene, wechose the combination of PXD101 plus docetaxel.

Our results show that this combination was alsoeffective in vivo against HRPC cells (fig. 5, A).

CONCLUSIONS

Results indicate that HDACI can be used to treat

patients with hormone independent and hormonedependent prostate cancer. Moreover, a sequential

combination of docetaxel and HDACIs resulted in asynergistic increase in the antiproliferative effects of

either drug given alone in DU145 HRPC cellsin vitro and in vivo. These preclinical findings sup-

port the clinical evaluation of HDACIs combinedwith docetaxel for HRPC.

ACKNOWLEDGMENTS

SAHA and PXD101 were synthesized at Crystal

Genomics, Seoul, Korea.

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histone deacetylase inhibitor potentiates anticancer effect of docetaxel via modulation of bcl-2...

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