Design, synthesis, and pharmacological characterization of 4-[4,4-Dimethyl-3-(4-hydroxybutyl)-5-oxo-2-thioxo-1-imidazolidinyl]-2-iodoben zonitrile as a high-affinity nonsteroidal androgen receptor ligand

Article abstract of DOI:10.1021/jm000163y

4-[4,4-Dimethyl-3-(4-hydroxybutyl)-5-oxo-2-thioxo-1-imidazolidinyl]-2-trifluo romethylbenzonitrile (RU 59063) is a prototype of a new class of high-affinity nonsteroidal androgen receptor (AR) ligands. The search for a radioiodinated AR ligand prompted us

Full text of DOI:10.1021/jm000163y

3344
J . Med. Chem. 2000, 43, 3344-3347
Brief Articles
Design , Syn th esis, a n d P h a r m a cologica l Ch a r a cter iza tion of
4-[4,4-Dim eth yl-3-(4-h yd r oxybu tyl)-5-oxo-2-th ioxo-1-im id a zolid in yl]-
2-iod oben zon itr ile a s a High -Affin ity Non ster oid a l An d r ogen Recep tor Liga n d #
Marcian E. Van Dort,*,† Diane M. Robins,‡ and Bess Wayburn‡
Division of Nuclear Medicine, Department of Internal Medicine and Department of Human Genetics,
The University of Michigan Medical School, Ann Arbor, Michigan 48109-0552
Received April 10, 2000
4-[4,4-Dimethyl-3-(4-hydroxybutyl)-5-oxo-2-thioxo-1-imidazolidinyl]-2-trifluoromethylbenzo-
nitrile (RU 59063) is a prototype of a new class of high-affinity nonsteroidal androgen receptor
(AR) ligands. The search for a radioiodinated AR ligand prompted us to synthesize 4-[4,4-
dimethyl-3-(4-hydroxybutyl)-5-oxo-2-thioxo-1-imidazolidinyl]-2-iodobenzonitrile (DTIB) wherein
the trifluoromethyl group of RU 59063 was substituted with the similarly hydrophobic iodine
atom. DTIB displayed subnanomolar binding affinity (Ki ) 0.71 ( 0.22 nM) for the rat AR in
competitive binding assays. Additionally, DTIB demonstrated potent agonist activity, compa-
rable to that of the natural androgen 5R-dihydrotestosterone (DHT), in a cell-based functional
assay (cotransfection assay). DTIB represents a new lead for the development of high-affinity
radioiodinated AR radioligands.
Ch a r t 1. Structures of AR Ligands
In tr od u ction
The androgen receptor (AR) is an important member
of the superfamily of nuclear hormone receptors that
function as ligand-dependent regulators of transcrip-
tion.1 ARs respond to signaling by the endogenous
steroidal androgens testosterone and 5R-dihydrotest-
osterone (DHT) and play a critical role in sexual
development and function in males.2
The development and progression of prostate cancer
is known to be androgen-dependent, and AR expression
is frequently observed in primary prostate tumors and
metastases.3 Consequently, AR-targeted radioligands
are under investigation for the noninvasive imaging of
tumor sites in prostate cancer using positron emission
tomography (PET) and single-photon emission com-
puted tomography (SPECT).4 The majority of these
studies to date have focused on steroid-based ligands,
including the naturally occurring steroid ligands (tes-
tosterone, DHT) and synthetic steroids [mibolerone,
metribolone (R 1881)].5 Successful PET imaging of
prostate and prostate tumor metastases has been
recently reported in a preliminary clinical study with a
fluorine-18-labeled derivative of DHT.6 In contrast, the
development of high-affinity radioiodinated steroid-
based AR ligands for SPECT imaging has had limited
success. This is attributed, in part, to the sensitivity of
the AR ligand binding domain to the bulky nature of
iodinated steroids.7
Nonsteroidal antiandrogens offer an alternative ap-
proach to the design of high-affinity radioiodinated AR
ligands. Since nonsteroidal AR ligands have greater
conformational flexibility and are more amenable to
structural modification than steroid-based ligands, they
could serve as templates for the design of iodinated
ligands that are better accommodated at the AR ligand
binding domain. RU 59063 (Chart 1) is a prototype
member of a novel class of nonsteroidal antiandrogens
which displays high AR affinity (Ka ) 5.4 nM for human
AR) and selectivity (>1000-fold selectivity for AR over
progestin, glucocorticoid, mineralocorticoid, and estro-
gen receptors).8 In addition, [3H]RU 59063 demon-
strated high specific binding and 3-8-fold greater AR
binding affinity as compared to [3H]testosterone in
radioligand binding assays conducted with rat, mouse,
hamster, and human AR tissue preparations.8
#
Presented in part at the 218th National Meeting of the American
Chemical Society, New Orleans, LA, Aug 22-26, 1999.
* To whom correspondence should be addressed. Mailing address:
Division of Nuclear Medicine, 3480 Kresge III Building, University of
Michigan Medical School, Ann Arbor, MI 48109-0552. Tel: (734) 763-
4647. Fax: (734) 936-1571. E-mail: mvandort@umich.edu.
† Division of Nuclear Medicine, Department of Internal Medicine.
‡ Department of Human Genetics.
The high AR selectivity, reduced lipophilicity, and
ease of synthesis of RU 59063 and its derivatives as
compared to steroid analogues make these compounds
attractive candidates for investigation as radioiodinated
AR radioligands. As a first step toward this goal, we
10.1021/jm000163y CCC: $19.00 © 2000 American Chemical Society
Published on Web 08/05/2000

Brief Articles
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 17 3345
Sch em e 1a
Ta ble 1. Inhibition Constants for DTIB (1) and Selected
Ligands at the Rat ARa
compd
Ki ( SEM (nM)b
DTIB (1)
RU 59063
DHT
testosterone
mibolerone
0.71 ( 0.22
2.23 ( 0.50
0.69 ( 0.20
1.4 ( 0.4
0.75 ( 0.08
a
Binding studies were conducted using rat prostate cytosol AR
and [3H]mibolerone as radioligand. Data are presented as mean
( SEM and are the average of at least three determinations each
conducted in duplicate.
b
a
Reagents and conditions: (a) 1. NaNO2, H2SO4, 0 °C, 2. NaI;
(b) 1. SOCl2, 60 °C, 2. NH4OH; (c) SOCl2, reflux; (d) SnCl2, EtOH,
reflux; (e) CSCl2, THF:H2O, rt; (f) 2-(4-hydroxybutyl)amino-2-
cyanopropane, Et3N, THF, reflux; (g) 2 N HCl, CH3OH, reflux.
selected the iodinated derivative 4-[4,4-dimethyl-3-(4-
hydroxybutyl)-5-oxo-2-thioxo-1-imidazolidinyl]-2-iodo-
benzonitrile (DTIB) (Chart 1) for initial evaluation based
on the similar hydrophobic properties of the trifluorom-
ethyl (π ) 0.88) and iodine (π ) 1.12) groups.9 We report
here the synthesis of DTIB and its initial in vitro
pharmacological characterization (AR binding, AR-
mediated transcriptional activation) as a novel high-
affinity, nonsteroidal AR ligand.
F igu r e 1. Comparison of AR agonist dose response of RU
59063, DTIB, and DHT in cotransfected CV-1 cells. Data points
represent the mean ( SEM of triplicate determinations;
chloramphenicol acetyltransferase (CAT) activity is reported
as % conversion/µg of protein; RU 59063 was tested at two
dose levels: 10-7 and 10-6 M.
Ch em istr y
DTIB (1) was prepared by a seven-step synthetic route
as outlined in Scheme 1. Commercially available 2-amino-
4-nitrobenzoic acid was converted to 2-iodo-4-nitroben-
zoic (2) in moderate yield (42%) via the Sandmeyer
diazotization reaction. Sequential reaction of 2 with
thionyl chloride and ammonia provided 2-iodo-4-ni-
trobenzamide (3). Dehydration of 3 was accomplished
with thionyl chloride to give the nitrile 4, which was
subsequently reduced to the aniline derivative 5 with
stannous chloride in EtOH. Synthesis of the isothio-
cyanate derivative 6 and subsequent steps were con-
ducted according to the general procedure of Teutsch
et al.8 Thus, treatment of 5 with thiophosgene gave the
isothiocyanate 6 which was condensed with 2-(4-hy-
droxybutylamino)-2-cyanopropane in the presence of
triethylamine to afford the imine intermediate 7. Acid
hydrolysis of 7 provided DTIB (1) as a white amorphous
solid in 16% overall yield. DTIB and intermediate
compounds were purified by silica gel flash chromatog-
and the binding affinity values obtained for other
routinely used AR ligands such as DHT, mibolerone,
and testosterone are presented in Table 1. As seen from
these data, DTIB (Ki ) 0.71 ( 0.22 nM) is one of the
highest-affinity AR ligands reported to date. DTIB
inhibited [3H]mibolerone binding to the rat AR as
efficiently as mibolerone and DHT, two tritiated AR
ligands currently in use. Furthermore, DTIB showed a
3-fold improvement in AR affinity over RU 59063
indicating that iodine substitution was well-tolerated
in this region of the molecule. Hill coefficients (data not
shown) were close to unity in all cases indicating that
the binding interaction was to a single site.
The AR functional activity of DTIB, RU 59063, and
DHT was evaluated using an in vitro cell-based assay
(cotransfection assay) system.11 DTIB and RU 59063
displayed a dose-dependent stimulation of AR-mediated
transcriptional activity in these assays with a potency
comparable to that of the natural hormone agonist DHT
(Figure 1). Similar agonistic effects (data not shown)
were observed for these ligands using a more complex
natural gene fragment (C′∆9, a transcriptional enhancer
of the mouse sex-limited protein gene12). The agonistic
behavior of RU 59063 seen in transfection experiments
is in contrast to the potent in vivo AR antagonist activity
reported for this analogue in rodent models.8 A similar
discrepancy between in vitro and in vivo functional
1
raphy and fully characterized by H NMR, mass spec-
tral, and elemental analysis.
Resu lts a n d Discu ssion
The binding affinity of DTIB and RU 59063 to the
rat prostate cytosolic AR was determined using a
competitive binding assay in the presence of the high-
affinity AR radioligand, [3H]mibolerone.10 These data

3346 J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 17
Brief Articles
2-Iod o-4-n itr oben za m id e (3). A mixture of 2 (6.60 g, 22.5
mmol) and SOCl2 (100 mL) was heated at 60 °C for 3 h under
argon and concentrated by rotoevaporation. Residual SOCl2
was removed from the crude product mixture by coevaporation
with dry CHCl3 (3 × 50 mL). Concentrated aqueous NH3 (100
mL) was then added and the mixture stirred overnight at
ambient temperature. The precipitate was filtered, rinsed with
H2O and dried in an oven at 60 °C. The crude product was
purified by flash chromatography (75% EtOAc in hexane) to
give 5.6 g (85%) of 3 as cream-colored crystals: mp 208-209
°C (EtOAc); 1H NMR (CD3OD) δ 8.70 (d, 1H, J ) 2.2, H-3),
8.29 (dd, 1H, J ) 8.5, 2.2, H-5), 7.59 (d, 1H, J ) 8.4, H-6).
Anal. (C7H5N2O3I) C, H, N.
activity has been observed for the structurally similar
antiandrogen RU 56187.13
Although our initial design strategy of DTIB was
based on the similar hydrophobicity of iodine and
trifluoromethyl, it must be noted that these functional
groups do not share similar electronic and steric proper-
ties. In particular, iodine is less electronegative (σm, σp
values are 0.35 and 0.18, respectively, for iodine versus
0.43 and 0.54, respectively, for CF3) and has a 3-fold
larger steric size than trifluoromethyl (molar refractivity
values for iodine and CF3 are 13.9 and 5.0, respec-
tively).9 Our binding affinity data therefore indicate that
the AR binding domain that interacts with the trifluo-
romethyl substituent in RU 59063 is sufficiently large
to accommodate the increased steric bulk of an iodine
atom. Taken together, these results suggest that steric
and hydrophobic interactions may be more important
than electronic interactions at this binding region for
high AR binding in this class of nonsteroidal ligands.
2-Iod o-4-n itr oben zon itr ile (4). A mixture of 3 (5.5 g, 18.8
mmol) and SOCl2 (35 mL) was refluxed for 3 h under argon
and concentrated under reduced pressure. The residue was
purified by flash chromatography (20% EtOAc in hexane) to
give 4.12 g (80%) of 4 as cream-colored crystals: mp 154-155.5
1
°C [EtOAc:hexane (1:4)]; H NMR (CDCl3) δ 8.76 (d, 1H, J )
2.2, H-3), 8.32 (dd, 1H, J ) 8.5, 2.2, H-5), 7.82 (d, 1H, J ) 8.6,
H-6). Anal. (C7H3N2O2I) C, H, N.
4-Cya n o-3-iod oa n ilin e (5). A mixture of 4 (2.3 g, 8.4 mmol)
and SnCl2‚2H2O (9.3 g, 41.4 mmol) in EtOH (35 mL) was
refluxed for 2 h. The mixture was concentrated under reduced
pressure and treated with H2O (100 mL) and the pH was
adjusted to 9 by treatment with 5% aqueous NaHCO3. The
mixture was extracted with EtOAc; the organic layers were
washed with saturated brine and H2O and dried. The residue
obtained after removal of solvent was purified by flash
chromatography (40% EtOAc in hexane) to afford 2 g (98%) of
5 as white fluffy crystals: mp 146-148 °C [EtOH:H2O (1:5)];
1H NMR (CDCl3) δ 7.33 (d, 1H, J ) 8.5, H-5), 7.14 (d, 1H, J )
2.3, H-2), 6.62 (dd, 1H, J ) 8.5, 2.2, H-6), 4.19 (br s, 2H,
exchangeable with D2O, NH2). Anal. (C7H5N2I) C, H, N.
2-Iod o-4-isoth iocya n a toben zon itr ile (6). A solution of 5
(0.54 g, 2.2 mmol) in THF (10 mL) was added dropwise at
ambient temperature to a well-stirred suspension of thiophos-
gene (0.30 g, 2.6 mmol) in H2O (5 mL). TLC analysis [silica;
hexane:EtOAc (6:1)] at 3 h indicated completeness of reaction.
The mixture was diluted with H2O (20 mL), extracted with
CHCl3 (2 × 20 mL) and dried. The residue obtained after
removal of solvent was dried under high vacuum to afford a
quantitative yield of a yellow-brown solid which was used
directly in the next step. A small portion of the crude product
was purified by flash chromatography (15% EtOAc in hexane)
to give an analytical sample of 6 as a white solid: mp 116-
118 °C; 1H NMR (CDCl3) δ 7.76 (d, 1H, J ) 2.1, H-3), 7.58 (d,
1H, J ) 8.3, H-6), 7.26 (dd, 1H, J ) 8.3, 2.1, H-5); HRMS (EI)
calcd for C8H3N2IS (M+) 285.9062, found 285.9058.
Con clu sion s
In summary, the present study has identified a new
iodinated nonsteroidal AR ligand, DTIB, which demon-
strates subnanomolar binding affinity and potent ago-
nist activity in in vitro AR binding and functional
assays, respectively. These studies also suggest the
existence of a large hydrophobic pocket in the AR
binding domain that interacts with the meta substituent
of this pharmacophore. Studies are currently underway
to evaluate the utility of radioiodinated analogues of
DTIB as radioligands for in vitro and in vivo studies of
AR.
Exp er im en ta l Section
Gen er a l Meth od s. Melting points were determined with
a Thomas-Hoover melting point apparatus and are uncor-
rected. 1H NMR spectra were obtained in either CDCl3 or CD3-
OD with a Bruker WM-360 (360 MHz) instrument using
tetramethylsilane (TMS) as internal standard. Chemical shifts
(δ) are reported in parts per million (ppm) relative to TMS,
and coupling constants (J ) are reported in hertz (Hz). Elemen-
tal analyses were performed by the Department of Chemistry,
University of Michigan, and were within (0.4% of the calcu-
lated values. RU 59063 and 2-(4-hydroxybutylamino)-2-cyano-
propane were synthesized as previously described.8 All other
chemical reagents were obtained from Aldrich Chemical Co.,
Milwaukee, WI, and were used without further purification.
Organic extracts were dried over anhydrous Na2SO4 and
concentrated to dryness by rotoevaporation under reduced
pressure.
4-[4,4-Dim eth yl-3-(4-h yd r oxybu tyl)-5-im in o-2-th ioxo-1-
im id a zolid in yl]-2-iod oben zon itr ile (7). A solution of 2-(4-
hydroxybutylamino)-2-cyanopropane8 (0.30 g, 1.94 mmol) in
anhydrous THF (2 mL) was added dropwise at ambient
temperature to a stirred solution of the crude isothiocyanate
6 (0.55 g, 1.94 mmol) and Et3N (25 mg, 0.25 mmol) in
anhydrous THF (2 mL). The reaction mixture was refluxed
for 1 h, at which point, TLC analysis [silica; hexanes:EtOAC
(6:1)] indicated completeness of reaction. The reaction mixture
was concentrated under reduced pressure and the residue
purified by flash chromatography (35% acetone in chloroform)
to provide 0.55 g (64%) of the title compound 7 as a straw-
colored viscous liquid: 1H NMR (CDCl3) δ 7.93 (s, 1H, Ar-H),
7.75 (d, 1H, J ) 8.3, Ar-H), 7.49 (d, 1H, J ) 8.2, Ar-H), 3.75-
3.66 (m, 4H, -NCH2- and -CH2O-), 1.92 (m, 2H, CH2), 1.66
(m, 2H, CH2), 1.58 (s, 6H, C(CH3)2); HRMS (EI) calcd for
2-Iod o-4-n itr oben zoic Acid (2). A vigorously stirred slurry
of 2-amino-4-nitrobenzoic acid (10 g, 55 mmol) and aqueous 9
N H2SO4 (64 mL) was diazotized at 0 °C (ice-salt bath) by
dropwise addition of a solution of NaNO2 (4.17 g, 60.4 mmol)
in water (50 mL). The mixture was stirred at 0 °C for 1 h and
treated with urea (1.1 g, 18.3 mmol) to destroy excess HNO2.
A solution of NaI (9.9 g, 66 mmol) in H2O (50 mL) was then
added dropwise at 0-5 °C. The reaction was allowed to warm
to ambient temperature, stirred a further 2 h, diluted with
1% aqueous NaHSO3 solution (200 mL) and filtered. The crude
product was rinsed with hot EtOAc (3 × 100 mL) to remove a
dark orange side product and the residue was purified by flash
chromatography [gradient elution with EtOAc:hexane:glacial
acetic acid (30:70:1 to 80:20:1)] to give 6.8 g (42%) of 2 as light
C
16H19N4ISO (M+) 442.0324, found 442.0320.
4-[4,4-Dim et h yl-3-(4-h yd r oxyb u t yl)-5-oxo-2-t h ioxo-1-
im id a zolid in yl]-2-iod oben zon itr ile (DTIB, 1). A solution
of the imine derivative 7 (0.48 g, 1.1 mmol) in CH3OH (8 mL)
was treated with aqueous 2 N HCl (1 mL) and refluxed for 1
h. The cooled reaction mixture was poured into H2O (80 mL)
and extracted with EtOAc (2 × 50 mL), and the organic layers
dried and concentrated under reduced pressure. The crude
product was purified by flash chromatography (70% EtOAc in
1
yellow needles: mp 143.5-145 °C [benzene:hexane (1:1)]; H
NMR (CD3OD) δ 8.77 (d, 1H, J ) 2.2, H-3), 8.30 (dd, 1H, J )
8.5, 2.2, H-5), 7.92 (d, 1H, J ) 8.5, H-6). Anal. (C7H4NO4I) C,
H, N.

Brief Articles
J ournal of Medicinal Chemistry, 2000, Vol. 43, No. 17 3347
(3) Hobisch, A.; Culig, Z.; Radmayr, C.; Bartsch, G.; Klocker, H.;
Hittmair, A. Androgen receptor status of lymph node metastases
from prostate cancer. Prostate 1996, 28, 129-135.
hexane) to give 0.44 g (90%) of DTIB (1) as a white amorphous
solid: H NMR (CDCl3) δ 7.96 (d, 1H, J ) 1.9, Ar-H3), 7.71
(d, 1H, J ) 8.3, Ar-H6), 7.52 (dd, 1H, J ) 8.3, 1.9, Ar-H5),
3.73 (m, 4H, -NCH2- and -CH2O-), 1.94 (m, 2H, CH2), 1.66
(m, 2H, CH2), 1.57 (s, 6H, C(CH3)2). Anal. (C16H18N3O2IS) C,
H, N.
1
(4) (a) Katzenellenbogen, J . A. Designing steroid receptor-based
radiotracers to image breast and prostate tumors. J . Nucl. Med.
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testosterone: a radiolabeled ligand for the androgen receptor.
J . Nucl. Med. 1997, 38, 402-409. (c) Ali, H.; Rousseau, J .; van
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An d r ogen Recep tor Bin d in g Assa ys. Androgen receptor
binding assays were conducted by a commercial laboratory
(MDS Panlabs, Bothell, WA) using [3H]mibolerone as radio-
ligand and rat prostate cytosol as a source of AR as previously
described.10 In brief, cytosol from the ventral prostate of
castrated male Wistar rats (175 ( 25 g) was prepared in
modified phosphate buffer (pH 7.2) containing protease inhibi-
tors and triamcinolone acetonide (5 mM). Competitive binding
of test ligands were determined by incubation of increasing
concentrations (0.3-100 nM) of each ligand and a saturating
concentration of [3H]mibolerone (2 nM) with cytosol (0.4 mg)
at 4 °C for 18 h. Separation of bound and free radioligand was
achieved by further incubation with a hydroxylapatite slurry
for 15 min and filtration. Radioactivity bound to the washed
filters was quantified by liquid scintillation counting. Binding
affinity data are reported as mean ( standard error of the
mean (SEM) and represent the average of at least three
separate experiments each conducted in duplicate.
Cotr a n sfection Assa ys. AR-mediated transcriptional ac-
tivation by ligands was studied in a cell-based bioassay system
(cotransfection assays) as previously reported.11 In brief,
mammalian CV-1 cells were transfected with the mouse AR
and an androgen-responsive reporter gene, consisting of three
tandem androgen response elements upstream of the thymi-
dine kinase promoter and the bacterial chloramphenicol
acetyltransferase (CAT) gene. Media containing the reference
compound (DHT) or test compound in concentrations ranging
from 1 nM to 1 µM were added to the cells. Three independent
transfection assays were performed. Following incubation, cells
were washed with phosphate buffered saline and lysed by
several cycles of freeze-thaw. Cell extracts were assayed for
CAT activity and the data reported as CAT conversion rates
(%).11
(9) Hansch, C.; Leo, A. The Hydrophobic Parameter: Measurement
and Calculation. In Exploring QSAR: Fundamentals and Ap-
plications in Chemistry and Biology; Heller, S. R., Ed.; American
Chemical Society: Washington, DC, 1995; Chapter 1, p 7,
Chapter 3, p 73, Chapter 4, pp 108-109.
(10) (a) Schilling, K.; Liao, S. The use of radioactive 7R,17R-dimethyl-
19-nortestosterone (mibolerone) in the assay of androgen recep-
tors. Prostate 1984, 5, 581-588. (b) Liao, S.; Witte, D.; Schilling,
K.; Chang, C. The use of a hydroxylapatite-filter steroid receptor
assay method in the study of the modulation of androgen
receptor interaction. J . Steroid Biochem. 1984, 20, 11-17.
(11) (a) Adler, A. J .; Scheller, A.; Hoffman, Y.; Robins, D. M. Multiple
components of a complex androgen-dependent enhancer. Mol.
Endocrinol. 1991, 5, 1587-1596. (b) Gorman, C. M.; Moffat, L.
F.; Howard, B. H. Recombinant genomes which express chloram-
phenicol acetyltransferase in mammalian cells. Mol. Cell. Biol.
1982, 2, 1044-1051. (c) Seed, B.; Sheen, J -Y. A simple phase-
assay for chloramphenicol acyltransferase activity. Gene 1988,
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(12) Robins, D. M.; Scheller A.; Adler, A. J . Specific steroid response
from a nonspecific DNA element. J . Steroid Biochem. Mol. Biol.
1994, 49, 251-255.
(13) Kemppainen, J . A.; Langley, E.; Wong, C.; Bobseine K.; Kelce,
W. R.; Wilson, E. M. Distinguishing androgen receptor agonists
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progesterone acetate and dihydrotestosterone. Mol. Endocrinol.
1999, 13, 440-454.
Ack n ow led gm en t . We thank the staff of the
Phoenix Memorial Laboratories for use of their facilities.
This research was supported by grants from the Na-
tional Institutes of Health (CA 77287 to M.V.D. and DK
56356 to D.M.R.) and the SPORE in Prostate Cancer
(P50 CA 69568 to M.V.D.).
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