Synthesis of the catechols of natural and synthetic estrogens by using 2-iodoxybenzoic acid (IBX) as the oxidizing agent

Article abstract of DOI:10.1016/j.steroids.2004.11.005

A method for the synthesis of 2-hydroxyestrone/estradiol, 4-hydroxyestrone/estradiol, 3′-hydroxydiethylstilbestrol, 3′-hydroxyhexestrol, and 3′-hydroxydienestrol is reported, in which 2-iodoxybenzoic acid (IBX) and the corresponding phenolic estrogen are reacted. Treatment of the natural estrogens, estrone/estradiol, with stoichiometric amounts of IBX in dimethylformamide initially yielded a mixture of estrone/estradiol-2,3- and -3,4-quinones, which were reduced in situ to the corresponding catechols by treatment with a 1 M aqueous solution of ascorbic acid. Chromatographic separation of the reaction products afforded 2- and 4-hydroxyestrone/estradiol in good overall yields (79%). In the case of the synthetic estrogens containing two identical phenolic rings, protection of one ring is a prerequisite for the synthesis of the monocatechol. Thus, diethylstilbestrol and dienestrol were protected at one phenol ring as their methyl ethers. The resulting monophenols were treated with stoichiometric amounts of IBX for 1 h, followed by treatment with 1 M aqueous ascorbic acid to obtain the corresponding catechols in more than 70% yield. Furthermore, the catechol of diethylstilbestrol, protected at one ring, was reduced by catalytic hydrogenation at the C3-C4 double bond to obtain 3′-hydroxyhexestrol in 90% yield. Removal of the protected methoxy groups of the synthetic estrogen catechols was carried out by treatment with a 1 M solution of boron tribromide in dichloromethane. This method is highly efficient for the preparative scale synthesis of catechols of both natural and synthetic estrogens.

Full text of DOI:10.1016/j.steroids.2004.11.005

Steroids 70 (2005) 173–178
Synthesis of the catechols of natural and synthetic estrogens
by using 2-iodoxybenzoic acid (IBX) as the oxidizing agent
Muhammad Saeed, Muhammad Zahid, Eleanor Rogan, Ercole Cavalieri∗
Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center,
986805 Nebraska Medical Center, Omaha, NE 68198-6805, USA
Received 7 June 2004; received in revised form 20 October 2004; accepted 2 November 2004
Abstract
A method for the synthesis of 2-hydroxyestrone/estradiol, 4-hydroxyestrone/estradiol, 3ꢀ-hydroxydiethylstilbestrol, 3ꢀ-hydroxyhexestrol,
and 3ꢀ-hydroxydienestrol is reported, in which 2-iodoxybenzoic acid (IBX) and the corresponding phenolic estrogen are reacted. Treat-
ment of the natural estrogens, estrone/estradiol, with stoichiometric amounts of IBX in dimethylformamide initially yielded a mixture of
estrone/estradiol-2,3- and -3,4-quinones, which were reduced in situ to the corresponding catechols by treatment with a 1 M aqueous solution
of ascorbic acid. Chromatographic separation of the reaction products afforded 2- and 4-hydroxyestrone/estradiol in good overall yields (79%).
In the case of the synthetic estrogens containing two identical phenolic rings, protection of one ring is a prerequisite for the synthesis of the
monocatechol. Thus, diethylstilbestrol and dienestrol were protected at one phenol ring as their methyl ethers. The resulting monophenols
were treated with stoichiometric amounts of IBX for 1 h, followed by treatment with 1 M aqueous ascorbic acid to obtain the corresponding
catechols in more than 70% yield. Furthermore, the catechol of diethylstilbestrol, protected at one ring, was reduced by catalytic hydrogenation
at the C3–C4 double bond to obtain 3ꢀ-hydroxyhexestrol in 90% yield. Removal of the protected methoxy groups of the synthetic estrogen
catechols was carried out by treatment with a 1 M solution of boron tribromide in dichloromethane. This method is highly efficient for the
preparative scale synthesis of catechols of both natural and synthetic estrogens.
© 2005 Elsevier Inc. All rights reserved.
Keywords: Estradiol; Estrone; Diethylstilbestrol; Hexestrol; Dienestrol; Catechol synthesis; 2-Iodoxybenzoic acid (IBX)
1. Introduction
synthetic estrogens, diethylstilbestrol (DES) and hexestrol
(HES), have been found to be associated with cancer in an-
imals and humans [4–10]. There is mounting experimental
evidence suggesting that tumor initiation arises from specific
metabolites of endogenous estrogens [11–13].
Both endogenous and synthetic estrogens are mainly me-
tabolized to their respective catechols [14–19]. Catechol
estrogens are metabolically oxidized to their electrophilic
quinones, which react with DNA to form specific depuri-
nating adducts that can lead to mutations and, eventually, to
the initiation of cancer [11–13]. To study the mechanism of
tumor initiation by endogenous and synthetic estrogens, it
was necessary to synthesize the catechol metabolites of these
estrogens on a preparative scale.
Several of the most common cancers in western societies
occur in hormonally responsive tissues, including breast, en-
dometrium, and ovary in women and prostate in men. These
cancershavebeencausallylinkedtoexposuretosyntheticand
endogenous steroidal hormones or their metabolites [1–3].
Endogenous estrogens, estradiol (E2) and estrone (E1), and
Abbreviations: CDCl3, deuterated chloroform; DES, diethylstilbestrol;
DIES, dienestrol; DMF, N,N-dimethylformamide; DMSO-d6, deuterated
dimethylsulphoxide; E1, estrone; E2, estradiol; ESI-MS, electron spray
ionization-mass spectrometry; HES, hexestrol; HMPA, hexamethylphos-
phoramide; HPLC, high performance liquid chromatography; IBX, 2-
iodoxybenzoic acid; NMR, nuclear magnetic resonance; TBDMS, tert-
butyldimethylsilyl; THF, tetrahydrofuran
The standard chemical methods for preparing 2- and
4-hydroxyestrogens are laborious multistep chemical re-
actions [20,21], giving low overall yields and practically
∗
Corresponding author. Tel.: +1 402 559 7237; fax: +1 402 559 8068.
E-mail address: ecavalie@unmc.edu (E. Cavalieri).
0039-128X/$ – see front matter © 2005 Elsevier Inc. All rights reserved.
doi:10.1016/j.steroids.2004.11.005

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M. Saeed et al. / Steroids 70 (2005) 173–178
no selectivity for the synthesis of 4-hydroxyestrogens.
Yoshizawa et al. [22] reported the chemical synthesis of
2-hydroxyestrogens using benzoyl peroxide; however, the
synthesis of 4-hydroxyestrogens was not achievable by this
method. A multistep method reported by Stubenrauch and
Knuppen [23] was not successful for the direct conversion of
estradiol to the 2- and 4-hydroxyestradiols, and the authors
needed to reduce the corresponding 17-oxo compound with
sodium borohydride for the preparation of estradiol catechols
[23]. Use of potassium nitrosodisulfonate (FREMY’s salt)
for the one-step chemical conversion of estrone/estradiol to a
mixture of the corresponding 2,3- and 3,4-quinones had been
reported earlier by Gelbke et al. [24]. The method involved
a laborious work-up and ended up with very low overall
yields of catechol estrogens. Regarding the catechols of syn-
thetic estrogens, DES and HES, our laboratory previously re-
ported the chemical synthesis of 3ꢀ-hydroxydiethylstilbestrol
(3ꢀ-OH-DES) and 3ꢀ-hydroxyhexestrol (3ꢀ-OH-HES) [25];
however, overall yields were low due to multi-step re-
actions, and the formation of a mixture of isomers was
observed.
We report here a new method for the synthesis of the cat-
echols of both natural and synthetic estrogens in good to
excellent yields. After oxidation of E1 (1a) or E2 (1b) with
2-iodoxybenzoic acid (IBX) to the 2,3- and 3,4-quinones,
subsequentreductionwitheitherKIora1 Maqueoussolution
of ascorbic acid gives the corresponding catechol estrogens
in good overall yield. This method was also used for the syn-
thesis of the catechols of synthetic estrogens, demonstrating
the general applicability of this method.
the phenol to a polar product. The red-colored solution was
treated with a 1 M solution of ascorbic acid in water until the
color of the reaction mixture turned light yellow. The reaction
product was extracted with ethyl acetate (3× 50 mL), and the
combined organic phases were evaporated after drying over
anhydrous sodium sulfate (Na2SO4). The residue was chro-
matographed over a column of silica gel impregnated with
ascorbic acid as described by Gelbke and Knuppen [28]. The
product was then purified by preparative HPLC using 10%
acetonitrile and 90% H2O (with 0.4% acetic acid) as a mo-
bile phase for the initial 5 min and then a linear gradient to
100% acetonitrile in 25 min, to afford the catechol. The spec-
troscopic data of 2- and 4-OHE1/E2, 3ꢀ-OH-DES and 3ꢀ-OH-
HES synthesized by this method agree with those reported
earlier [24,25].
2.2.1. 3ꢀ-OH-4ꢀꢀ-OCH3-DES (8)
Compound 8 was synthesized from 4ꢀ-OCH3-DES (7) ac-
cording to the general method described above. Yield 68%;
1H NMR (ppm, DMSO-d6): δ 8.80 (s, 1H, Ar-OH, ex-
changeable with D2O), 8.75 (s, 1H, Ar-OH, exchangeable
with D2O), 7.08 (d, J = 8.3 Hz, 2H, H-2ꢀꢀ, H-6ꢀꢀ), 6.92 (d,
J = 8.8 Hz, 2H, H-3ꢀꢀ, H-5ꢀꢀ), 6.70 (d, J = 7.8 Hz, 1H, H-5ꢀ),
6.56 (d, J = 1.9 Hz, 1H, H-2ꢀ), 6.42 (dd, J = 7.8, 1.9 Hz, 1H,
H-6ꢀ), 3.75 (s, 3H, OCH3), 2.09 (q, J = 7.8 Hz, CH2), 2.01 (q,
J = 7.8 Hz, CH2), 0.70 (t, J = 7.3 Hz, 2 × CH3).
2.2.2. 3ꢀ-OH-4ꢀꢀ-OCH3-DIES (14)
Compound 14 was synthesized from 4ꢀ-OCH3-DIES (13)
according to the general method described above. Yield 75%;
1H NMR (ppm, CDCl3): δ 8.79 (bs, 2H, Ar-OH, exchange-
able with D2O), 7.01 (d, J = 8.3 Hz, 2H, H-2ꢀꢀ, H-6ꢀꢀ), 6.91 (d,
J = 8.3 Hz, 2H, H-3ꢀꢀ, H-5ꢀꢀ), 6.70 (d, J = 7.8 Hz, 1H, H-5ꢀ),
6.50 (d, J = 1.9 Hz, 1H, H-2ꢀ), 6.36 (dd, J = 7.8, 1.9 Hz, 1H,
H-6ꢀ), 5.29 (q, J = 6.8 Hz, CH), 5.15 (q, J = 6.8 Hz, CH), 3.75
(s, 3H, OCH3), 1.43 (dd, J = 6.8, 6.8 Hz, 2 × CH3).
2. Experimental
2.1. Chemicals and instrumentation
DES, E2, E1, DIES and ascorbic acid were purchased
from Sigma (St. Louis, MO) and used as such without fur-
ther purification. 2-Iodobenzoic acid, oxone and dimethylfor-
mamide (DMF) were purchased from Aldrich Chemical Co.
(Milwaukee, WI). IBX was synthesized from 2-iodobenzoic
acid and oxone as previously described [26]. Preparative
HPLC was performed on a Waters (Milford, MA) 600E sol-
vent delivery system equipped with a 996 photodiode ar-
ray detector and a Phenomenex (Torrance, CA) Luna (2) C-
2.3. General method for the synthesis of
monomethylated synthetic estrogens
Sodium hydride (NaH) (358 mg, 60% suspension) was
suspended in DMF (25 mL) at 0 ◦C in a 50-mL three-neck
round-bottomed flask and mechanically stirred under argon
for 5 min. DES or DIES (2 g, 7.46 mmol) was added slowly,
and the ice bath was removed. A thick slurry of the salt of
DES/DIES was formed after 1 h, which was thinned with
a small amount of DMF to facilitate stirring. A solution of
iodomethane (CH3I, 566 ␮L) in DMF (20 mL) was added
slowly into the mixture from a dropping funnel. After the
formation of a clear solution, thin-layer chromatography of
the reaction mixture was conducted, which indicated almost
complete consumption of the starting material. The reaction
mixture was plunged into crushed ice containing 5% HCl,
and the reaction product was extracted with ethyl acetate
(3× 50 mL). The combined organic layers were washed with
brine and water, dried over sodium sulfate (Na2SO4), and the
1
˚
18 column (10 ␮m, 100 A, 21.2 mm × 250 mm). H NMR
spectra were recorded in DMSO-d6 or CDCl3 on a Varian-
Inova 500 instrument at 499.835 MHz at 25 ◦C. Chemical
shifts are reported relative to DMSO-d6 (2.49 ppm) or CDCl3
(7.26 ppm).
2.2. General procedure for the synthesis of catechols
IBX (1 mmol) was added to a stirred solution of phe-
nol (1 mmol) in DMF (50 mL) in the dark. After 1 h, thin-
layer chromatography indicated the complete conversion of

M. Saeed et al. / Steroids 70 (2005) 173–178
175
Scheme 1. Synthesis of 2-OHE1/E2 (4a and 4b) and 4-OHE1/E2 (5a and 5b).
solvent was removed under low pressure. The residue was
purified by flash column chromatography with 5% ethyl ac-
etate in hexane to yield monomethylated estrogens as a white
solid.
ent starting from 30% methanol in 70% water (0.4% TFA) to
100% methanol in 30 min at a flow rate of 7 mL/min.
2.4.1. dl-3ꢀ-OH-4ꢀꢀ-OCH3-HES (10a)
Yield (38.6 mg, 72 %). 1H NMR (ppm, DMSO-d6): δ 8.48
(bs, 2H, Ar-OH, exchangeable with D2O), 6.82 (d, J = 8.8 Hz,
2H, H-2ꢀꢀ, H-6ꢀꢀ), 6.69 (d, J = 8.8 Hz, 2H, H-3ꢀꢀ, H-5ꢀꢀ), 6.47
(d, J = 7.8 Hz, 1H, H-5ꢀ), 6.32 (d, J = 1.5 Hz, 1H, H-2ꢀ), 6.17
(dd, J = 7.8, 1.5 Hz, 1H, H-6ꢀ), 3.66 (s, 3H, OCH3), 2.59–2.47
(m, 2H, 2CH), 1.76 (m, 2H, CH2), 1.48-1.36 (m, 2H, CH2),
0.64 (t, J = 7.3 Hz, 3H, CH3), 0.62 (t, J = 7.3 Hz, 3H, CH3).
2.3.1. 4ꢀꢀ-OCH3-DES (7)
1
Yield (1.47 g, 70%). H NMR (ppm, DMSO-d6): δ 9.30
(s, 1H, Ar-OH, exchangeable with D2O), 7.09 (d, J − 8.3 Hz,
2H, H-1ꢀ,H-6ꢀ), 6.97 (d, J = 8.3 Hz, 2H, H-1ꢀꢀ, H-6ꢀꢀ), 6.93
(d, J = 8.3 Hz, 2 H, H-3ꢀ, H-5ꢀ), 6.75 (d, J = 8.3 Hz, 2 H, H-
3ꢀꢀ, H-5ꢀꢀ), 3.75 (s, 3 H, OCH3), 2.06 (m, 4 H, 2 × CH2),
0.69 (t, J = 7.32 Hz, 6 H, 2 × CH3). 13C NMR 157.9, 156.0,
138.5, 137.9, 134.4, 132.5, 129.6, 129.5, 115.1, 113.7, 55.1,
28.2, 13.5, 13.4. Electronsprayionization-massspectrometry
(ESI-MS) m/z: 283.1 (M + H)+.
2.4.2. Meso-3ꢀ-OH-4ꢀꢀ-OCH3-HES (10b)
Yield (9.7 mg, 18%). 1H NMR (ppm, DMSO-d6): δ 8.63
(bs, 2H, Ar-OH, exchangeable with D2O), 7.10 (d, J = 8.8 Hz,
2H, H-2ꢀꢀ, H-6ꢀꢀ), 6.87 (d, J = 8.8 Hz, 2H, H-3ꢀꢀ, H-5ꢀꢀ), 6.66
(d, J = 7.8 Hz, 1H, H-5ꢀ), 6.58 (d, J = 1.5 Hz, 1H, H-2ꢀ), 6.46
(dd, J = 7.8, 1.5 Hz, 1H, H-6ꢀ), 3.73 (s, 3H, OCH3), 2.42-
2.30 (m, 4H, 2CH, CH2), 1.32-1.08 (m, 2H, CH2), 0.45 (t,
J = 7.3 Hz, 3H, CH3), 0.44 (t, J = 7.3 Hz, 3H, CH3).
2.3.2. 4ꢀꢀ-OCH3-D1ES (13)
1
Yield (1.50 g, 71%). H NMR (ppm, CDCl3): δ 7.10 (d,
J = 8.3 Hz, 2H, H-1ꢀ,H-6ꢀ), 7.05 (d, J = 8.3 Hz, 2H, H-1ꢀꢀ, H-
6ꢀꢀ), 6.93 (d, J = 8.3 Hz, 2 H, H-3ꢀ, H-5ꢀ), 6.86 (d, J = 8.3 Hz,
2 H, H-3ꢀꢀ, H-5ꢀꢀ), 5.30 (m, 2 H, CH), 3.85 (s, 3 H, OCH3),
1.48 (d, J = 6.35 Hz, 6 H, 2 × CH3). 13C NMR 158.0, 154.0,
144.9, 144.8, 132.3, 132.2, 131.1, 130.9,125.1, 114.9, 113.4,
55.2, 15.2. ESI-MS m/z: 281.1 (M + H)+.
2.5. General procedure for deprotection of methyl
groups
Boron tribromide (BBr3, 3 mL) was added over 15 min
to a stirred solution of methylated catechol (1 mmol) in
dichloromethane (50 mL) at −78 ◦C under argon. The re-
action mixture was stirred at the same temperature for 1 h
and then at room temperature for 2 h. The reaction mixture
was plunged into a beaker containing crushed ice and sodium
bicarbonate (NaHCO3, 50 mg). The reaction products were
extracted with ethyl acetate (3× 100 mL). The combined or-
ganic layers were washed with brine and water, dried over
sodium sulfate (Na2SO4), and the solvent was removed un-
der low pressure. The residue was purified by preparative
HPLC as described above.
2.4. Synthesis of 3ꢀ-OH-4ꢀꢀ-OCH3-HES (10)
A solution of 8 (50 mg, 0.17 mmol), 5% palladium on car-
bon (Pd/C, 3 mg) and ethyl acetate (3 mL) was stirred for
10 h under a hydrogen atmosphere (using a balloon filled
with H2 gas). The Pd/C was removed by filtration thor-
ough a pad of Celite. The filtrate was dried to afford 10
as a diastereomeric mixture of dl- (10a) and meso-3ꢀ-OH-
4-OCH3-HES (10b) in 8:2 ratio, which were separated by
preparative HPLC. The column [Phenomenex, Luna C-18(2),
250 mm × 21.2 mm i.d.] was eluted by using a linear gradi-

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M. Saeed et al. / Steroids 70 (2005) 173–178
Table 1
Outcome of the reaction of E1 with IBX in different solvents
Solvent
Time (h)
Yield (%)
4a:5a
Recovered 1a (%)
39
33
0
5.1
0
0
Acetone
Acetonitrile
DMF
Dioxane
DMF:HMPA
THF
10
10
3
3
3
50
55
79
65
40
76
2:1
1:1
1:1.2
1:1.1
1:1
3
1.2:1
3. Results and discussion
the recovery of starting material were observed, even after the
reaction mixture was stirred for a longer time. The low stabil-
ity of the quinones rendered the reaction mixtures complex,
thus making a longer reaction time practically nonfeasible.
Use of dioxane as the solvent (Table 1) gave a 65% reaction
yield as a 1:1 mixture of both catechols; however, some of
the starting E2 was also observed after 3 h. No starting E2
was observed when DMF, DMF:hexamethylphosphoramide
(HMPA) or tetrahydrofuran (THF) was used as solvent, and
the overall reaction yields were found to be 79, 40, and 76%,
respectively. Although the yield of reaction varied with differ-
ent solvents, there was no practical selectivity in the synthesis
of 4-OHE1/E2 over 2-OHE1/E2, and we always obtained a
nearly 1:1 mixture of both catechols.
In light of a recently reported method for the preparation of
o-quinones by the oxidation of the corresponding monophe-
nolic compounds with IBX [27], the synthesis of catechol
estrogens was carried out according to Scheme 1. Treatment
of E1 (1a) or E2 (1b) with a stoichiometric amount of IBX in
DMF at room temperature resulted in the formation of a mix-
ture of the corresponding o-quinones (2a/b and 3a/b). The
quinones were easily reduced to the catechols 2-OHE1/E2
(4a/b) and 4-OHE1/E2 (5a/b) by treating with a 1 M solution
of ascorbic acid or KI/acetic acid [24]. The separation of 2-
and 4-OHE1/E2, thus formed, was achieved by column chro-
matography on silica gel impregnated with ascorbic acid as
described by Gelbke and Knuppen [28].
Synthesis of 3ꢀ-OH-DES (9) and 3ꢀ-OH-DIES (15) was
initiated from the commercially available DES (6) and DIES
(12), respectively (Schemes 2 and 4). Since both DES (6)
and DIES (12) contain two phenolic rings, one of them must
be protected with some easily removable protective group.
Different solvents were tested to increase the overall yield
of the reaction and to observe any selectivity in the formation
of4-OHE1/E2 over2-OHE1/E2. Useofacetoneoracetonitrile
as solvent was not successful, and low reaction yields with
Scheme 2. Synthesis of trans- (9a) and cis-3ꢀ-OH-DES (9b). (a) NaH (1.2 eq), DMF, CH3I (1.1 eq); (b) IBX, DMF, rt, in the dark for 1 h and then add 1 M
ascorbic acid; (c) BBr3, CH2Cl2, −78 ◦C.

M. Saeed et al. / Steroids 70 (2005) 173–178
177
Scheme 3. Synthesis of dl-3ꢀ-OH-HES (11a) and its meso-isomer (11b). (a) 5% Pd/C, H2, ethyl acetate, rt; (b) BBr3, CH2Cl2, −78 ◦C.
Scheme 4. Synthesis of 3ꢀ-OH-DIES (15). (a) NaH (1.2 eq), DMF, CH3I (1.1 eq); (b) IBX, DMF, rt, in the dark for 1 h and then add 1 M ascorbic acid; (c)
BBr3, CH2Cl2, −78 ◦C.
We used both tert-butyldimethylsilyl (TBDMS) and methyl-
ether as protective groups; however, we preferred methyl-
ether groups over TBDMS ethers, because of the highly
volatile nature of the latter. After careful monomethylation of
DES (6) and DIES (12) with iodomethane (CH3I) and sodium
hydride (NaH) in DMF, methylated phenols 7 and 13 were
achieved in 70 and 71% yields, respectively. Methylether-
phenol 7 or 13 was then converted to the catechol 8 or
14, respectively, by IBX in DMF, followed by treatment
with ascorbic acid. Removal of the protective group in 8 or
14 was achieved by treating each compound with BBr3 in
dichloromethane to produce the target catechols 3ꢀ-OH-DES
(9) or 3ꢀ-OH-DIES (15), respectively, in almost quantitative
yields (Schemes 2 and 4).
Reduction of the C3-C4 double bond in 8 with 5% palla-
dium on carbon (Pd/C) under a hydrogen atmosphere pro-
duced the HES derivative, 10. We have observed that 3ꢀ-
OH-4ꢀꢀ-OCH3-DES (8) was present as an equilibrium mix-
ture of trans- (8a) and cis-isomers (8b) in an 8:2 ratio, re-
spectively [25], if stored at room temperature for a longer
time. Therefore, it may be desirable to separate both iso-
mers before reduction, which can be achieved by using
preparative HPLC. Alternatively, reduction of the mixture as
such yielded both the dl-3ꢀ-OH-4ꢀꢀ-OCH3-HES (10a) and its
meso-isomer (10b) in 8:2 ratio, respectively, as determined
by HPLC and NMR analyses (Scheme 3). Separation of the
diastereomers was achieved conveniently by preparative re-
verse phase HPLC. Deprotection of the methyl group by re-
acting 10a or 10b with BBr3 in CH2Cl2 yielded the corre-
sponding 3ꢀ-OH-HES (11a or 11b) in good yield from 6.
In conclusion, we report a one-step method for the syn-
thesis of catechols of both natural and synthetic estrogens,
starting from the corresponding monophenolic compounds.
Acknowledgements
This research was supported by U.S. Public Health Ser-
vice grant P01 CA-49210 from the National Cancer Institute.
Core support to the Eppley Institute is provided by grant P30
CA36727 from the National Cancer Institute.
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