Aromatase inhibitors

This invention provides for a method of inhibiting aromatase and treating or preventing estrogen-dependent diseases in mammals by administering certain pyrimidine derivatives. Pharmaceutical formulations of the pyrimidine derivatives are also provided.

BACKGROUND OF THE INVENTION 
Estrogens are synthesized from androgenic steroids. In the biosynthetic 
pathway for estrogen formation, aromatization is an essential step. It is 
generally believed that if the aromatase enzyme could be effectively 
inhibited, a useful treatment for estrogen dependent disorders could be 
obtained (see Cancer Research, Vol. 42, Suppl. 8:3261s (1982)). 
Several estrogen dependent diseases exist which could be treated with 
aromatase inhibitors. These include breast cancer, endometriosis, 
polycystic ovarian disease, benign breast disease, and endometrial cancer. 
A beneficial effect of antiestrogens in the treatment of breast cancer has 
been well established (see Br. J. Cancer, 25, 270 (1971)). Two of the 
known aromatase inhibitors, testolactone and aminoglutethimide, have shown 
a beneficial effect in treatment of breast cancer. See Cancer Research, 
supra. 
Endometriosis is characterized by an abnormal proliferation of the 
endometrium of the uterus. Since the endometrium is dependent on estradiol 
for its growth, an inhibitor of estrogen production should stop the 
progression of the disease. 
Benign breast disease, or often called fibrocystic breast disease, appears 
to be dependent on ovarian steroids. See Cancer, 49, 2534 (1982). 
Aromatase inhibitors have not been tried in this disease, but 
antiestrogens seem to be of benefit. See Obstet. Gynecol., 54, 80 (1979). 
Polycystic ovarian disease is one of the most common causes of infertility 
in women. The disease appears to result from an abnormality in steroid 
metabolism, and the major form of therapy in this disease is the 
antiestrogen, clomiphene. See Clin. Endocrinol., 12, 177 (1980). 
It is the purpose of this invention to provide compounds which inhibit the 
enzyme aromatase in mammals and are therefore useful in the treatment or 
prevention of breast cancer and other estrogendependent diseases. 
SUMMARY OF THE INVENTION 
This invention provides for a method of inhibiting the enzyme aromatase in 
mammals with an effective amount of a compound of the formula I 
##STR1## 
and pharmaceutically acceptable salts thereof, wherein R is 
##STR2## 
2-chloroethyl, C.sub.3 -C.sub.8 cycloalkyl, phenoxy-substituted C.sub.1 
-C.sub.4 alkyl, or norbornan-2-yl, R.sub.1 is methyl, trifluoromethyl, 
methoxy, fluoro, chloro, bromo, or nitro; 
R.sub.2 is hydrogen, methyl, trifluoromethyl, methoxy, fluoro, chloro, 
bromo, or nitro; 
each of R.sub.3 and R.sub.4 is independently hydrogen, methyl, methoxy, 
fluoro, or chloro; and 
X is hydrogen, hydroxy, methyl, or halo, with the proviso that if R.sub.1 
is methoxy, R must be substituted phenyl with at least one of R.sub.3 and 
R.sub.4 being other than hydrogen. 
By virtue of their ability to inhibit the enzyme aromatase, the compounds 
of formula I are useful in the treatment and prevention of 
estrogen-dependent diseases, especially breast cancer, in mammals. 
A further aspect of this invention is a pharmaceutical formulation 
comprising one or more of the compounds of formula I in combination with a 
suitable pharmaceutical carrier, diluent, or excipient therefor. The 
formulations provided by this invention are particularly useful in 
treating mammals suffering from estrogen-dependent diseases such as breast 
cancer.

DETAILED DESCRIPTION AND PREFERRED EMBODIMENT 
The term "C.sub.1 -C.sub.4 alkyl" refers to branched and straight chain 
aliphatic radicals of one to four carbon atoms such as methyl, ethyl, 
propyl, isopropyl, butyl, tert-butyl, and the like. The term "C.sub.3 
-C.sub.8 cycloalkyl" refers to the saturated alicyclic rings of three to 
eight carbon atoms such as cyclopropyl, methylcyclopropyl, cyclobutyl, 
cyclopentyl, cyclohexyl, cyclooctyl, and the like. 
A preferred group of compounds useful in this invention are the compounds 
of formula I wherein: 
(a) R.sub.1 is fluoro or chloro, 
(b) R.sub.2 is hydrogen or chloro, especially at the 3-position, 
(c) X is hydroxy, hydrogen, or fluoro, and 
(d) R is phenyl (R.sub.3 and R.sub.4 are both hydrogen) or substituted 
phenyl, especially where one of R.sub.3 and R.sub.4 is chloro or fluoro, 
especially in the 4'-position. 
Especially preferred compounds are those wherein X is hydrogen or hydroxy, 
and 
(a) R.sub.1 and R.sub.2 are both chloro, especially where R.sub.2 is at the 
3-position, and R is unsubstituted phenyl, or 
(b) R.sub.1 is chloro or fluoro, R.sub.2 is hydrogen, and R is 
mono-substituted phenyl, especially where one of R.sub.3 and R.sub.4 is 
chloro or fluoro, preferably at the 4'-position, and the other of R.sub.3 
and R.sub.4 is hydrogen. 
The most preferred compound is 5-bis(4-chlorophenyl)methylpyrimidine and 
its pharmaceutically acceptable salts. 
A preferred method of treatment according to this invention comprises 
administering a dose effective for inhibiting the enzyme aromatase of one 
of the preferred compounds of this invention. Similarly, a preferred 
formulation according to this invention comprises one of the preferred 
compounds of this invention in combination with a pharmaceutical carrier 
therefor. 
Most of the compounds used in this invention and methods of making the 
compounds are disclosed in U.S. Pat. No. 3,818,009. A preferred and novel 
method of preparing the pyrimidinemethanols in this invention (I, X is 
hydroxy) is taught in U.S. Pat. No. 3,869,456. Both patents are expressly 
incorporated in this application by reference. The compounds as disclosed 
in the patents are described as being useful as fungicides, bactericides, 
herbicides, and plant growth regulators. The methods of using the 
compounds of Formula I as fungicides and as plant growth regulators are 
claimed in U.S. Pat. Nos. 3,887,708 and 3,868,244, respectively. The 
patents do not disclose any utility for use in humans or any utility 
related to the inhibition of aromatase. The compounds used in this 
invention wherein X is methyl are claimed in U.S. Pat. No. 3,818,009 and 
can be prepared from the corresponding compounds where X is hydrogen by 
alkylation with a methyl halide following the general liquid 
ammonia/alkali metal amide procedure as described in U.S. Pat. No. 
2,727,895. 
The compounds of Formula I wherein R is 2-chloroethyl can be prepared from 
the corresponding vinyl compounds taught in the above patents by any of a 
number of methods known in the art such as the addition of hydrogen 
chloride under anti-Markovnikov conditions. Alternatively, the 
2-chloroethyl compounds may be prepared in the same manner as taught in 
the above references utilizing the appropriate .beta.-chloropropiophenone 
derivative. 
As will be recognized by those skilled in the art, except when R is a 
phenyl group substituted identically to the substitution of R.sub.1 and 
R.sub.2, the compounds of Formula I contain an asymmetric carbon atom. 
This invention is not limited to any particular isomer but includes the 
individual enantiomers as well as the racemates of the compounds of 
Formula I. 
The pharmaceutically acceptable acid addition salts of the bases 
represented by Formula I can be prepared employing those acids of 
sufficient acidity to form acid addition salts with the weakly basic 
pyrimidine group. These include both inorganic and organic acids such as 
hydrochloric, hydrobromic, hydriodic, sulfuric, phosphoric, oxalic, 
methanesulfonic, benzenesulfonic, p-toluenesulfonic, maleic, and the like 
acids. Preferred acids for salt formation are the inorganic acids, 
especially hydrochloric acid. 
The compounds may be administered by any number of routes, including the 
oral, subcutaneous, intramuscular, intravenous, transdermal, and rectal 
routes, usually employed in the form of a pharmaceutical composition. Such 
compositions are prepared in a manner well known in the pharmaceutical art 
and comprise at least one active compound. 
Accordingly, the invention includes a pharmaceutical composition comprising 
as active ingredient a compound of formula I associated with a 
pharmaceutically acceptable carrier. In making the compositions of the 
present invention, the active ingredient will usually be mixed with a 
carrier, or diluted by a carrier, or enclosed within a carrier which may 
be in the form of a capsule, sachet, paper or other container. When the 
carrier serves as a diluent, it may be a solid, semi-solid or liquid 
material which acts as a vehicle, excipient or medium for the active 
ingredient. Thus, the composition can be in the form of tablets, pills, 
powders, lozenges, sachets, cachets, elixirs, emulsions, solutions, 
syrups, suspensions, aerosols (as a solid or in a liquid medium), 
ointments containing for example up to 10% by weight of the active 
compound, soft and hard gelatin capsules, suppositories, sterile 
injectable solutions, and sterile packaged powders. 
Some examples of suitable carriers are lactose, dextrose, sucrose, 
sorbitol, mannitol, starches, gum acacia, calcium phosphate, alginates, 
calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, 
cellulose, tragacanth, gelatin, syrup, methyl cellulose, methyl- and 
propylhydroxybenzoates, talc, magnesium stearate, water, or mineral oil. 
The formulations can additionally include lubricating agents, agents, 
emulsifying and suspending agents, preserving agents, sweetening agents or 
flavoring agents. The compositions of the invention may, as is well known 
in the art, be formulated so as to provide quick, sustained, or delayed 
release of the active ingredient after administration to the patient. 
For oral administration, a compound of this invention is admixed with 
carriers and diluents molded into tablets or enclosed in gelatin capsules. 
The mixtures can alternatively be dissolved in liquids such as ten percent 
aqueous glucose solution, isotonic saline, sterile water, or the like, and 
administered intravenously or by injection. Such solutions can, if 
desired, be lyophilized and stored in a sterile ampoule ready for 
reconstitution by the addition of sterile water for ready intramuscular 
injection. 
Preferably the compositions are formulated in a unit dosage form, each 
dosage containing from about 1 to 500 mg., more usually about 5 to 300 
mg., of the active ingredient. The term "unit dosage form" refers to 
physically discrete units suitable as unitary dosages for human subjects 
and other mammals, each unit containing a predetermined quantity of active 
material calculated to produce the desired therapeutic effect, in 
association with the required pharmaceutical carrier. 
The active compounds are effective over a wide dosage range. For example, 
dosages per day will normally fall within the range of about 0.05 to 300 
mg./kg. In the treatment of adult humans, the range of about 0.1 to 50 
mg./kg., in single or divided doses, is preferred. However, it will be 
understood that the amount of the compound actually administered will be 
determined by a physician, in the light of the relevant circumstances 
including the condition to be treated, the choice of compound to be 
administered, the age, weight, and response of the individual patient, the 
severity of the patient's symptoms, and the chosen route of 
administration, and therefore the above dosage ranges are not intended to 
limit the scope of the invention in any way. 
The compounds used in this invention are useful in preventing or 
therapeutically treating estrogen-dependent diseases, including breast 
cancer, in mammals by virtue of their ability to inhibit the enzyme 
aromatase. The ability to inhibit aromatase was demonstrated by employing 
a modification of the isolated rat ovarian microsome method of Brodie et 
al. in J. Steroid Biochem., 7, 787 (1976). In this test system, ovarian 
microsomes are obtained from rats treated with pregnant mares serum 
gonadotropin. Test compounds are added to reaction vials containing 0.1 
.mu.M 4-androstene-3,17-dione, 100,000 dpm 1,2[.sup.3 H]-androstenedione, 
the microsomes and a NADPH generating system. The concentrations of the 
inhibitors tested ranged between 0.005 and 10 .mu.M. In this assay, 
aromatization of androstenedione results in the production of [.sup.3 
H]-H.sub.2 O which is isolated by extracting the samples with chloroform 
and treating the aqueous phase with charcoal to remove the free steroid. 
Samples are counted in a liquid scintillation spectrometer and the percent 
inhibition determined by comparing the results with the samples incubated 
without inhibitor. Potency is determined based on the concentration of 
inhibitor in .mu.M required to produce a 50% inhibition of enzyme activity 
(EC.sub.50) when the concentration of substrate (androstenedione) is 0.1 
.mu.M . The EC.sub.50 's of certain of the compounds of formula I are 
summarized in Table 1. 
TABLE 1 
______________________________________ 
Aromatase Inhibition in the Rat 
Ovarian Microsome Assay 
Compound of Formula I EC.sub.50 * 
______________________________________ 
.alpha.-cyclohexyl-.alpha.-(4-methyl- 
6.4 
phenyl)-5-pyrimidine- 
methanol 
.alpha.-(3-chlorophenyl)-.alpha.- 
0.35 
(4-chlorophenyl)-5- 
pyrimidinemethanol 
.alpha.-(4-trifluoromethyl- 
1.95 
phenyl)-.alpha.-phenyl-5- 
pyrimidinemethanol 
.alpha.-(4-methoxy-3-methyl- 
1.4 
phenyl)-.alpha.-(3,4-dimethyl- 
phenyl)-5-pyrimidine- 
methanol 
.alpha.-(2-chlorophenyl)-.alpha.- 
4.1 
(4-chlorophenyl)-5- 
pyrimidinemethanol 
.alpha.-(2-chloro-4-methoxy- 
4.3 
phenyl)-.alpha.-(4-chloro- 
phenyl)-5-pyrimidine- 
methanol 
.alpha.-(4-chlorophenyl)-.alpha.- 
7.8 
(norbornan-2-yl)-5- 
pyrimidinemethanol 
.alpha.-(4-chlorophenyl)-.alpha.- 
0.18 
(4-fluorophenyl)-5- 
pyrimidinemethanol 
.alpha.-(4-methoxyphenyl)-.alpha.- 
** 
(2-chlorophenyl)-5- 
pyrimidinemethanol 
.alpha.-(2-chloroethyl)-.alpha.- 
1.8 
(4-fluorophenyl)-5- 
pyrimidinemethanol 
.alpha.-phenyl-.alpha.-(3,4-di- 
3.2 
methylphenyl)-5-pyrimidine- 
methanol 
5-[(2-chlorophenyl)fluoro- 
1.2 
(4-fluorophenyl)methyl]- 
pyrimidine 
5-[(2-chlorophenyl)(4- 
0.5 
chlorophenyl)methyl] 
pyrimidine 
.alpha.-(3-chlorophenyl)-.alpha.- 
4.1 
(4-methoxyphenyl)-5- 
pyrimidinemethanol 
.alpha.-cyclopropyl-.alpha.-(4-fluoro- 
4.2 
phenyl)-5-pyrimidine- 
methanol 
.alpha.-(4-chlorophenyl)-.alpha.- 
5.4 
(4-phenoxy-n-butyl)-5- 
pyrimidinemethanol 
.alpha.-(2-chlorophenyl)-.alpha.- 
5.6 
(4-fluoro-2-methoxy- 
phenyl)-5-pyrimidine- 
methanol 
.alpha.-(3,4-dichlorophenyl)- 
1.0 
.alpha.-phenyl-5-pyrimidine- 
methanol 
.alpha.-(4-methoxy-3-methyl- 
8.3 
phenyl)-.alpha.-(3-methylphenyl)- 
5-pyrimidinemethanol 
.alpha.-(4-chlorophenyl)-.alpha.-(3- 
1.65 
fluorophenyl)-5-pyrimidine- 
methanol 
5-[(3,4-dichlorophenyl)- 
1.35 
fluorophenylmethyl]- 
pyrimidine 
.alpha.-(2,4-dichlorophenyl)-.alpha.- 
8.0 
phenyl-5-pyrimidine- 
methanol 
.alpha.-(4-nitrophenyl)-.alpha.-phenyl- 
0.27 
5-pyrimidinemethanol 
.alpha.-cyclohexyl-.alpha. -(2,4-dichloro- 
13.8 
phenyl)-5-pyrimidinemethanol 
.alpha.-(3-chlorophenyl)-.alpha.-(4-fluoro- 
1.2 
phenyl)-5-pyrimidinemethanol 
.alpha.-(4-fluorophenyl)-.alpha.-phenyl- 
2.2 
5-pyrimidinemethanol 
5-bis(4-chlorophenyl)methyl- 
0.055 
pyrimidine 
5-[(3-chlorophenyl)(4-chloro- 
0.68 
phenyl)fluoromethyl]pyrimidine 
.alpha.,.alpha.-bis(4-chlorophenyl)-5- 
0.071 
pyrimidinemethanol 
.alpha.-(2,4-dimethylphenyl)-.alpha.- 
3.5 
phenyl-5-pyrimidinemethanol 
.alpha.,.alpha.-bis(4-methylphenyl)-5- 
0.41 
pyrimidinemethanol 
.alpha.-(4-chloro-2-methoxyphenyl)- 
3.6 
.alpha.-phenyl-5-pyrimidinemethanol 
.alpha.-(4-chlorophenyl)-.alpha.-cyclo- 
4.0 
hexyl-5-pyrimidinemethanol 
.alpha.-(2-chlorophenyl)-.alpha.-(4- 
4.1 
fluorophenyl)-5-pyrimidine- 
methanol 
.alpha.-(4-bromophenyl)-.alpha.-cyclo- 
5.4 
propyl-5-pyrimidinemethanol 
.alpha.-(2,4-difluorophenyl)-.alpha.- 
5.6 
phenyl-5-pyrimidinemethanol 
.alpha.-(4-chlorophenyl)-.alpha.-cyclo- 
5.7 
propyl-5-pyrimidinemethanol 
.alpha.-(4-fluorophenyl)-.alpha.-(3- 
1.25 
methoxyphenyl)-5-pyrimidine- 
methanol 
5-[(4-chlorophenyl)fluoro(4- 
0.195 
fluorophenyl)methyl]pyrimidine 
5-[bis(4-chlorophenyl)fluoro- 
0.078 
methyl]pyrimidine 
5-[1,1-bis(4-chlorophenyl)- 
0.082 
ethyl]pyrimidine 
.alpha.,.alpha.-bis(4-fluorophenyl)-5- 
0.33 
pyrimidinemethanol 
.alpha.-(3-fluorophenyl)-.alpha.-(4- 
1.15 
fluorophenyl)-5-pyrimidinemethanol 
______________________________________ 
*Concentration of compound in .mu.M required to achieve 50% inhibition of 
aromatase activity when substrate concentration is 0.1 .mu.M. 
**17.6% inhibition at 10.0 .mu.M. 
By virtue of their ability to inhibit the enzyme aromatase, the compounds 
of this invention are able to inhibit the synthesis of estrogens in 
mammals, thereby making the compounds useful in the treatment of 
estrogen-dependent diseases, such as breast cancer. This in vivo activity 
was demonstrated in the following test systems. 
Estrogen Synthesis Inhibition in Rats 
Immature female Wistar rats (45-55 grams) were divided into control and 
test groups of 2-8 animals each. Test compounds were administered for 
seven days either daily by gavage in corn oil or as a component of the 
diet. Control animals received either corn oil or diet without the test 
compound. Beginning on the fourth day of the test, all animals treated 
with the test compound and one half of the control animals were given a 
subcutaneous injection of 1.0 mg. of testosterone propionate in corn oil. 
The remaining control animals received only an equivalent volume of corn 
oil. On the seventh day of the test, rats treated with testosterone 
propionate were injected subcutaneously with 100 .mu.Ci of [.sup.3 
H]-testosterone in 50 .mu.1. of saline-ethanol (3:1). 
After two hours, the animals were killed by decapitation. Uteri were 
isolated, trimmed of extraneous connective tissue, and weighed. As 
summarized in Table 2 below, the corn oil treated animals exhibited low 
uterine weight and represent unstimulated or negative controls. In the 
control animals treated with testosterone propionate, estrogens produced 
by aromatization stimulated the uterus resulting in an increase in weight. 
Compounds which inhibit aromatization produced uterine weights 
significantly lower than those of the testosterone treated controls. 
Ovaries from rats treated with [.sup.3 H]-testosterone were excised, 
cleaned of extraneous tissue, and homogenized in 2.5 ml. of a 1.0 mM 
potassium phosphate buffer containing 3.0 mM MgCl.sub.2.6H.sub.2 O, 320 mM 
sucrose, and 0.25% Triton X-100 (polyethylene glycol p-isooctyl phenyl 
ether, Rohm and Haas) at pH 6.5. The ovarian steroids were extracted with 
1.5 ml. of 9:1 toluene/ethanol to which had been added 25 to 100 mcg. each 
of unlabelled estradiol, estriol, and estrone, and approximately 1000 dpm 
of [.sup.14 C]-estradiol. The samples were vortexed, centrifuged at 
500.times.g for 10 minutes, and the organic phase was transferred to a 
conical vial. Two additional extractions were performed on the residue in 
the same way. The pooled organic extracts were evaporated for subsequent 
thin-layer chromatography 
Ovarian proteins were precipitated by the addition of 5.0 ml. of ethanol to 
the remaining aqueous phase. After an overnight incubation at 4.degree. 
C., the samples were centrifuged at 1500.times.g for 10 minutes. The 
supernatant was discarded and the pellet was dissolved in 0.3 N potassium 
hydroxide. Protein was determined according to the method of Bradford, 
Analytical Biochemistry, 72, 248 (1976). 
The organic residue from each above extraction was redissolved in 9:1 
dichloromethane/methanol. The solution of each sample was applied to 
separate silica gel thin layer chromatography plates which contained a 
fluorescent indicator. The plates were developed the first dimension with 
160:38:1.5:0.5 dichloromethane/ethyl acetate/methanol/acetic acid to 
within 3 cm. of the top of the plate. After air-drying, the plate was 
developed in the second dimension with 180:19:1 
dichloromethane/methanol/ammonium hydroxide. The plate was air-dried and 
viewed under 254 nm. UV light. 
The visible spots were marked and the plates were sprayed with primulin 
(0.001% in 4:1 acetone/water) according to the method of Wright, J. 
Chromatography, 59, 220 (1971) which allowed for the identification of 
additional steroids under 365 nm. UV light. The spots were scraped from 
the plate using a glass wool plugged Pasteur pipet attached to a vacuum 
line. The steroids were eluted directly into scintillation vials by the 
addition of 0.2 ml. of dichloromethane followed by two washes each of 2.0 
ml. of methanol. The organic solvent was evaporated and 10.0 ml. of 
scintillation fluid (Beckman Ready Solv-NA) was added to the vials. 
Samples were analyzed by liquid scintillation spectrometry and the 
corrections were made based on the recoveries of the [.sup.14 C]-steroid. 
Steroid concentrations are expressed as femtomoles per milligram protein. 
TABLE 2 
__________________________________________________________________________ 
Effects of Compounds of Formula I on 
estrogen levels and uterine weight 
Test Mean Uterine 
Mean Steroid Concentration** 
No. 
Compound Dose* 
Animals 
Weight (mg.) 
estradiol 
estrone 
estriol 
__________________________________________________________________________ 
I .alpha.,.alpha.-bis(4-chlorophenyl)- 
30 3 130.67 0.31.sup.+ 
0.34 1.24 
5-pyrimidinemethanol 
300 2 75.00.sup.+ 
0.14.sup.+ 
0.70 2.74 
testosterone-treated control 
-- 6 176.67 2.07 0.64 1.98 
Corn oil control 
-- 4 51.75.sup.+ 
-- -- -- 
II 5-bis(4-chlorophenyl)- 
30 4 92.25.sup.+ 
0.14.sup.+ 
0.14 0.59 
methylpyrimidine 
300 5 80.20.sup.+ 
0.15.sup.+ 
0.12 0.29 
Testosterone-treated control 
-- 8 179.13 0.97 0.20 0.36 
Corn oil control 
-- 5 79.80.sup.+ 
-- -- -- 
__________________________________________________________________________ 
*ppm in feed. 300 ppm corresponds to approximately 30 mg./kg./day; 30 ppm 
corresponds to approximately 3 mg./kg./day. 
**femtomoles per milligram of protein. 
.sup.+ significantly different from testosteronetreated control, p &lt; 0.05 
 
DMBA-induced Mammary Tumor Inhibition 
Mammary tumors were produced in female Sprague-Dawley rats which were 50-60 
days old by the gavage administration of 20 mg. of 
7,12-dimethylbenz[a]anthracene (DMBA). About six weeks after DMBA 
administration, the mammary glands were palpated at weekly intervals for 
the appearance of tumors. Whenever one or more tumors appeared and were 
measurable in an animal, that animal was selected for experimentation. An 
attempt was made to uniformly distribute the various sizes of tumors in 
the treated and control groups such that one group did not start with rats 
having tumors which, on the average, were significantly larger than those 
of any other group. Each control and test group contained 8 animals. The 
test compound was administered either mixed into the food at a 
concentration of 300 ppm (corresponding to an appropriate daily dose of 30 
mg./kg.) or the compounds were dissolved or suspended in corn oil and 
administered once daily by gavage. Every experiment included a group of 
control rats having tumors and were either given food without the compound 
admixed or corn oil vehicle by gavage, depending on how the test compound 
was administered. The tumors were measured at the start of the experiments 
and generally had an area of approximately 15-100 mm.sup.2. The area of 
each tumor was calculated by multiplying the shortest and longest 
diameters of the tumor. The treatment and measurement of animals continued 
for 4-8 weeks at which time the final areas of the tumors were determined. 
For each compound (and control) at each dose level, the change in the mean 
tumor area was determined. The mean change was analyzed for its 
significance using Dunnett's t-test. The results of these tests are shown 
in Table 3 below. 
TABLE 3 
__________________________________________________________________________ 
Anti-Tumor Activity 
Duration 
Mean Tumor Area (mm.sup.2) 
Test No. 
Compound Dose* of Test 
Start Finish 
__________________________________________________________________________ 
I Control -- 5 weeks 
72.2 924.3 
.alpha.,.alpha.-bis(4-chlorophenyl)- 
300 
ppm 60.7 10.0.sup.+ 
5-pyrimidinemethanol 
II Control -- 5 weeks 
48.4 1116 
5-bis(4-chlorophenyl)- 
300 
ppm 78.6 154 
methylpyrimidine 
III Control -- 8 weeks 
144.5 1371 
.alpha.,.alpha.-bis(4-chlorophenyl)- 
300 
ppm 160.4 355.sup.+ 
5-pyrimidinemethanol 
IV Control -- 4 weeks 
54.8 913.6 
.alpha.,.alpha.-bis(4-chlorophenyl)- 
5 mg./kg. 56.3 123.5 
5-pyrimidinemethanol 
30 mg./kg. 71.5 34.3 
V Control -- 5 weeks 
59 1381 
5-bis(4-chlorophenyl)- 
1 mg./kg. 66 210.sup.+ 
methylpyrimidine 
7.5 
mg./kg. 63 159.sup.+ 
15 mg./kg. 50 9.sup.+ 
30 mg./kg. 69 146.sup.+ 
__________________________________________________________________________ 
*when dosed in the diet, reported as ppm. 300 ppm corresponds to 
approximately 30 mg./kg./day. Doses reported in mg./kg. given daily by 
gavage. 
.sup.+ statistically different from control, p &lt; 0.05. 
In order to more fully illustrate the operation of this invention, the 
following formulation examples are provided. The examples are illustrative 
only and are not intended to limit the scope of the invention. The 
formulations employ as active compounds any of the pharmaceutical 
compounds of formula I. 
EXAMPLE 1 
Hard gelatin capsules are prepared using the following ingredients: 
______________________________________ 
per capsule 
______________________________________ 
Active compound 250 mg. 
Starch dried 200 mg. 
Magnesium stearate 
10 mg. 
Total 460 mg. 
______________________________________ 
The above ingredients are mixed and filled into hard gelatin capsules in 
460 mg. quantities. 
EXAMPLE 2 
Capsules each containing 20 mg. of medicament are made as follows: 
______________________________________ 
per capsule 
______________________________________ 
Active ingredient 20 mg. 
Starch 89 mg. 
Microcrystalline cellulose 
89 mg. 
Magnesium stearate 
2 mg. 
Total 200 mg. 
______________________________________ 
The active ingredient, cellulose, starch and magnesium stearate are 
blended, passed through a No. 45 mesh U.S. sieve and filled into hard 
gelatin capsules in 200 mg. quantities. 
EXAMPLE 3 
Capsules each containing 100 mg. of active ingredient are made as follows: 
______________________________________ 
per capsule 
______________________________________ 
Active ingredient 100 mg. 
Polyoxyethylenesorbitan monooleate 
50 mcg. 
Starch powder 250 mg. 
______________________________________ 
The above ingredients are thoroughly mixed and are placed in an empty 
gelatin capsule. 
EXAMPLE 4 
Tablets each containing 10 mg. of active ingredient are made up as follows: 
______________________________________ 
per tablet 
______________________________________ 
Active ingredient 10 mg. 
Starch 45 mg. 
Microcrystalline cellulose 
35 mg. 
Polyvinylpyrrolidone 
4 mg. 
(as 10% solution in water) 
Sodium carboxymethyl starch 
4.5 mg. 
Magnesium stearate 0.5 mg. 
Talc 1 mg. 
Total 100 mg. 
______________________________________ 
The active ingredient, starch and cellulose are passed through a No. 45 
mesh U.S. sieve and mixed thoroughly. The solution of polyvinylpyrrolidone 
is mixed with the resultant powders which are then passed through a No. 14 
mesh U.S. sieve. The granules so produced are dried at 
50.degree.-60.degree. C. and passed through a No. 18 mesh U.S. sieve. The 
sodium carboxymethyl starch, magnesium stearate and talc, previously 
passed through a No. 60 mesh U.S. sieve, are then added to the granules 
which, after mixing, are compressed on a tablet machine to yield tablets 
each weighing 100 mg. 
EXAMPLE 5 
A tablet formula is prepared using the ingredients below: 
______________________________________ 
per tablet 
______________________________________ 
Active compound 250 mg. 
Cellulose microcrystalline 
400 mg. 
Silicon dioxide fumed 
10 mg. 
Stearic acid 5 mg. 
Total 665 mg. 
______________________________________ 
The components are blended and compressed to form tablets each weighing 665 
mg. 
EXAMPLE 6 
Suppositories each containing 25 mg. of active ingredient are made as 
follows: 
______________________________________ 
per suppository 
______________________________________ 
Active ingredient 25 mg. 
Saturated fatty acid glycerides to 
2,000 mg. 
______________________________________ 
The active ingredient is passed through a No. 60 mesh U.S. sieve and 
suspended in the saturated fatty acid glycerides previously melted using 
the minimum heat necessary. The mixture is then poured into a suppository 
mold of nominal 2 g. capacity and allowed to cool. 
EXAMPLE 7 
Suspensions each containing 5 mg. of medicament per 5 ml. dose are made as 
follows: 
______________________________________ 
per 5 ml. of suspension 
______________________________________ 
Active ingredient 5 mg. 
Sodium carboxymethyl cellulose 
50 mg. 
Syrup 1.25 ml. 
Benzoic acid solution 
0.10 ml. 
Flavor q.v. 
Color q.v. 
Purified water to 5 ml. 
______________________________________ 
The medicament is passed through a No. 45 mesh U.S. sieve and mixed with 
the sodium carboxymethylcellulose and syrup to form a smooth paste. The 
benzoic acid solution, flavor and color is diluted with some of the water 
and added, with stirring. Sufficient water is then added to produce the 
required volume. 
EXAMPLE 8 
An aerosol solution is prepared containing the following components: 
______________________________________ 
Weight % 
______________________________________ 
Active ingredient 0.25 
Ethanol 29.75 
Propellant 22 70 
(Chlorodifluoromethane) 
______________________________________ 
The active compound is mixed with ethanol and the mixture added to a 
portion of the propellant 22, cooled to -30.degree. C. and transferred to 
a filling device. The required amount is then fed to a stainless steel 
container and diluted further with the remaining amount of propellant. The 
valve units are then fitted to the container.