3-(2-Haloalkyl)-1,4-oxathiins and 2-(2-haloalkyl)-1,4-dithiins, and treatment of leukemia and tumors therewith

Novel 3-(2-haloalkyl)-1,4-oxathiins or 2-(2-haloalkyl)-1,4-dithiins, useful for regressing or inhibiting the growth of leukemia and tumors in mammals. The compounds have the formula.: ##STR1## wherein R.sub.1 is an alkyl group containing up to 4 carbon atoms, cyclohexyl or phenyl; PA1 R.sub.2 is hydrogen or ethyl; PA1 R.sub.3 and R.sub.4 are each hydrogen, methyl or ethyl, and PA1 when either R.sub.3 and R.sub.4 is methyl or ethyl, the other is hydrogen; PA1 X is halogen; and PA1 Y is oxygen or sulfur and PA1 when Y is sulfur, R.sub.3 and R.sub.4 are both hydrogen. and pharmaceutical compositions comprising said compounds in admixture with a pharmaceutically acceptable substantially non-toxic carrier.

TECHNICAL FIELD 
This invention relates to new 3-(2-haloalkyl)-1,4-oxathiins and 
2-(2-haloalkyl)-1,4-dithiins. More particularly, the invention relates to 
new 3-(2-haloalkyl)-1,4-oxathiin and 2-(2-haloalkyl)-1,4-dithiin analogs 
which have anti-leukemia and anti-tumor activity, to pharmaceutical 
compositions containing such analogs as the therapeutically effective 
constituents thereof, and to a method utilizing the same for inducing the 
regression of leukemia and/or the inhibition of growth of tumors in 
mammals. 
BACKGROUND OF THE INVENTION 
2-haloalkyl analogs of oxathiins and dithiins have not previously been 
described in the chemical literature. Some haloethyl analogs of various 
5-member heterocyclic systems are known, i.e., those of the type: 
##STR2## 
One such compound is chlorethiazol, viz., 
5-(2-chloroethyl)-4-methylthiazole: 
##STR3## 
This compound has been tested and found inactive as an anti-cancer agent. 
Nor has any anti-cancer activity been reported in connection with the 
other compounds of types (I) and (II) noted in the literature. 
SUMMARY OF THE INVENTION 
In accordance with the present invention, there are provided 
3-(2-haloalkyl)-1,4-oxathiins and 2-(2-haloalkyl)-1,4-dithiins of the 
formula: 
##STR4## 
wherein: R.sub.1 is an alkyl group containing up to 4 carbon atoms, 
cyclohexyl or phenyl; 
R.sub.2 is hydrogen or ethyl; 
R.sub.3 and R.sub.4 are each hydrogen, methyl, or ethyl, and when R.sub.3 
or R.sub.4 is methyl or ethyl, the other is hydrogen; 
X is halogen (preferably chloro); and 
Y is oxygen or sulfur and, when Y is sulfur, R.sub.3 and R.sub.4 are both 
hydrogen. 
In particular, the compounds of the invention include the 3-(2-halo lower 
alkyl)-1,4-oxathiins of the formula: 
##STR5## 
the 2-(2-halo lower alkyl)-1,4-dithiins of the formula: 
##STR6## 
wherein R.sub.1, R.sub.2, R.sub.3, R.sub.4 and X are as defined above. 
In the following description, the preparation and testing of the compounds 
of the invention is described in connection with the preferred 
chloro-substituted compounds (X.dbd.Cl). It will, however, be understood 
that the invention embraces the analogous halo-substituted oxathiins and 
dithiins as well. 
The oxathiin and dithiin derivatives of the invention may be readily 
prepared in three sequential steps. In the first step, an appropriate 
2-acylbutyrolactone or 2-acyl-4-ethylbutyrolactone of the formula: 
##STR7## 
is reacted with halogen (e.g., chlorine gas) in the presence of base. The 
reaction is carried out at temperatures of from about 5.degree. C. to 
30.degree. C., preferably about 10.degree. to b 20.degree. C. The 
resulting 2-acyl-2-halobutyrolactone or 
2-acyl-4-ethyl-2-halobutyrolactone: 
##STR8## 
is reacted with acid, e.g., HCl, to open the ring, forming a haloketone: 
EQU (VIII) R.sub.1 COCH(X)CH.sub.2 CH(R.sub.2)X 
The latter is then recovered by steam distillation, extraction, and 
re-distillation. 
The final step in the synthesis is the reaction of a mixture of the 
haloketone with an appropriate mercaptoethanol: 
EQU (IX) HS(R.sub.3)CHCH(R.sub.4)OH, 
followed by cyclization with an acid catalyst to compound (III). The 
haloketone and mercaptoethanol are desirably reacted in approximately 
equimolar ratios, and at temperatures of from 5.degree. to 60.degree. C. 
PTSA (p-toluenesulfonic acid) may be utilized as the acid catalyst, the 
cyclization being effected under reflux with the removal of water. 
The compounds of the invention are cytotoxic agents useful to induce the 
regression of malignancies such as lymphoid and lymphocytic leukemia, as 
well as to inhibit the growth of various cancers, e.g, melanocarcinoma, 
sarcoma, and mammary xenograft tumors. They may be used alone or in 
combination with other chemotherapeutic agents active for these purposes. 
As used herein, the terms "regression" and "inhibition" comprehend 
arresting or retarding the growth of the malignancy or other manifestation 
of the disease, as compared with the course of the disease in the absence 
of treatment. 
Administration of the compounds of the invention to mice in amounts ranging 
from about 50-800 mg./kg., preferably from about 200-400 mg./kg., of body 
weight has been found effective to induce the regression of leukemia and 
to inhibit the growth of tumors. The interrelationship of dosages for 
mammals of other sizes and species is described by Freidreich, E. J., et 
al., Quantitative Comparison of Toxicity of Anti-Cancer Agents in Mouse, 
Rat, Hamster, Dog, Monkey and Man, Cancer Chemotherapy, Reg. 50, No. 
4,219-244, May 1966. 
The dosage level may, of course, be adjusted to provide optimum response. 
For example, several divided doses may be administered daily, or the dose 
may be proportionally reduced, as indicated by the exigencies of the 
situation. 
The active compounds may suitably be administered parenterally, 
intraperitoneally, intravenously or orally. Solutions or dispersions of 
the acitve compounds can be prepared in water, suitably mixed with a 
surfactant such as hydroxypropylcellulose. Dispersions can also be 
prepared in glycerol, liquid polyethylene glycols, and mixtures thereof, 
and in oils. Under ordinary conditions of storage and use, these 
preparations contain a preservative to prevent the growth of 
microorganisms. 
The forms suitable for injectable use include sterile aqueous solutions or 
dispersions and sterile powders for the extemporaneous preparation of 
sterile injectable solutions or dispersions. For such uses the form must 
be sterile and must be fluid to the extent necessary to provide easy 
syringability. It must be stable under the conditions of manufacture and 
storage and must be preserved against the contaminating action of 
microorganisms such as bacteria and fungi. 
The carrier can be a solvent or dispersing medium containing, for example, 
water, ethanol, a polyol (for example, glycerol, propylene glycol, and 
liquid polyethylene glycol, or the like), suitable mixtures thereof, and 
vegetable oils. The proper fluidity can be maintained, for example, by the 
use of a coating such as lecithin, by the maintenance of the required 
particle size in the case of a dispersion, and by the use of surfactants. 
Prevention of the action of microorganisms can be insured by various 
anti-bacterial and anti-fungal agents, for example, paraben, 
chlorobutanol, phenol, sorbic acid, thimerosal, or the like. In many cases 
it may be preferable to include isotonic agents, for example sugars or 
sodium chloride, in the dosage form. Prolonged absorption of the 
injectable formulations can be brought about by incorporating agents 
delaying absorption, for example, aluminum monostearate and gelatin, 
therein. 
Sterile injectable solutions are prepared by incorporating the active 
compound in the appropriate solvent, in admixture with various of the 
other ingredients enumerated above, as required, followed by filter 
sterilization. Generally, dispersions are prepared by incorporating the 
sterilized active ingredient in a sterile vehicle which contains the 
dispersing medium and any other required ingredients. When, on the other 
hand, sterile powders are used to prepare sterile injectable solutions, it 
is preferred to subject a sterile, filtered solution of the desired 
ingredients to vacuum drying or freeze-drying, yielding a powder of the 
active ingredient plus any additional desired ingredients. 
As used herein, "pharmaceutically acceptable, substantially nontoxic 
carrier or excipient" includes solvents, dispersing media, coatings, 
anti-bacterial and anti-fungal agents, isotonic and absorption delaying 
agents, and the like. The use of such media and agents as carriers or 
excipients for pharmaceutically active substances is well known in the 
art. Except insofar as any conventional medium or agent is incompatible 
with the active ingredient or toxic in admixture therewith, its use in the 
formulations of the invention is contemplated. Supplementary active 
ingredients can also be incorporated in the therapeutic compositions. 
It may be advantageous to formulate the compositions of the invention in 
unit dosage forms for ease of administration and uniformity of dosage. A 
unit dosage form, as used herein, refers to a physically discrete unit 
suitable for use as a unitary dosage for the mammalian subjects to be 
treated; each unit contains a predetermined quantity of active material 
calculated to produce the desired therapeutic effect, in association with 
the required pharmaceutically acceptable carrier. Specifications for unit 
dosage forms are dictated by and directly depend on (a) the unique 
characteristics of the active material and the particular therapeutic 
effect to be achieved, and (b) the limitations inherent in the art of 
compounding such an active material for the treatment of disease in living 
subjects having a diseased condition, without excessive cytotoxic side 
effects. 
Regression of leukemia and inhibition of tumor growth may be attained, for 
example, by the use of daily dosing for up to 5 or 10 days, or longer. 
Multiple dosing, or dosing on any desired periodic basis, may also be 
utilized. The therapeutically active ingredient is thus administered in 
amounts sufficient to aid regression and inhibition of further growth of 
the leukemia or tumor, in the absence of excessive deleterious side 
effects of a cytotoxic nature. 
PREFERRED EMBODIMENTS OF THE INVENTION 
Particularly preferred among the compounds of the invention are 
3-(2-chloroethyl)-5,6-dihydro-2-methyl-1,4-oxathiin: 
##STR9## 
the analogous 2-(2-chloroethyl)-3-methyl-5,6-dihydro-1,4-dithiin: 
##STR10## 
The invention will be described in greater detail in connection with the 
preparation and pharmacological testing of the preceding and other 
preferred compounds, having the following chemical structures: 
TABLE I 
______________________________________ 
Illustrative Compounds of the Invention 
##STR11## 
Example R.sub.1 R.sub.2 
R.sub.3 R.sub.4 
Y 
______________________________________ 
1 CH.sub.3 H H H O 
2 CH.sub.3 H H CH.sub.3 
O 
3 C.sub.3 H.sub.7 
H H H O 
4 C.sub.6 H.sub.5 
H H H O 
5 CH.sub.3 H H H S 
6 CH.sub.3 C.sub.2 H.sub.5 
H H O 
______________________________________

EXAMPLE 1 
Preparation of 3-(2-chloroethyl)-5,6-dihydro-2-methyl-1,4-oxathiin 
2-acetylbutyrolactone (256 g., 2 mol.), and anhydrous sodium acetate (170 
g.) in acetic acid (600 ml) were mixed and cooled in an ice bath with 
stirring. Chlorine gas (144 g.) was bubbled into the reaction mixture 
while maintaining the reaction temperature below 35.degree. C. Upon 
completion, the resulting precipitate was removed and acetic acid was 
removed under reduced pressure. The oil was taken up in toluene and was 
washed with water and aqueous sodium bicarbonate, and dried before 
removing the solvent. The residue, 2-acetyl-2-chlorobutyrolactone, bp. 
76.degree.-78.degree./0.25 mm, was isolated in 81% yield. 
The 2-acetyl-2-chlorobutyrolactone (234 g., 1.44 mole) was mixed with water 
(350 ml) and 12N hydrochloric acid (300 ml), and distilled. When 300 ml of 
distillate had been collected, additional water (300 ml) was added and the 
steam distillation continued until no more product was obtained. The 
product was extracted into methylene chloride, dried (magnesium sulfate) 
and the solvent removed. Distillation of the residue gave 
3,5-dichloro-2-pentanone (bp. 70.degree.-73.degree./12 mm 142 g., 57%). 
A mixture of the 3,5-dichloro-2-pentanone (39 g., 0.24 mole) and 
2-mercaptoethanol (20 g., 0.26 mole) in toluene (250 ml) was stirred, and 
triethylamine (27 g., 0.27 mole) added dropwise thereto. After stirring 
overnight at ambient temperature, the mixture was washed with dilute 
hydrochloric acid and subsequently refluxed with p-toluenesulfonic acid 
(0.5 g.) with azeotropic removal of water for 4 hours. Upon cooling, the 
reaction mixture was washed with aqueous sodium bicarbonate, dried 
(magnesium sulfate), and the toluene removed to leave the product 
3-(2-chloroethyl)-5,6-dihydro-1,4-oxathiin (bp. 
62.degree.-70.degree./0.025 mm) in 60% yield and 98% purity (G.C.). N.M.R. 
spectrum (Deuterchloroform) gave lambda values: 1.85 (3H,S) 2.3-2.65 
(2H,t); 2.9-3.5 (2H,t); 3.95-3.7 (2H,t); 4.1-4.25 (2H,t). 
EXAMPLE 2 
Preparation of 2,6-dimethyl-3-(2-chloroethyl)-5,6-dihydro-1,4-oxathiin 
A mixture of 0.1 mole (15.4 g.) 3,5-dichloro-2-pentanone, and 0.1 mole (9.2 
g.) 1-mercapto-2-propanol was stirred in 100 ml toluene; 0.1 mole (10.4 
g.) triethylamine was added dropwise. The mixture was stirred at room 
temperature overnight, then washed with dilute hydrochloric acid and 
reluxed with 0.5 g. PTSA, using a Dean-Stark trap to remove water for 
about 7 hours. The PTSA solution was washed with sodium bicarbonate, dried 
over magnesium sulfate and the solvent was removed. The product was 
distilled; bp 80.degree.-82.degree./0.1 mm. Yield 47%. 
EXAMPLE 3 
Preparation of 2-propyl-3-(chloroethyl)-5,6-dihydro-1,4-oxathiin 
A mixture of 0.056 mole (10 g.) of 1,2-dichloro-4-heptanone and 0.06 mole 
(5 g.) 2-mercaptoethanol was stirred in 200 ml toluene; 0.06 mole (6 g.) 
triethylamine was added dropwise. The mixture was stirred at room 
temperature overnight, then washed with dilute hydrochloric acid and 
refluxed with 0.5 mole PTS, using a Dean-Stark trap to remove water for 7 
hours. The PTSA solution was washed with 5% sodium bicarbonate, dried, 
filtered and the solvent was removed. The product was distilled; bp 
80.degree.-85.degree./0.05 mm. Yield 23%. 
EXAMPLE 4 
Preparation of 2-phenyl-3-(2-chloroethyl)-5,6-dihydro-1,4-oxathiin 
4-chlorobutyrophenone (36.4 g., 0.2 mole) was stirred in 100 ml methylene 
chloride at room temperature. Bromine (32 g., 0.2 mole) was added 
dropwise. After complete addition the solution was washed with aqueous 
sodium bicarbonate, dried with magnesium sulfate, filtered and the solvent 
removed. The residue was taken up in 300 ml toluene, and 2-mercaptoethanol 
(18 g., 0.23 mole) was added. Triethylamine (22 ml) was added slowly and 
the mixture stirred overnight at ambient temperatures. The mixture was 
then washed with dilute hydrochloric acid and subsequently refluxed with 
PTS (0.5 g.) with azeotropic removal of water for 5 hours. Upon cooling, 
the reaction mixture was washed with aqueous sodium bicarbonate and water, 
dried (magnesium sulfate), filtered, and the toluene removed. The product 
was purified by preparative liquid chromatography in 17% yield. NMR 
spectrum (deuterochloroform) gave ppm values; 2.50-2.75 (2H,t); 3.0-3.15 
(2H,t); 3.45-372 (2H,t); 4.28-4.42 (2H, 5); 7.33 (5H,s). 
EXAMPLE 5 
Preparation of 2-(chloroethyl)-3-methyl-5,6-dihydro-1,4-dithiin 
A mixture of 0.1 mole (15.4 g.) 3,5-dichloro-2-pentanone, 0.1 mole (9.4 g.) 
ethanedithiol and 0.3 mole PTSA was stirred at room temperature overnight. 
The product was taken up in toluene, washed with 5% sodium bicarbonate and 
water, dried over magnesium sulfate, filtered and the solvent removed. The 
product was distilled; bp. 114.degree.-116.degree./0.7 mm. Yield 32%. 
EXAMPLE 6 
Preparation of 3-(2-chlorobutyl)-5,6-dihydro-2-methyl-1,4-oxathiin 
2-acetyl-4-ethyl butyrolactone was prepared from ethylacetoacetate and 
1,2-epoxybutane according to the method described in Johnson U.S. Pat. No. 
2,443,827. A mixture of 40 g sodium hydroxide (1.0 mole), 270 ml water and 
90 ml ethanol was cooled to 0.degree., stirred, and 130 g 
ethylacetoacetate (1.0 mole) and 72 g 1,2-epoxybutane (1.0 mole) added. 
Stirring was continued at 0.degree., and the mixture was thereafter left 
at 4.degree. C. for 48 hours. The reaction mixture was neutralized with 80 
ml acetic acid, extracted with toluene, washed with water, sodium 
bicarbonate and finally with water. The mixture was then dried (magnesium 
sulfate), filtered and the solvent removed, leaving a product, bp 
86.degree.-96.degree.-/0.1 mm, in 45% yield. 
The 2-acetyl-4-ethyl butyrolactone, 70 g (0.45 mole), prepared above was 
stirred in 135 ml acetic acid with 38 g sodium acetate. 32 g. of Cl.sub.2 
was bubbled in with stirring in ice. The precipitate was filtered off, 
acetic acid removed and product distilled, bp 65.degree.-77.degree./0.05 
mm. 2-acetyl-2-chloro-4-ethyl butyrolactone was obtained in 84% yield. 
The above product, 72 g (0.38 mole), was added to 90 ml hydrochloric acid 
and 105 ml water. The product was steam distilled, a further 100 ml water 
added and distillation continued until 250 ml was collected. The 
distillate was extracted with methylene chloride, dried (magnesium 
sulfate) filtered and solvent removed. The product was distilled at about 
10 mm, bp 84.degree.-97.degree., to give 26 g (38% yield) of 
3,5-dichloro-2-heptanone. 
A mixture of 26 g (0.14 mole) of 3,5-dichloro-2-heptanone, 12 g (0.15 mole) 
of 2-mercaptoethanol, and 15 g (0.15 mole) of triethylamine in 200 ml 
toluene, was stirred at ambient temperature overnight. The mixture was 
then washed with dilute hydrochloric acid and subsequently refluxed with 
0.1 g. p-toluenesulfonic acid with azeotropic removal of water for six 
hours. Upon cooling, it was washed with aqueous sodium bicarbonate, dried 
(magnesium sulfate), and the solvent was removed to leave the product. 
3-(3-chlorobutyl)-5,6-dihydro-2-methyl-1,4-oxathiin, bp 
82.degree.-85.degree./0.05 mm, was obtained in 35% yield. N.M.R on the 
product was satisfactory. 
IN VIVO PHARMACOLOGICAL TESTING OF COMPOUNDS OF THE INVENTION 
Subrenal Capsule Human Mammary Carcinoma MX-1 Xenograft 
Samples of various test compounds were tested in accordance with National 
Cancer Institute Protocol (Cancer Chemotherapy Reports Part 3 Vol. 3, No. 
2, September 1972). Each test (NCI 3MBG5) involved implantation of a tumor 
fragment (surgical explant in 1974 from the primary mammary tumor of a 29 
year old woman with no previous chemotherapy; Reference: Tumor Bank 
information) under the membranous covering of the kidney of either an 
athymic Swiss or athymic random bred mouse, 6 animals per test group and 
12 per control, one sex per experiment. The male mice weighed a minimum of 
18 grams and the female mice weighed a minimum of 17 grams, and all of the 
test animals were within a four gram weight range. The test compound was 
administered by intraperitoneal injection, commencing one day after tumor 
implant and was repeated every fourth day for a total of three injections. 
The test animals were weighed and the implanted tumor was measured and 
recorded on day 0 and day 11--the final evaluation day. The parameter 
measured is the change in tumor weight for the treated (T) and control (C) 
animals. An initial T/C&lt;20% is considered necessary to demonstrate 
moderate activity. A reproducible T/C&lt;10% is considered significant 
activity. 
The results of this test with the compounds of each of the above examples, 
and with various control compounds, are tabulated in Table II. 
TABLE II 
__________________________________________________________________________ 
Subrenal Capsule Human Mammary Carcinoma 
MX-1 Xenograph Test 
Test Compounds 
##STR12## 
Dose T/C % 
T/C % 
T/C % 
TC % 
Example 
R.sub.1 
R.sub.2 
R.sub.3 
R.sub.4 
Y (mg/kg) 
T/C % 
(repeat) 
(repeat) 
(repeat) 
(repeat) 
__________________________________________________________________________ 
1 CH.sub.3 
H H H O 1200 Toxic 
Toxic 
Toxic 
600 3 84 Toxic 
300 47 59 -60(5).sup.1 
150 35 68 4(5).sup.1 
Cold 800 129 Toxic 
Toxic 
Sample.sup.2 600 Toxic 
Toxic(1).sup.1 
300 -100(4).sup.1 
Toxic 
150 2(5).sup.1 
- 87 
75 23(2).sup.1 
-10 
2 CH.sub.3 
H H CH.sub.3 
O 1200 Toxic 
600 Toxic 
300 Toxic 
150 -40(4).sup.1 
3 C.sub.3 H.sub.7 
H H H O 300 Toxic(2).sup.1 
Toxic 
150 -44(3).sup.1 
1(1).sup.1 
75 2(1).sup.1 
2(1).sup.1 
37.5 32 
4 C.sub.6 H.sub.5 
H H H O 400 9(1).sup.1 
300 17 49 
150 58 50 
75 50 
37.5 66 
5 CH.sub.3 
H H H S 1200 Toxic 
600 -50(4).sup.1 
300 32 
150 71 
6 CH.sub.3 
C.sub.2 H.sub.5 
H H O 400 23 16 
300 58 18 14 
150 58 43 38 
75 74 
37.5 86 
Control A 
CH.sub.3 
H H C.sub.3 H.sub.7 
O 1200 Toxic 
600 Toxic 
300 51 79 
150 62 81 
75 89 
37.5 98 
Control B 
CH.sub.3 
H CH.sub.3 
CH.sub.3 
O 600 Toxic 
300 Toxic 
150 Toxic 
75 101 
Control C 
CH.sub.3 
H H/CH.sub.3 
H/CH.sub.3 
S 300 68 
150 76 
75 93 
37.5 113 
Control D 
CH.sub.3 
H H/C.sub.6 H.sub.5 
C.sub.6 H.sub.5 /H 
O 300 106 
150 73 
75 99 
37.5 92 
__________________________________________________________________________ 
.sup.1 Animals cured indicated by 
.sup.2 Second purer, refrigerated sample tested. 
From the data appearing in Table II, it may be seen that the compounds of 
each of Examples 1-4 exhibited significant activity at one dosage or 
another. 
The preferred compound of Example 1 was subjected to a number of further in 
vivo tests. The results are summarized in Table III, and set forth in 
detail in Table II (above), and in Tables IV-VII; 
TABLE III 
__________________________________________________________________________ 
SUMMARY OF TUMOR PANEL TEST RESULTS 
RESULTS 
TEST SYSTEM STATUS TABULATED 
__________________________________________________________________________ 
3MBG5 (MAMMARY XENOGRAPH) 
ACTIVE (DN2 LEVEL) 
TABLE II 
3PS31 (P388 LYMPHOCYTIC LEUKEMIA) 
ACTIVE Table IV 
2LE31 (L-1210 LYMPHOID LEUKEMIA) 
ACTIVE Table V 
3B131 (B16 MELANOCARCINOMA) 
ACTIVE (DN2 LEVEL) 
Table VI 
3M531 (M5076 SARCOMA) 
ACTIVE Table VII 
3C872 (COLON 38) INACTIVE (1 TEST)* 
3LE32 (L-1210 LYMPHOID LEUKEMIA) 
INACTIVE (1 TEST)* 
3CDJ2 (MAMMARY TUMOR) 
INACTIVE (1 TEST)* 
__________________________________________________________________________ 
*Results inconclusive to date. 
REGRESSION OF INTRAPERITONEALLY-IMPLANTED LYMPHOCYTIC LEUKEMIA P388 
The compound of Example 1 was tested by the standard National Cancer 
Institute Lymphocytic Leukemia P388 primary screen (NCI Protocol 1.200, 
Cancer Chemotherapy Reports Part 3 Vol. 3, No. 2, September 1972). Each 
test (NCI 3PS 31) involved implantation of the leukemia cells (American 
Journal of Pathology, 33, No. 3, page 603, 1957) in six DBA/2 mice, one 
sex per experiment, the male mice weighing a minimum of 18 grams and the 
female mice weighing a minimum of 17 grams, and all of the test animals 
being within a three gram weight range. The test compound was administered 
by intraperitoneal injections, in 0.1 ml. doses of diluted ascitic fluid 
(10.sup.6 cells per dose), commencing one day after the tumor implant and 
continuing daily for nine days. 
The test animals were weighed and survivors recorded on a regular basis 
during a thirty day test period. The ratio of survival time for the 
treated (T) and control (C) animals was determined at varying dosages. The 
experiments were repeated to assess reproducibility. The results are 
tabulated in Table IV: 
TABLE IV 
______________________________________ 
Lymphocytic Leukemia P388 Test 
Dose T/C % 
(mg/kg) T/C % repeat 
______________________________________ 
400 -- 117 
200 138 130 
100 107 114 
50 88 107 
______________________________________ 
An initial T/C equal to or greater than 125% is considered necessary to 
demonstrate activity, a reproducible T/C equal to or in excess of 125% is 
considered worthy of further study and a reproducible T/C equal to or 
greater than 175% is considered significant in this protocol. From the 
data appearing in Table IV, it may be seen that the compound of Example 1, 
when used at a dosage of 200 mg/kg, demonstrated activity worthy of 
further study. 
REGRESSION OF INTRAPERITONEALLY-IMPLANTED LYMPHOID LEUKEMIA L1210 
Samples of the test compound of Example 1 were tested in accordance with a 
further National Cancer Institute protocol (NCI Protocol 1.100, Cancer 
Chemotherapy Reports Part 3 Vol. 3, No. 2, September 1972) to determine 
the effects of the compounds on intraperitoneally-implanted L1210 leukemia 
(J. Nat'l. Cancer Inst. 13(5): 1328, 1953). The test protocol (NCI 3LE 31) 
was similar to Protocol 1.200 above, save that 10.sup.5 L1210 leukemia 
cells were implanted in the test animals. The test compound was 
administered daily for a period of nine days. The tests were carried out 
at varying dosage leves and with varying numbers of repetitions. It has 
been statistically determined that an intial T/C value in this test at 
least equal to 125% is necessary to demonstrate activity, while a 
reproducible T/C equal to or greater than 125% warrants further study. A 
reproducible T/C of 150% or higher is considered significant activity in 
this test. 
The results are tabulated in Table V: 
TABLE V 
______________________________________ 
Intraperitoneally-Implanted L1210 
Lymphoid Leukemia Test 
Dose T/C % T/C % T/C % 
(mg/kg) T/C % repeat repeat 
repeat 
______________________________________ 
800 147 Toxic 155 151 
400 no deaths 
162 130 133 
recorded 
200 114 120 113 127 
100 107 106 111 111 
50 111 102 
______________________________________ 
As may be seen from Table V, the compound of Example 1 exhibited 
significant activity in the i.p.-implanted lymphoid leukemia test at 
dosage levels of both 800 mg/kg and 400 mg/kg (reproducible T/C of 150% or 
higher). 
INTRAPERITONEALLY-IMPLANTED B16 MELANOMA 
The compound of Example 1 was further tested against an 
intraperitoneally-implanted B16 melanoma.sup.1, in accordance with 
National Cancer Institute melanotic Melanoma B16 Protocol 1.300.sup.2 [NCI 
3B131]. 
FNT .sup.1 Handbook on Genetically Standardized Jax Mice. Roscoe B. Jackson 
Memorial Laboratory, Bar Harbor, Maine, 1962. See also Ann. NY Acad. Sci., 
Vol. 100, Parts 1 and 2 (Conference on the Biology of Normal and Typical 
Pigment Cell Growth of 1961), 1963. 
FNT .sup.2 Cancer Chemotherapy Reports, Part 3, Vol 3, No. 2, September 1972. 
A 1:10 tumor brei was implanted intraperitoneally in B6C3F1 mice, employing 
test groups complying with the criteria described above in connection with 
NCI Protocol 1.200, except that ten animals were utilized per test group. 
The test compound was administered intraperitoneally at various doses. The 
animals were weighed and survivors recorded on a regular basis for 60 
days. The T/C values were then calculated, the results obtained being 
illustrated in Table VI. 
TABLE VI 
______________________________________ 
Melanocarcinoma B16 Test 
Dose T/C % T/C% 
(mg/kg) T/C % repeat repeat 
______________________________________ 
800 150 Toxic Toxic 
400 155 176 120 
200 137 140 157 
100 129 115 134 
50 124 106 
25 111 
______________________________________ 
A T/C value in excess of 125% is considered necessary to demonstrate 
moderate activity, and a reproducible T/C value equal to or in excess of 
150% is considered significant activity, in the above test. From the data 
tabulated in Table VI above, it can be seen that the compound of Example 1 
exhibited significant activity in the melanocarcinoma B16 test at dosage 
levels as low as 200 mg/kg. 
INTRAPERITONEALLY-IMPLANTED M5076 ASCITIC SARCOMA 
The compound of Example 1 was additionally tested against an 
intraperitoneally-implanted M5076 sarcoma in accordance with National 
Cancer Institute 3M531 Protocol. 
1.times.10.sup.6 cells of ascitic fluid were implanted in the test mice, 
the test compound being administered beginning one day after implant and 
every fourth day thereafter for a total of four injections. The median 
survival times as percentages of the control survival time were as 
follows: 
TABLE VII 
______________________________________ 
M5076 Sarcoma Test 
Dose T/C % T/C % T/C % 
mg/kg T/C % repeat repeat 
repeat 
______________________________________ 
800 -- 129 Toxic Toxic 
400 98 117 Toxic 144 
200 98 102 118 120 
100 100 109 96 128 
50 98 98 100 110 
25 111 -- -- -- 
Vehicle.sup.1 
99 
______________________________________ 
.sup.1 Vehicle: 5% EtOH, 5% Cremophor, saline. 
The compound of Example 1 of the invention has also been employed in 
further in vivo testing in accordance with the following National Cancer 
Institute Test protocols: 
______________________________________ 
Protocol Test 
______________________________________ 
3C872 Subcutaneously-Implanted 
Colon 38 Carcinoma 
3LE32 Subcutaneously-Implanted 
L1210 Leukemia 
3CDJ2 Subcutaneously-Implanted 
Staged Mammary 
Adenocarcinoma CD8F1 
______________________________________ 
The following results were obtained 
TABLE VIII 
______________________________________ 
Protocol Dose T/C % 
______________________________________ 
3C872.sup.1 600 50 
300 46 
150 118 
75 109 
37.5 92 
3L32.sup.2 800 Toxic 
400 108 
200 104 
100 109 
50 94 
3CDJ2.sup.3 1000 22 
500 128 
250 81 
125 114 
62.5 101 
31.25 104 
______________________________________ 
.sup.1 In this test an initial T/C .gtoreq.42 is considered necessary to 
demonstrate moderate activity. 
.sup.2 There are currently no specific standards for this protocol. 
Activity is measured in accordance with the 3LE31 protocol, in which an 
initial T/C .gtoreq.125 indicates moderate activity. 
.sup.3 In this test a median tumor weight change of .ltoreq.20 
demonstrates activity. 
The compound of Example 1 did not exhibit activity in the 3C872, 3LE32, or 
3CDJ2 screens. 
In accordance with the present invention, a novel class of 
3-(2-haloalkyl)-1.4-oxathiin and 2-(2-haloalkyl)-1.4-dithiin derivatives 
is provided, which exhibits pharmacological activity in the regression 
and/or inhibition of the growth of leukemia and a number of malignant 
tumors in mammals. 
The preceding disclosure should be construed as illustrative only. The 
scope of the invention should be interpreted in accordance with the 
following claims.