Note: Descriptions are shown in the official language in which they were submitted.
1 157774
BACKGROUND OF THE INVENTION
Field of the Invention:
The present invention is directed to a method for
reducing mucin viscosity in mammals.
Description of the Prior Art:
me prior art techniques for treating mucus conditions
have depended upon where the mucus was located in the body.
Mucus impaction of the gastrointestinal tract, biliary and pan-
creatic ducts generally required surgical techniques for its
removal since its viscosity was too high for the body to rid
itself of the mucus by its normal path. Mucus in the respira-
tory tract - often called sputum - has been treated by a
variety of techniques including cough syrup and the like. One
typical technique for respiratory obstructions has involved the
aerosolization of N-acetylcysteine. While N-acetylcysteine is
very effective in reducing sputum viscosity, it is extremely
difficult, if not impossible, to obtain adequate delivery of
this agent to the small airways of the lungs. Hence, its
effectiveness in reducing the sputum viscosity is variable
depending upon whether or not adequate delivery has been
attained. In addition, N-acetylcysteine is extremely irritat-
ing to gastrointestinal and respiratory mucosa.
The problem of elevated sputum viscosity is particu-
larly aggravated in those individuals suffering from cystic
fibrosis wherein the high viscosity of the sputum leads to
clinical difficulties in clearance of the respiratory and
gastrointestinal tracts and in delivery of gas and nutrients to
their respective sites. Furthermore, in newborns, the cystic
fibrosis may be present as meconium ileus, an intestinal
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1 157774
obstructive syndrome due to the secretion of abnormally viscid
gastrointestinal mucus. At the present time, the only method
of treating these intestinal obstructions is through surgical
techniques to physically remove the blockage.
Accordingly, there exists a need for a method of
treating mucus impactions and for the reduction of sputum
viscosity which does not involve either surgical intervention
or the delivery of drugs in aerosol form.
SUMMARY OF THE INVENTION
Accordingly, it is an object of the present invention
to provide a treatment technique which provides for the reduction
in the viscosity of mucin solutions.
It is a further object of the present invention to
provide a treatment for respiratory and gastrointestinal
obstructions through the reduction of sputum and mucus viscosities.
It is yet another object of the present invention
to achieve the reduction of sputum viscosity by a treatment
involving either the oral or intravenous injection of a drug
into the mammal.
These and other objects of the present invention which
will become apparent have been attained through the oral or
intravenous administration of a compound which is metabolized
in vivo to produce compounds containing sulfhydryl groups.
BRIEF DESCRIPTION OF THE DRAWINGS
A more complete appreciation Gf the invention and
many of the attendant advantages thereof will be readily
obtained as the same becomes better understood by reference
to the following detailed description when considered in
connection with the accompanying drawings, wherein:
Figure 1 demonstrates the change in viscosity of
G_
1 157774
component I (~ ~ ~ ). Component IA (0-0-0), whole sputum
(--0-0-0-), and fibrinogen (~ ) with protein concentration
(A)
Figure 2 demonstrates the change in viscosity of
components I, IA and whole sputum with DTE additron (B),
Figure 3 demonstrates the change in viscosity of whole
sputum after addition of 5 mM MDP, and
Figure 4 demonstrates the (A) appearance of rree
sulfhydryls in the blood of mice after i.p. administration of
50 (~ ~ ), 100 (0-0-0), and 200 (~-0-0) mg/kg WR 2721. (B)
Appearance of free sulfhydryls in the blood of mice after
p.o. administration of 400 mg~kg. (C) Appearance of free
sulfhydryl in the lungs of mice after i.p. administration
of 200 mg~kg.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
The present invention is based on the isolation of
a mucin having a high molecular weight of 425,000 D from the
sputum of a patient with cystic fibrosis. This protein may
be responsible for the increased viscosity of the sputum since
increasing its con-entration results in a rise in viscosity.
Also, as in the case of the crude sputum, the viscosity of the
purified mucin can be decreased by treatment with sulfhydryl
reducing agents. Similar results were also obtained with
sputum obtained from patients suffering from pneumuccocal
pneumonia, chronic bronchitis and the like. m e biochemical
effect of the reducing agents on sputum mucin viscosity is
to split the molecule into a component which retains most of
the carbonydrate and ~t least two small peptides having mole-
cular weights of 65,000 Daltons and 27,000 Daltons. This
structural change leads to a dramatic alteration of the rheolo-
gical properties of the molecùle. This alteration of the
rheological properties of the molecule would allow the body to
1 157774
rid itself of the mucus secretions via the normal body
processes. Although we did find the use of sulfhydryl
compounds to be very effective in reducing the mucus viscosity,
the administration of sulfhydryl compounds to mammals has not
proven to be a satisfactory method for treating mucin with
sulfhydryl compounds to be administered either intravenously
or orally. In particular, fairly large doses of the sulfhydryl
compound must be given to compensate for the reaction of the
free thiol with plasma and gastrointestinal proteins while
enroute to the lungs and other organs where mucus impaction
may have occured. At the dosages which must be given, the
sulfhydryl compounds can lead to toxic side effects. We have
now discovered the problem which we encountered with the
administration of sulfhydryl compounds to reduce mucin visco-
sity may be overcome by administering the compounds which
; are converted to sulfhydryl groups in vivo. Suitable
compounds include pharmaceutically acceptable thiosulfates,
thiophosphates, disulfides and the like. ~he compounds which
are suitable for use in the present invention are all
characterized by containing a blocked sulfhydryl group wherein
the blocking agent is removed in vivo to form a sulfhydryl
group. Suitable compounds include aminoalkylthiosulfuric acids,
aminoalkylphosphorothioates, thiosulfatoalkylamines, phenal-
kylaminoalkylthiosulfuric acids, hydroxyalkylaminoalkylthio-
sulfuric acids, hydroxyaminoalkylphosphorothioates, alkoxy-
alkylaminoalkylthiosulfuric acids, cycloalkyloxyaminoalkyl-
thiosulfuric acids, phenoxyalkylaminoalkylthiosulfuric acids,
cycloalkylaminoalkylthiosulfuric acids, cycloalkylalkylamino-
alkylphosphorothioates, cycloalkylalkylaminoalkyldisulfides,
phenoxyalkylaminoethyldisulfides, hydroxyalkylaminoalkyldi-
1 157774
sulfides, alkylamidiniumthiosulfates, acetamidine derivativescontaining a blocked sulfhydryl group, arylalkylamidiniumthio-
sulfates, aminoalkylaminoalkylphosphorothioates, quinolyloxy-
alkylaminoalkylthiosulfuric acids, pyridyloxyalkylaminoalkyl-
thiosulfuric acids, phenoxy- and phenylthioalkylamidinium-
thiosulfates, cycloalkylamidiniumthiosulfates, and the like.
In general, any compound containing a blocked sulfhydryl group
which is pharmaceutically acceptable may be employed in the
present invention provided its blocked sulhydryl group is
0 converted to the free sulfhydryl group in vivo.
m e following compounds are preferred for use in the
present invention:
1. (H2NCOCH2CH2NHCH2CH2S-)2
2 H ~COCH2CH2~HCH2CH2SS03H
L
1 157774
I :
I'
,'
3. H2NcocH2cHzN~cH2cH2spo3 2
4. H2N(CH2)3NHCH2CH2SS03H
5, (H2N(CH2)3NHCH2CH2s-)2- '
6. CH3(CH2)9NHCH2CH2SS03H
7. CH3(CH2)9NHcH2cH2sPo3ff2 ,
¦ 8- (CH3(CH2)9NHCH2CH2S )2
l 9. H2NCH2CH2SP03H2
l 10. H2N(C~2)sNH(CH2)2SP03~2
11. H2N(CH2)3NH(C~2)sPo3H2
¦ Of these compounds, (11) is particularly preferred. This compound has
¦ been designated W~ 2721 in toxological tests conducted at the Walter Reed
Army Medical Center. This compound has been proposed as a novel antiradiation ¦
¦ drug and is the subject of United States Patent 3,892,824.
Il, .
In the treatment of the present invention, the compound containing ~he
¦ blocked sulfhydryl group is administered to the patient at a dosage rate
¦ sufficient to reduce the mucin viscosity. Dosage rates ranging from l.~g/kg/
¦ day to an excess of 100 mg/kg/day have proven satisfactory. Dosage rates
ir, the der of from 5 to 50 mg/kgJday are preferrbly employed. A
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1 157774
particularly preferred dosage rate involves the administration of 5 mg/kg
four times a day. The compound may be administered either orally or
intravenously in conjunction with a suitable pharmaceutical carrier.
Intravenous administration may involve the use of physiological saline
solutions which may or may not contain a sodium carboxy methyl cellulose
and if desired TWEEN 80 R. Obviously, simple physiological saline
solutions may be employed; water alone can be used, or the like.
In the preferred method the compound is administered orally with a
suitable solid carrier. If desired, adjuvants such as buffers and the like
may be employed. It may be embodied in suitable tablet form, such as
a WR 2721 containing tablet provided with an enteric coating, MR 2721
being present in major or minor amount. It may also be administered in
a suitable capsùle, of geletin or the like. Suitable solid carriers include
starch powder, lactose, glucose powder, sucrose, dextrose, manitol powder,
avicel (microcrystalline cellulose), calcium phosphate anhydrous, calcium
phosphate dihydrate, kaolin, talc, magnesium carbonate, magnesium hydroxide,
calcium carbonate, sodium bicarbonate, sorbitol and the like.
U.S. Patent 3,892,824 reports on the toxicity of the compounds falling
within formula I above. Animal studies conducted on these compounds
revealed LD 50's ranging from 450 mg/kg up to 1,300 mg/kg. For the
compound identified as WR 2721 no toxicity problems were encountered at
regular doses of up to 100 mg/kg/day in these toxological tests. In tests
on WR 2721 for its possible side effects, dosages were given to human
volunteers at rates of up to 30 mg/kg/day without any adverse effects
being noted. For this reason compound WR 2721 is the preferred compound
in view of the extensive pharmaceutical tests in connection with its use
as an antiradiation drug which have been performed to date.
Having generally described this invention, a further understanding can
57774~
: '
.
be obtained by reference to certain specific examples which are provided
herein for purposes of il1ustration only and are not intended to be limiting
unless otherwise specif,ied.
. ~I
EXAMPLE 1 ,
In the following examples, the following abbreviations have been
used:
GSSG - reduced glutathione
, GSH - oxidized glutathione
SDS - sodium dodecyl sulphate
DTE - dithioerythritol
DMS - dimercaptosuccinic acid
~P - N-2-mercaptoethyl-1,3-d;am;nopropane
MDP was prepared by boiling WR 2721 in 1 M HCl under nitrogen for
. -fiYe minutes. The solution was neutralized with sodium ~icarbonate.
Sputa were obtained by postural drainage from a 7-year old male
patient wlth a confirmed diagnosis of cystic fibrosis. .
. . ''.
! Samples were collected in jars containing ~.S ml of injectable
. gentamycin sulfate. Upon receipt the samples were added to 0.1 ml of a 1 M
sodium a~ide solution and processed within 24 hours of collection.
9 _
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1157774
Isolation and Electrophoretic Characterization of Sputum Components:
A 10-ml aliquot of whole cystic fibros1s sputum was applied to a
column (4 x 70 cm) of Bioge)~A5M (BioRad Corporation, Richmond, CA~ ¦
equilibrated with 0.01 M phosphate buffer (pH 7.0) containin~ 0.1 M NaCl
and 1 mM sodium azide and eluted with the same buf~er. The material .
appearing in the void volume was pooled and designated component I.
After isolation on Biogel A5M, component I was passed successively
through 0.8, 0.5 and 0.22 ~I filters (Millipore Corporation, Bedford, MA) to
el;minate any possible bacterial contamination.
Component I was concentrated to 5 ml by volume dialysis in tubing o~
5/8" diameter (SGA Scientific, Bloomfield, NJ) and added to a final
r~
concentration of 4% SDS. This sample was washed through an Arnicon~X~llOOA
filter (Amicon, Lexington, MA) under N2 pressure with 500 ml of 0.01 M
¦ phosphate buffer (pH 7.0) containing 0.1 M NaCl and 4% SDS. The material
retained by the filter was designated component IA and was reconcentrated
by vacuum dialysis for use in subsequent experiments.
. SDS-Polyacrylamide gradient gel electrophores;s (5-16% acrylamide)
was performed by the method of Maizel, J.V. (1971) Methods Virol. 5:179.246.
Gels were stained with either Coomassie blue; Fairbanks, G;, T.L. Steck,
D.F.H. Wallach, (1971) Biochem. 10:2606-2617; or periodic acid-Schiff.
reagent; Zacharias, R.J., T.E. Zell, J.H. Morrison, J.J. Woodlock, (1969)
Anal, Biochem. 31:148-1~2.
.'
~ ' I
. ' I
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1 1S7774
Purification and characterization of Component IA:
SDS-polyacrylamide gel electrophoresis was performed
on component IA. The major constituent of component IA has
an apparent molecular weight of 425KD. A number of other
constituents, one of which (60KD) is the main contaminant,
remain after the Amicon-SDS diafiltration, Efforts to remove
these contaminants have not been successful.
Following treatment of component IA with reducing agents
gel electrophoresis revealed several new Coomassie blue staining
bands at 65KD (65,000 Daltons) and 27 KD. These peptides were
apparently split from the large molecular weight mucin (425 KD)
- since PAS staining of another gel revealed a PAS positive band
in the same area as the original mucin.
VISCOMETRY:
The viscosity of crude sputum specimens before and after
the addition of sulfhydryl agents was measured in a 0.2 ml
pipette by determining the time required for 0.08 ml to run
out. Purified mucin samples were concentrated by vacuum
dialysis and dialyzed against 0.01 M phosphate buffer (pH 7.0)
for several days. Their viscosity was then measured using a
Beckman Low-Shear Rotary Viscometer (Model 250010). The
viscosity of fibrinogen solutions of various concentrations
was similarly measured. All viscometric studies were performed
at room temperature. Changes in viscosity with the addition
of 5mM DTE were measured by adding 50 ~ of a 500 mM aqueous
solution of DTE to 5 ml of sample.
d~ 11 -
1 157774
Viscosity of Sputum, Component I and Component IA:
The relationship of protein concentration to viscosity
is shown in Figure I for components I, IA and fibrinogen, a
large molecular weight glycoprotein. The effect of ~mM DTE
on whole sputum, component I and component IA is shown in
Figure 2. Both purified mucin fractions behave like sputum
with respect to viscosity changes upon addition of DTE.
Changes in sputum vicosity upon the addition of various
agents are shown in Table I. Of the sulfhydryl compounds
tested, DTE was the most efficacious in vitro. me oxidized
forms of glutathione and lipoic acid were very much less
active than their corresponding thiols, oxidized lipoic
acid being totally ineffective in reducing sputum viscosity
in vitro. Since DTE might be acting as a metal chelator,
and as a result of early reports the high concentration of
EDTA could reduce sputum viscosity, nonsulfhydryl chelators
were assayed in this system and found to be without effect.
Addition of 5 mM MDP to cystic fibrosis sputum reduced
the viscosity by 70% in 15 minutes as shown in Figure 3. The
parent thiophosphate (WR 2721) had no effect on sputum viscosity.
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X
I 157774
TABLE I
SHORT-TERM VISCOSITY CHANGES OF
WHOLE CYSTIC FIBROSIS SPUTUM
1 mM AgentMinimum Rel. Time
Added Viscosity * (min.)
DTE 21% 10
Antabuse 50% 10
Dimercaptosuccinate 50% 15
GSH 33% 40
GSSG 67% 15
Lipoate 100% 60
Dihydrolipoate 36% 10
D-penicilllamine 32% 20
WR 2721 30% 15
EDTA 100% 60
EGTA 100% 60
H202 100% 60
* Expressed as % of control viscosity.
X - 13 -
1 157774
Example 2
Tissue concentrations of MDP were determined at
various times after intraperitoneal and oral administration of
WR 2721 to mice. Lung and liver homogenates obtained by 5 minutes
of hand bouncing in 1 ml saline, or 1 mm thick tissue slices,
were incubated with 1 mg/ml of WR 2721 at 37C. Aliquots were
taken at various times for assessment of conversion to MDP.
Blood was obtained by retro-orbital puncture, the mice sacri-
ficed with CO2, and the lungs removed. The lung and liver
homogenates (0.9% in saline w/v) and blood samples were treated
with trichloroacetic acid to a final concentration of 10%,
and allowed to stand at 4C for ten minutes. Samples were
centrifuged at 3000 rpm for ten minutes and the supernatant
neutralized with sodium bicarbonate. These neutralized super-
natants were assayed for the presence of sulfhydryl groups by
the method of Ellman et al, (1959) Arch. Biochem. Biophys. 82:
70-77.
,Metabolic Studies of WR 2721:
As is shown in Figure 4, WR 2721 appears in the blood
of mice as the free thiol after both oral and i.p. administra-
tion. The kinetics of appearance of MDP in the lungs of mice
are similar to those for blood after oral (not shown) and i.p.
administration.
Oral administration of WR 2721 leads to a rapid
appearance of MDP in the blood and lungs, whereas parenteral
administration is followed by a rise in MDP concentration 24
hours later.
Homogenates of mouse lung, liver and small intestine
incubated with 1 mg/ml of WR 2721 converted all of the compound
; 30 to MDP within 30 minutes:
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1 157774
.-
1.
. '',
thus, demonstrating in v;vo conversion of WR 2721 to its active~ free
thiol derivative, for the reduction of mucin viscosity
The administration of compounds containing protected sulfhydryl
groups may be used in the treatment of any condition ~herein excessive
mucin viscosity is present. Such conditions include cystic fibrosis,
pneumonia, bronchitis, the common cold, mucin impaction of gastrointestinal
tract, pancreas, liver, and the like.
,
. . '.
. . ''(;~
.
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~ 157774
TABLE II
2-Aminoalkylthiosulfuric Aci~s
R
H2NCCH2SS03H
R2
.,
WR No. Rl R2 ALD
mq/kg
361 H H 625
1484 CH3 H 450
1488 CH3 CH3 800
1826l9 C2H5 H
2108 C2H5 CH3 700
1903 CH3(CH2)2 (DL) H 325
1904 CH3(CH2)2 (L) H 250
2085 CH3(CH2)2 (D) H 250
2176 CH3(CH2)2 CH3 300
4619 (CH3)2CH H 500
2293 (CH3)2CH CH3 >400
1663 CH3~CH2)3 H 175
2177 CH3~CH2)3 CH3 300
1661 (CH3)2cHcH2 H 200
2178 (CH3)2CHCH2 ~ CH3 240
1662 CH3CH2CH(CH3) H 250
2649 -(CH2~4- 800
2579 -(CH2)5- 300
2381 CH3(CH2)5 CH3 180
1112 C6 5CH2 H 250
- 16 -
I
1 157774
TABLE III
2-Aminoalkylphosphorothioates
IR~
2N ICCH2S 03H2
R2
WR No. Rl R2 ALD50
mg/kg
~,
638 H H 525
4468 CH3 H 700
1490 CH3 CH3 750
4470 C2H5 H 320
1961 CH3CH2 2 H 375
5137 (CH3)2CH H 6vû
2378 (CH3)2CHCH2 H 250
3311 (CH2)5 300
2819 CH3 2 2 CH3 >400
. . I
1 1S777~
TABLE IV
2-Thiosulfatoalkylamines
. R
H2NCH2CHSSP3H
WR No. R ALD50
mg/kg
. -- ~
361 H 625
2516 CH3 >800
1036 C2H5 500
1090 CH3CH2CH2 . 188
1081 CH3(CH2~3 150
1144 CH3(CH2)4 150
1250 CH3~CH2)7 300
1734 C6H11 150
769 C6H5
1446 C6H5CH2 88
. I
.
I
. ~ i 157774
TABLE V
2-Aminoethylphosphorothioates
R~
H2NCCH2SP3H2
R2 ''
WR No. Rl R2 ALD50
mg/kg
11439 H2NCH2 H 43
176241 CH20H CH2~H >1250
194423 H2NCH2CH2 (L) H >1250
179209 H2N(CH2)3 ~D~) H >1000
187094 H2N~CH2)3 (L) H > 800
176542 H2N(CH2)4 (DL) H 900a
187093 H2N(CH2)4 (L+) H 275
183977 H2N(CH2)5 H 500
. I
a Oral ad~inistration
~ _19_
I _ ~,
I
~ 15777~
TABLE YI
Al kylaminoethanethiosulfates
RNHCH2CH2SSOH3
WR No. R ALD50
mg/kg
361 H 625
258 CH3 350
1095 C2H5 S25
1033 CH3(CH2)2 350
2244 CH3(CH2)3 300
2341 CH3(CH2)4 200
2137 CH3(CH2)5 100
2078 . CH3(CH2)6 125
1618 CH3(CH2)7 125
t818 - CH3(CH2)8 40
1607 CH3(CH2)9 13
2111 CH3( CH2)10 8
2080 CH3(CH2)11 10
2236 CH3(CH2)12 10
Z281 CH3(CH2)13 . 10
2345 CH3(CH2)14 40
241 2 ~ C~3(CH2?15 . ` ; . 150
23 56 CH3(CH2)16 ~ 200
2362 CH3(CH2?17 . 400
. ...
. - 20 -
1 157774
TABLE YII
Branched - Alkylaminoethanethiosul~uric Acids
RNHCH2CH2SS03H ,
. ',
WR No. R ALD50
mg/kg
259 (CH3)2CH 270
874 (CH3)3C 250
8165 (CH3)3CCH2 475
2856 (CH3)2CHCH2CHCH3 175
2231 CH3(CH2)4CHCH3 175
1606 CH3(CH2)5CHCH3 138
2079 CH3(cH2)3cH(c2Hs)cH2 180
2246 CH (CH ) CHCH CH 125
2402 CH3(CH2~3CHCH2CH2CH3 140
2690 (C2H5)3CCH2 121J
(CH3)3CcH2c(cH3)2
2077 (CH3)2CH(cH2)5cH2 140
2386 - CH3(CH2)4CHCH2CH2CH3 200
2926 CH3(CH2)5CHCH2CH3 125
9528 (CH3)3CCH2CH(CH3)CH2CH2 100
2245 CH3(CH2)7CHCH3 25
2709 . CH3(CH2)6cHcH3 50
2390 CH3(CH2)6CHCH2CH3 100
3358 (CH3)2CH(CH2)3CH(CH3)CH2CH3 125
2411 CH3(CH2)8 3 20
21 -
Il ` .
. I
157774
TABLE VIII
Phenalkylami~noethanethiosulfuric Acids
A ~ (cH2)n-NHcH2cH2sso3H
W~ No. n A
3552 -- H ~800
2230 CH2 H 250
4701 CH2 4-oCH3 180
2456 (CH2)2 H 150
2229 (CH2)3 H 125
3300 (CH2)3 2-OCH3 200
2144 (CH2)3 3-OCH3 300
454?. (CH2)3 4-OCH3 200
2754 (CH2)4 H 300
71462 (CH2)4 4-Br 43
3086 (CH2)4 4-CH3 15
. 3337 (CH2)4 2,5-(CH3)2 20
3340 (CH2)4 2,4-(CH3)2 38
3342 (CH2)4 4-C2~5 15
4102 (CH2)4 2-OH 300
3050 (CH2)4 4-oCH3 175
4109,(CH2)4 3-OCH3 150
3338 (Ch2)4 3,4-(oCH3)2 350
4103 (CH2)4 4-oC2H5 22
3611 (CH2)4 4-C6Hll 15
6138CH(c2Hs)cH2 3-OC~3 100
.
- 22 -
I 1 157774
TABLE V I I I
(Conti nued )
WR No. n A
5983 CH(C2H5)CH2 4-OCH3 110
120767 CH2CH ( C2Hs ) 4 -OCH3 130
4114 CH(c2H5)cH2 H 125
7468 CH(c2H5)cH2 4-C1 140
3427 (CH2)5 H . 140
4548 (CH2)S 4-OCH3 45
3313 (CH2)6 H 38
4783 (CH2)6 4-OCH3~ 40
976~ (C~2)~ 5
':
.
. ),
- 23 -
l ~
1 157774
TABLE IX
Hydroxyalkylam~noethanethiosulfuric Acids
RNHcH2c~l2s~io3H
WR N0. R
3124 HOCH2CH2 1200
3095 HOCH2CH2CH2 - 700
2997 CH3CHOHCH2 - 600
2g86 HocH2cHoHcH2 1750
3569 (HCH2)2cH 1750
2905 HOCH2CHOHCHCH20H1500
3273 HocH2cHoHcHoHcH21800
3093 (CH3)2C(OH)CHOHCH21400
2970 H0CH2 (CH2)6 550
2971 ! H0CH2(CH2)7 625
2865 CH3~CH2)5CHCH2~ 140
2908 HocH2~cH2)8 3
. 2907 H0CH2 (CH2) 9 > 200
2909 C~3 (CH2) 7C~C~20H25
3024 HOCH2 (CH2)8CHOHCH2
27748 HOCH2CHOH(CH2)9 400
,,
.. ,
. '.
.. ` I ~ :
1 157774
TABLE X
Hydro~yalkylaminoethylphosphorothioates
. RNHCH2CH2sPo3~2
WR NO.
8186 HOCH2CH2 950
8192 CH3CE~OHCH2 1250
8187 HOCH2CHOHCH2 2000
27736 CH3CH2cH2(cHOH)2cH2 450
27077 (CH3)2CH~CHOH)2CH2 275
42597 (cH3cH2)2cHcHoHcH2 240
25077 CH3(CH2)4(CHOH)2CH2
36978 (cH3)3ccH2cH(cH3)cHoHcH2 125
52114 CH3(cN2)4cHoNcH2cNoNcN2 1800
., ~ ,
Ij 1 157774
~ - '
TABLE XI
Hydroxyalkylaminoethyldisulfldes
RNHcH2cH2sscH2cH2NHR
WR N0. R
3123 HOCH2CH2 800
3097 HOCH2CH2CH2 500
29563 HOCH2CHOHCH2 1250
3568 (H0CH2)2CH 1250
39748 HOCH2(CH2)4 600
36951 HOCH2(CH2)5 220
4853 CH3(cH2)4cHoHcHoHcH2(rac. 1) 75
5027 CH3(CH2)4cHOHc~oHcH2(rac. 2) 75
30462 H0CH2CHOH(CH2)g 175
3331 CH3(CH2)7(CHOH)2(CH2)8 133
~ ~¦ 3333 CU3(CH2)sCU03(C~2~11 133
''.
. ''`,
~`~ ~
11S7774
TABLE XII
Alkoxyalkylaminoethanethiosulfuric Acids
R-0-(CH2)nNHCH2CH2~S03H
WR No. R n ALD50
mg/kg
3604 HOCH2CH 2 900 .
3607 HOCH2CHOHCH2 2 . 2000
983 CH2 = CH 2 63
CH3 4 375
159244 CH3(CH2)~ 5
3070 CH3 8 150
2991 CH3 9 80
. - i
.1
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II -~
. I ,
1 157774
TABLE XIII
Cycloalkyloxyaminoethanethiosulfuric Acids
R-0 -(cH2)nNHcHzcH2sso3H
ALD50 ORAL DATA
K N0. R n .
3069 (CH2)5CH 3 lS0
4111 (CH2)4CH 4 150
2972 (CH2)5CH 4 150 1500
4444 (CH2)6CH 4 lS0
4446 (CH2)7C~ 4 250
4541 (CH2)4C~ 5 150 1300
2973 (CH2)5CH 5 lS0 >800
4538 2-CH3C6Hlo . 75 1100
~781. 1-CH3C6Hlo lS0 1000
4540 4-CH3C6Hlo 5 18 1300
_~8415 2-(CH3)2CCH 5 CP3 60 >900
4542 ~CH2)6CH 5 35
4545 (CH2)4CH 6 200 1800
2990 (CH2)5CH 6 150
4544 (CH2)5CH 8 15
3610 (cH2)scHcH2cH2 4 38
5331 .(CH2)5CHCH(Cz~5~ 96 >600
3567 (C~2)5CHCH2 5 18
3716 (CH2)7CH 5 180
.. __.
. - 28 -
1 157774
TABLE XIV
Phenoxyalkylaminoethanethiosulfuric Acids
. A ~ -(CH2)nNHCH2CH~SSO3H
.INTRAPERITONEAL DAT~ ~ ORAL DATA
WR_NO _ A n _............. _
2146 H 2175
4381a H 2200
3071 3-C1 2lS0
3240 2-C2H5O 2150
4543b H 2700
3237 H 3150
5146a H 3175
~301 3-CH3 3125
5979 2-CH3 3150
3305 H 4>250
3431a H 4 75
. 3087 2-CH3 4150
2941 3-CH3 4175 1050
3121 4-CH3 . 4 35 1400 .
3565 4-C2H5 4 40
33712 2-(CH3)2CN .4 320
4537 4-C6Hll j 4 50
3830 4-CH30 4200 ~2000
3828 4-C2H5O 4 95 ~ 100
1 157774
TABLE XIV
( Con ti nued )
. INT~A~ERITONEAL DATA ORAL D~TA
,;R No. A n - ALD50 ~ ALD50
5981 4-Br 4 >150
4106 4-C1 4 175 _
3819 3-C1 4 150 ~1250
4792 2-C1 4 150 >3200
111489 3-(C2Hs)2N 4 175 > 900
4447 4-N02 4 150 ~2400
3335 3-CF3 4 45 ~1200
4769 2,3-(CH3)2 4 22
3336 2,4-(CH3)2 4 20 '1000
4805 2,5-(CH3)2 4 40 >2000
478.8 2,6-(CH3)2 4 250 >1500
3343 3,4-(CH3)2 4 45 >1200
124940 3-CH3-4-(CH3)2N - 4 150
75234 3-(CH3)2N-4-~H3 4 . 180 > 900
6137 3-CH3-4-CH3S 4 187
150640 3-NH2-4-CH3 4 150 . > 300
3122 2,4-C12 4 125 >2000
43899 2,4-~(CH3)2CH)2 4 75
3832 2-CH30-4-C2H5 4 125 ~2400
36958. 2~3,6-(CH3~3 4 ~ >100 .
39750 2-Cl-4-Br 475
40640 2-(CH3)3C 438 _
3822 H 5 ~ 50 >2000
- 30 -
_ . ,
1 157774
TABLE XIV - .
( Conti nued )
. -INTRAPERITONE~L VATA ORAL DATA
~LD50 - ALD ~~~
NO. A n mg/kg
3826a H 5 300 _
4S46 2-CH3 5 200 >3200
¦ 136181 3-(CH3)2N-4-CH3 5 135 _
3821 H 6 500
4777 2-CH3 6 150 >1500
~'
. ,1
. The phenylthio analog
'~. 2-Naphthyloxyethylaminoethanethiosulfuric acid
. . i.
~'`
1 157774
TABLE XV
Phenoxyalkylaminoethyldisulfides
[ ~o--(C112)nNHC~2c1l2s~2
WR N0. A n _
6368 H ` 3 120
5141a H 3 lS0
33895 4-CH3 4 430
36159 2,5-(CH3)3 4 45 '.
33714 2~6-(cH3)3 4 S60
. 40644 2,3,6-(CH3)3 4 56
43893 2-(C~3)2CH 4 100
40643 4-CH30 4 430
33896 2-CH3 6 74
. a. The phenylthio analog .
1 157774
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1 157774
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TABLE XYI I
Cycloalkylalkyla~inoethyldisulfides
. ~ ' . I
~R-(CH2)n-NHCH2cH2s-12
AIJU50
WR N0. R n
692 (CH2)5CH 0 92
48680 2-bornyl . 0 180 I,
148667 l-adamantyl 1 75
196267 4 H2NCH2C6H10 1 60
3023 (CH2)5CH 4 22
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, ',
1 157774
.
. .
TABLE XVIII
Cycloalkylalkylaminoethylphosphorothioates
R-(CH2)n-~cH2cH2sPo3H2
ALDso
WR No. R n D~/kg
40335 2-bornyl 0 430
35979 l-adamantyl 0 240
12;696 2-adamantyl 0 150
133567 (CH2)11CH 0 70
74171 2-nrobornyl 0 240
142076 l-adamantyl 2 70
4481 4-CH3-2(=0)c6H8 C(CH3)2 375
151321 l-adamantyl 3 115
159742 (CH2)5CH 4. 45
117909 (CH2)5CH CH(C2H5)CH2 125
4480 2(=O)C6Hg CH(n-C3H7)88
: 75235 4-C6Hll-C6Hll >1800
.
. , i,
I1 157774
TABLE XIX
2-Pyri~3yloxyalkylaminoethanethiosulfuric Acids
A
O (cH2)nNHcH2cH2sso3~
.INTRAPERITONEAL DATAORAL DATA
17R NO. _ A n
37679 ~l ,2
104704 5-C1 2150
99590 S-Br . 2175
98080 H 3l9S
80859 3-~1 3160
122961 5-C1 3150 800
~4203 5-Br 3125 ~ 900
: 91493 3-N02 3175
;91489 5-N02 3180
80865 6-OC2H5 3105
108504 5-C1 4190 >1200
.104695 5-Br 4180 > 900
82422 5-Cl- 5225 ~1000
138412 3,5-C12- 5175 900
99582 3-Br 5170
43900 5-Br 590
. 50480 6-Br 5140
99581~ 3,5-Br2 S160 ~1200
99575 5-I 5100
7931 5-CN 5175
. - 40 -
,_ ,
1 157774
TABLE XIX ~continued)
i
2-Pyridyloxyalkylaminoethanethiosulfuric Acids
~ ~ 0(CH2)nNHCH2SSO3H
INTRAPERITONEAL DATA ORAI DATA
WR NO. A n mg/kq
104696 5-NO2 5 150 ~ 900
85565 4-CH3 5 185
94533 5-CH3 5 125 ~900
106162 5-C1 6 120 >800
91494 5-Br 6 225
1 124950 3,5-Br2 6 250
i 145723 6-Br 7 50
146105 5-1 7 100
94531 5-Br 10 160
- 41 -
1 157774
TABLE XX
2-QuinolyloxyalkylaminoetHanethiosulfuric Acids
A ~ (cH2)n~cH2c~2sso3H
INTRAPERITONEAL DATA ORAL DATA
.._
WR No. A n ~ - ..
104700 4-CH3 2280
122960 4-CH3,6-OCH3 2100
106156 H . 3220
¦25930a H 34SO
9463b H 3125
122968 4-OCH3 3175 . .
54204 4-CH3 3180 >900
134784 4-CF3 3160
120769 4-CH3,8-C1 3300
.108502 4-CH3,6-C1 3 75 >600
. 111490 4-CH3,6-OCH3 3100
122970 4,6-(CH3)2 3125 >900
126456 4-C3H7 3125 >600
104706 4-CH3 4225 >750
111486 4-Cl. 5275
122959 4-OCH3 5225 >900
42761 .4-CH3 5740
123953 4,6-(CH3)2 5115
132199 4-CF3 5250
.1492 4-CH3,6-C1 5 ¦ 175
- 42 -
'
~1 ` ` 1 157774
TABLE XX
( Conti nued )
. INTRAPERITONEAL DATA
. _ .
ALD50
W~ No. A n mg/kg
120768 4-CH3,8-C1 5325
i~4951 4-C3H7 5120
369S5c 7-Cl S250
1~;1491d H 5175
106157 4-CH3 6 90
._ _ !
2 5-Quinolyloxypropylaminoethanethiosulfu-ic acid.
b 8-Quinolyloxypropylaminoethanethiosulfuric acid.
7-Chloro-4-quinolyloxypen~ylaminoethanethiosulfuric acid. I
d 1 ~oquino1yloxypenty1a=iooethanethio~u1furic acid.
.
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1 157774
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TABLE XXIII
Parent Acetamidine Derivatives
H2NC ~ =1~ ) CH25Y
INTRAPERITONEAL DATA ORAL DATA
ALD50 ALD50
WR NO. Y mq/kq mq/kq
76842 H 52 150
¦166817 SCH2C(----NH)NH2 60 125
1551 SO3H 87 300
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