Note: Descriptions are shown in the official language in which they were submitted.
~35 1/~
:~L321~72r~
PHARMACEUTICAL COMPOSITIONS CONTAINING
CiEMlNAL DIPHOSPHONATES
James J. BenedTct
Chris~opher M. Perkins
,
TECH~NiCAL FIELD
This invention relates to pharmaceutical compositions con-
10 taining compounds which are useful in treating or preventingdiseases characterized by abnormal calcium and phosphate metab-
olism, in particular those which are characterized by abnormal
bone metabolism. This invention further relates to a method of
treating or preventing diseases characterized by abnormal calcium
and phosphate metabolism using pharmaceutical compositions of the
present invention.
BACKGROUND OF THE INVENTION
A number of pathological conditions which can afflict warm-
blooded animals involve abnormal calcium and phosphate metab-
20- olism. Such conditions may be divided into two broad categories.
1. Conditions which are characterized by anomalous mohi-
lization of calcium and phosphate leading to general or specific
bone loss or excessively high calciurn and phosphate levels in the
fluids of the body. Such conditior s are sometimes referred t~
25 herein as pathological hard tissue demineralizations.
2. Conditions which cause or result from deposition of
calcium and phosphate anomalously in the body~ These conditions
are sometimes referred to herein as pathological calcifications.
The first category includes osteoporosis, a condition in
30 which bone hard tissue is lost disproportionateiy to the devel-
opment of new hard tissue. Marrow and bone spaces become
larger, fibrous binding decreases, and compact bone becomes
fragile. Osteoporosis can be subclassified as menopausal, senile,
drug induced (e.g., adrenocorticoid, as can occur in steroid
35 therapy), disease induced te.g., arthritic and ~umor), etc.,
however, the manifes~ations are essentially the same. Another
condition in the first category is Pagetls disease (osteitis de-
~3~ 2~
-- 2 --
formans). In this disease, dissolution of normal bone occurs
which is then haphazardly replaced by soft, poorly mineralized
tissue such that the bone becomes deformed from pressures of
weight bearing, particularly in the tibia and ~emur. Hyperpara-
thyroidism, hypercalcemia of malignancy, and os~eolytic bone
metastases are conditions also inclu~ed in the first category.
The second category, involving conditions manifested by
anomalous calcium and phosphate deposition, includes myositis
ossificans progressiva, calcinosis universalis, and such afflictions
as arthritis, neuri~is, bursitis, tendonitis and other inflammatory
conditions which predispose involved tissue to deposition of
calcium phosphates.
Polyphosphonic acids and their pharmaceutically-acceptable
salts have been proposed for use in the treatment and prophyl-
axis of such conditions. In particular diphosphonates, like
ethane-1-hydroxy~ diphosphonic acid (EHDP), propane-3-
amino-1-hydroxy-1,1-diphosphonic acid (APD), and dichloro-
methane diphosphonic acid ~CI2MDP) have been the subject of
considerable research efforts in this area. Paget's disease and
heterotopic ossification are c urrently success~ully treated with
EHDP. The diphosphonates tend to inhibit the resorption of bone
tissue, which is beneficial to patients suf~ering from excessive
bone loss. However, EHDP, APD and many other prior art
diphosphonates have the propensity of inhibiting bone mineral-
ization when administered at high dosage levels.
It is believed that mineralization inhibition is predominantly a
mass related physico-chemical effect, whereas resorption inhibition
results from a biological interaction with the cells. It is therefore
desirable to develop more biologically potent diphosphonate com-
pounds that can be administered at low dosage levels which cause
little or no mineralization inhibition, thereby resulting in a wider
margin of safety. Low dosage levels are also desirable to avoid
the gastro-intestinal discomfort ~like diarrhea) sometimes associ-
ated with oral administration of large quantities of ciiphospho-
nateS~
It is therefore an object of this invention to provide high
potency compositions for the treatment and prophylaxis of ab-
1 3 ~
normal calcium and phosphate metabolism. It is a still further
object of this invention to provide an improved method for
treating diseases characterized by abnormal calcium and phosphate
metabol ism .
BACKGRO_ND ART
U.S. Patent 3,683,080, issuecl August 8, 1972, to Francis,
discloses compositions comprising polyphosphonates, in particular
diphosphonates, and their use in inhibiting anomalous deposition
and mobilization of calcium phosphate in animal tissue.
Japanese Patent 80-98,193, issued July 25, 1980, to Nissan
Kygaku Kagyo K . K. discloses pyridyl ethane diphosphonic acid,
S-(pyridyl)-thiomethane diphosphonic acid, and the derivatives
with halogen or alkyl group substitution on the pyridyl ring.
These compouncls are used as post-emergence herbicides.
Japanese Pa~ent 80-98,105, issued July 25, 1980, to Nissan
Chemical Industries, discloses N-t3-pyridyl)-aminomethane di-
phosphonic acid, and the derivatives with halogen or alkyl group
substitution on the pyridyl ring, for use as herbicides. Various
N-(pyridyl)-aminomethane diphosphonates are also disclosed in
West German Patent 2,831,578, issued February 1, 1979 to Fumio,
for use as herbicides.
European Patent Application 100,718 ~Sanofi SA), published
February 15, 1984, discioses various alkyl diphosphonates which
are -substituted by a sulfide attached to a 5- or 6-membered
nitrogen- or sulfur-containing heterocycle. These compounds are
used as anti-inflammatory and anti-rheumatic drugs.
British Patent Application 2,004,888, published April 11,
1979, discloses N-(3-methyl-2-picolyl)-aminomethane and related
compounds for use in herbicidal compositions.
W. Ploger et al., Z. Anorg. Allg. Chem., 389, 119 (1972),
discloses the synthesis of N- (4-pyridyl)-aminomethane diphos-
phonic acid. No properties or utility of the compound are dis-
closed .
SUMMARY OF THE INVENTION
The present invention relates to pharmaceutical compositions
comprising:
(a) from about 0. 001 mg P to about 600 mg P of a geminal
~32~7~,~
diphosphonic acid compound, or its pharmaceutically-acceptable
salt or ester, in which the diphosphonic acid-containing carbon is
Iinked directly, or via a chain of length from 1 to about 5 atoms,
to a 6-membered aromatic ring containing one or more nitrogen
5 atoms with the parts of said compouncl being comprised as ~ollows:
- said ring may be unsubstituted or substituted with one or
more substituents selected from the group consisting of substi-
tuted and unsubstituted alkyl (saturated or unsaturated) having
from 1 to about 6 carbon atoms, substituted and unsukstituted
10 aryl, substituted and unsubstituted benzyl, hydroxy, halogen,
carbonyl, alkoxy, nitro, amido, amino, substituted amino, car-
boxylate, and combinations thereof;
- said linking chain may be all carbon atoms, a nitrogen atom
or nitrogen-containing chain, an oxygen atom or oxygen-contain-
15 ing chain, or a selenium atom or selenium-containing chain, with
said chain being unsubstituted or substituted on the nitrogen
and/or carbon atoms, independently, with one or more substituted
or unsubstituted alkyl (saturated or unsaturated) having from 1
to about 4 carbon atoms ,and said nitrogen atom also may be
20 substituted with an acyl group;
- said diphosphonate--containing carbon may be unsubstituted
or substituted with substituted or unsubstituted alkyl ( saturated
or unsaturated) having from 1 to about 6 carbon atoms, sub-
stituted or unsubstituted aryl, substituted or unsubstituted
~5 benzyl, amino, substituted amino, amido, hydroxy, alkoxy, halo-
gen or carboxylate, except where said diphosphonate-containing
carbon is directly bonded to a nitrogen, selenium, or oxygen atom
in the linking chain, then the substituents may be substituted or
unsubstituted alkyl (saturated or unsaturated) having from 1 to
30 about ~ carbon atoms, substituted or unsubstituted aryl, or
substituted or unsubstituted benzyl; and
(b) a pharmaceutical carrier.
The invention further encompasses a method of treating
diseases characterized by abnormal calcium and phosphate metabo-
35 lism, comprising administering to a human or animal in need ofsuch treatment a safe and effective amount of a diphosphonic
acid-containing composition of the present invention.
~ 3 2 ~ ~ 2 r~
DETAILED DESCRIPTION OF THE INVENTION
This invention relates to pharmaceutical compositions, pref-
erably in unit dosa~e form, comprising a pharmaceutical carrier
and a safe and effective amount of geminal diphosphonic acid
5 compounds, or their pharmaceutically-acceptable salts and esters,
in which the diphosphonic acid-containing carbon is linked to a 6
membered aromatic ring containing one or more nitrogen atoms.
Preferred rings are pyridine, pyridazine, pyrimidine, and
pyrazine. Most preferred are pyriMidine, and especially pyri-
10 dine. The rings may be unsubstituted or substituted with one ormore substituents selected from the group consisting of substitut-
ed and unsubstituted alkyl (saturated or unsaturated) having
from 1 to about 6 carbon atoms, substituted and unsubstituted
aryl (e.g., phenyl and naphthyl), substituted and unsubstituted
15 benzyl, hydroxy, halogen, carbonyl ~e.g., -CHO and -COCH3),
alkoxy (e.g., methoxy and ethoxy), nitro, amido (e.g.,
-NHCOCH3)~ amino, substituted amino (e.g., dimethylamino,
methylamino, and diethylamino), carboxylate (e.g., -OCOCH3),
and combinations thereof. The rings may be fused with other
20 rings, e.g., benzene fused with pyridine (e.g., quinoline), and
cyclohexane fused with pyridine (e.g., 5,6,7,8-tetrahydro-
quinoline). Additional substituents could be substituted or
unsubstituted sulfide, sulfoxide, sulfate, or suifone.
The linkage from the diphosphonic acid-containing carbon to
25 the ring may be direct through a single bond, or by a chain of
length of from 1 to about 5 atoms. The chain may be all carbon
atoms, a nitrogen atom or nitrogen-containing chain, an oxygen
atom or oxygen-containing chain, or a selanium atom or selenium-
containing chain. The carbon and nitrogen atoms in the chains
30 may, independently, be unsubstituted or substituted with one ~or
one or two in the case of carbon atoms) substituted or unsub-
stituted alkyl (saturated or unsaturated) having from 1 to about 4
carbon atoms (methyl and ethyl being preferredl. The nitrogen
atoms in the chains may also be substituted with an acyl group
35 te.g., -COCH3). Unsubstituted carbon and nitrogen atoms in the
chain are preferred. Also preferred are chains one atom in
p~
-- 6 --
length, i.e., -CH2-, -NH-, and -O-.
The carbon atom which has the phosphonate groups attached
to it may be unsubstituted (i.e., 3 hydrogen atom~, or sub-
stituted with amino, substituted amino, amido, hydroxy, alkoxy,
halogen, carboxylate, substituted or unsubstituted alkyl (satu-
rated or unsaturated) having from l to about 6 carbon atoms,
substituted or unsubstituted aryl, or substituted or unsubstituted
benzyl. For the compounds in which the phosphonate-containing
carbon is linked to the ring via an oxygen, selenium, or
nitrogen-containing chain, and that oxygen, selenium, or nitrogen
atom is bonded directly to the phosphonate containing carbon,
then the substituent on the phosphonate-containing carbon may be
substituted or unsubstituted alkyl (saturated or unsaturated)
having from l to about 6 carbon atoms, substituted or
unsubstituted aryl, or substituted or unsubstituted benzyl.
Thus, diphosphonic acid compounds to be included in the
pharmaceutical compositions of the present invention have the
structure: -
/R2 \ R2~ P~3H2
R3--Z _ -C--Q---C~C-PO H
R2 m R2 j7nRl
wherein Q is oxygen, -NR4-, selenium, or a single bond,
preferred being oxygen, -NR4-, or a single bond; m + n is an
integer from 0 to about 5, with m + n = 0 or 1 pref~rred for Q
25 being oxygen, selenium, or -N R4-, and m ~ n = l or 2 preferred
otherwise; Z is a ring selected from the group consisting of
pyridine, pyrida~ine, pyrimidine, and pyrazine, with preferred
being pyrimidine, and especially pyridine; Rl is hydrogen, sub-
stituted or unsubstituted amino, amido, hydroxy, alkoxy, halo-
30 gen, carboxylate, substituted or unsubstituted alkyl lsaturated orunsaturated) having from l to about 6 carbon atoms, substituted
or unsubstituted aryl, or substituted or unsubstituted benzyl,
except that when n = 0 and Q is oxygen, selenium, or -NR4-
then Rl is hydrogen, substituted or unsubstituted alkyl (satu-
35 rated or unsaturated) having from l to about 6 carbon atoms,substituted or unsubstituted aryl, or substituted or unsubstituted
132~ 127
-- 7 --
benzyl, with R1 being hydrogen, chloro, amino, methyl, or
hydroxy preferred; each R2 is, independently, hydrogen, or
substituted or unsubstituted alkyl (saturated or unsaturated)
having from 1 to about 4 carbon atoms, with R2 being hydrogen
5 preferred; R3 is one or more substituents selected from the group
consisting of hydrogen, substituted or unsubstituted alkyl (satu-
rated or unsaturated) having from 1 to about 6 carbon atoms,
substituted and unsubstituted aryl, substituted and unsubstituted
benzyl, hydroxy, halogen, carbonyl, alkoxy, nitro, amido, amino,
10 substituted amino, carboxylate, and combinations thereof, with
preferred being hydrogen, methyl, amino, chloro, methoxy, nitro,
hydroxy and combinations thereof; R4 is hydrogen, substituted or
unsubstituted alkyl (saturated or unsaturated) having from 1 to
about 4 carbon atoms, or acyl (i.e., the amide of the nitrogen),
15 with preferred being hydrogen, methyl, or ethyl; and pharma-
ceutically-acceptable salts and esters of these compounds. Fi-
nally, for any of the R1, R2, R3, or R4 substituents which are
themselves substituted, the substitution on these substituents may
be any one or more of the above substituents, preferred being
20 methyl, ethyl, amino, chloro, nitro, metho~y, hydroxy, acet-
amido, and acetate.
More specifically, the diphosphonic acid compounds, and
their pharmaceutically-acceptable salts and esters, to be included
in the pharmaceutical compositions of the present invention are of
25 the structure:
/ R2 ~ l03H2
3 t R ~ R 3 2
/ R2\ / 12\ ~3 2
t R2~ R4t R2~ R1
IR2\ 1 IR2~ 1 3~12
t ~1 t 27~ 1
132~
-- 8 --
wherein m + n, Z, R1, R2, R3, and R4 are as described above.
Generally preferred diphosphonic acid compounds, and their
pharmaceutically acceptable salts and esters, to be inçluded in the
pharmaceutical compositions of the pnesent invention are of the
S structure:
N~H~ C--PO3H2 or
R3~ Ct Cl--PO3H~ ;
wherein for both structures above m + n = 1 or 2; R1 is hydro-
15 gen, chloro, amino, or hydroxy; R3 is one or more substituents
selected from the group consisting of hydrogen, methyl, amino,
chloro, nitro, methoxy, hydroxy, and combinations thereof; or
~ tC~ )3H2 or
R4 H R1
R3~R4tH~ 3H2 r
H ~3H2
R~_ Q--~C --C r P03H2 ; or
R3~_ o ~c~ C _ PO3H2; or
wherein for the four preceding structures n = 0 or 1; Rl is
hydrogen, chloro, amino, or hydroxy when
132~ ~2~
n = 1, and R1 is hydrogen when n = 0; R3 is one or more substitu-
ents setected from the group consisting olF hydrogen, methyl,
amino, chloro, methoxy, nitro, hydroxy, and combinations there-
of and R4 is hydrogen, methyl, or ethyl.
Specific examples of compounds which may be utilized in
compositions o~ the present invention include;
N-(2-pyridyl)-aminomethane diphosphonic acid;
N-(2-~5-amino)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(5-chloro)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(5-nitro)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(3,5-dichloro)-pyridyl)-aminomethane diphosphonic acid;
N-(4-pyridyl)-N-ethyl-aminomethane diphosphonic acid;
N-(2-(3-picolyl))-aminomethane diphosphonic acid;
N-(2-(4-picolyl))-aminomethane diphosphonic acid;
N-(2-(5-picolyl))-aminomethane diphosphonic acid;
N-(2-(6-picolyl))-aminomethane diphosphonic acid;
N-(2-(3,4-lutidine))-aminomethane diphosphonic acid;
N-(2-14,6-lutidine))-aminomethane diphosphonic acid:
N-(2-pyrimidyl)-aminomethane diphosphonic acid;
N-(4-(2,6-dimethyl)-pyrimidyl)-aminomethane diphosphonic acid;
N-(2-(4,6-dihydroxy)-pyrimidyl)-aminomethane diphosphonic acid;
N-(2-(5-methoxy)-pyridyl)-aminomethane diphosphonic acid;
N-~2-pyridyl)-2-aminoethane-1,1-diphosphonic acid;
N-(2-(3-picolyl))-2-aminoethane-1,1-diphosphonic acid;
N-(3-pyridylj-2-amino-1-chloroethane-1,1-diphosphonic acid;
N-~2-(4-picolyl))-2-amino-1-hydroxy-ethane-1 ,I-diphosphonic
acid;
(2-pyridyl)-methane diphosphonic acid;
(3-pyridyl)-aminomethane diphosphonic acid;
(2-pyridyl)-chloromethane diphosphonic acid;
(4-pyridylj-hydroxymethane diphosphonic acid;
2-(2-pyridyi)-ethane-1,1-diphosphonic acid;
2-(3-pyridyl)-ethane-1,1-diphosphonic acid;
2-(4-pyridyl)-ethane-1,1-diphosp~Dnic acid;
2-(2-pyridyl)-1-amino-ethane-1,1-diphosphonic acid;
2-(2-pyrimidyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
132~727
-- 10 --
2-(2-(3-picolyl))-1-chloro-ethane-1,1-diphosphonic acid
2-(2-(4-methoxy)-pyridyl)-ethane-1,1-diphosphonic acid;
1-(2-pyridyl)-propane-2,2-diphosphonic acid:
2-(2-pyridyl)-1-chloro-ethane-1,1-diphosphonic acid;
5 2-(2-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
2- ( 3-py ridy I ) -1 -hyd roxy-ethane- 1 ,1 -di phosphon ic ac id
2-(4-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
3-(3-pyridyl)-1-hydroxy-propane-1,1-diphosphonic acid;
0-(2-pyridyl)-2-oxa-ethane-1,1-diphosphonic acid;
10 0-(2-pyridyl)-oxamethane diphosphonic: acid;
0-(2-pyrimidyl)-oxamethane diphosphonic acid;
0-(2-(4-amino)-pyridyl)-oxamethane diphosphonic acid;
0-(2-pyrimidyl)-2-oxa-ethane-1,1-diphosphonic acid;
0-(2-(3-picolyl))-2-oxa-ethane-1,1-diphosphonic acid;
15 0-(2-(3-picolyl))-oxamethane-diphosphonic acid;
0-(2-pyridyl)-1-hydroxy-2-oxa-ethane-1,I-diphosphonic acid;
0-(4-pyridyl)-1-amino-2-oxa-ethane-1,1-diphosphonic acid; and
pharmaceutically-acceptable salts and esters thereof.
Preferred compounds are
20 N-(2-(5-amino)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(5-chloro)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(3-picolyl))-aminomethane diphosphonic acid;
N-(2-(4-picolyl))-aminomethane diphosphonic acid;
N-(2-(5-picolyl))-aminomethane diphosphonic acid;
25 N-(2-(6-picolyl))-aminomethane diphosphonic acid
N-(2-(3,4-iutidine~)-aminomethane diphosphonic acid;
N-(2-pyrimidyl)-aminomethane diphosphonic acid;
N-(2-pyridyl)-2-aminoethane-1,1-diphosphonic acid;
2-(2-pyridyl)-ethane-1,1-diphosphonic acid;
30 2-(3-pyridyl)-ethane-1,1-diphosphonic acid;
2-(4-pyridyl)-ethane-1,1-diphosphonic acid,
2-(2-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
2- ( 3-pyridyl ) -1 -hydroxy-ethane-1 ,1 -diphosphonic acid;
2-(4-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
35 0-t2-(3-picolyl)) oxamethane-diphosphonic acid; and
pharmaceutically-acceptable salts and esters thereof.
- 132072 1
The diphosphonate compounds to be included in the pharma-
ceutical compositions of the present inventlon can be made using
- the synthetic methods disclosed in Japanese Patent 80-g8,193
(July 25, 1980, to Nissan Kygaku Kagyo K.K.), Japanese Patent
5 80-98,105 (July 25, 1980, to ~lissan ChemTcal Industries), West
German Patent 2,831,578 (February 1, 1979, to Fumio), and W.
Ploger et al., Z. A~. A~ Chem., 389, 119 (1972), The
amlnoethane diphosphonie acid compounds, however, are best
prepared as follows:
Synthesis of N-(2-(3-picolyl)?aminoethane DP
The above-named compound is prepared via a typical Michael
reaction between tetraethyl vinyldiphosphonate and
2-amino-~-picoline. (See H.O. House, Modern Synthetic Reaction
2nd Ed. W.A. BenJamin Inc. p. 595-6~3.
To a solution of 1.62 g (15 mmol) of 2-amino-3-picoltne in
tetrahydrofuran at 5C was added 4. 50 g ( 15 mmol ) tetraethyl
vinyldiphosphonate. The reaction mixture was stlrred at room
temperature for 16 hours. Evaporation of the solvent and
chromatography (acetone/hexane, 4/1~ of the product on silica gel
gave pure tetraethyi N-(2-(3-picolyl~)-2-aminoethane
diphosphonate. P-31 NMR of the pure tetraethyl ester in CDCI3
shows a resonance at 22.1 ppm. The ester was hydrolyzed in
refluxlng 6N HCI overnight. The product ~howed a P-31 NMR
signal in D2O at pH = 12 of 19.0 ppm.
N-(2-pyridyl)-2-aminoethane DP and N-(2-(5-picolyl))-2-
aminoethane i)P were prepared in an identical manner.
Compounds having the general formula
R3_ z ~ ~2~H3HP~3H2
~wherein n is an ineeger of from 1 to about 5, preferably n = 1;
and Z, R2 and R3 are as described hereinbefore, with preferred
Z being pyrimidine and especially pyridine, preferred R2 being
hydrogen, and preferred R3 ~eing one or more substituents
132~7
- 12 -
selected from the group consisting of hydrogen, methyl, amino,chloro, nitro, methoxy, hydroxy, and combinations thereof) are
best prepared as follows:
Synthesis of 2-(2-pyridyl)-1-hydroxy-ethane-1 ,1-diphosphonic
5 acid:
A 3-neck round-bottom flask fitted with a re~lux condenser
and a magnetic stir bar is charged with 6.94 grams (0.04 mole)
2-pyridine acetic acid, 9.84 grams (0.14 mole1 phosphorus acid,
and 150 ml of chlorobenzene. This reaction mixture is heated on
a boiling water bath, and 16.5 grams (0.12 mole) phosphorus
trichloride is added dropwise with stirring. This reaction mixture
is heated for 2-1/2 hours during which time a viscous yellow oil
forms. The reaction mixture is then cooled in an ice bath and
the chlorobenzene solution is decanted off from the solidified
product. The reaction flask containing this solidified product is
charged with 150 ml of water and heated in a boiling water bath
for several hours. The hot solution is then filtered through
Celite 545R. 300 ml of methanol is added to the warm filtrate
solution, and a precipitate develops. After cooling in ice for 1
hour, the precipitate is filtered off and then washed with
methanol/water (1/1 volume/volume), methanol, and ether, and air
dried. The product may be recrystallized from hot water. Yield
is approximately 5.9 grams (52%~. The sample is characterized by
P-31 and C-13 NMR.
By "pharmaceutically-acceptable salts and esters" as used
herein is meant hydroly~able esters and salts of the diphos-
phonate compounds whi~h have the same general pharmacological
properties as the acid form from which they are derived, and
which are acceptable from a toxicity viewpoint. Pharmaceuti-
cally-acceptable salts include alkali metal ~sodium and potassium),
alkaline earth metal (calcium and magnesium), non-toxic heavy
metal ~stannous and indium), and ammonium and low molecular
weight substituted ammonium (mono-, di- and triethanolamine)
salts. Preferrecl compounds are the sodium, potassium, and
ammonium saltsO
By "pharmaceutical carrier" as used herein is meant one or
more compatible solid or liquid filler diluents or encapsulating
~32~7~,7
-- 13 --
substances. By "compatible" as used herein is meant that the
components of the composition are capable of being commingled
without interacting in a manner which would substantially de-
crease the pharmaceutical efficacy of the total cornposition under
ordinary use situations.
Some examples of substances which can serve as pharma-
ceutical carriers are sugars such as lactose, ~31ucose and sucrose:
starches such as corn starch and potato starch; cellulose and its
derivatives, such as sodium carboxymethylcellulose, ethylcellulose,
cellulose acetate; powdered tragacanth; malt; ~elatin, talc; stearic
acid; magnesium stearate; calcium sulfate; vegetable oils, such as
peanut oil, cottonseed oil, sesame oil, olive oil, corn oil and oil of
theobroma; polyols such as propylene glycol, glycerin, sorbitol,
mannitol, and polyethylene glycol; agar; alginic acid; pyrogen-
free water; isotonic saline; and phosphate buffer solutions, as
well as other non-toxic compatible substances used in pharma-
ceutical formulations. Wetting ayents and lubricants such as
sodium lauryl sulfate, as well as coloring agents, flavoring
agents, lubricants, excipients, tableting agents, stabilizers,
anti-oxidants and preservatives, can also be present. Other
compatible pharmaceutical additives and actives (e.g., vitamin D
or vitamin D metabolites, and mineral supplements) may be
included in the pharmaceutical compositions of the present
invention .
The choice of a pharmaceutical carrier to be used in con-
junction with the diphosphonates of the present composi~ions is
basically determined by the way the diphosphonate is to be ad-
ministered . I f the compound is to be injected, the preferred
pharmaceutical carrier is sterile, physiological saline, the pH of
which has been adjusted to about 7. ~1. However, the preferred
mode of administering the diphosphonates of the present invention
is oraliy, and the preferred unit dosage form is therefore tablets,
capsules and the like, comprising from about 0.1 mg P to about
600 mg P of the diphosphonic acid compounds described herein.
Pharmaceutical carriers suitable for the preparation of unit dosage
forms for oral administration are weli known in the art. Their
~ ~ 2 ~ P~
-- 14 --
selection will depend on secondary considerations like taste, cost,
shelf stability, which are not critical for the purposes of the
present invention, and can be made without difficuity by a person
skilled in the art. The pharma~:eutical carrier employed in con-
junction with the diphosphonates of the present invention is used
at a concentration sufficient to provide a practical size to dosage
relationship. Preferably, the pharmaceutical carrier comprises
from about 0. 01% to about 99. 99% by weight of the total
composition .
EXAMPLE I
Capsules are prepared by conventional methods, comprised
as follows:
ingredient Mg per capsule
N-(2-(3-picolyl)) AMDP 100 (as mg P)
Starch 55 . 60
Sodium lauryl sulfate 2 . 90
The above capsules administered orally twice daily for 6
months substantially reduce bone resorption in a patient weighing
approximately 70 kilograms afflicted with osteoporosis. Similar
results are obtained when the N-(2-(3-picolyl))-aminomethane
diphosphonic acid, or its pharmaceutically-acceptable salt or
ester, in the above-described capsules is replaced with
N-(2-(5-amino)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(5-chloro)-pyridyl)-aminomethane diphosphonic acid
N-(2-(4-picolyl))-aminomethane diphosphonic acid;
N-(2-(5-picolyl))-aminomethane diphosphonic acid;
N-(2-(6-picolyl))-aminomethane diphosphonic acid;
N-(2-(3,4-lutidine) )-aminomethane diphosphonic acid;
N-(2-pyrimidyl)-aminomethane diphosphonic acid;
N-(2-pyridyl)-2-aminoethane-1,1-diphosphonic acid;
2- ( 2-pyridyl )-ethane-l ,1 -diphosphonic acid;
2-(3-pyridyl)-ethane-1,1-diphosphonic acid;
2-(4-pyridyl)-ethane-1 ,1-diphosphonic acid;
2-(2-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
2-(3-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
~ 3 2 ~
2- ( 4-py ridyl ) -1 -hydroxy-ethane-1 ,1 -diphosphonic acid;
0-(2-(3-picolyl) )-oxamethane-diphosphonic acid or the
pharmaceuticalty-acceptable salts or esters thereof.
EXAMPLE N
Tablets are prepared by conventional methocls, formulated as
fol I ows:
.Ingredient mg per tablet
N-12-pyrimidyl) AMDP 25.00
Lactose 1~0 . 00
10 Starch 2 . 50
Magnesium stearate 1 . 00
The above tablets administered orally twice daily for 6
months substantially reduce bone resorption in a patient weighing
approximately 70 kilograms afflicted with osteoporosis. Similar
15 results are obtained when the N-(2-pyrimidyl) AMDP, or its
pharmaceutically-acceptable salt or ester, in the above-described
tablets is replaced with
N-(2-(5-amino)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(5-chloro)-pyridyl)-aminomethane diphosphonic acid
20 N-(2-13-picolyl~)-aminomethane diphosphonic acid
N- ( 2- ~ 4-picolyl ) ) -aminomethane diphosphonic acid;
N-12-[S-picolyl))-aminomethane diphosphonic acid;
N-12-(6-picolyl))-aminomethane diphosphonic acid;
N-12-(3,4-lutidine))-aminomethane diphosphonic acid;
25 N-(2-pyridyl~-2-aminoethane-1,1-diphosphonic acid;
2-[2-pyridyl~-ethane-1,1-diphosphonic acid;
2-(3-pyridyl)-ethane-1,1-diphosphonic acid
2-(4-pyridyl~-ethane-1 ,l-diphosphonic acid;
2-12-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
30 2-13-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
2- ( 4-pyridyl ) -1 -hydroxy-ethane-1 ,1 -diphosphonic acid;
0-(2-(3-picolyl) )-oxamethane-diphosphonic acid; or the
pharmaceutically-acceptable salts or esters thereof.
~.32~2
-- 16 --
EXAMPLE l l i
Injectable solutions are prepared ~y conventional methods
using 1. 0 ml of either physiological saline or water solution and
3.5 mg o~ 2-(2-pyridyl)-ethane~ diphosphonic acid, adjusted to
5 pH = 7.4.
One injection, one time daily for 4 days results in appreci-
able alleviation of hypercalcemia of malignancy in patients weigh-
ing approximately 70 kilograms.
Similar results are obtained when the 2-(2-pyridyl)-ethane-1,1-
10 diphosphonic acid in the above-described treatment is replaced
with
N-~2-(5-amino)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(5-chloro)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(3-picolyl))-aminomethane diphosphonic acid;
15 N-(2-(4-picolyl))-aminomethane diphosphonic acid;
N-(2-(5-picolyl))-aminomethane diphosphonic acid;
N-(2-(6-picolyl))-aminomethane diphosphonic acid;
N-(2-(3,4-lutidine))-aminomethane diphosphonic acid;
N-(2-pyrimidyl)-aminomethane diphosphonic acid,
20 N-(2-pyridyl)-2-aminoethane-1 ,1-diphosphonic acid;
2-(3-pyridyl)-ethane-1,1-diphosphonic acid;
2-(4-pyridyl)-ethane-1,1-diphosphonic acid;
2- ( 2-pyridyl ) -1 -hydroxy-ethane-1 ,1 -diphosphonic acid;
2-(3-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
25 2-(l~-pyridyl)-l-hydroxy-ethane-l~l-diphosphonic acid;
0-(2-(3-picolyi))-oxamethane-diphosphonic acid; or
pharmaceutically-acceptable salts or esters thereof.
The compositions of the present invention are useful in the
treatment of abnormal caicium and phosphate metabolism. Other
30 diphosphonis acids and their pharmaceutically-acceptable salts
have been proposed for use in the treatment and prophylaxis of
such conditions. In particular, ethane-1-hydroxy-1,1-diphos-
phonic acid (EHDP), propane-3-amino-1-hydroxy-1,1-diphosphonic
acid (APD), and dichloromethane diphosphonic acid (Cl2MDP) have
35 been the subject of considerable research efforts in this area.
However, the compositions of the prese~t invention are
generally more biologically potent in inhibiting bone resorption
than the art-disclosed diphosphonates. Thus, the compositions of
~32~2P~
-- 17 --
the present invention may provide one or more of the following
advantages over the art-disclosed diphosphonates of ( 1 ) being
more potent in inhibiting bor~e resorption: (2) po$sessing less
potential ~or inhibition of bone mineralization, since mineralization
inhibition is believed to be predominantly a mass related
physico-chemical effect; ~3) having ~enerally a wider margin of
safety ( i . e ., wider dosing interval between the lowest effective
antiresorptive dose and the lowest dose producing mineralization
inhibition~ (4) allowing lower oral dosages to be administered,
thereby avoiding the gastro-intestinal discomfort (like diarrhea)
sometimes associated with higher dosages of diphosphonates; and
(5) having potential for flexibility of dosing methods.
Another aspect of this invention is a method for treating or
preventing diseases characterized by abnormal calcium and phos-
phate metabolism, in particutar those which ara characterized by
abnormal bone metabolism, in persons at risk to such disease,
comprising the step of administering to persons in need of such
treatment a safe and effective amount of a diphosphonic acid-
containing composition of the present invention.
The preferred mode of administration is oral, but other
modes of administration include, without limitation, transdermal,
mucosal, sublingual, intramuscular, intravenous, intraperitoneal,
and subcutaneous administration, as well as topical application.
By "abnormal calcium and phosphate metabolism" as used
herein is meant (1 ) conditions which are characterized by anom-
alous mobilization of calcium and phosphate leading to general or
specific bone loss, or excessively hi~h calcium and phosphate
levels in the fluids of the body; and (2) conditions which cause
or result from deposition of calcium and phosphate anomalously in
the body. The first category includes, but is not limited to~
osteoporosis, Pagets disease, hyperparathyroidism, hypercalcemia
of malignancy, and osteolytic bone metastases. The second
category includes, but is not limited to, myositis ossificans
progressiva, calcinosis universalis, and such afflictions as
arthritis, neuritis, bursitis, tendonitis and other inflammatory
13 ~ r~
- 18 -
conditions which predispose involved tissue to deposition of
calcium phosphates.
By "person at risk", or "person in need of such treatment",
as used herein is meant any human or lower animal which suffers
S a significant risk of abnormal calcium and phosphate metabolism if
left untreated, and any human or lower animal diagnosed as being
afflicted with abnormal calcium and phosphate metabolism. For
example, postmenopausal women; plersons undergoing certain
steroid therapy; persons on certain anti-convulsant drugs; per-
sons diagnosed as having Pagets disease, hyperparathyroidism,
hypercalcemia of malignancy, or osteolytic bone metastases;
persons diagnosed as suffering from one or more of the various
forms of osteoporosis; persons belonging to a population group
known to have a significantly higher than average chance of
developing osteoporosis, e.g., postmenopausal women, men over
age 65, and persons being treated with drugs known to cause
osteoporosis as a side effect; persons diagnosed as suffering from
myositis ossificans progressiva or calcinosis universalis; and
persons afflicted with arthritis, neuritis, bursitis, tendonitis and
other inflammatory conditions which predispose involved tissue to
diposition of calcium phosphate.
By "human or lower animal afflicted with or at risk to osteo-
porosis" as used herein is meant a subject diagnosed as suffering
from one or more of the various forms of osteoporosis, or a
subjec~ belonging to a group known to have a significantly higher
than average chance of developing osteoporosis, e.g., postmeno-
pausal women, men over the age of 65, anà persons being treated
with drugs known to cause osteoporosis as a side effect ( such as
ad~enocorticoid ) .
By "safe and effective amount" as used herein is meant an
amount of a compound or composition high enough to significantly
positively modify the condition to be treated, but low enough to
avoid serious side effects (at a reasonable benefit/risk ratio),
within the scope of sound medical judgment. The sa~e and effec-
tive amount of diphosphonates will vary with the particular con-
dition being treated, the age and physical condition of the patient
~ 3 ~ 7
- 19 --
bein~3 treated, the severity of the condition, the duration of
treatment, the nature of concurrent therapy, and the specific
diphosphonate employed. However, single dosages can range from
about 0. 001 mg P to about 3500 mg P, or from about 0.1 micro-
grams P/kg of body weight to about 500 mg P/kg of body weight.
Preferred single ciosages are from about 0.1 mg P to about 600 mg
P, or from about 0.01 to about 50 mg P/kg of body weight. Up
to about four single dosages per day may be administered. Daily
dosages greater than about 2000 rng P/kg are not requlred to
produce the desired effect and may produce undesirable side
effects. The hlgher dosages within this range are, of course,
required in the case of oral administration because of limited
absorption .
Schenk Model
The compounds were evaluated for 'n vivo bone resorption
inhibition and mineralization inhibition in an animal model system
known in the field of bone metabolism as the Schenk Model. The
general principles of this model system are disclosed in Shinoda et
al., Calcif. Tissue Int., 35, 87-99 (1983); and in Schenk et al.,
Caicif. Tissue Res. 11, 196-214 (i973).
Materials and Methods
Animals
Preweaning 17-day-old 130 gms) male Sprague Dawley rats
25 (Charles River Breeding Laboratories~ were shipped with their
mothers and placed in plastic cages with their mothers upon
arrival. At 21 days of age, pups receiving Rat Chow and water
ad libitum were randomly allocated into treatment groups com-
prislng five animais per group, except for control animals re-
30 ceiving saline vehicle which had 10 rats per group. On day 0and again on day 1 all animals were given a subcutaneous in-
jection of Calcein (Sigma~ as a 196 soiution in 0.9~ NaCI solution to
label the skeleton.
Dose Solutions and Dosing Procedure
. . . _ .
All solutions were prepared for subcutaneous injection in
0.9~ normal saline and adjusted to pH 7.4 using NaOH and/or
132~72~
-- 20 --
HCI. Dose solution calculation was made by considering the mass
of powder (based on molecular weight, hydration) of the active
material in mg/kg (body weight) that corresponds to mgP/kg.
Concentrations were based on dosing 0.2 ml/100 g body weightO
Initially, all compounds were administered at 0.1, 1.0 and 10.0 mg
- Plkglday for 7 days. Compounds showing activity at 0.1 mg
P/kglday were then tested at logarithmic decrements down to
0.001 mg Plkg/day. Adjustments in dosage based on changes in
body weight were made on a daily basis.
Necropsy, Tissue Processing and Histomorphometry
On day 8 after the start of dosing, all animals were sac-
rificed by CO2 asphyxiation. Tibias were dissected free and
placed in 70% ethyl alcohol. One tibia was dehydrated in graded
ethanol solutions and embedded in methyl methacrylate using a
rapid procedure described in Boyce et al., Lab. Investig., 48,
683-689 ( 1 983) .
IThe tibla was sectioned longitudlnally through the
metaphyseal area ( LeitzR saw microtome at 150 ,~L ) . Specimens
were stained on one surface with silver nitrate and mounted on
2û microscope slides for evaluation with a Quantlmet Image Analyzer
(Cambridge Instruments, Inc.) using both incandescent and
ultraviolet illumination. Metaphyseal trabecular bone content was
measured in the region between the fluorescent label and the
growth plat~: expressed as percent of total area (bone + mar-
row). Epiphyseal growth plate wldth was obtained as the mean
value of 10 equally-spaced measurements across the section.
Statistical evaluation of data was made using parametrlc and
non-parametric analysis of varlance and Wilcoxons rank sum test
to determine a statistically significant effect compared to control
animals.
The Schenk model provided data for in vivo bone resorption
inhibition by the compounds. The lowest effective [antiresorp-
tive) dose ( LED ~ for representative compounds tested, as
determined by the Schenk model, are provided in Table 1.
1~2~2~
- 21 -
TAB LE
Lowest Effective (Antiresorptive) Dose
Schenk
Diphosphonate Compound LED (mg P/kg~
EHDP 1. 0
Cl2MDP 1. 0
APD 0.1
N-(2-pyridyl) AMDP 0.01
N-(2-(5-chloro)-pyridyl) AMDP 0.01
N-(2-~3-picolyl~) AMDP 0.001
N-(2-(4-picolyl)) AMDP 0.001
N-(2-t5-picolyl)) AMDP 0.001
N-(2-~6-picolyl)) AMDP 0.001
N-(2-pyrimidyl) AMDP 0.001
N-(4-pyridyl~-N-ethyl AMDP 0.1
2-(2-pyridyl) EDP 0.01
2-(3-pyridyl) EDP 0.01
1-(2-pyridyl) propyl DP 10
EHDP = ethane-1-1hydroxy-1,1-DP
Ci2MDP = dichloromethane DP
APD = 3-aminopropane-1 hydroxy-1 ,l-DP
AMDP = aminomethane diphosphonic acid, where the ring is at-
tached to the amine.
* = Compounds inciuded in pharmaceutical compositions of the
25 present invention.
EDP = ethane-1 ,1-c!iphosphonic acid, where the ring is attached
at the 2 position of the ethane.
Propyl DP = propane-2,2-diphosphonic acid
Diphosphonate compounds which have a bone mineralization
30 inhibiting effect cause widening of the epiphyseal growth plate,
since matrix production continues but mineralization is impeded.
The widening of the epiphyseal growth plate as observed in the
Schenk rrodel is, therefore, a measure of the mineralization in-
hibiting effect of the diphosphonate compound tested.
132~7~
-- 22 --
The lowest tested dosages producing a statistically signifi-
cant widening of epiphyseal growth plate for compounds tested
are given in Table ll.
TABLE I I
Mineralization Inhibition (Schenk Mode!l)
- Lowest tested dosage
producing a statistically
significant widening of
Diphosphonateepiphyseal growth plate
10 Compound (m~ P/Kgl
EHDP 10
APD 10
C12MDP .. _
N-12-pyridyl) AMDP 0.1
15 N-(4-pyridyl)-N-ethyl AMDP 1)
N-[2-(3-picolyl) ) AMDP -- 1 )
N-(2-(4-picolyl)) AMDP 0.1
N-(2-(5-picolyl)) AMDP 0.1
N-(2-(6-picolyl) ) AMDP -- 1 )
20 N-(2-pyrimidyl) AMDP 1.0
N-(2-15-chloro)-pyridyl) AMDP__ 1 )
2- ( 3-pyridyl ) ED P --
2-(2-pyridyl ) EDP __ t )
- = No plate widening obserYed at highest dose tested lhighest
25 dose tested is 10 mg P/kg/day unless otherwise indicated~
1 ) = Highest dose evaluated is 1 mg P/kg/day lcompound lethally
toxic at 10 mg P/kg/day)
EHDP = ethane-1-hydroxy-1,1-DP
APD = 3-aminopropane-1-hydroxy-1,1-DP
30 C12MDP = Dichloromethane DP
AMDP = aminomethane diphosphonic acid, where the ring is at-
t3~hed to the amine
l--DP -- etharie-1,1-diphosphonic acid, where the ring is attached
at the 2 position of the ethane
35 * = Compounds included in pharmaceutical compositions of the
present invention
1 3 ~ 7
-- 23 --
Thyroparathyroidectomized ~TPTX?_Rat Model
The compounds were evaluated for 'n vlvo bone resorption
inhibition potency by an animal model system known as the thyro-
parathyroidectomized (TPTX) rat model. The general principles
of this model system are dlsclosed in Russell et al., Calclf.
Tissue_ Research, 6, 183-196 (1970), and in Muhlbauer and
.Fleisch, Mineral Electrolyte Metab., 5, 296-303 (1~81 ), The
basic biochemlcal concept of the TPTX system is inhibltion of the
parathyroid hormone (PTH) - induced rise in serum and ionized
calcium levels by the respective bone active polyphosphonates.
Materials and Methods:
Materials
Low calcium and low phosphorous diets used were prepared
by TekladR Test Diets ( Harlan Industries, Madison, Wisconsin
53711; Order #TD82195) in a pellet form of approximately 0.18%
calcium and 0. 22~ phosphorous. The diets contained all the
essential vitamins and mlnerals required for the rat, with the
exception of calcium and phosphorous. The calcium and phos-
phorous levels of the pellets were verified analytically ( Procter
Gamble Co., Miami Valley Laboratories, Cincinnati, Ohio).
PTH was acquired as a powdered bovine extract (Sigma
Chemical Co., P. O. Box 14508, St. Louis, Missouri, order
#P-0892, Lot #72F-9650) at an activity of 138 USP units per mg.
PTH was prepared in 0.~% saline such that the final concentration
was 100 U.S.P./ml. All solutions were filtered through a #4
Whatman Filter Papsr and refiltered through a 0.45 Jum Metricel R
fTlter .
Dose Solutions and Dosing Procedure
All solutions of compounds ts be tested for bone resorption
inhibition potency were prepared for subcutaneous injectlon in
0.996 normal saline and adjusted to pH 7. ll using NaOH and/or
HCI. Dose solution calculation was made by considering the mass
of powder (based on molecular weight, hydration) of the active
material in mg/kg (body weight) that corresponds to mg P/kg.
Concentrations were based on dosing 0.2 mlllO0 grams of body
~' .
i32~2~
- 24 -
weight. Initially, all compounds were administered at 0.01, 0.1,
and 1. 0 mg P/kg/day for 4 days. Where necessary the test was
repeated, whereby the animals were administered with 0. 5 LED in
order to refine the determination of l ED. Adjustments in dosage
based on changes in body weight were made on a daily basis.
Animals
In this study 50 male Wistar rats weighing approximately
150-1 6û grams were thyroparathyroidectomized surgically by the
breeder (Charles River Breeding Laboratories). All rats were
double housed on arrival in suspende~d cages with Purina Labora-
tory Rodent ChowR and tap water _ libitum. After acclimation
to the laboratory environment for 3-5 days, the rats were placed
on a low calcium, low phosphorous (0.18~/0.22%) diet (TekladR)
and given 2~ (W/V) calcium gluconate supplemented deionized
water via water bottles.
Method
On day four of low-calcium diet all rats were anesthetized
with KetasetR (Ketamine Hydrochloride, 100 mg/ml, Bristol
Myers), 0.10 ml/100 grams of body weight, weighed and then bled
from the retro-orbital venous plexus for serum total calcium
analysis using Flame Atomic Absorption ( FAA) . Ail rats weighing
less than 180 grams were eliminated from the study. Animals
were then randomized statistically such that the mean total serum
calcium for each group was the same. Only rats deemed hypo
calcemic (total serum calcium ~8.0 mg/dl) were placed in study
groups comprising six animals per group.
Treatments with the various experimental compounds com-
menced on day 6 and laste~ through day 9 of the study (at 1:00
P . M . each day) . Dose solutions were prepared to be given at a
constant rate of 0.2 ml/100 grams o~ body weight subcutaneously
in the ventral skin flap where the hind leg meets the torso. All
rats were weighed and dosed daily. A 25 gauge 5/8" needle was
used to administer drug, alternating dose sites daily. On day 8,
animals were changed to deionized, distilled water via water
bottles. On day 9 all rats were fasted in the afternoon at ap-
proximately 4:00 P.M. On day 10 of study no treatment was
132~12~7
- 25 --
given. In the morning a 600 ,ul sample of whole ~lood was col-
lected from each rat in Microtainer (B-D#5060) serum separater
tubes for serum total calcium ( F~A) . Two 125 ~JI samples of
heparinized whole blood were also collected to be used for ionized
5 calcium analysis. Immediately following blood collection all rats
- were weighed and injected with bovine parathyroid hormone
subcutaneously at a rate of 75 USP (filteredl per 100 grams of
body weight. Blood sampling for total and ionized calcium was
repeated three and one-half hours post-PTH injection.
10All pre- and post-PTH total and ionized calciums were stat-
istically analyzed for significance compared to PTH alone t~ontrol)
using Students t-test, analysis of variance, and their non-
parametric equivalents. The post minus pre-change and ~ change
were also determined on calcium levels and pre-drug vs post-drug
15 body weights.
The physiological effect of the PTH challenge is a rise in
serum calcium level, with peak activity observed at three and
one-half hours. Since the hormonal and dietary controls of
calcium metabolism are minimized in the TPTX model, an observed
20 increase in serum calcium level is presumably the result of re-
sorption of bone material. Since polyphosphonates tend to inhibit
resorption of bone materials, the animals pretreated with poly-
phosphonate showed a rise in serum calcium level after PTH
challenge which was less than that found in control animals which
25 had been treated with saline vehicle instead. The lowest dose at
which the polyphosphonate is capable of inhibiting bone
resorption, as evidenced by a decreased rise in serum calcium
upon PTH challenge, is a measure of the bone resorption inhibi-
tibn potency of the polyphosphonate. The LED values of the
30 bone resorption inhibition potency of representative compounds as
determined by the TPTX rat model are presented in Table lll.
~ 32~ l%7
-- 26 --
TABLE 111
Lowest Effective ~Antiresorptive) Dose
TPTX
Diphosphonate Compound LED (m~ Plkg~
5 EHDP 1.0
C~2MDP 1 . O
APD 0.1
N-l2-pyridyl) AMDP 0.01
N-[2-[5-amino)-pyridyl) AMDP 0.01
N-(2-(5-chloro)-pyridyl) AMDP 0.01
N-(2-(5-nitro)-pyridyl) AMDP 0.1
N-12-(5-carboxy)-pyridyl) AMDP N
N-(2-13,5-dichloro)-pyridyl) AMDP 1.0
N-14-pyridyl)-N-ethyl AMDP 0.1
N- 12-13-picolyl ) ) AMDP 0. 002
N- ( 2-1 4-picolyl ) ) AMDP 0.001
N- 12-15-picolyl ) ) AMD P 0 . 001
N-[2-[6-picolyl ) ) AMDP 0 . 01
N-[2-[3~4-lutidine)) AMDP 0.01
20 . N-[2-(4,6-lutidine) AMDP 0.01 1 )
N-~2-pyrimidyl) AMDP 0.01
N- ( 4-(2, 6-dimethyl ) -pyrimiiyl ) AMDP 1.0
N- ( 2- ( 4, 6-dihydroxy ) -pyrimidyl ) AMD P 0 . 01 1 )
N-(2-pyridyl ) AEDP 0. 01
N-(2-t3-picolyl) AEDP 10
2- ( 2-pyridyl ) EDP 0.01
2- ( 3-pyridy 1 ) EDP 0.01
2-(4-pyridyl) EDP 0.1
1-12-pyridyl) propyl DP 1.0
2-~2-pyridyl)-1-chloroethane DP 0.1
0-(2-pyridyi)-oxamethane DP 1.0
0-(2-(3-picolyl))-oxamethane DP 0.1
N = no activity at any of the dosage levels tested
EHDP = ethane-l-hydroxy-l,l-DP
35 C12MDP = dichloromethane DP
APD = 3-aminopropane-1-hydroxy-1,1-DP
132~r~ %r
-- 2~ -
AMDP = aminomethane diphosphonic acid, where the ring i5 at-
tached to the amine
AEDP = 2-aminoethane-1 ,1-diphosphonic acid, where the ring is
attached to the amine
5 EDP = ethane-1 ,1-diphosphonic acid, where the ring is attached
at the 2 position of the ethane
propyl DP = propane-2,2-diphosphonic acid
* = Compounds included in pharmaceutical compositions of the
present invention0 1) = activity level questionable due to lack of dose response
EXAMPLE IV
Patients weighing approximately 70 kilograms who are clin-
ically diagnosed as sufferin~3 from hypercalcemia of malignancy are
administered 0.7 mg P of 2-(2-pyridyl~-ethane-1,1-diphosphonic
15 acid, or its pharmaceutically-acceptable salt or ester, by a 2-1/2
hour intravenous infusion one time daily for 4 days. This
treatment results in an appreciable alleviation of the hypercalcemia
of malignancy.
Similar results are obtained when the 2-~2-pyridyl)-ethane-
20 1,1-diphosphonic acid in the above-described treatment is re-
placed with
N-(2-(5-amino)-pyridyl)-aminomethane diphosphonic acid;
N-(2-l5-chloro)-pyridyl)-aminomethane diphosphonic acid;
N-(2-(3-picolyl))-aminomethane diphosphonic acid
25 N-(2-l4-picolyl))-aminomethane diphosphonic acid;
N-(2-(5-picolyl))-aminomethane diphosphonic acid;
N-12-(6-picolyl))-aminomethane diphosphonic acid;
N-~2-(3,4-lutidine~)-aminomethane diphosphonic acid;
,N-(2-pyrimidyl)-aminomethane diphosphonic acid;
30 N-(2-pyridyl)-2-aminoethane-1,1-~iphosphonic acid;
2-(3-pyridyl)-ethane-1,1-diphosphonic acid;
2- ( 4-pyridyl ) -ethane-1 ,1 -diphosphonic acid;
2-(2-pyridyl)-1-hydroxy-ethane-1,1-diphosphonic acid;
2- ( 3-pyridyl ) -1 -hydroxy-ethane-1 ,1 -diphosphonic acid;
~2~72~
- 28 --
2-(4-pyridyl)-1-hydroxy-ethane-1 ,1-diphosphonic acid;
O- ~ 2 - ( 3-picolyl ) ) -oxame~hane-diphosphon ic ac id; or
pharmaceutically-acceptable salts or esters thereof.
WHAT IS CLAIMED IS:
