CA2199251A1 - Phosphonates and parathyroid hormone for osteoporosis - Google Patents

Abstract

The present invention provides methods of treating a human or other animal subject having a bone metabolism disorder, comprising the steps of: (a) administering to said subject a safe and effective amount of an antiresorptive compound during a period of from about 2 weeks to about 6 months; (b) administering to said subject a safe and effective amount of a parathyroid hormone, during a period of from about 3 to about 12 months. Preferred antiresorptive compounds include bisphosphonates, estrogen compounds, and antiestrogen compounds.

Description

WO96/07418 0 2 11 ~ ~ 2 5 ~ PCT/US95/11336 Phosphonates and Parathyroid Hormone for Osteoporosis BACKGROUND OF THE INVENTION
This invention relates to methods of increasing bone mass in humans and other ~nim~ls, i.e., for the treatment of osteoporosis and related bone metabolic disorders. In particular, this invention relates to such methods of treatment by the administration of an antiresorptive compound and parathyroid hormone.
The most common metabolic bone disorder is osteoporosis. Osteoporosis can be generally defined as the reduction in the quantity of bone, or the atrophy of skeletal tissue. In general, there are two types of osteoporosis: primary and secondary. "Secondary osteoporosis" is the result of an identifiable disease process or agent. However, approximately 90% of all osteoporosis cases is "primary osteoporosis". Such primary osteoporosis includes postmenopausal osteoporosis, age-associated osteoporosis (affecting a majority of individuals over the age of ~0 to 80), and idiopathic osteoporosis affecting middle-aged and younger men and women.
For some osteoporotic individuals the loss of bone tissue is sufficiently great so as to cause mechanical failure of the bone structure. Bone fractures often occur, for example, in the hip and spine of women suffering from postmenopausal osteoporosis. Kyphosis (abnormally increased curvature of the thoracic spine) may also result.
The mech~nism of bone loss in osteopo,olics is believed to involve an imbalance in the process of "bone remodeling". Bone remodeling occurs throughoutlife, renewing the skeleton and ...~ n;ng the sl~el~glh of bone. This remodelinginvolves the erosion and filling of discrete sites on the surface of bones, by an olga~ ed group of cells called "basic multicellular units" or "BMUs". BMUs primarily consist of"osteoclasts", "osteoblasts". and their cellular precursors. In the remodeling cycle, bone is resorbed at the site of an "activated" BMU by an osteoclast, folll~ng a resorption cavity. This cavity is then filled with bone by osteoblasts.
Normally, in adults, the remodeling cycle results in a small deficit in bone, due to incomplete filling of the bone resorption cavity. Thus, even in healthy adults, age-related bone loss occurs. However, in many people, particularly in post-menopausal osteoporotics, there is an increase in the number of BMUs that are activated. This increased activation accelerates bone remodeling, resulting in abnormally high bone loss.

WOg6/07418 0 2 1 99 2 ~ ~ PCT/USg5/11336 Although its etiology is not fully understood, there are many risk factors thought to be associated with osteoporosis. These include low body weight, low calcium intake, physical inactivity, and estrogen deficiency.
Many compositions and methods are described in the medical literature for the "treatment" of osteoporosis. Many of these compositions and methods attempt to either slow the loss of bone or to produce a net gain in bone mass. See, for example, R. C. Haynes, Jr. et al., "Agents affecting Calcification", The Pharmacological Basis of Therapeutics~ 7th Edition (A. G. Gilman, L. S. Goodman et al., Editors, 1985); G. D. Whedon et al., "An Analysis of Current Concepts and Research Interest in Osteoporosis", Current Advances in Skeleto~enesis (A. Ornoy et al., Editors, 1985); and W. A. Peck, et al., Physician's Resource Manual on Osteoporosis (1987), published by the National Osteoporosis Foundation.
Among the treatments for osteoporosis suggested in the literature is the administration of bisphosphonates or other bone-active phosphonates. See, for example, Storm et al., "Effect of Intermittent Cyclical Etidronate Therapy on Bone Mineralization and Fracture Rate in Women with Post-Menopausal Osteoporosis", 322 New Fn~l~nd Journal of Medicine 1265 (1990); and Watts et al., "IntermittentCyclical Etidronate Treatment of Post-Menopausal Osteoporosis", 323 New F.ngl~n~Journal of Medicine 73 (1990). Such tre~trn~nts using a variety of bisphosphonates are described in U.S. Patent 4,761,406, Flora et al., issued August 2, 1988; U.S.
Patent 4,812,304, Anderson et al., issued March 14, 1989; U.S. Patent 4,812,311,Uchtman, issued March 14, 1989; and U.S. Patent 4,822,609, Flora, issued April 18, 1989. The use of such phosphonates for the lle~LIllenL of osteoporosis, and other disorders involving abnormal calcium and phosphate metabolism, is also described in U.S. Patent 3,683,080, Francis, issued August 8, 1972; U.S. Patent 4,330,537, Francis, issued October 28, 1980; U.S. Patent 4,267,108, Blum et al., issued May 12, 1981; European Patent Publication 298,553, Ebetino, published Januaryll, 1989;
and Francis et al., "Chemical, Biochemical, and Medicinal P~opc.Lies of the Diphos-phonates", The Role of Phosphonates in Living Systems. Chapter 4 (1983).
A~ lion of estrogen is also used as a means to prevent osteoporosis in postmenopausal women. This therapy typically involves daily a~mini.~tration of from about 0.625 milligrams to about 1.25 milligrams of conjugated estrogens, or equivalent amounts of other estrogen hormones. Estrogen may also be used to treat osteoporosis (i.e., actual building of bone in osteoporotics), although this has not been fully established. See, for example, Barzel, "Estrogens in the Prevention and Treatment of Post-Menopausal Osteoporosis: a Review", 85 American Journal of Medicine 847 (1988); Barzel, "Estrogen Therapy for Osteoporosis: Is it Effective?", WO96/07418 0 2 1 9~ 2 9 1 PCT/US95/11336 Hospital Practice 95 (1990); Ettinger, et al., "Post-Menopausal Bone Loss is Prevented by Treatment with Low-Dosage Estrogen with Calcium", 106 Annals in Internal Medicine 40 (1987); Lindsay, et al., "The Minimnm Effective Dose of Estrogen for Prevention of Post-Menopausal Bone Loss", 63 Obstetrics and Gynecolo~y 759 (1984); "Estrogen", D~ug Information 1765 (1990); and World Patent Publication 92 14474, McOsker, published September 3, 1992. Furtherrnore,the use of estrogen has been associated with certain side effects, such as uterine bleeding. See, Rudy, "Hormone Replacement Therapy - How to Select the Best Preparation and Regimen," 88 Post~raduate Medicine 157 (1990).
Parathyroid hormone has also been suggested as a therapy for osteoporosis.
Treatments using parathyroid hormone are disclosed in the following references:
Hefti, et al., "Increase of Whole-Body Calcium and Skeletal Mass in Normal and Osteoporotic Adult Rats Treated with Parathyroid Hormone", 62 Clin. Sci. 389-396(1982); Hock, et al., "Resorption Is Not Essential for the Stimulation of Bone Growth by hPTH-(1-34) in Rats In Vivo", 4(3) Jnl. of Bone and Mineral Res. 449-458 (1989); German Patent Publication DE 39 35 738, Fo~s-nan, published May 8, 1991; U.S. Patent 4,698,328, Neer, et al., issued October6, 1987; U.S. Patent 4,833,125, Neer, et al., issued May 23, 1989; U.S. Patent 5,118,667, Adams et al., issued June 2, 1992; World Patent Publication 93 11786, Geddes and Boyce, published June 24, 1993; U.S. Patent 4,822,609, Flora, issued April 18, 1989; U.S.
Patent 4,812,304, Anderson, et al., issued March 14, 1989; German Patent Publication DE 32 43 358, Hesch, published May 24, 1984; Hesch, et al., "Resultsof a Stim~ ting Therapy of Low Bone Metabolism in Osteoporosis with (1 -38h PTH
and Diphosphonate EHDP" 66(19) IClin. Wschr. 976-984 (Oct 1988); German Patent Publication DE 32 43 358, Hesch, published May 24, 1984; Delling, et al.,"Morphologic Study of Pelvic Crest Spongiosa in Patients with Osteoporosis during ADFR Therapy with Parathyroid Horrnone and Diphosphonates", 128(1) Z. Orthop.
1-5 (1990) (hereinafter "Delling, et al."); and Delmas, et al., "The In Vivo Anabolic Effect of hPTH-(1-34) Is Blunted When Bone Resorption Is Blocked By A
Bisphosphonate" 6(1) J. Bone Mineral Res. S136 (#214) (Aug. 1991).
Applicant has found, su~ ingly, that the ~rlministration of parathyroid hormone after a~lmini~tration of an antiresorptive compound provides benefits not recognized in the art. Acco~dingly, the methods of this invention provide effective methods of preventing and treating osteoporosis, with improved efficacy and reduced side effects UJIll~lal ed to methods among those known in the art.
SUMMARY OF THE INVENTION
The present invention provides methods of treating a human or other animal subject having a bone metabolism disorder. comprising the steps of:
(a) administering to said subject a safe and effective amount of an antiresorptive compound, during a period of from about 2 weeks to about 6 months;
(b) adminictering to said subject a safe and effective amount of a parathyroid hormone, during a period of from about 3 to about 12 months.
DESCR~PTION OF THE INVENTION
The methods of the present invention comprise the administration of an antiresorptive compound and parathyroid hormone compound to a human or other animal subject. Specific compounds and compositions to be used in these processes must, accordingly, be pharmaceutically-acceptable. As used herein, such a "pharmaceutically-acceptable" component is one that is suitable for use with humans and/or animals without undue adverse side effects (such as toxicity, irritation, and allergic response) commensurate with a reasonable benefit/risk ratio.

Antiresorptive Compounds The methods of this invention comprise a~minictering one or more antiresorptive compounds. As used herein, an "a~ eso~ re compound" is a compound that, when adminictPred to a human or other animal subject, prevents bone loss by direct or indirect effect on the number of osteoclasts and/or theirmetabolism. Preferred antiresorptive compounds include those selected from the group consisting of bone-active phosphonates, estrogen compounds, antiestrogen compounds, calcitonin compounds, and mixtures thereof. Particularly p~c~-~ed antiresorptive compounds include bisphosphonates, estrogen compounds, and antiestrogen compounds.

~one-Active Phosphonates:
P-er~ d antiresorptive compounds useful in the methods of this invention include bone-active phosphonates. As referred to herein, a "bone-active phosphonate" includes one or more compounds of the general formula Pl 03H2 A--C--B
(1) l O=P--R
I

OH

WO96/07418 ~D 2 ~ ~ g 2 ~ ~ PCT/US95/11336 and pharmaceutically-acceptable salts and esters thereof, wherein A, B, and R are as defined hereinafter.
In Formula (1), "R" is hydroxy (for bisphosphonates), or hydrogen or alkyl (for phosphonoalkylphosphinates). In the phosphonoalkylphosphinates, R is preferably unsubstituted alkyl, especially lower alkyl. When R is substituted alkyl, preferred substituents include halogen, unsubstituted or substituted phenyl, unsubstituted or substituted pyridinyl, unsubstituted amino, amino substituted with one or two lower alkyl groups, hydroxy, or carboxy. More l)refel . ed substituents are fluoro, phenyl, unsubstituted amino, and hydroxy; most pl erel I ~d are fluoro (especially when present as trifluoromethyl) and phenyl.
Particularly prefel I ed R moieties in the phosphonoalkylphosphinates are unsubstituted lower alkyl groups, especially unsubstituted, straight-chain, saturated lower alkyl groups. Also pr~Çel ~ ed R moieties are methyl, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl, tert-butyl, and n-hexyl. More preferably, R is methyl, ethyl, n-propyl, or n-butyl. Most preferably, R is methyl.
In Formula (1), "A" is hydrogen; halogen; nitro; alkyl; heterocycle; aryl;
heteroaryl; unsubstituted amino, or the amide thereof derived from a caboxylic acid of a substituent group; amino substituted with one substituent group, or the amide thereof derived from a carboxylic acid of a substituent group; amino substitutédindependently with one alkyl group and one substituent group; hydroxy, or the ester thereof derived from a carboxylic acid of a substituent group; ether having a substituent group; thiol, or the thiol ester thereof derived from a carboxylic acid of a substituent group; thioether having a substituent group, or the sulfoxide and sulfone derivative thereof; -S03H, the pharm~ceutically-acceptable salts thereof, the ester thereof derived from an alcohol of a substituent group, the unsubstituted amide thereof, or the amide thereof substituted with one or two alkyl groups; -C02H, the pharmaceutically-acceptable salts thereof, the ester thereof derived from an alcohol of a substituent group, the unsubstituted amide thereof, or the amide thereof substituted with one or two alkyl groups; aldehyde; ketone having a substituent group; callJal~,ate, unsubstituted or substituted with one or two alkyl groups;
peptides having from about 1 to about 100 amino acid moieties; or the A and B
moieties are covalently linked to form a ring having from 3 to 7 atoms with from O to 3 heteroatoms selected from the group consisting of nitrogen, sulfur, phosphorus and oxygen, the ring being unsubstituted or substituted with one or more of the above substitl1ents of A; or the A and B moieties are replaced by an unsubstituted or substituted alkyl moiety ~tt~ched to the geminal carbon (the carbon shown in structure ( I ) hereinabove) by a double bond.

-WO 96/07418 ~ PCT/US95/11336 Preferably, A is one of the following moieties.
( I ) hydrogen (2) halogen (preferably fluoro or chloro, more preferably fluoro) (3) substituted or unsubstituted alkyl having the general structure Y f Rl ' n wherein:
(a) n is an integer from 1 to 10, preferably from 1 to 5, more preferably 1 or 2, more preferably 1;
(b) each Rl is, independently, hydrogen, halogen, lower alkyl, unsubstituted amino or the amide thereof derived from a carboxylic acid of a lower alkyl group, amino substituted with one lower alkyl group or the amide thereof derived from a carboxylic acid of a lower alkyl group, amino substituted independently with two lower alkyl groups, hydroxy or the ester thereof derived from a carboxylic acid of a lower alkyl group, -C02H or the pharmaceutically-acceptable salts thereof or the ester thereof derived from an alcohol of a lower alkyl group or the unsubstituted amide thereof or the amide thereof substituted with one or two lower alkyl groups, ether having a lower alkyl group, -P03H2 or the pharmaceutically-acceptable salts thereof, and nitro, or two Rl s on the same carbon atom are =O or =NR9 (where R9 is lower alkyl or may be hydrogen when there is another nitrogen atom ~tt~ched to the same carbon atom as the =NR9 moiety), or two R1's on adj~cçnt carbon atoms may be replaced by an additional bond between the carbon atoms; or an Rl on the first-carbon atom (from the right side of structure (2) hereinabove) and B (see structure (1) hereinabove) may be replaced by an additional bond;
and (c) Y is halogen; nitro; cyano; heterocycle; aryl; heteroaryl; unsubstituted amino, and the amide thereof derived from a carboxylic acid of an alkyl, heterocycle, aryl or heteroaryl group; amino substituted with one alkyl, heterocycle, aryl or heteroaryl group and the arnide thereof derived from a carboxylic acid of an alkyl group; amino substituted independently with one alkyl group and one alkyl, heterocycle, aryl or heteroaryl group; hydroxy, and WO96/07418 ~ ~ ~ 9 9 2 9 11 PCT/US9~/11336 the ester thereof derived from a carboxylic acid of an alkyl, heterocycle, aryl or heteroaryl group; ether having an alkyl, heterocycle, aryl or heteroaryl group;
thiol, and the thiol ester thereof derived from a carboxylic acid of an alkyl, heterocycle, aryl or heteroaryl group; thioether having an al'Kyl, heterocycle, aryl or heteroaryl group, and the sulfoxide and sulfone derivatives thereof; -S03H, the pharrnaceutically-acceptable salts thereof, the ester thereof derived from an alcohol of an alkyl group, the unsubstituted amide thereof, and the amide thereof substituted with one or two alkyl groups; -C02H, the pharmaceutically-acceptable salts thereof, the ester thereof derived from an alcohol of an alkyl group, the unsubstituted amide thereof, and the amide thereof substituted with one or two alkyl groups; P03H2, the pharmaceutically-acceptable salts thereof, the ester thereof derived from an alcohol of an alkyl group, the unsubstituted amide thereof, and the amide thereof substituted with one or two alkyl groups; -(R8)P02H (where R8 is hydrogen or unsubstituted lower alkyl), the pharm~se~ltis~lly-acceptable salts thereof, the ester thereof derived from an alcohol of an alkyl group, the unsubstituted amide thereof, and the amide thereof substituted with one or two alkyl groups; aldehyde; ketone having an alkyl group; ca.l,a,..dle, unsubstituted or substituted with one or two alkyl groups; or peptidyl. For bisphosphonates, Y is preferably a heterocycle (preferably 5 to 7 membered heterocycles having one or two nitrogen atoms); amino; and substituted amino. Particularly pl~.-ed Y moieties include pyridyl, amino, and amino substituted with one or two lower alkyl groups. Plerel~bly, for phosphonoalkylphosphinates, Y is halogen (p.erelably fluoro); trifluolonle~l.yl; ether having a lower alkyl group;
unsubstituted amino, and the amide thereof derived from a carboxylic acid of a lower alkyl group, amino substituted with one lower alkyl group and the amide thereof derived from carboxylic acid of a lower alkyl group; arnino substituted independently~with two lower alkyl groups; or peptidyl having from one to .
about slx ammo acld moletles.

(4) cycloalkyl having from 4 to 10 carbon atoms, pl erel ~bly 5 or 6 carbon atoms (5) heterocycle having 5 or 6 atoms in the ring; more preferably one or two nitrogen atoms in the ring, more preferably having one nitrogen atom in the ring.
Particularly pl ert;l I ed heterocycles are unsubstituted or substituted piperidinyl, pyrrolidinyl, pipela~llyl~ and morpholinyl.

(6) unsubstituted and substituted phenyl and naphthyl (7) unsubstituted and substituted 5 and 6 membered ring heteroaryls having one or two heteroatoms (especially nitrogen heteroatoms), preferably pyridinyl w096/07418 q~ 5 11 PCT/US95/11336 (8) an amine-containing moiety having the general structure:

Rl R2 Y C N

Rl ~ m wherein (a) m is an integer from 0 to I 0, preferably from 0 to 5, more preferably 0 or 1, more preferably 0;
(b) Rl and Y are as described hereinbefore; and (c) R2 is hydrogen, lower alkyl or acyl derived from a carboxylic acid of a lower alkyl (9) an oxygen-containing moiety having the general structure:

Rl I

Y C O
I

R
wherein (a) m is an integer from 0 to 10, preferably from 0 to S, more preferably 0 or 1, more prefel~bly 0; and (b) Rl and Y are as described hereinbefore (10) sulfur-co..~ ;..g moiety having the general structure:

Rl I

Rl ~ m wherein (a) m is an integer from 0 to 10, preferably from 0 to S, more preferably 0 WO 96/07418 ~ 9 Z 5~ ~ PCT/US95/11336 or 1, more preferably 0; and (b) Rl and Y are as described hereinbefore In Formula (1), "B" is hydrogen; halogen; unsubstituted or substituted lower alkyl; unsubstituted or substituted cycloalkyl having from 3 to 7 atoms in the ring;
unsubstituted or substituted heterocycle having from 3 to 7 atoms in the ring;
unsubstituted or substituted phenyl, hydroxy, or the ester thereof derived from a carboxylic acid of a lower alkyl group; thiol, unsubstituted amino, or the amidethereof derived from a carboxylic acid of a lower alkyl group; amino substituted with one lower alkyl group, or the amide thereof derived from a carboxylic acid of a lower alkyl group; amino substituted independently with two lower alkyl groups; or -CO2H, the pharmaceutically-acceptable salts thereof, the ester thereof derived from an alcohol of a lower alkyl group, the unsubstituted amide thereof, or the amidethereof substituted with one or two lower alkyl groups.
To maintain chemical stability of these compounds, the A and B moieties preferably do not both have heteroatoms (nitrogen, oxygen or sulfur), or a heteroatom and a halogen, bonded to the phosphonate moiety (i.e., the carbon atom geminally substituted with the phosphorous atoms). Thus, when the A moiety has an oxygen, sulfur, nitrogen, or halogen atom bonded to the phosphorous-substituted methylene carbon, then B is selected from hydrogen; unsubstituted or substituted-lower alkyl, cycloalkyl, heterocycle (where a carbon atom of the heterocycle is bonded to the geminal carbon atoms), or phenyl, -COOH, the pharm~ceutically-acceptable salts thereof, the ester thereof derived from an alcohol of a lower alkyl group, the unsubstituted amide thereof, and the amide thereof substituted with one or two lower alkyl groups.
Preferably B is hydrogen, halogen, unsubstituted or substituted lower alkyl, unsubstituted or substituted phenyl, unsubstituted or substituted benzyl, hydroxy or the ester thereof derived from a carboxylic acid of a lower alkyl group, thiol, unsubstituted amino or the amide thereof derived from a carboxylic acid of a lower alkyl group, amino substituted with one lower alkyl group or the amide thereof derived from a carboxylic acid of a lower alkyl group, amino substituted independently with two lower alkyl groups, or -COOH or the pharm~ceutically-acceptable salts thereof and the ester thereof derived from an alcohol of a lower alkyl group and the unsubstituted amide thereof or the amide thereof substituted with one or two lower alkyl groups.
More plefe-ably, B is hydrogen, chloro, methyl, ethyl, hydroxy, thiol, unsubstituted amino, (N-methyl)amino, (N,N-dimethyl)amino, -COOH or the pharm~ceutically-acceptable salts thereof, -COOCH2, or -CONH2. More p. e~el ably, WO96/07418 ~ 2 11 ~ 9 2 5 1 PCT/US9Stll336 B is hydrogen, methyl, chloro, amino, or hydroxy; more preferably hydrogen, or hydroxy, or amino, or thiol; more preferably hydroxy. Particularly plefe~led bone-active phosphonates include those wherein A is a moiety of groups (3) or (8) above, and B is hydroxy Particularly preferred bisphosphonates useful herein are of the formula:
3 P~3H2 / ~ \ I
R2_ X C C R

\ R / n PO3H2 wherein: n is an integer from O to 7 (preferably from O to 3, more preferably 1); Rl is hydrogen, chloro, amino, or hydroxy (preferably hydrogen or hydroxy); X is -NH-, quaternary amine, oxygen, sulfur, or a single bond (preferably -NH- or single bond);
R2 and is a substituted or unsubstituted 5- to 7-membered carbocycle (preferably 6-to 7- membered, more preferably benzene or cycloheptyl), a substituted or unsubstituted 5- to 7-membered heterocycle having from 1 to 3 heteroatoms (preferably a 6-membered heterocycle having 1 or 2 nitrogen atoms, wherein a ring nitrogen may be quaternarized), -NH2, amino substituted with one alkyl or two alkyl (preferably Cl-Cs) groups to give a secondary or tertiary amine, respectively, quaternary amino, or hydrogen; wherein if R2 is a substituted 5- to 7-membered carbocycle or heterocycle, the substituent is one or more substinlentc selected from the group consisting of substituted and unsubstinlte~, saturated or unsaturated alkyl having from 1 to about 6 carbon atoms, substituted and unsubstituted aryl, substituted and unsubstituted benzyl, hydroxy, halogen, carbonyl, alkoxy, nitro,amido, amino, substituted amino, carboxylate, and co",b;,lations thereof, hydrogen being plerelled; each R3 is, independently, hydrogen, or substituted or unsubstituted alkyl (saturated or unsaturated) having from 1 to about 4 carbon atoms; and their pharrnaceutically-acceptable salts and esters.
The term "ph~-. ~eeutically-acceptable salts and esters", as used herein, means hydrolyzable esters and salts of the bone-active phosphonates which have the same general pharmacological p~ope.Lies as the acid form from which they are derived, and which are pharm~ceutically acceptable. Pharm~ceutically-acceptable salts include, for example, alkali metals (e.g., sodium and potassium), alkaline earth metals (e.g., calcium and m~sillm), non-toxic heavy metals (e.g., stannous and indium), and allulloluum and low molecular weight substituted ammonium (e.g., mono-, di- and triethanolamine) salts. Preferred compounds are the sodium, pot~csillm, and ammonium salts. Pharmaceutically-acceptable esters include WO 96/07418 ~ 9 ~ 5 1I PCT/US95/11336 unsubstituted and substituted alkyl, aryl and phosphoryl esters. Nonlimiting examples of pharmaceutically-acceptable esters include, for example, isopropyl, tertiarybutyl, 2-chloroethyl, 2,2,2-trichloroethyl, 2,2,2-trifluoroethyl, p-toluenesulfonylethyl, glycyl, sarcosyl, benzyl, phenyl, 1,2-hexanoylglyceryl, p-nitrophenyl, 2,2 dimethyl-1,3-dioxolene-4-methyl, isopentenyl, o-carbomethoxyphenyl, piraloyloxymethylsalicylyl, diethylamidophosphoryl, pivaloyloxymethyl, acyloxymethyl, propionyloxymethyl, isobutyryloxymethyl, dodecyl, octadecyl, and isopropyloxymethyl.
Specific examples and definitions for substituents useful in the compounds of Formulas (1) through (6) are described in European Patent Publication 298,553, Ebetino, published January 11, 1989 (incorporated by reference herein). That application also describes phosphonoalkylphosphinates useful in the methods of this invention (wherein R is hydrogen or alkyl), and methods for making such compounds. Methods of making phosphonoalkylphosphinates are also described in European Patent Publication 298,555, Ebetino, published January 11, 1989 (incorporated by reference herein).
Bisphosphonates useful in the methods of this invention (wherein R is hydroxy), and methods for making such compounds, are described in the following patent doc~ments, all incorporated by ~rellce herein: U.S. Patent 3,553,314,-Francis, issued January 5, 1971; U.S. Patent 3,683,080, Francis, issued August 8, 1972; U.S. Patent 3,846,420, Wollmann et al., issued November 5, 1974; U.S. Patent 3,899,496, Schindler et al., issued August 12, 1975; U.S. Patent 3,941,772, Ploger et al., issued March 2, 1976; U.S. Patent 3,957,160, Ploger et al., issued May 18, 1976;
U.S. Patent 3,962,432, Schmidt-Dunker, issued June 8, 1976; U.S. Patent 3,979,385, Wollmann et al., issued September7, 1976; U.S. Patent 3,988,443, Ploger et al., issued October 26, 1976; U.S. Patent 4,054,598, Blum et al., issued October 18, 1977; U.S. Patent 4,113,861, Fleisch et al., issued September 12, 1978; U.S. Patent 4,117,090, Ploger, issued September26, 1978; U.S. Patent 4,134,969, Schmidt-Dunker, issued January 16, 1979; U.S. Patent 4,267,108, Blum et al., issued May 12, 1981; U.S. Patent 4,304,734, Jary et al., issued Decenlber8, 1981; U.S. Patent 4,330,537, Francis, issued May 18, 1982; U.S. Patent 4,407,761, Blum et al., issued October 4, 1983; U.S. Patent 4,469,686, Andrews, issued September 4, 1984; U.S.
Patent 4,578,376, Rosini, issued March 25, 1986; U.S. Patent 4,608,368, Blum et al., issued August26, 1986; U.S. Patent 4,621,077, Rosini et al., issued November4, 1986; U.S. Patent 4,687,767, Bosies et al., issued August 18, 1987; U.S. Patent 4,687,768, Benedict et al., issued October 18, 1987; U.S. Patent 4,711,880, Stahl et al., issued December 8, 1987; U.S. Patent 4,719,203, Bosies et al., issued WOg6/07418 ~ 9 2 5 ~ PCTIUSg5/11336 January 12, 1988; U.S. Patent 4,927,814, Gall et al., issued May22, 1990; U.S.
Patent 4,990,503, Isomura et al., issued February 5, 1991; German Offenlegungsschri~ 2,104,476, Worms, published August 17, 1972; German OffenlegungsschriPt 2,343,147, Ploeger et al., published April 3, 1975; German Offenlegungsschrift 2,360,798, Worms et al., published June 26, 1975; German Offenlegungsschrift 2,513,966, Schmidt-Dunker, published October 7, 1976; GermanOffenle~lngcschrift 2,541,981, Eimers et al., published March24, 1977; German Offenle~lngcschrift 3,334,211, Blum, published April 4, 1985, Japanese Patent Publication 78/59,674, Suzuki et al., published May 29, 1978; Japanese Patent Publication 79/135,724, Suzuki et al., published October 22, 1979; Japanese Patent Publication 80/98193, Suzuki et al., published July 25, 1980; European Patent Publication 88,359, Blum et al., published September 14, 1983; European Patent Publication 100,718, Breliere et al., published February 15, 1984; European Patent Publication 186,405, Benedict et al., published July 2, 1986; European Patent Publication 197,478, Bosies et al., published October 15, 1986; European Patent Publication 230,068, Benedict et al., published July 29, 1987; European Patent Publication 273,514, Ebetino et al., published July 6, 1988; European Patent Publication 274,158, Ebetino et al., published July 13, 1988; European Patent Publication 282,309, Sakamoto et al., published September 14, 1988; European Patent Publication 282,320, Isomura et al., published Seplenlbel 14, 1988; PCT
Patent Publication 87/03598, Binderup et al., published June 18, 1987; and PCT
Patent Publication 88/00590, Gall et al., published January 28, 1988.
Ple~el~ed bone-active phosphonates useful in the methods of this invention include: N-(2'-(3'-methyl)-pyridinyl)aminomethane phosphonomethylphosphinic acid;
N-(2'-(5'-methyl)-pyridinyl)amino meth~ne phosphonomethylphosphinic acid; N-(2'-(3'-methyl)-piperidinylidene)aminomethane phosphonomethylphosphinic acid; N-(2'-(5'-methyl)-piperidinylidene)a"l"~omethane phosphonomethylphosphinic acid; 2-(2'-pyridinyl)ethane- I -phosphono- 1 -methylphosphinic acid; 2-(2'-piperidinyl)ethane- 1-phosphono- 1 -methylphosphinic acid; 2-(p-aminophenyl)- 1 -hydroxy-ethane- 1-phosphono- 1 -methylphosphinic acid; 2-(m-aminophenyl)- 1 -hydroxy-ethane- 1-phosphono-l-methylphosphinic acid; N-(1-(5-amino-2-methyl-1-oxo)-pentyl)aminomethane phosphonomethylphosphinic acid; N-(2'-(3'-methyl)-piperidinylidene)amino",~,lhal1e phosphonobutylphosphinic acid; S-(2~-pyridinyl)thio, . ~el h~ ne phosphonomethylphosphinic acid; 2-(2-pyridyl)- 1-hydroxyethane-1-phosphono-1-methyl phosphinic acid; 2-(3-pyridyl)-1-hydroxyethane- 1 -phosphono- I -methylphosphinic acid; 2-(N-imidazoyl)- 1-hydroxyethane- 1 -phosphono- 1 -methylphosphinic acid; 3-(N-pentyl-N-methylamino)-WO 96/07418 ~ 9 ~ 5 ~ PCT/US95/11336 1 -hydroxypropane- 1 -phosphono- 1 -methylphosphinic acid; 4-amino- 1 -hydroxybu-tane- 1 -phosphono- 1 -methylphosphinic acid; 3 -(N-pyrollidino)- 1 -hydroxypropane- 1-phosphono- 1 -methylphosphinic acid; N-cycloheptyl aminomethanephosphonomethylphosphinic acid; S-(p-chlorophenyl) thiomethanephosphonomethylphosphinic acid; (7-dihydro- 1-pyrindine)methanephosphonomethylphosphinic acid;(7-dihydro- 1-pyrindine)hydroxymethanephosphonomethylphosphinic acid; (6-dihydro-2-pyrindine)hydroxymethanephosphonomethylphosphinic acid; 2-(6-pyrolopyrindine)-1 -hydroxyethane- 1 -phosphono- 1 -methyl phosphinic acid; 1 -hydroxyethane- 1,1-bisphosphonic acid; 1 -hydroxy pentane- 1,1 -bisphosphonic acid; methane bisphosphonic acid; dichloromethanebisphosphonic acid;
hydroxymethanebisphosphonic acid; 1 -aminoethane- 1,1 -bisphosphonic acid; 2-aminoethane- 1,1 -bisphosphonic acid; 3-aminopropane- 1,1 -bisphosphonic acid; 3-aminopropane- 1 -hydroxy- 1,1 -bisphosphonic acid; 3-(dimethylamino)- 1-hydroxypropane-l,l-bisphosphonic acid; 3,3-dimethyl-3-amino-1-hydroxypropane-1, l-bisphosphonic acid; phenylaminomethane bisphosphonic acid; N,N-dimethylaminomethane bisphosphonic acid; N-(2-hydroxyethyl) aminomethane-bisphosphonic acid; 4-amino-1-hydroxybutane-1,1-bisphosphonic acid; 5-amino-1-hydroxypentane- 1,1 -bisphosphonic acid; 6-amino- 1 -hydroxyhexane- 1,1-bisphosphonic acid; indan-2,2-bisphosphonic acid; hexahydroindan-2,2-bisphos-phonic acid; 2-methylcyclobutane-1,1-bisphosphonic acid; 3-chlorocyclopentane-1,1-bisphosphonic acid; cyclohexane-1,1-bisphosphonic acid; 2-(2-pyridyl)-1-hydroxyethane- 1,1 -bisphosphonic acid; N-(2-(5-amino)-pyridyl)-aminometh~ne bisphosphonic acid; N-(2-(5-chloro)-pyridyl)-aminomethane bisphosphonic acid; N-(2-(3-picolyl))-amino.,le~l,ane bisphosphonic acid; N-(2-(4-picolyl))-aminor"elh~l-e bisphosphonic acid; N-(2-(5-picolyl))-aminometh~ne bisphosphonic acid; N-(2-(6-picolyl))-aminometh~ne bisphosphonic acid; N-(2-(3,4-lutidine))-aminom~ll-al-e bisphosphonic acid; N-(2-pyrimidyl)-aminomethane bisphosphonic acid; N-(2-pyridyl)-2-aminoethane- 1,1 -bisphosphonic acid; 2-(2-pyridyl)-ethane- 1, I -bisphosphonic acid; 2-(3-pyridyl)-ethane-1,1-bisphosphonic acid; 2-(4-pyridyl)-ethane-l,l-bisphosphonic acid; 2-(2-(3-picolyl))-ox~eth~ne-1,1-bisphosphonic acid;
2-(3 -pyridyl)- 1 -hydroxyethane- 1,1 -bisphosphonic acid; 2-(N-imidazoyl)- 1-hydroxyethane- 1,1 -bisphosphonic acid; 3 -(N-pentyl-N-methylamino)- 1-hydroxypropane- 1,1 -bisphosphonic acid; 3-(N-pyrollidino)- I -hydrox~, opane- 1,1-bisphosphonic acid; N-cycloheptylal,linolllt;lhane bisphosphonic acid; S-(p-chlorophenyl) thiometh~nebisphosphonic acid; (7-dihydro-1-pyrindine)meth~nebis-phosphonic acid; (7-dihydro-1-pyrindine)hydrox~ l.anebi~l~hosphonic acid; (6-WO96/07418 0 a ~ ~ ~ 2 9 ~ PCT/US9~/11336 dihydro-2-pyrindine)hydroxymethanebisphosphonic acid; 2-(6-pyrolopyridine)- 1-hydroxyethane- I, I -bisphosphonic acid; and pharmaceutically-acceptable salts and esters thereof.
Particularly preferred bone-active phosphonates useful in the methods of this invention include: l-hydroxyethane-l, I-bisphosphonic acid; dichloromethane bisphosphonic acid; 3-amino-1-hydroxypropane-1,1-bisphosphonic acid; 6-amino-1-hydroxyhexane- I ,1 -bisphosphonic acid; 4-amino- 1 -hydroxybutane- 1,1 -bisphosphonic acid; 2-(3-pyridyl)-1-hydroxyethane-1,1-bisphosphonic acid; 2-(N-imidazoyl)-l-hydroxyethane- I, I -bisphosphonic acid; 3-~N-pentyl-N-methylamino)- I -hydroxypropane- I, I -bisphosphonic acid; 3-(N-pyrollidino)- 1 -hydroxypropane- I ,1-bisphosphonic acid; N-cycloheptylaminomethanebisphosphonic acid; S-(p-chlorophenyl) thiomethanebisphosphonic acid; (7-dihydro-1-pyrindine)methane bis-phosphonic acid; (7-dihydro- 1 -pyrindine)hydroxymethane bisphosphonic acid; (6-dihydro-2-pyrindine)hydroxymethanebisphosphonic acid; 2-(6-pyrolopyridine)-1-hydroxyethane- 1,1 -bisphosphonic acid; and pharmaceutically-acceptable salts and esters thereof.

Estro~en Compound:
Among the antiresorptive compounds useful in this invention are estrogén compounds. As referred to herein, an "estrogen compound" refers to naturally occurring hormones, synthetic steroidal compounds, and non-steroidal compounds, and conjugates, metabolites and derivatives thereof, which having estrogenic activity.
Naturally-occurring estrogen compounds are steroids which contain a cyclopentanoperhyd~ol)hena~hrene ring system. Such naturally-occurring estrogen compounds are obtained from plegndnL mares' urine or prepared synthetically, using methods well-known in the art. See: "Estrogens", Drug Inro~,aLion 1765 (1990);
and Rudy, "Hormone Repl~cçm~nt Therapy - How to Select the Best Pr~pa.~lion and Rçgimlon," 88 Postgraduate Medicine 157 (1990); and C. Chricti~ncçn et al., "Estrogens, Bone Loss and Prevention," I Osteoporosis Int. 7 (1990); all of which are inco"Jol aled by reference herein.
Estrogen compounds useful in the methods of this invention include, for example, estradiol, estrone, estriol, equilin, equilenin, estradiol cypionate, estradiol valerate, ethinyl estradiol, polyestradiol phosphate, e~L~ upipdle, diethylstilbestrol, dienestrol, chlorotri~nicpne~ and mixtures thereo~ A p.~relled estrogen hormone useful herein is "conjugated estrogen", which is a mixture of sodium salts of the water-soluble sulfate esters of estrone and equilin. Such conjugated estrogens may also contain other estrogenic substances found in pleg"an~ mares' urine, such as 17-a-dihydroequiline, 17-a-estradiol, equilenin, and 17-a-dihydroequilenin Calcitonin Compounds:
~ mong the antiresorptive compounds useful in this invention are calcitonincompounds. As referred to herein, calcitonin compounds are calcium reg~ ting horrnones whose essential biological activity is to oppose the bone and renal effects of parathyroid hormone, i.e., to inhibit bone resorption and reduce urinary calcium excretion. Natural calcitonin is a 32 amino acid polypeptide horrnone secreted form the parafollicular cells of the thyroid gland in mammals and by the ultimobranchial gland of birds and fish. The linear amino acid sequence varies between species as does the potency but when the various calcitonins are ~ministered at similar International Unit equivalents, all such calcitonins will provide equivalent efficacy as an antiresorptive compound.
Natural calcitonin can be isolated from m~mm~ n glands, i.e., from pig, cow, human, etc., or from salmon, eel and other sources by a procedure similar to that described in Behrens, G.il~ans Ann Rev Biochem 38, 83, 1969. In addition tothe isolation of the peptide from glands, any of the calcitonin sequences may bem~nllf~ctured by synthetic processes such as by solid phase synthesis ( J Hirt, et al., Rec Trav Chim 98, 143, 1979), or produced by recolllbinant methods ( J W Jacobs et al., J Biol Chem 254, 10600, 1979). Calcitonin compounds also include synthetic-natural hybrids, and analogues, mixtures, peptidomimetics and other variants of the natural calcitonin molecule.

Antiestrogens Among the anliresol ~ /e compounds useful in this invention are the antiestrogen compounds. As referred to herein, an "antiestrogen compound" is a compound which has the estrogen agonist activity of inhibition of bone resorption, but which has estrogen antagonist activity on other tissues, notably breast and uterus.
Antiestrogens may be steroids or non-steroids. Steroidal antiestrogens are exemplified by tamoxifen and related compounds. Many compounds that have been discovered for reasons of other activities, such as anti-progesterones or anti-androgens, are also antiestrogens useful in the methods of this invention. Non-steroidal antiestrogens also include, for example, raloxifene, and related compounds disclosed in U.S. Patent 4,418,068, issued November 29, 1983 (incorporated by reference herein).

Parathyroid Hormone WO 96/07418 'g 2 1 ~ ~ ~ g ~ PCT/US95/11336 The methods of this invention also comprise administration of a parathyroid hormone. As referred to herein, "parathyroid horrnone" refers to the naturally occurring human parathyroid hormone, synthetic analogs thereof, parathyroid hormone and parathyroid hormone fragments manufactured by recombinant DNA
technology, and parathyroid hormone fragments and parathyroid hormone fragment analogs. Parathyroid hormone useful in the methods of this invention includes, for example hPTH (1-38), hPTH (1-34), hPTH (1-37), hPTH (2-34), and hPTH (2-38).
Detailed descriptions of the types of parathyroid hormones available and methods for manufacturing parathyroid hormone are disclosed in the following references, allincorporated by reference herein, U.S. Patent 4,105,602, Colescott, et al., issued August 8, 1978; U.S. Patent 4,698,328, Neer, et al., issued October6, 1987; U.S.Patent 4,833,125, Neer, et al., issued May 23, 1987; DE 32 43 358, Hesch, publication date May 24, 1984; and DE 39 35 738, Forssmann, et al., publication date May 8, 1991.

Methods of Treatment This invention provides methods for treating a human or other animal subject having a bone metabolism disorder, comprising the steps of:
(a) administering to said subject a safe and effective amount of an antiresorptive compound, during a period of from about 2 weeks to about 6 months;
(b) adminict~ring to said subject a safe and effective amount of a parathyroid hormone, during a period of from about 3 to about 12 months.
Preferably, in step (a), said antiresorptive compound is ~rlministçred for from about 2 months to about 6 months. Also preferably, in step (b), said parathyroid hormone is ;slçred for from about 4 months to about 8 months, more preferably for about 6 months. Preferably steps (a) and (b) are repeated from I to 5 times (i.e, so the entire method comprises pelro~ 'ce of each step, in sequence, 2 to 6 times). In a plefel,ed method of this invention, the allL;lesol~ /e compound is ~lminictered during the t~ea~lllent period of step (b); i.e., said anLilesoly~i~/e compound is ~ministered concurrently with the parathyroid hol---one.
A pleft--ed method ofthis invention co--ll,lis;l-g the steps of:
(a) admini.ctçring to said subject a safe and effective amount of an antiresorptive compound, during a period of from about 2 weeks to about I
month;
(b) arlminictering to said subject a safe and effective amount of a parathyroid hormone compound, during a period of from about 3 to about 6 months.

WO96/07418 1~ 2 ~ ~ 9 ~ 5 ~ PCT/US95/11336 (c) administering to said subject a safe and effective amount of an antiresorptive compound, during a period of from about 3 months to about 6 months;
(d) a~lministering to said subject a safe and effective amount of a parathyroid hormone compound, during a period of from about 3 to about 6 months.
Preferably, in step (c) said period during which said antiresorptive compound is~(imini~tçred is from about 2 months to about 6 months. Also preferably, in step (b) and in step (d), said period during which said parathyroid hormone is a~mini~tered is from about 4 months to about 8 months, more preferably for about 6 months.
Preferably steps (c) and (d) are repeated from 1 to 4 times (i.e, so the method comprises pe~rol~llance of step (a) and (b), and each of steps (c) and (d) are then performed, in sequence, 2 to 4 times).
The antiresorptive compound and parathyroid hormone are administered in a "safe and effective amount", which, as referred to herein, is the quantity of a material which is sufficient to yield a desired therapeutic response without undue adverse side effects (such as toxicity, irritation, or allergic response) co~",ensurate with a reasonable benefit/risk ratio when used in the manner of this invention. The specific "safe and effective amount" will, obviously, vary with such factors as the particular condition being treated, the physical condition of the patient, the duration of thé
tre~tm~nt, the nature of concurrent therapy (if any), and the specific formulations employed.
The specific amount and dosage regimen of a particular antiresorptive compound ~-iminict~red during the methods of this invention is a function of thepotency of the compound, as well as other factors. Preferably, the antiresorptive compound is ~rlmini~tered in amounts and in reg;.~e-..c recognized in the art of useful for treating osteoporosis in accG, dance with sound medical practice.
Potency of an antiresorptive compound can be ~ylessed in terms of its "LED" or "least effective dose", which is the minim.lm dose of compound that is effective, by itself, to cause a significant inhibition of bone resorption. Preferably, in the methods of this invention, the antiresorptive compound is ~dmini~tered at a level of at least about 0.8 LED of the compound, more preferably from about .8 LED to about 5 LED, more preferably from about .8 to about 3 LED.
The specific LEDs of a given antiresorptive compound will vary depending upon its chemical composition, and method of ~mini~tration (i.e., oral or parenteral). The lower the LED, the more potent the compound. Generally, it is desirable to ~iminicter higher potency antiresorptive compounds in lower doses and on a fewer number of days in the period of l~e~ ..1 Likewise, the higher the LED, W096/07418 0 2 ~ ~ 9 2 9 ~ PCT/US9SI11336 the less potent the compound. Accordingly, then, in general, it is desirable to atlmini~ter a lower potency antiresorptive compound in higher doses and on a greater number of days in the period of tre~tment.
In particular, the LEDs for the bone-active phosphonates may be determined using any of several art-recognized in vivo models. One such model is the thyroparathyroidectomized ("TPTX") rat model. In this model, compounds are evaluated for in vivo bone resorption inhibition potency, by measuring their ability to inhibit the increase of serum calcium levels caused by administration of parathyroid hormone in rats whose parathyroid gland has been removed. This model is described in Russell et al., 6 Calcified Tissue Research 183 (1970); Muhlbauer et al., 5 Mineral Electrolite Metabolism 296 (1981); U.S. Patent 4,761,406, Flora et al., issued August 2, 1988; and European Patent Publication 298,553, Ebetino, published January 11, 1989; all of which are incorporated by reference herein.
Another model is the "Schenk Model", which measures the effects of bone-active phosphonates on bone growth in young rats. This model is described in Schenk et al., 11 Calcif. Tissue Res. 196 (1973); Shinoda et al., 35 Calcif. Tissue Int.
87 (1983); U.S. Patent 4,761,406, Flora et al., issued August 2, 1988; and European Patent Publication 298,553, Ebetino, published January 11, 1989; all of which are incorporated by reference herein.
Another model is the "ovariectomized" or "OVX" rat model, which measures the ability of bone-active phosphonates to prevent loss of bone in female rates induced by ovariectomy. This model is described in Wronski et al., 125 Endocrinology 810 (1989), inco,yo~ d by reference herein.
The LEDs for bone active phosphonates are conveniently expressed in "mgP/kg", which, as referred to herein, is the amount of compound, expressed as milligrams phosphorus in the compound, per kilogram weight of the subject to be treated. Because the bone active phosphonates vary in molecular weight, eA~,es~;"g the amount ~dnlinictçred in mg P/kg normalizes the co"",alison between compoundsof varying potel1c;es. In order to d~telllline the mg P/kg ~ lçled to a patient according to the methods of this invention, the following conversion formula is used:

mg/kg compound admir~istered = (m~ ph~kSgPh~rUS) X ~nobcular wei~ht of compound) (For example, 2-(3-pyridinyl)-1-hydroxyethane-1,1-bisphosphonate has a molecularweight of 350. Two phosphorus atoms have a molecular weight of 62. Thus, if a patient is dosed at 0.01 mg/kg of the compound, then about 0.002 mg P/kg was WO96/07418 1~ ~ ff ~ ~ ~ 9 ~ ~CT/US95/11336 ~dmini.ctered.
The LEDs for parenteral dosing of pl efel ~ ed bone-active phosphonates useful herein are: 1.0 mg P/kg, for l-hydroxyethane-l,l-bisphosphonic acid; 0.5 mg P/kg, for dichloromethane bisphosphonic acid; 0.03 mg P/kg, for 3-amino-1-hydroxypropane-l, l-bisphosphonic acid; 0.001 mg P/kg, for 4-amino-1-hydroxybutane-l,l-bisphosphonic acid; 0.1 mg P/kg, for 6-amino-1-hydroxyhexane-l,l-bisphosphonic acid; 0.01 mg P/kg, for N-(2-pyridyl) aminomethane-l,l-bisphosphonic acid; 0.0003 mg P/kg, for 2-(3-pyridyl)-1-hydroxyethane-1,1-bisphosphonic acid; 0.0001 mg P/kg, for N-cycloheptyl-aminomethanebisphosphonic acid; 0.0001 mg P/kg, for 3-(N-pentyl-N-methylamino)-l-hydroxypropane-l,l-bisphosphonic acid; 0.01 mg P/kg, for 3-(dimethylamino)-1-hydroxypropane-1,1-bisphosphonic acid; 0.01 mg P/kg, for 3-(N-pyrollidino)-l-hydroxypropane-l, l-bisphosphonic acid; 0.03 mg P/kg, for N-cycloheptylaminomethanebisphosphonic acid, and 0.3 mg P/kg for S-(p-chlorophenyl)thiomethanebisphosphonic acid. (The LEDs for oral dosing would be higher, depending upon the systemic absorption of the phosphonate. Typically, absorption from oral a~lminictration is from about 1% to about 10%. Thus, oral LEDs are typically about ten- to one hundred-fold higher than the parenteral LEDs.) Sirnilarly, the LED of the estrogen horrnone is that level of the hormone which, by itself, is effective to prevent bone loss in subjects having osteoporosis.
That level is generally recognized to be about 0.625 mg per day of conjugated estrogen or an equivalent dose of other estrogen horrnones (for example, 25 mg per day of ethinyl estradiol; or 2 mg per day of 17-b-estradiol). Conjugated estrogen is pr~rel~bly adminictered at a level of from about 0.3 mg to about 1.25 mg per day, preferably about 0.63 mg per day. See, Barzel, "Estrogens in the Prevention and Tre~tment of Post-Menopausal Osteoporosis: a Review", 85 American Journal of Medicine 847 (1988); Lindsay, et al., "The ~inimllm Effective Dose of Estrogen for Prevention of Post-Menopausal Bone Loss", 63 Obstetrics and Gynecology 759 (1984); Genant et al., "Effect of Estrone Sulfate on Po~.nenopd~sal Bone Loss", 76 Obstetrics and Gynecology 529 (1990); all of which are incoll,ol~ted by reference herein.
~ Calcitonin compounds are preferably dosed at levels recognized in the art as suitable for the l,~~ -l of osteoporosis. Pre~.,ed dosages include 100 International Units (I. U.) per day for the subcutaneous and interrnoscular injections and appro~iJl,dlely twice that for nasal a~mini.ctration. }~e~in~el~ of this invention also include reduced dosages, such as 25-50 I.U, and injections given for 5 days out of every week, every other day, or in cycles.

WO96/07418 ~1 a ~ 4 9 2 ~ ~ PCT/US95111336 Parathyroid hormone is routinely dosed in International Units (IU). ~n the methods of this invention, parathyroid hormone is preferably administered at levels of from about 100 to about 700 IIJ per day, more preferably from about 200 to about600 IU per day, more preferably from about 400 to about 500 IU per day.
During the treatment period of step (a), and (optionally) the treatment period of step (b), the antiresorptive compound can be administered daily, or in a cyclical fashion. Such cyclical regimens are generally described in U.S. Patents 4,761,406, Flora et al., issued August 2, 1988, U.S. Patent 4,812,304, Anderson et al., issued March 14, 1989; U.S. Patent 4,822,609, Flora, issued April 18, 1989; World Patent Publication 93 11786, Geddes and Boyce, published June 29, 1993; and World Patent Publication 92 11474, McOsker, published September 3, 1992; all of which are incorporated by reference herein. For methods using a bisphosphonate, the bisphosphonate must be given at least one day of every thirty(30)-days of said treatment period. Preferably, the bisphosphonate may be given every day, every second day, every third day, every fourth day, every fifth day, or every sixth day, of said treatment period.
During the treatment period of step (b), the parathyroid hormone must be given at least one day every seven days of every thirty(30)-days. Preferably, the parathyroid hormone is a-lmini~tered at least about 50% of the days during the period of step (b) The methods of this invention comprise llt;"l"l~;lll of osteoporosis at all stages of the disorder. Since osteoporosis is an ongoing process of bone loss, rather than a disorder having a discrete beginning- or end-point, "treatment", as referred to herein, consists of any method which stops, slows, or reverses the process of bone loss which occurs in osteoporosis.
Plerelled methods of this invention comprise ll~ .,f ~1 of osteoporosis in subjects who have already lost skeletal mass (herein rerellèd to as "establishedosteoporosis"). Such methods of this invention for the tre~tm~nt of established osteoporosis plerèu~bly comprise ~ministering the actives for a period of time sufficient to achieve an increase in the net skeletal mass of said subject. The increase in mass may be in cortical bone, trabecular bone, or both. Preferably, the net skeletal mass is increased by at least about 5% per year, prerelably by at least about 10% per year.
The specific period of time sufficient to achieve an increase in the net skeletal mass of the subject may depend on a variety of factors. Such factors include, for example, the specific actives employed, the amount of actives a~mini~tered, the age and sex of the subject, the specific disorder to be treated, concollulalll therapies g6/07418 ~ 2 ~ 1I PCT/US95/11336 employed (if any), the general physical health of the subject (including the presence of other disorders), the extent of bone loss in the individual, and the nutritional habits of the individual.
The therapeutic regimen ~Itili7in~; the methods of this invention are preferablycontinued for at least about twelve months. Of course, a therapeutic regimen may be continued indefinitely, according to sound medical practice. Preferably the subject is treated until a net skeletal mass is obtained commensurate with reduced fracture risk as assessed by the patient's physician.
Also, preferably, the subject is ~lministered nutritional and other therapeutic agents to aid in the increase of bone mass. Such optional agents include, for example, Vitamin D and calcium.
In the methods of this invention, "~ministering" refers to any method which, in sound medical practice, delivers the actives used in this invention to the subject to be treated in such a manner so as to be effective in the building of bone. The actives may be adminictçred by any of a variety of known methods of a~mini~tration, e.g., orally, dermatomucosally (for example, dermally, sublingually, intranasally, andrectally), parenterally (for example, by subcutaneous injection, intr~ml'sc~ r injection, intra-articular injection, intravenous injection), and by inhalation. Thus, specific modes of a~ministration include, but are not limited to, for example, oral, transdermal, mucosal, sublingual, intrAml'scul~r~ intravenous, inll~peli~oneal, subcutaneous administration, and topical application.
A prefcl,ed method for the tre~tmPnt of osteoporosis in~llldes an initial diagnostic step, to determine the presence of the disorder. Thus, a prt;re- . ed method of this invention comprises the steps of pe.r~"..ln~g a diagnostic on a human subject for the detection of osteoporosis and, upon obtahhng a positive result from saiddiagnostic, ~.iminictering the actives according to the methods of this invention. For such methods for lteAU~ of posl-,l.,nopa.lsal female subjects prior to signific~nt bone loss, said initial diagnostic step complises pelrolll.il-g a diagnostic ford~ ,ng menopause. Such methods are well known in the art, and include, for example, dele:-lluna~ion of the bone mass and rate of bone r~mod~ling The rate of bone remodeling can be d~;L~l",ned by measurement of bioc~ernic~l markers. See, Hui, et al., "The Contribution of Bone Loss to Po~t,l,enopd.lsal Osteopolosis,"
1 Osteoporosis Int. 30 (1990), incol~G,~ed by reference herein.
Suitable diagnostics for the detection of established osteoporosis are also well known in the art. Such methods include the measurement of the radiodensity of skeletal radiographs, ql~ \re computerized tomography, single energy photon absorptiometry, dual-energy photon absol ~"iometry, dual energy X-ray WO96107418 ~ ~ ~ 9 ~ 2 5 1 PCT/US95/11336 absorptiometry, and quantitative digital radiography Diagnostic techniques amongthose useful herein are described in W. A. Peck et al., Physician's Resource Manual on Osteoporosis ( 1987), published by the National Osteoporosis Foundation (incorporated by reference herein).
Dosage Forms:
The antiresorptive compound and parathyroid hormone may be a~mini~tered in any of a variety of pharmaceutically-acceptable compositions. Such compositions may comprise an active and a pharm~ceutically-acceptable carrier. Pharmaceutically-acceptable carriers include solid or liquid filler diluents or encapsulating substances, and mixtures thereof, that are suitable for a~mini~tration to a human or lower animal.
The term "compatible", as used herein, means that the components of the pharmaceutical composition are capable of being commingled with the actives, andwith each other, in a manner such that there is no interaction which would substantially reduce the pharrn~ceutical efficacy of the pharmaceutical composition under ordinary use situations. Pharrn~ceutically-acceptable carriers must, of course, be of sufficiently high purity and sufficiently low toxicity to render them suitable for ~mini~tration to the human or lower animal being treated.
Some examples of the substances which can serve as pharrn~ceutical carriers are: sugars, such as lactose, glucose and sucrose; starches, such as corn starch and potato starch; cellulose and its derivatives, such as sodium carboxymethylcellulose, ethylcellulose, cellulose acetate; powdered trag~c~ntll, malt; gelatin; talc; stearic acid;
magnesium stearate; 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; phosphate buffer solutions; wetting agents and lubricants such as sodium lauryl sulfate; coloring agents; flavoring agents; and preservatives. Other compatible pharm~ceutic~l additives and actives may be included in the pharmaceutically-acceptable carrier for use in the compositlons of the present mventlon.
The choice of a pharmaceutically-acceptable carrier to be used in conjunction with the active is determined by the way the active is to be a~lmini~tered. If the active is to be injected, the plcfcl-cd pharrn~ceutic~l carrier is sterile water, physiological saline, or mixtures thereof. The pH of such parenteral composition is preferablyadjusted to about 7.4. Suitable pharmaceutically-acceptable carriers for topicalapplication include those known in the art for use in creams, gels, tapes, patches, and similar topical delivery means.
The pharmaceutically-acceptable carrier employed in conjunction with the WO 96/07418 ~ ~ 2 ~ ~ PCT/US95/11336 actives is used at a concentration sufficient to provide a practical size to dosage relationship. The pharmaceutically-acceptable carriers, in total, may comprise from about 0.1% to about 99.9% by weight of the pharm~ceLltical compositions of the present invention, preferably from about 5% to about 80%, and most preferably from about 10% to about 50%.
A prere~led method of administering bisphosphonates is orally, in a unit-dosage form (i.e., a dosage form containing an amount of active suitable for ?~minictration in one single dose, according to sound medical practice). Preferred unit dosage forms for bisphosphonate include tablets, capsules, suspensions, andsolutions, comprising a safe and effective amount of active. Pharmaceutically-acceptable carriers suitable for the p,epara~ion of unit dosage forms for oral ~minictration are well known in the art. Their 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 difficulty by a person skilled in the art. Preferably, oral unit dosage forms of the bone-active phosphonate comprise from about 0.0005 mgP/kg oral per day to about 1.0 mgP/kg oral per day of the phosphonate.
A prefe"ed method of ~mini~tçrjng calcitonin compounds and parathyroid horrnone is via subcutaneous injection in a unit dosage form. Plere-led unit dosagé
forms for injection include sterile solutions of water, physiological saline, or mixtures thereof. The pH of said solutions should be adjusted to about 7.4. Preferably, unit dosage forms of parathyroid hormone comprise from about 4 IU to about 15 IU per kg per day.
Other p.er~. . ed dose forms for parathyroid and calcitonin compounds include nasal, trandsermal, rectal, sublingual, and oral. Plere--ed oral forms include, for example, lipQsomes, lipid emulsions, and proteinaceous cages.

Kits:
This invention also provides kits for conveniently and effectively implementing the methods of this invention. Such kits comprise one or more unit doses of antiresorptive compound, one or more unit doses of parathyroid hormone,'!I and a means for f~cilit~tin~ compliance with methods of this invention. Such kits provide a convenient and effective means for assuring that the subject to be treated takes the applop.iate active in the correct dosage in the correct manner. The compliance means of such kits includes any means which f~ it~tes admini~tering the actives according to a method of this invention. Such compliance means includçs instructions, p~c~ing and dispensing means, and col,lbind~ions thereof. Examples WO96/07418 al 2 1 ~ ~ 2 5 ~ PCT/US95/11336 of packaging and dispensing means are well known in the art, including those described in U.S. Patents 4,761,406, Flora et al., issued August 2, 1988; and U.S.
Patent 4,812,311, Uchtman, issued March 14, 1989 and U.S. 4,833,125, Neer et al., issued May 23, 1989, all incorporated by reference herein.
The following non-limiting examples illustrate the compositions, processes and uses of the present invention.

An Asian male human patient weighing approximately 65 kg and diagnosed with idiopathic osteoporosis is treated by a method of this invention. Specifically, for a period of four months, the patient is administered the bisphosphonate, 2-(3-pyridyl)-l-hydroxyethane-l,l-bisphosphonic acid. The patient is orally admini.~tered one tablet per day, with each tablet cont~ining 0.002 mgP/kg per day of the bisphosphonate. The bisphosphonate treatment is discontinued. Then, for the nextsix months, the patient is atlminictçred parathyroid hormone (human synthetic fragment 1-34, or h PTH 1-34). The hormone is subcutaneously ~tlminictered at a dose of 13 IU/kg via insulin syringe to the anterior thigh for five days out of every week during the six-month period.
A biopsy of iliac crest bone is taken and reveals an increase in mean wall thickness of the remodeling units (BMU) compared to her baseline biopsy. The activation frequency and depth of resorption cavities on cancellous, cortical and endocortical surfaces are not significantly increased above the values observed at baseline. Bone mineral density is measured, indicating an increase of 11%.

A human Callc~ci~n female patient weighing app,o~",ately 60 kg and diagnosed with postmenopausal osteoporosis is treated by a method of this invention.
Specifically, the patient is ~rlminictered conjugated estrogen for a period of four months. The estrogen is a~ministered via a transdermal patch, delivering estrogen at a level of 0.625 mg per day. A~er the four month period, the estrogen ~minictration is contin~ed However, the patient is then also ~minictered parathyroid horrnone (human synthetic fragment 1-34, or hPTH 1-34) for six months, as a daily nasal spray delivering 5 IU/kg.
A blood sample is then obtained and analyzed for the bone specific marker, osteocalcin, and bone-derived and total alkaline phosphatase. Osteocalcin values are increased by 57% and both bone and total alkaline phosphatase are slightly elevated compared to pl~ values. Base mineral density is measured, indicating an WO96/07418 ID 2 ~ ~ ~ 2 5 1 PCTIUS95/11336 increase of 10%.

Claims (13)

WHAT IS CLAIMED IS:

1. A method of treating a human or other animal subject having a bone metabolism disorder, comprising the steps of:
(a) administering to said subject a safe and effective amount of an antiresorptive compound, during a period of from about 2 weeks to about 6 months.
(b) administering to said subject a safe and effective amount of a parathyroid hormone, during a period of from about 3 to 12 months.

2. A method of treating a human or other animal subject, according to Claim 1, wherein, in step (b), said parathyroid hormone is administered for about 6 months.

3. A method of treating a human or other animal subject, according to Claim 1, wherein said steps (a) and (b) are repeated from 1 to 6 times.

4. A method of treating a human or other animal subject, according to Claim 3, additionally comprising administration of an antiresorptive compound during step (b).

5. A method of treating a human or other animal subject, according to Claim 4, wherein, in step (a), said antiresorptive compound is administered for from about 2 months to about 6 months.

6. A method of treating a human or other animal subject, according to Claim 5, wherein said antiresorptive compound is an estrogen compound.

7. A method of treating a human or other animal subject, according to Claim 6, wherein said estrogen compound is a conjugated estrogen, and is administered at a level of from about 0.3 mg to about 1.25 mg per day.

8. A method of treating a human or other animal subject, according to Claim 5, wherein said antiresorptive compound is a calcitonin compound.

9. A method of treating a human or other animal subject, according to Claim 5, wherein said antiresorptive compound is a phosphonate compound.

10. A method of treating a human or other animal subject, according to Claim 9, wherein said phosphonate compound is a bisphosphonate selected from the group consisting of oxyethane- 1,1-bisphosphonic acid; dichloromethane bisphosphonic acid;
3-amino- 1 -hydroxypropane- 1,1-bisphosphonic acid; 6-amino- 1 -hydroxyhexane- 1, 1-bisphosphonic acid; 4-amino-1-hydroxybutane-1,1-bisphosphonic acid; 2-(3-pyridyl)-1 -hydroxyethane- 1, 1 -bisphosphonic acid; 2-(N-imidazoyl)- 1 -hydroxyethane-1,1 -bisphosphonic acid; 3-(N-pentyl-N-methylamino)- 1 -hydroxypropane- 1, 1 -bisphosphonic acid; 3-(N-pyrollidino)-1-hydroxypropane-1,1-bisphosphonic acid;
N-cycloheptylaminomethanebisphosphonic acid; S-(p-chlorophenyl) thiomethanebisphosphonic acid; (7-dihydro-1-pyrindine)methane bisphosphonic acid;
(7-dihydro- 1 -pyrindine)hydroxymethane bisphosphonic acid; (6-dihydro-2-pyrindine)hydroxymethanebisphosphonic acid; 2-(6-pyrolopyridine)-1-hydroxyethane- 1,1 -bisphosphonic acid; and pharmaceutically-acceptable salts and esters thereof.

11. A method of treating a human or other animal subject, according to Claim 10,wherein said bisphosphonate is 2-(3-pyridyl)- 1 -hydroxyethane- 1,1 -bisphosphonic acid.

12. A method of treating a human or other animal subject having a bone-metabolism disorder, comprising the steps of:
(a) administering to said subject a safe and effective amount of an antiresorptive compound, during a period of from about 2 weeks to about 1 month;
(b) administering to said subject a safe and effective amount of a parathyroid hormone compound, during a period of from about 3 to about 6 months.
(c) administering to said subject a safe and effective amount of an antiresorptive compound, during a period of from about 3 months to about 6 months;
(d) administering to said subject a safe and effective amount of a parathyroid hormone compound, during a period of from about 3 to about 6 months.

13. A method of treating a human or other animal subject, according to Claim 12,wherein said steps (c) and (d) are repeated from 1 to 5 times.