WO1995024207A1 - Composition for the treatment of lung disease - Google Patents

Abstract

Disclosed are pharmaceutical compositions for the treatment of lung disease, and in particular Cystic Fibrosis, comprising a substituted azetidinone of general formula (I), which have been found to be potent elastase inhibitors and a (F)-actin shortening protein, such as gelsolin.

Description

TITLE OF THE INVENTION

COMPOSITION FOR THE TREATMENT OF LUNG DISEASE

BACKGROUND OF THE INVENTION

This invention is directed to pharmaceutical compositions comprising an elastase inhibitor and an (F)-actin shortening protein such as gelsolin or vitamin D binding protein (DBP), for use in the treatment of Cystic Fibrosis and related diseases. In particular, this invention is directed to pharmaceutical compositions comprising elastase inhibitors of Formula I

and (F)-actin shortening proteins, Gelsolin being preferred.

We have found that a group of new substituted azetidinones are potent elastase inhibitors and therefore are useful anti-inflammatory and antidegenerative agents.

Proteases from granulocytes and macrophages have been reported to be responsible for the chronic tissue destruction mechanisms associated with inflammation, including rheumatoid arthritis and emphysema. Accordingly, specific and selective inhibitors of these proteases are candidates for potent anti-inflammatory agents useful in the treatment of inflammatory conditions resulting in connective tissue destruction, e.g., rheumatoid arthritis, emphysema, bronchial

inflammation, chronic bronchitis, glomerulonephritis, osteoarthritis, spondylitis, lupus, psoriasis, atherosclerosis, sepsis, septicemia, shock, myocardial infarction, reperfusion injury, periodontitis, cystic fibrosis and acute respiratory distress syndrome. The role of proteases from granulocytes, leukocytes or macrophages are related to a rapid series of events which occurs during the progression of an inflammatory condition:

(1) There is a rapid production of prostaglandins (PG) and related compounds synthesized from arachidonic acid. This PG synthesis has been shown to be inhibited by aspirin-related nonsteroidal anti-inflammatory agents including indomethacin and phenylbutazone. There is some evidence that protease inhibitors prevent PG production;

(2) There is also a change in vascular permeability which causes a leakage of fluid into the inflamed site and the resulting edema is generally used as a marker for measuring the degree of inflammation. This process has been found to be induced by the proteolytic or peptide cleaving activity of proteases, especially those contained in the

granulocyte, and thereby can be inhibited by various synthetic protease inhibitors, for example, N-acyl benzisothiazolones and the respective

1,1-dioxides. Morris Zimmerman et al., J. Biol. Chem., 255, 9848 (1980); and

(3) There is an appearance and/or presence of lymphoid cells, especially macrophages and polymorphonuclear leukocytes

(PMN). It has been known that a variety of proteases are released from the macrophages and PMN, further indicating that the proteases do play an important role in inflammation.

In general, proteases are an important family of enzymes within the peptide bond cleaving enzymes whose members are essential to a variety of normal biological activities, such as digestion, formation and dissolution of blood clots, the formation of active forms of hormones, the immune reaction to foreign cells and organisms, etc., and in pathological conditions such as the degradation of structural proteins at the articular cartilage/pannus junction in rheumatoid arthritis etc.

Elastase is one of the proteases. It is an enzyme capable of hydrolyzing the connective tissue component elastin, a property not contained by the bulk of the proteases present in mammals. It acts on a protein's nonterminal bonds which are adjacent to an aliphatic amino acid. Neutrophil elastase is of particular interest because it has the broadest spectrum of activity against natural connective tissue

substrates. In particular, the elastase of the granulocyte is important because, as described above, granulocytes participate in acute

inflammation and in acute exacerbation of chronic forms of

inflammation which characterize many clinically important

inflammatory diseases.

Proteases may be inactivated by inhibitors which block the active site of the enzyme by binding tightly thereto. Naturally

occurring protease inhibitors form part of the control or defense mechanisms that are crucial to the well-being of an organism. Without these control mechanisms, the proteases would destroy any protein within reach. The naturally occurring enzyme inhibitors have been shown to have appropriate configurations which allow them to bind tightly to the enzyme. This configuration is part of the reason that inhibitors bind to the enzyme so tightly (see Stroud, "A Family of Protein-Cutting Proteins" Sci. Am., July 1974, pp. 74-88). For example, one of the natural inhibitors, αi-Antitrypsin, is a glycoprotein contained in human serum that has a wide inhibitory spectrum covering, among other enzymes, elastase both from the pancreas and the PMN. This inhibitor is hydrolyzed by the proteases to form a stable acyl enzyme in which the active site is no longer available. Marked

reduction in serum αi-antitrypsin, either genetic or due to oxidants, has been associated with pulmonary emphysema which is a disease

characterized by a progressive loss of lung elasticity and resulting respiratory difficulty. It has been reported that this loss of lung elasticity is caused by the progressive, uncontrolled proteolysis or destruction of the structure of lung tissue by proteases such as elastase released from leukocytes. J. C. Powers, TIBS, 211 (1976).

Rheumatoid arthritis is characterized by a progressive destruction of articular cartilage both on the free surface bordering the joint space and at the erosion front built up by synovial tissue toward the cartilage. This destruction process, in turn, is attributed to the protein-cutting enzyme elastase which is a neutral protease present in human granulocytes. This conclusion has been supported by the following observations:

( 1 ) Recent histochemical investigations showed the accumulation of granulocytes at the cartilage/pannus junction in rheumatoid arthritis; and

(2) a recent investigation of mechanical behavior of cartilage in response to attack by purified elastase demonstrated the direct participation of granulocyte enzymes, especially elastase, in rheumatoid cartilage destruction. H. Menninger et al., in Biological Functions of Proteinases, H. Holzer and H. Tschesche, eds. Springer-Verlag, Berlin, Heidelberg, New York, pp. 196-206, 1979.

(F)-actin shortening proteins including gelsolin, plasma gelsolin (brevin), vitamin D binding protein (DBP), villin, fragmin, and severin, and their use in shortening (F)-actin and treating (F)-actin mediated diseases is described in U.S. 5,260,224, issued to Stossel et al, on November 9, 1993, which patent is hereby incorporated by

reference.

As discussed therein, Gelsolin has been cloned (Kwialkowski, D. J. et al, Nature, 323:455-458 (1986); Kwialkowski, D. J. et al, J. Cell Biol., 106:375-384 (1988)) and fragments of the native protein which retain the ability to bind actin have been identified (Bryan, J., J. Cell Biol., 106:1553-1562 (1988); Yin, H. L. et al., J. Cell Biol., 107:465a (1988), abst. no. 2616); Kwialkowski, D. J. et al., J. Cell Biol., 108:1717-1726 (1989); Way, M. et al., J. Cell Biol.,

109:593-605 (1989)).

Gelsolin's primary function in the plasma is to sever actin filaments. If gelsolin is present in excess of actin, only gelsolin-actin complexes result; if actin is in excess, there are free actin oligomers and gelsolin-actin complexes. The actin severing occurs by way of a non-proteolytic cleavage of the noncovalent bond between adjacent actin molecules.

Efficacious levels of actin-binding compounds may be administered so as to provide therapeutic benefits against the secondary toxic effects of excessive extracellular actin. By "efficacious levels" of actin-binding compounds is meant levels in which the toxic effects of free extracellular actin are, at a minimum, ameliorated. By "excessive" extracellular actin is meant an amount of extracellular actin which exceeds the ability of the plasma proteins to bind and clear the actin from extracellular fluids without secondary tissue damage or toxic effects. By "secondary" tissue damage or toxic effects is meant the tissue damage or toxic effects which occur to otherwise healthy tissues, organs, and the cells therein, due to the presence of excessive

extracellular actin in the plasma, usually as a result of a "primary" tissue injury elsewhere in the body.

Stossel et al., recently presented a paper and abstract entitled Filamentous (F)-Actin Is Abundant In Cf Sputum, And F-Actin-Shortening Proteins Diminish Sputum Viscosity In Vitro. Stossel et al., found actin in CF sputum by immunoblotting, and CF sputum samples accelerated the nucleation of monumeric actin, consistent with the presence of F-actin. Stossel et al., speculated that F-actin therefore might contribute importantly to the mechanics of CF sputum. In a test model F-actin severing proteins instantaneously and substoichiometrically shorten actin filaments, and nM concentrations of one such protein, human plasma gelsolin, rapidly reduced the viscosity of CF sputum by up to 70% in a concentration-dependent manner. Gelsolin also diminished the elastic modulus of CF sputum by 50% in shear deformation over a frequency range of 0.001-10 Hz. Over ten times more bovine pancreatic DNAse I (which binds actin subunits in addition to its DNA hydrolyzing activity) was required to diminish CF sputum viscosity by the same extent as gelsolin.

More recently, Vasconcellos et al., have reported that concentrations of 100 to 500 nM, gelsolin, purified from human plasma, rapidly diminished the viscosity of sputum form cystic fibrosis patients an average of 64% from 3321199 Pa-s over a 60 minute incubation period. On the other hand, at a concentration of 250 nM, bovine pancreatic DNase I and Gc golbulin had no effect on viscosity in this time period, however, they did appear to enhance gelsolin activity. See Science, Vol. 263, pp 969-971 (February 18, 1994). BRIEF DESCRIPTION OF THE INVENTION

This invention relates to pharmaceutical compositions and methods for the treatment of lung disease, and in particular Cystic

Fibrosis comprising the a non-toxic effective amount of an elastase inhibitor such as the compounds of Formula (I),

a non-toxic effective amount of a (F)-actin shortening protein, such as gelsolin, and a pharmaceutically acceptable carrier.

DETAILED DESCRIPTION OF THE INVENTION

This invention relates to pharmaceutical compositions and methods for the treatment of lung disease, and in particular Cystic

Fibrosis comprising the a non-toxic effective amount of an elastase inhibitor such as the compounds of Formula (I),

a non-toxic effective amount of a (F)-actin shortening protein, such as gelsolin, and a pharmaceutically acceptable carrier.

With regard to the compounds of Formula I, said compounds being further detailed in the Schemes, Examples and Claims, the instant invention is more particularly directed to the compounds of the

and pharmaceutically acceptable salts thereof wherein:

R is C_1-6alkyl;

R¹ isC_1-6alkyl orC_1-6alkoxy-C_1-6alkyl; M is

(1) hydrogen,

(2) C_1-6alkyl,

(3) hydroxy C_1-6alkyl,

(4) halo C_1-6alkyl,

(5) C_2-6alkenyl, or

(6) C_1-6alkoxy-C_1-6alkyl;

Ra and Rb are each individually

(1) hydrogen,

(2) C_1-6alkyl,

(3) halo,

(4) carboxy,

(5) C_1-6alkoxy, (6) phenyl,

(7) C_1-6alkylcarbonyl,

(8) di-(C_1-6alkyl)amino,

(9) hydroxy;

R² and R³ are each independently

(1) hydrogen,

(2) C_1-6alkyl,

(3) halo,

(4) carboxy,

(5) C_1-6alkoxy, (6) phenyl,

(7) C_{1 -6}alkylcarbonyl,

(8) aminoC_2-3alkyIoxy carbonyl wherein the

amino is optionally mono or di substituted with C_{1 -6}alkyl,

(9) aminoC_2-3alkylamino carbonyl wherein the amino is optionally mono or di substituted with C_1-6alkyl,

(10) hydroxy,

(11) aminocarbonyl wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl,

(12) hydroxymethyl,

(13) aminocarbonyloxy C_{1 -3}alkyloxy wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl,

(14) cyano,

(15) morpholinocarbonylphenyl,

(16) amino wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl,

with the proviso that R² and R³ may be joined together to form a methylenedioxy group or a furan ring,

(17) morpholinocarbonyl;

R₄ is (a) , or

(b)

where R_x is carboxy

C_1-6 alkyl, benzyloxycarbonylC_1-3alkyl, or

t-butoxycarbonylC_1-3alkyl, wherein

Q is a covalent bond or

wherein R₅ and R₆ are each individually C_1-3alkyl or hydrogen,

Y is

or

or a covalent bond;

R₁₂ is hydrogen orC_1-3alkyl;

R₇ and R₈ are each individually

(a) hydrogen,

(b) C_1-6alkyl,

(c) C_1-6alkyloxy C_2-3alkyl,

(d) hydroxy C_2-6alkyl,

(e) polyhydroxyC_2-6alkyl, (f) carboxamido C_{1 -6}alkyl,

(g) polyacyloxyC_{2 -6}alkyl,

(h) C_{1 -6}alkanoyl,

(i) substituted phenyl or phenyl C_{1 -6}alkyl,

wherein the substituent is X₁ as defined immediately below,

(j) C_{2 -6}alkenyl,

(k) C_6-10cycloalkenyl,

(l) heteroaryl C_{1 -6}alkyl wherein the hetero aryl includes pyridinyl, imidazolyl, triazolyl, benzylimidazolyl, and furyl,

(m) carboxy C_{1 -6}alkyl,

(n) carbo C_{1 -6}alkoxy C_{1 -3}alkyl,

(o) phenylsulfonyl,

(p) C_{1 -6}alkylsulfonyl,

(q) benzyloxy,

(r) morpholinyl C_{1 -3}alky lsulfonyl,

(s) tetrahydropyranyl,

(t) aminoC_{1 -3}alkylsulfonyl wherein the amino is optionally mono or di substituted with

C_{1 -6}alkyl,

(u) aminocarbonyl wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl,

(v) aminocarbonyloxyC_{2 -6}alkyl wherein the

amino is optionally mono or di substituted with C_{1 -6}alkyl,

(w) azabicyclo of 7 to 12 atoms,

(x) di C_{1 -3}alkylamino C_{2 -6}alkyl wherein the

amino is optionally mono or di substituted with C_{1 -6}alkyl,

(y) bicycloalkyl of 7 to 12 atoms,

(z) C_3-10cycloalkyl optionally substituted with C_{1 -6}alkyl,

(aa) pyrazolidinyl, (bb) substituted piperidinyl or prrolidinyl wherein the substituent is hydrogen, C_{1 -3}alkyl, hydroxyC_{1 -3}alkylbenzyl, carboxamido or amino wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl,

(cc) substituted pyrrolidinyl wherein the substituent is carboxamido or amino wherein the amino is optionally mono or di substituted with C₁- 6alkyl,

(dd) pyrimidinyl,

(ee) N-cyano-N'-phenylamidino,

(ff) phosphonoC_{1 -6}alkyl, or

(gg) α-C_{1 -3}alkyl benzyl or mono or di substituted benzyl or mono or di substituted pyridylmethyl, wherein the substituents are X₁ and

X₂,

wherein

X₁ is

(1) hydrogen,

(2) halo,

(3) C_{1 -6}alkyl,

(4) halo-C_{1 -6}alkyl,

(5) C_{2 -6}alkenyl,

(6) hydroxy-C_{1 -6}alkyl,

(7) C_{1 -6}alkylcarbonyl,

(8) C_{1 -6}alkylcarbonylamino,

(9) CN,

(10) CF₃,

(11) CH₃O,

(12) amino wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl,

(13) carboxy, or

(14) phenylsulfonylaminocarbonyl;

X₂ is hydrogen, halo or C_{1 -6}alkyl; n is 1, 2, 3, 4 or 5;

R₉ is selected from hydrogen, C_{1 -4} alkyl, and C_{1 -3}alkoxyC_{1 -3}alkyl; or phenyl, phenyl C_{1 -3}alkyl, pyridyl, and pyridyl C_{1 -3}alkyl;

R₁₀ and R_{1 1} are each independently selected from

hydrogen, C_{1 -4}alkyl, and C_{1 -3}alkoxy C_{1 -3}alkyl, or aryl as defined above, or are together O=; or wherein R₇ and R₈ are joined together to form mono or di substituted ring of 4, 5, 6, or 7 atoms or 7 to 12 atoms such as

(1) piperidinyl or homopiperdinyl,

(2) piperazinyl,

(3) morpholinyl, thiomorpholinyl or 1,1-dioxo-4- thiomorpholinyl,

(4) pyrroylidinyl,

(5) pyrryl,

(6) imidazolyl,

(7) triazolyl,

(8) saturated azabicyclo of 7 to 12 atoms,

(9) azaspiro having 3 to 9 carbon atoms, said ring being saturated,

(10) tetrazolyl,

(11) pyrazolidinyl,

(12) dihydodimethoxyisoquinolyl,

(13) azetidinyl, or

(14) diazabicyclo ring of 7-12 atoms,

wherein the substituents are each selected from the group consisting of hydrogen and C_{1 -3}alkyl, benzyloxycarbonyl, carboxy, phenyl C_{1 -3}alkyl amino carbonyl, pyrrolidinylmethyl, hydroxy C_{1 -3}alkyl, C_{1 -6}alkyloxy, C_{1 -4}alkyloxy carbonyl, aminocarbonyl wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl, and oxo; or

-N(R7)R8 may be an amino acid residue including natural amino acids such as lysine; or R₈ and R₉ are joined together to form a mono or di substituted saturated monocyclic ring of 6 to 7 atoms and having two hetero atoms which are the nitrogens to which R₈ and R₉ are attached; said rings to include piperazinyl and homopiperazinyl; or R₉ and R₁₀ are joined together to form a mono or di substituted monocyclic saturated ring of 5 to 7 atoms and having one hetero atom which is the nitrogen to which

R₉ is attached; or

wherein R₉ and R₁₂ are joined together to form a mono or di substituted saturated monocyclic ring of 5, 6, or 7 atoms, said ring having one hetero atom which is the nitrogen to which R₉ is attached; or

wherein R₁₀ and R₁₂ are joined together to form a mono or di substituted saturated monocyclic ring of 5, 6, or 7 carbon atoms; or wherein R₈ and R₁₁ are joined together to form a mono or di substituted saturated monocyclic ring of 5, 6, or 7 atoms, said ring having one hetero atom which is the nitrogen to which R₈ is attached; and the substituents are independently selected from Hydrogen and C_{1 -3}alkyl.

As appreciated by those of skill in the art the term "alkyl" such as in C_{1 -6}alkyl, includes, methyl, ethyl, propyl, butyl, pentyl, and hexyl, and where appropriate, branched chained forms including isopropyl and tert-butyl.

As may also be appreciated by those of skill in the art, the

spacer in definition Y, may, in the alternative be placed to the right of CR₁₀R₁₁.

As may also be appreciated, the group

may also be oxidized to the corresponding oxide

In one Class the instant invention is directed to the compounds of the Formula (I)

and pharmaceutically acceptable salts thereof wherein:

R is C_1-6alkyl;

R¹ is C_1-6alkyl or C_1-6alkoxy-C_1-6alkyl;

M is

(1) hydrogen,

(2) C_1-6alkyl,

(3) hydroxy C_1-6alkyl,

(4) halo C_1-6alkyl,

(5) C_2-6alkenyl, or

(6) C_1-6alkoxy-C_1-6alkyl;

Ra is

(1) hydrogen,

(2) C_1-6alkyl,

(3) halo,

(4) carboxy,

(5) C_1-6alkoxy,

(6) phenyl,

(7) C_1-6alkylcarbonyl,

(8) amino wherein the amino is optionally mono or di substituted with C_1-6alkyl;

Rb is hydrogen, or C_1-6alkyl,

R² and R³ are each independently

(1) hydrogen,

(2) C_1-6alkyl,

(3) halo,

(4) carboxy, (5) C_{1 -6}alkoxy,

(6) phenyl,

(7) C_{1 -6}alkylcarbonyl,

(8) amino wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl, or with the proviso that R² and R³ may be joined together to form a methylenedioxy group or a furan ring;

R ₄ is or

where R_x is carboxy-C_{1 -6}alkyl,

benzyloxycarbonylC_{1 -3}alkyl, or

t-butoxycarbonylC_{1 -3}alkyl, wherein

Q is a covalent bond or

wherein R₅ and R₆ are each individually C_{1 -3}alkyl or hydrogen Y is

or

or a covalent bond; R₁₂ is hydrogen or C_1-3alkyl;

R₇ and R₈ are each individually

(a) hydrogen,

(b) C_1-6alkyl,

(c) C_1-6alkyloxyC_2-3alkyl,

(d) hydroxy C_2-6alkyl,

(e) carboxamido C_1-6alkyl,

(f) C_1-6alkanoyl,

(g) substituted phenyl or phenyl C_1-6alkyl

wherein the substituents are X₁, and X₂,

(h) C_2-6alkenyl,

(i) C_6-10cycloalkenyl,

G) heteroaryl C_1-6alkyl wherein the hetero aryl includes pyridinyl, imidazolyl, triazolyl, benzylimidazolyl, and furyl,

(k) carboxy C_1-6alkyl,

(l) C_l-6alkylsulfonyl,

(m) carboC_1-6alkyloxyC_2-3alkyl,

(n) morpholinyl C_1-3alkylsulfonyl,

(o) aminoC_1-3alkylsulfonyl wherein the amino is optionally mono or di substituted with

C_{1 -6}alkyl,

(p) aminocarbonyl wherein the amino is optionally mono or di substituted with C_{1 -6}alkyl,

(q) aminocarbonyloxyC_{1 -6}alkyl wherein the

amino is optionally mono or di substituted with C_{1 -6}alkyl,

(r) di C_{1 -3}alkylamino C_{1 -6}alkyl wherein the

amino is optionally mono or di substituted with C_{1 -6}alkyl,

(s) pyrazolidinyl,

(t) substituted piperidinyl as defined above, (u) substituted pyrrolidinyl as defined above, (v) pyrimidinyl,

(w) benzyloxy,

(x) C_3-10cycloalkyl,

(z) α-C_{1 -3}alkyl benzyl or mono or di substituted benzyl or mono or di substituted

pyridylmethyl, wherein the substituents are X₁ and X₂,

wherein

X₁ is

(1) hydrogen,

(2) halo,

(3) C_{1 -6}alkyl,

(4) halo-C_{1 -6}alkyl,

(5) C_{2 -6}alkenyl,

(6) hydroxy-C 1 -6alky 1,

(7) C_{1 -6}alkylcarbonyl,

(8) C_{1 -6}alkylcarbonylamino,

(9) di-C_{1 -3}alkylamino, or

(10) carboxy;

X₂ is hydrogen, halo or C_{1 -6}alkyl; n is 1, 2, 3, 4 or 5;

R₉ is selected from hydrogen, C_{1 -4} alkyl, and

C_{1 -3}alkoxyC_{1 -3}alkyl;

R₁₀ and R_{1 1} are each independently selected from hydrogen, C_{1 -4}alkyl, and C_{1 -3}alkoxy C_{1 -3}alkyl; or wherein R₇ and R₈ are joined together to form mono or di substituted ring of 4, 5, 6, or 7 atoms such as

(1) piperidinyl,

(2) piperazinyl,

(3) morpholinyl,

(4) pyrroylidinyl,

(5) pyrryl,

(6) imidazolyl,

(7) triazolyl,

(8) tetrazolyl,

(9) pyrazolidinyl,

(10) azetidinyl,

wherein the substituents are each selected from the group consisting of hydrogen and C_{1 -3}alkyl, benzyloxycarbonyl, carboxy, phenyl C_{1 -3}alkyl amino carbonyl, pyrrolidinyl, methyl, hydroxy C_{1 -3}alkyl,

C_{1 -6}alkyloxy, C_{1 -4}alkyloxy carbonyl, and oxo; or

R₈ and R₉ are joined together to form a saturated ring of 5 to 7 atoms and having two hetero atoms; or

R₉ and R₁₀ are joined together to form a saturated ring of 5 to 7 atoms and having one hetero atom; or

wherein R₉ and R₁₂ are joined together to form a ring of 5, 6, or 7 atoms, said ring being saturated; or wherein R₁₀ and R₁₂ are joined together to form a ring of 5, 6, or 7 atoms, said ring being saturated; or

wherein R₈ and R_{1 1} are joined together to form a ring of 5, 6, or 7 atoms, said ring being saturated and having one hetero atom. In one subclass, the invention concerns compounds of

Formula I

wherein

R is C_1-3alkyl;

R₁ is C_1-3alkyl;

M is

(a) C_1-6alkyl, or

(b) C_2-6alkenyl;

R² is

(a) hydrogen,

(b) C_1-6alkyl, or C_1-6alkoxy, and

R³ is hydrogen, or

R² and R³ are joined together to form a methylenedioxy group or a furan ring;

R₅ and R₆ are each individually hydrogen or C_1-3alkyl;

R₇ and R₈ are each independently selected from

(a) hydrogen,

(b) C_1-3alkyl,

(c) C_1-3alkoxyC_2-3alkyl,

(d) C_3-7cycloalkyl,

(e) hydroxyC_2-3alkyl,

(d) carbo C_1-4alkyloxymethyl,

(g) substituted benzyl wherein the substituents are

X₁ and X₂

wherein X₁ is hydrogen and X₂ is

(1) hydrogen,

(2) halo, or

(3) C_1-3alkyl; n is 1, 2 or 3, and

R₉, R₁₀ and R₁₁ are each independently selected from hydrogen, C_1-4alkyl, and C_1-3alkoxy

C_1-3alkyl; or R₇ and R₈ are joined together to form a substituted ring selected from

(a) piperidinyl,

(b) piperazinyl, and

(c) morpholinyl;

or

R₈ and R₉ are joined together to form a ring of 6 to 7 atoms and having two hetero atoms;

R₉ and R₁₀ are joined together to form a saturated ring of 5 to 7 atoms and having one hetero atom; or

wherein R₉ and R₁₂ are joined together to form a ring of 5, 6, or 7 atoms, said ring being saturated; or

wherein R₁₀ and R₁₂ are joined together to form a ring of 5, 6, or 7 atoms, said ring being saturated;

or

wherein R₈ and R_{1 1} are joined together to form a ring of 5, 6, or 7 atoms, said ring being saturated and having one hetero atom.

In a narrower sub-class are the compounds wherein

Q is a covalent bond;

R is methyl or ethyl;

R₁ is methyl or ethyl;

M is

(a) C_{1 -4}alkyl, or

(b) C_{2 -3} alkenyl;

R² is

(a) hydrogen,

(b) C_{1 -3}alkyl, or C_{1 -3}alkoxy, and

R³ is hydrogen, or

R² and R³ are joined together to form a furan or

dioxacyclopentane ring;

n is 1 or 2;

R₉ and R₁₀ are each independently selected from (a) C_{1 -3}alkyl,

(b) C_{1 -3}alkoxy C_{1 -3}alkyl,

(c) hydrogen,

R₇ and R₈ are each independently selected from

(a) C_{1 -3}alkyl,

(b) C_{1 -3}alkoxyC_{2 -3} alkyl,

(c) hydrogen,

(d) hydroxy ethyl,

(e) carboethoxymethyl,

(f) cyclopropyl,

or

R₇ and R₈ are joined together to form a substituted ring selected from

(a) piperidinyl, and

(b) morpholinyl, or

R₈ and R₉ are joined together to form a piperazine ring.

As is defined above, various rings are formed when R₈, R₉, R₁₀ and R₁₂ are joined. The following is a non-limiting description of some of the preferred rings that are formed when these various substituents are joined.

R₈ and R₉ are joined

R₉ and R₁₀ are joined

R₉ and R₁₂ are joined

R₁₀ and R₁₂ are joined

R₁₀ and R₁₂ are joined

As discussed above, (F)-actin shortening proteins are effective in reducing the viscosity of sputum, particularly cystic Fibrosis sputum. As shown by the Vasconcellos et al., reference cited above, liquefaction is not instantaneous. However, once a sufficient degree of viscosity reduction is achieved, a patient can more easily expectorate or otherwise rid himself of excess sputum.

One problem that has not been appreciated is that liquefaction, considered alone, may affect the patient adversely. For example, viscosity reduction may provide the destructive proteases within the sputum greater access to lung and related tissues. Thus, the applicant has found it to be of surprising importance to inhibit these destructive proteases during the transition period from the onset of viscosity reduction until the sputum is expectorated. Moreover, the high proportion of protease destruction caused by the elastase in the sputum has also gone unappreciated.

The compounds of Formula I are surprisingly, highly active in viscous sputum as well as liquified sputum.

As a result of the above factors, treatment with applicants composition is capable of returning a patent with lung disease to substantially normal lung function, as measured, for example, by FEV₁ (forced expiration volume). In particular, treatment with applicants composition is capable of returning a patent with lung disease to at least 60-75% of normal lung function, as measured, for example, by FEV₁. Moreover, treatment with applicants composition is capable of returning a patent with lung disease to 75-90% of normal lung function or greater, as measured, for example, by FEV₁.

Accordingly, in one aspect the invention encompasses a method of treating a patient with a lung disease, comprising:

administration to a patient in need of sputum viscosity reduction, a therapeutically effective non-toxic amount of an (F)-actin shortening protein and a therapeutically effective non-toxic amount of compound of Formula I as described herein.

Within this aspect the invention encompasses a method of treating a patient with a lung disease, comprising: administration to a patient in need of sputum viscosity reduction, a therapeutically effective non-toxic amount of an (F)-actin shortening protein and a therapeutically effective non-toxic amount of compound of Formula I as described herein, said amounts effective to return the lung function of said patients to at least 60-75% of normal as measured by FEV₁.

Within this class the invention encompasses a method of treating a patient with a lung disease, comprising:

administration to a patient in need of sputum viscosity reduction, a therapeutically effective non-toxic amount of an (F)-actin shortening protein and a therapeutically effective non-toxic amount of compound of Formula I as described herein, said amounts effective to return the lung function of said patients to 75 to 90% of normal as measured by FEV₁.

The compounds of the invention are prepared by known methods or are prepared among other methods by the following representative schemes. For example, methods for making such compounds are disclosed in EP O 337 549, published October 18, 1989, which is hereby incorporated by reference.

This invention also relates to a method of treating

inflammation in patients using a compound of Formula (I), particularly a preferred compound as the active constituent.

It has been found that the following compound are effective inhibitors of the proteolytic function of human neutrophil elastase as shown below in Table 1 to 10.

1

Enzyme Assays for the Inhibition of Human Polymorphonuclear Leukocyte Elastase Via Hydrolysis of N-t-Boc-alanyl-alanyl- prolylalanine-p-nitroanilide (Boc-AAPAN) or N-t-Boc-alanyl- prolylvaline-p-nitro-anilide fBoc-AAPVN) Reagent:

0.05M TES (N-tris[hydroxymethyl]methyl-2-minoethanesulfonic acid) Buffer, pH 7.5.

0.2 mM Boc-AAPAN or Boc-AAPVN.

To prepare substrate, the solid was first dissolved in 10.0 ml DMSO. Buffer at pH 7.5 was then added to a final volume of 100 ml.

Crude extract of human polymorphonuclear leukocytes (PMN) containing elastase activity. Inhibitors (azetidinones) to be tested dissolved in DMSO just before use.

To 1.0 ml of 0.2 mM Boc-AAPAN in a cuvette, 0.01-0.1 ml of DMSO with or without inhibitor was added. After mixing, a measurement was taken at 410 mμ to detect any spontaneous hydrolysis due to presence of test compound. 0.05 Milliliters of PMN extract was then added and the ΔOD/min at 410 mμ was measured and recorded. Beckman model 35 spectrophotometer was used.

Results were expressed to the nearest thousand Kobs/I which is the second order rate constant in per mole per second for inactivation of the enzyme.

The elastase activity in the crude PMN extract may vary from one preparation to another. A control of each new batch is run, and the volume added in the assay procedure is adjusted according to activity.

This invention also relates to a method of treating

inflammation in patients using a compound of Formula (I), particularly a preferred compound as the active constituent.

It has been found that the compounds of Formula (I) are effective inhibitors of the proteolytic function of human neutrophil elastase.

Accordingly, the compounds of Formula (I), can be used to reduce inflammation and/or relieve pain in diseases such as emphysema, rheumatoid arthritis, osteoarthritis, gout, bronchial inflammation, chronic or acute bronchitis, cystic fibrosis, adult respiratory distress syndrome, atherosclerosis, sepsis, septicemia, shock, periodontitis, glomerular nephritis or nephosis, myocardial infarction, reperfusion injury, infectious arthritis, rheumatic fever and the like, and may reduce hemorrhage in acute promyelocytic leukemia and the like.

For each of the uses, the compounds of Formula (I) and (F)-actin shortening proteins, may be administered orally, topically, parenterally, by inhalation spray or rectally in dosage unit Formulations containing conventional non-toxic pharmaceutically acceptable carriers, adjuvants and vehicles. The term parenteral as used herein includes subcutaneous injections, intravenous, intramuscular, intrasternal injection or infusion techniques. In addition to the treatment of warmblooded animals such as mice, rats, horses, dogs, cats, etc., the compounds of the invention are effective in the treatment of humans.

The pharmaceutical compositions containing the active ingredient may be in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs. Compositions intended for oral use may be prepared according to any method known to the art for the manufacture of pharmaceutical compositions and such compositions may contain one or more agents selected from the group consisting of sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide pharmaceutically elegant and palatable preparation. Tablets contain the active ingredient in admixture with non-toxic pharmaceutically acceptable excipients which are suitable for the manufacture of tablets. These excipients may be, for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, for example, corn starch, or alginic acid; binding agents, for example starch, gelatin or acacia, and lubricating agents, for example, magnesium stearate, stearic acid or talc. The tablets may be uncoated or they may be coated by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period. For example, a time delay material such as glyceryl monostearate or glyceryl distearate may be employed.

Formulations for oral use may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example, peanut oil, liquid paraffin, or olive oil.

Aqueous suspensions contain the active materials in admixture with excipients suitable for the manufacture of aqueous suspensions. Such excipients are suspending agents, for example, sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethylcellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia; dispersing or wetting agents may be a naturally-occurring phosphatide, for example, lecithin, or condensation products of an alkylene oxide with fatty acids, for example, polyoxyethylene stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example, heptadecaethyleneoxycetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids and a hexitol such as polyoxyethylene sorbitol monooleate, or

condensation products of ethylene oxide with partial esters derived from fatty acids and hexitol anhydrides, for example, polyoxyethylene sorbitan monooleate. The said aqueous suspensions may also contain one or more preservatives, for example, ethyl, or n-propyl, p-hydroxy-benzoate, one or more coloring agents, one or more flavoring agents, and one or more sweetening agents, such as sucrose or saccharin.

Oily suspension may be formulated by suspending the active ingredient in a vegetable oil, for example, arachis oil, olive oil, sesame oil or coconut oil, or in a mineral oil such as liquid paraffin. The oily suspensions may contain a thickening agent, for example, beeswax, hard paraffin or cetyl alcohol. Sweetening agents such as those set forth above, and flavoring agents may be added to provide a palatable oral preparation. These compositions may be preserved by the addition of an antioxidant such as ascorbic acid.

Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, suspending agent and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified by those already

mentioned above. Additional excipients, for example, sweetening, flavoring and coloring agents, may also be present.

The pharmaceutical compositions of the invention may also be in the form of oil-in-water emulsions. The oily phase may be a vegetable oil, for example, olive oil or arachis oils, or a mineral oil, for example, liquid paraffin or mixtures of these. Suitable emulsifying agents may be naturally-occurring gums, for example, gum acacia or gum tragacanth, naturally-occurring phosphatides, for example, soy bean, lecithin, and esters or partial esters derived from fatty acids and hexitol anhydrides, for example, sorbitan monooleate, and condensation products of the said partial esters with ethylene oxide, for example, polyoxyethylene sorbitan monooleate. The emulsions may also contain sweetening and flavoring agents.

Syrups and elixirs may be formulated with sweetening agents, for example, glycerol, propylene glycol, sorbitol or sucrose. Such formulations may also contain a demulcent, a preservative and flavoring and coloring agents. The pharmaceutical compositions may be in the form of a sterile mjectable aqueous or oleagenous suspension. This suspension may be formulated according to the known art using those suitable dispersing or wetting agents and suspending agents which have been mentioned above. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent, for example, as a solution in 1,3-butane diol. Among the acceptable vehicles and solvents that may be employed are water, Ringer's solution glucose in water and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed including synthetic mono- or

diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.

The pharmaceutical compositions of this invention may also be administered in the form of suppositories for rectal administration of the drug. These compositions can be prepared by mixing the drug with a suitable non-irritating excipient which is solid at ordinary

temperatures but liquid at the rectal temperature and will therefore melt in the rectum to release the drug. Such materials are cocoa butter and polyethylene glycols.

For topical use, creams, ointments, jellies, solutions or suspensions, etc., containing the anti-inflammatory agents are employed. The amount of active ingredient(s) that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of

administration. For example, a formulation intended for the oral administration of humans may contain from 5 mg to 2000 mg or 5000 mg of each active agent(s) compounded with an appropriate and convenient amount of carrier material which may vary from about 5 to about 95 percent of the total composition. For purposes of this specification, this broad dosage range is specifically intended to include, but is not limited to, range of 5 mg to 2000 mg; 25 mg to 2000 mg; 5 mg to 1000 mg; 25 mg to 1000 mg; 5 mg to 500 mg; and 25 mg to 500 mg. Dosage unit forms will generally contain between from about 25 mg to about 500 mg of active ingredient(s).

Furthermore, it is also possible that most effective treatment may warrant administration of an initial dosage of one range (e.g., 1-5 mg of active agent per kg of patient weight) followed by administration of a second range (e.g., 0.1 to 1 mg of active agent per kg of patient weight).

It will be understood, however, that the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, rate of excretion, drag combination and the severity of the particular disease undergoing therapy.

With specific regard to (F)-actin shortening proteins, in particular gelsolin, the dosage can be calculated in the following manner. The normal blood gelsolin concentration is 2.4 μM (2.4 μmol/L), and the normal blood DBP concentration is 5 μM (5 μmol/L). Thus, the total blood actin-binding capacity (ABC) is approximately 7.5 μmol/L. The blood volume if 6% of the body weight, hence a 70 Kg person has 4.2 liters of blood and thus (4.2 L × 7.5 μmol/L) 31.5 μmols ABC. Since DBP and gelsolin are distributed throughout the

extracellular space (which is 10% of the body weight, the body contains (7.5 × 7) 52.5 μmols ABC. It may be desired to administer between 32 and 53 μmols of an actin binding (herein actin shortening) compound (or 0.46 μmol/kg body weight) to cover total complexing or depletion of endogenous ABC. Since 0.425 mg of actin is equal to 1 μmol, and since there is 4.86 mg actin per gram of skeletal muscle, each gram of muscle contains 11.3 μmol actin or 4.6 grams of muscle destruction could neutralize total body ABC. However, because the toxic effects of actin are presumably local (e.g., inhibition of clot lysis), sequestered or kinetically determined (e.g., actin permeates an organ faster than binding proteins neutralize it), it is likely that a theoretically minimum dose will have to be adjusted upward in order to achieve kinetically favorable therapeutic effects. The kinetic effect can be important, for example, since hemolysis of about half of erythron, which should liberate only 4.2 μmol of actin, reduces the plasmia gelsolin

concentration by half acutely (Smith, et al., Blood 72:214-2181 (1988)), suggesting slow equilibration between extravascular and blood

compartments. Conversely, a therapeutically effective state, capable of breaking up local deposits of actin, may be achievable only by a transient pulse of a high concentration of actin-binding molecules.

The compounds of the invention can be administered in any appropriate pharmacological carrier for administration. They can be administered in any form that effects prophylactic, palliative,

preventative or curing conditions of tissue injury in humans and animals.

The following example illustrates the preparation of the compounds of Formula I useful in the method of treatment of the present invention, but does not limit the scope of the invention. Starting materials may be optionally prepared as disclosed in EPO 337 549 published.

October 18, 1989 which is hereby incorporated by reference. Where appropriate, compounds may be produced and used in the form of pharmaceutically acceptable salts. For example, the basic compounds may be used in the form of a hydrochloride or mesylate or other acceptable salt. See Preformulation in Remington's Pharmaceutical Sciences, Mack Publishing, Easton PA.

The glycolic acid derivatives described herein can be prepared according to the following scheme. The starting acid (as carboxy late anion) may be alkylated (Ex 1A) with a suitably protected α-halo acetic acid derivative to give the glycolate ester 4 which can be deprotected (Ex 1B) to the glycolic acid ester 5. Treatment of 5 with an amine utilizing a condensing agent such as dicyclohexylcarbodumide or carbomyldiinidazole (Ex 2) affords the deserved amide 6. Alternately, the starting acid 1 may be converted to its acid chloride 2 (Ex 3A) and treated with a suitably protected α-hydroxyalkanoic acid (Ex 3B) in the presence of base to give the protected ester 7. Deprotection (Ex 4), followed by conversion to the acid chloride and treatment with the appropriate amine (Ex 5) affords the desired amide 9. EXAMPLE 1

A. t-Butoxycarbonylmethyl [S-(R*, S*)]4-((3,3-diethyl-1- (((1-(4-methylphenyl)butyl)amino)-carbonyl)-4-oxo-2- azetidinyl)-oxy)benzoate

To a solution of 0.806 gm of [S-(R*, S*)] 4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)-carbonyl)-4-oxo-2-azetidinyl)oxy)-benzoic acid in ~3ml DMF is added 0.23 gm. triethylamine followed by 0.50 gm of t-butyl bromoacetate and the mixture stirred overnight at room temperature. Ethyl acetate (25 ml) is then added and the resultant mixture is washed with 2 x 10 ml water, 10 ml saturated sodium bicarbonate, and 20 ml brine. The organic layer is dried through sodium sulfate and concentrated in vacuo. Chromatography on Silica gel 60 (350 ml column) and elution with 10% ethyl acetate in hexanes gave 0.67 gm of the t-Butoxycarbonylmethyl [S-(R*, S*)]4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoate.

In a similar manner can be prepared 2-(dimethylamino)-2-oxoethyl, (S-(R*,S*))-4-((3,3-Diethyl-1-(((1-(4-methylphenyl)butyl)-amino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoate (Compound 6), and 2-(N-methylacetamido)ethyl. {2-(R*,S*)}- 4-{ {3,3-Diethyl-1-{ { { 1-(4-methylphenyl)butyl}amino}carbonyl}-4-oxo-2-azetidinyl}oxy}-benzoate, (Compound 7).

B. Carboxymethyl [S-(R*,S*)]4-((3,3-diethyl-1- (((1-(4- methyl-phenyl)butyl)amino)carbonyl)-4-oxo-2- azetidinyl)-oxy)benzoate

To the above ester is added 2 ml of anisole and the resulting mixture is cooled in an ice bath and 5 ml of ice cold

trifhioroacetic acid is added. The reaction mixture is stirred cold for three hours then allowed to come to room temperature. After 30 minutes, the reaction mixture is concentrated in vacuo and the residue chromatographed on silica gel 60. Elution with 20% ethyl acetate in hexanes containing 1 % acetic acid gives 0.53 gm of desired carboxymethyl [S-(R*,S*)]4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoate.

Compound 2

Analysis: C₂₈H₃₄N₂O₇+0.3H₂O

Calc: C, 65.19; H, 6.75; N, 5.43

Found: C, 65.30; H, 6.76; N, 5.23

Compound 6

Analysis: C₃₀H₃₉N₃O₆+0.5H₂O

Calc: C, 65.91; H, 7.38; N, 7.60

Found: C, 65.71; H, 7.63; N, 7.50

Compound 7

Analysis: C₃₁H₄₁N₃O₆+0.6EtOAc

Calc: C, 66.35; H, 7.64; N, 6.95

Found: C, 66.52; H, 7.89; N, 6.83,

EXAMPLE 2

2-(bis(2-hydroxyethyl)amino)-2-oxoethyl(S-(R*,S*))-4-((3,3-diethyl-1-(((1-(4-methyl-phenyl)butyl)amino)-carbonyl)-4-oxo-2-azetidinyl)oxy) benzoate

To a solution of 0.125 gm of the acid from 1B in 2-3 ml of methylene chloride is added 0.050 gm of carbonyldiimidazole. The mixture is stirred for 30 minutes at room temperature at which time 0.060 gm of diethanolamine is added along with 1 ml of DMF and 2 ml of methylene chloride. The resulting mixture is stirred overnight at room temperature then concentrated in vacuo. Silica gel

chromatography of the residue using 2.5 to 5.0% methanol in methylene chloride gives 0.123 gm of the desired Compound 3, 2-(bis(2-hydroxyethyl)amino)-2-oxoethyl(S-(R*,S*))-4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoate. Compound 3

Analysis: C₃₂H₄₃N₃O₈, +0.4H₂O

Calc: C, 63.53; H, 7.30; N, 6.95

Found: C, 63,51; H, 7.45; N, 6.95.

Similarly were prepared

Compound 4

Analysis: C₃₁H₄₀N₄O₇

Calc: C, 64.12; H, 6.94; N, 9.65

Found: C, 64.12; H, 7.18; N, 9.44.

Compound 5

Analysis: C₃₂H₄₃N₃O₉ +0.3H₂O

Calc: C, 62.08; H, 7.09; N, 6.79

Found: C, 61.89; H, 7.39; N, 6.88

Compound 8

Analysis: C₄₀H₄₇N₃O₈

Calc: C, 68.85; H, 6.79; N, 6.02

Found: C, 68.79; H, 7.06, N, 5.88

EXAMPLE 3

Preparation of 1-Methyl-2-oxo-2-(phenylmethoxy)ethyl(2S-(1(S*),R*,-(R)))-4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)carbonyl)-4-oxo-2-azetidinyl)oxy) benzoate, Compound 10

A.

To a solution of 1.0 gm [S-(R*, S*)]4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoic acid in 10 ml methylene chloride is added 2 ml of oxalyl chloride followed by a catalytic amount of DMF. The reaction is stirred 1 hour at room temperature then concentrated in vacuo to yield the acid chloride which is used as is in the next step. B.

A solution of the above acid chloride in 10 ml of methylene chloride is cooled in an ice bath and a solution of 1.25 gm benzyl L-lactate and 2.0 gm of triethylamine in 10 ml of methylene chloride is added. The mixture is stirred at room temperature overnight then concentrated in vacuo. Chromatography of the residue on silica gel using methylene chloride as the eluent yields 0.795 of the desired 1-Methyl-2-oxo-2-(phenylmethoxy)ethyl (2S-(1(S*),R*,(R)))-4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoate, Compound 10.

Analysis: C₃₆H₄₂N₂O₇

Calc: C, 70.34; H, 6.89; N, 4.56

Found: C, 70.45; H, 7.05; N, 4.48.

EXAMPLE 4

Preparation of 1-carboxyethyl [S-(R*,S*)] 4-((3,3-diethyl-1-(((1-(4-methyl-phenyl)butyl)amino)carbonyl)-4-oxo-2-azetidin-yl)oxy)benzoate

A mixture of 0.69 gm of the benzylester prepared in Example 3 and 0.2 gm 10% Pd/C in 10 ml of EtOAc is treated with hydrogen at 40 psi. When the reaction is complete the mixture is filtered and concentrated in vacuo to yield 0.56 gm of 1-carboxyethyl [S-(R*,S*)]4-((3,3-diethyl-1-(((1-(4-methyl-phenyl)butyl)amino)-carbonyl)-4-oxo-2-azetidinyl)oxy)benzoate, Compound 11.

Analysis: C₂₉H₃₆N₂O₇

Calc: C, 66.40; H, 6.92; N, 5.34

Found: C, 66.66; H, 7.26; N, 5.05. EXAMPLE 5

2-(diethylamino)-1-methyl-2-oxoethyl[S-(R*,S*)]-4-((3,3-diethyl-1-(((1-(4-methyl-phenyl)butyl)amino)-carbonyl)-4-oxo-2-azetidinyl)oxy)benzoate

The acid (.250 gm) from Example 4 is treated with oxalyl chloride according to the procedure of Example 3A and the

corresponding acid chloride is obtained. This material is dissolved in 5 ml methylene chloride and 0.4 ml of diethylamine added. After 1 hour the reaction mixture is concentrated in vacuo and the residue taken in ethyl acetate and washed with saturated sodium bicarbonate solution. The organic layer is dried through sodium sulfate, concentrated and the residue chromatographed on silica gel. Elution with 5% of ethyl acetate in methylene chloride gives Compound 12.

Analysis: C₃₃H₄₅N₃O₆

Calc: C, 68.37; H, 7.82, N, 7.25

Found: C, 68.40; H, 7.93, N, 7.40.

EXAMPLE 6

(S(R*,S*))-1-(((4-((3,3-diethyl-1-(((1-(4-methyl-phenyl)butyl)amino)-carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl)oxy)acetyl) L-proline

A.

When benzyl L-lactate is replaced by L-proline benzyl ester hydrochloride and triethylamine in the procedure of Example 3 the corresponding amide with L-proline benzyl ester, Compound 8, is obtained.

Analysis: C₄₀H₄₇N₃O₈

Calc: C, 68.85; H, 6.79, N, 6.02

Found: C, 68.79; H, 7.06, N, 5.88. B.

Reduction of the material obtained in Example 6A

according to the procedure of Example 4 affords Compound 9.

Analysis: C₃₃H₄₁N₃O₈+0.5H₂O

Calc: C, 64.27; H, 6.86; N, 6.81

Found: C, 64.49; H, 6.90; N, 6.68.

EXAMPLE 7

[S-(R*,S*)] 1-(((4-((3,3-diethyl-1-(((1-(4-methyl- phenyl)butyl)amino)carbonyl-4-oxo-2-azetidinyl)oxy)benzoyl)oxy)acetyl-N-benzyl-L-prolinamide

Treatment of the acid obtained in Example 6B, Compound 9, with oxalyl chloride according to Example 3A gives the

corresponding acid chloride which when treated with benzylamine gives the desired benzyl amide, Compound 19.

Analysis: C₄₀H₄₈N₄O₇

Calc: C, 68.95; H, 6.94; N, 8.04

Found: C, 68.93, H, 7.02; N, 7.96.

EXAMPLE 8

To a solution of the acid chloride (prepared from 0.55 gm of [S-(R*, S*)] 4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)-carbonyl)-4-oxo-2-azetidinyl)oxy)benzoic acid according to the procedure of Example 3 A) in 3 ml of methylene chloride is added 0.15 gm of N,N-dimethylaminoethanol. The reaction mixture is stirred overnight at room temperature, concentrated in vacuo, then taken up in ethyl acetate (25 ml) and washed with saturated sodium bicarbonate solution. The organic layer is dried through sodium sulfate and concentrated in vacuo. Silica gel chromatography of the residue using 2.5% methanol in methylene chloride gives 0.59 gm of Compound 1, 2-(dimethylamino)ethyl (S-(R*,S*))-4-((3,3-diethyl-1-(((1-(4-methylphenyl)butyl)amino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoate Analysis: C₃₀H₄₁N₃O₅

Calc: C, 68.81; H, 7.89; N, 8.02

Found: C, 68.85; H, 8.09; N, 7.97.

When N,N-dimethylaminoethanolamine is replaced by the appropriate amino alcohols the corresponding esters are obtained.

Compound 13 1-Dimethylamino-2-propyl [S-(R*,S*)]-4-[[3,3-diethyl- 1-[[[1-(4-methyl-phenyl)butyl]amino]carbonyl]-4-oxo-2- azetidin-yl]oxy]benzoate

Analysis: C₃₁H₄₃N₃O₅

Calc: C, 69.25; H, 8.06; N, 7.82

Found: C, 68.97; H, 8.01; N, 7.80.

Compound 14 3-Dimethylamino-1-propyl [S-(R*,S*)]-4-[[3,3-diethyl- 1-[[[1-(4-methyl-phenyl)butyl]amino]carbonyl]-4-oxo-2- azetidin-yl]oxy]benzoate

Analysis: C₃₁H₄₃N₃O₅

Calc: C, 69.25; H, 8.06; N, 7.81

Found: C, 68.85; H, 8.19; N, 7.72.

Compound 16 2-Diethylaminoethyl [S-(R*,S*)]-4-[[3,3-diethyl-1-[[[1- (4-methyl-phenyl)butyl]amino]carbonyl]-4-oxo-2- azetidinyl]-oxy]benzoate

Analysis: C₃₂H₄₅N₃O₆

Calc: C, 69.66; H, 8.22; N, 7.62

Found: C, 69.37; H, 8.41; N, 7.51.

Compound 17 2-(1-[4-morpholino]ethyl) [S-(R*,S*)]-4-[[3,3-diethyl-1- [[[1-(4-methyl-phenyl)butyl]amino]carbonyl]-4-oxo-2- azetidinyl]oxy]benzoate Analysis: C₃₂H₄₃N₃O₆

Calc: C, 67.94; H, 7.66; N, 7.43

Found: C, 67.67; H, 7.90; N, 7.26.

Compound 18 4-dimethylaminobutyl [S-(R*,S*)]-4-[[3,3-diethyl-1-[[[1- (4-methyl-phenyl)butyl]amino]carbonyl]-4-oxo-2- azetidinyl]oxy]benzoate

Analysis: C₃₂H₄₅N₃O₅ +0.2 H₂O

Calc: C, 69.21; H, 8.24; N, 7.56

Found: C. 69.35; H, 8.24; N, 7.29.

Compound 20 2-dimethylamino-2-methyl-1-propyl [S-(R*,S*)]-4-[[3,3- diethyl-1-[[[1-(4-methyl-phenyl)butyl]amino]carbonyl]- 4-oxo-2-azetidinyl]oxy]benzoate

Analysis: C₃₂H₄₅N₃O₅

Calc: C, 69.66; H, 8.22; N, 7.62

Found: C, 69.52; H, 8.47; N, 7.59.

Compound 21 2-(diisopropylamino)ethyl [S-(R*,S*)]-4-[[3,3-diethyl-1- [[[1-(4-methyl-phenyl)butyl]amino]carbonyl]-4-oxo-2- azetidinyl]oxy]benzoate

Analysis: C₃₄H₄₉N₃O₅

Calc: C, 70.44; H, 8.52; N, 7.25

Found: C, 70.28; H, 8.76; N, 7.13.

Compound 22 Benzyl [S-(R*,S*)]-4-[2-[[4-[[3,3-diethyl-1-[[[1-(4- methylphenyl)butyl]amino]carbonyl]-4-oxo-2- azetidinyl]oxy]-benzoyl]oxy]-ethyl]-1-Piperazine- carboxylate

Analysis: C₄₀H₅₀N₄O₇

Calc: C, 68.75; H, 7.21; N, 8.02

Found: C, 68.39; H, 7.30; N, 7.84. Compound 23 2-(dibutylamino)ethyl [S-(R*,S*)]-4-[[3,3-diethyl-1-[[[1- (4-methyl-phenyl)-butyl]amino]carbonyl]-4-oxo-2- azetidin-yl]oxy]benzoate

Analysis: C₃₆H₅₃N₃O₅

Calc: C, 71.14; H, 8.79; N, 6.91

Found: C, 71.00; H, 9.03; N, 6.81.

Compound 24 [S-(R*,S*)]-6-(dimethylamino)hexyl-4-[[3,3-diethyl-1- [[[1-(4-methyl-phenyl)butyl]amino]carbonyl]-4-oxo-2- azetidin-yl]oxy]-benzoate

Analysis: C₃₄H₄₉N₃O₅+1H₂O

Calc: C, 68.31; H, 8.60; N, 7.03

Found: C, 68.34; H, 8.29; N, 6.86.

Compound 26 2-(4-methyl-1-piperazinyl)ethyl[S-(R*,S*)]-4-[[3,3- diethyl-1-[[[1-(4-methyl-phenyl)butyl]amino]carbonyl]- 4-oxo-2-azetidinyl]oxy]benzoate,

Analysis: C₃₃H₄₆N₄O₅+0.8H₂O

Calc: C, 66.82; H, 8.09; N, 9.44

Found: C, 67.28; H, 8.10; N, 8.96.

Compound 28 2-(diphenylamino)ethyl[S-(R*,S*)]-4-[[3,3-diethyl-1- [[[1-(4-methylphenyl)-butyl]amino]carbonyl]-4-oxo-2- azetidin-yl]oxy]benzoate

Analysis: C₄₀H₄₅N₃O₅ +1.4H₂O

Calc: C, 71.40; H, 7.16; N, 6.25

Found: C, 71.62; H, 6.99; N, 5.99.

Compound 29 2-(di-2-propenylamino)ethyl [S-(R*,S*)]-4-[[3,3-diethyl- 1-[[[1-(4-methylphenyl)-butyl]amino]carbonyl]-4-oxo-2- azetidin-yl]oxy]benzoate,

Analysis: C₃₄H₄₅N₃O₅

Calc: C, 70.93; H, 7.88; N, 7.30

Found: C, 71.18; H, 8.06; N, 7.34. Compound 30 2-(dimethylamino)-2-phenylethyl [S-(R*,-S*)]-4-[[3,3- diethyl-1-[[[1- (4-methyl- phenyl)butyl]amino]carbonyl]- 4-oxo-2-azetidinyl]oxy]benzoate

Analysis: C₃₆H₄₅N₃O₅

Calc. C, 72.09; H, 7.56; N, 7.00.

Found: C, 71.75; H, 7.67; N, 6.70.

Compound 31 2-[methyl(phenylmethyl)amino]ethyl [S-(R*,S*)]-4- [[3,3-diethyl-1-[[[1-(4-methyl-phenyl)butyl]- amino]carbonyl]-4-oxo-2-azetidinyl]oxy]benzoate

When [S-(R*,S*)]-4-[[3,3-diethyl-1-[[[1-(4-methyl¬phenyl) butyl]amino]carbonyl]-4-oxo-2-azetidinyl]oxy]-2,6-dimethyl benzoic acid is used in place of [S(R*,S*)]-4-[[3,3-diethyl-1-[[[1-(4-methyl-phenyl)butyl]amino]carbonyl]-4-oxo-2-azetidinyl]oxy]benzoic acid in the procedure of Example 3A and allowed to react with the appropriate amino alcohols the following esters are obtained.

Compound 304 2-(dimethylamino)ethyl [S,(R*,S*)]-4-[[3,3-diethyl- 1-[[[1-(4-methylphenyl)-butyl]amino]carbonyl]-4- oxo-2-azetidin-yl]oxy]-2,6-dimethyl-benzoate

Compound 305 2-(diethylamino)ethyl [S-(R*,S*)]-4-[[3,3-diethyl-1- [[[1-(4-methylphenyl)-butyl]amino]carbonyl]-4-oxo- 2-azetidin-y1]oxy]-2,6-dimethyl-benzoate

Compound 306 2-[bis(1-methylethyl)amino]-ethyl [S-(R*,S*)]-4- [[3,3-diethyl-1-[[[1-(4-methylphenyl)butyl]amino]- carbonyl]-4-oxo-2-azetidinyl]oxy]-2,6-dimethylbenzoate

Treatment of the acid [S-(R*,S*)]-4-[[3,3-diethyl-1-[[[1-(4-methylphenyl)butyl]amino]carbonyl]-4-oxo-2-azetidinyl]oxy]-benzene-acetic acid with oxalyl chloride according to the procedure of Example 3A affords the corresponding acid chloride which when allowed to react with the appropriate amino alcohol according to the procedure of Example 8 gives the following amino esters:

Compound 32

Analysis: C₃₁H₄₃N₃O₅

Calc: C, 69.25; H, 8.06; N, 7.82

Found: C, 69.02; H, 7.86; N, 7.74.

Compound 33

Analysis: C₃₂H₄₅N₃O₅

Calc: C, 69.66; H, 8.22; N, 7.62

Found: C, 69.10; H, 8.17; N, 7.50.

Compound 34

Analysis: C₃₃H₄₇N₃O₅

Calc: C, 70.06; H, 8.38; N, 7.43

Found: C, 69.70; H, 8.41; N, 7.05.

Compound 35

Analysis: C₃₃H₄₅N₃O₆

Calc: C, 68.37; H, 7.82; N, 7.25

Found: C, 68.55; H, 8.19; N, 7.08.

Compound 36

Analysis: C₃₂H₄₅N₃O₅

Calc: C, 69.66; H, 8.22; N, 7.62

Found: C, 69.60; H, 8.49; N, 7.55.

EXAMPLE 9

To a solution of 0.247 g of Compound 1 in 2 ml of ethyl acetate is added 0.125 gm of m-chloroperoxy benzoic acid. After 30 minutes at room temperature the reaction mixture is concentrated in vacuo. Chromatography of the residue on silica gel using methylene chloride/methanol/water 85/15/1.5 gives the desired N-oxide Compound

15.

Analysis: C₃₀H₄₁N₃O₈+1.4H₂O

Calc: 63.79; H, 7.82; N, 7.44

Found: 63.89; H, 7.85; N, 7.27.

EXAMPLE 10

[S-(R*,S*)] 2-[4-[[[2-(dimethylammo)-emyl]amino]-carbonyl]phenoxy]-3,3-diethyl-N-[1-(4-methylphenyl)-butyl]-4-oxo-1-azetidinecarboxamide

To a solution of 0.104 g carbonyldiimidazole in 2 ml methylene chloride is added a solution of 0.227 g of [S-(R*,S*)]-4-((3,3-diethyl-1-((-1-(4-methylphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)-oxy)benzoic acid in 3 ml methylene chloride. The

mixture is stirred at ambient temperature for 30 minutes at which time 0.100 g of N,N-dimethyl-ethylenediamine is added. After stirring overnight at room temperature the reaction mixture is poured into benzene (50 ml) and washed with water. The organic layer is separated, dried through sodium sulfate and concentrated in vacuo. Silica gel chromatography using 5% methanol in methylene chloride yields 0.160 g of 2-[4-[[[2-(dimethylamino)ethyl]amino]-carbonyl]phenoxy]-3,3-diethyl-N-[1-(4-methylphenyl)butyl]-4-oxo-1-azetidinecarboxamide.

(Compound 73).

Analysis: C₃₀H₄₂N₄O₄+0.4H₂O

Calc: C, 68.00; H, 8.14; N, 10.57

Found: C, 68.01; H, 8.18; N, 10.62. EXAMPLE 11

A. (S-(R*,S*))-4-((3,3-diethyl-1-((1-(4-methylphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)-oxy)-benzoyl chloride

To a solution of 0.150 g of [S-(R*,S*)] 4-((3,3-diethyl- 1-(((1-(4-methylphenyl)butyl)amino)-carbonyl)-4-oxo-2-azetidinyl)-oxy)benzoic acid in 5 ml methylene chloride containing a catalytic amount of dimethylformamide is added 0.5 ml of oxalyl chloride. The mixture is stirred at room temperature for 30 minutes and then concentrated in vacuo to yield (S-(R*,S*))-4-((3,3-diethyl-1-((1-(4-methylphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)-benzoyl chloride.

B. [S-(R*,S*)]-2-[4-[[[2-(dimethylamino)ethyl]methylamino]carbonyl]-phenoxy]-3,3-diethyl-N-[1-(4- methylphenyl]butyl]-4-oxo-1-azetidinecarboxamide

The above acid chloride is dissolved in 3 ml methylene chloride and a solution of 0.20 gm of N,N,N'-trimethylethylenediamine in 2 ml methylene chloride is added. The mixture is stirred overnight and then concentrated in vacuo. The residue is extracted between ethyl acetate and saturated sodium bicarbonate solution.

The organic layer is dried through sodium sulfate and concentrated in vacuo. Silica gel chromatography of the residue (5% methanol in methylene chloride) affords 0.117 g of [S-(R*,S*)]-2-[4-[[[2-(dimethylamino)emyl]methylamino]carbonyl]phenoxy]-3,3-diethyl-N-[1-(4-methylphenyl]butyl]-4-oxo-1-azetidinecarboxamide.

(Compound 75).

Analysis: C₃₁H₄₄N₄O₄+0.5H₂O

Calc: C, 68.23; H, 8.31; N, 10.27

Found: C, 68.20; H, 8.41; N, 10.10. When N,N,N'-trimethylethylenediamine of Example 11b is replaced by the appropriate amines, there is obtained the

corresponding amides.

1) Compound 76

Analysis: C₃₂H₄₆N₄O₄

Calc: C, 69.79; H, 8.42; N, 10.17

Found: C, 69.02; H, 8.60; N, 9.54.

2) Compound 77

Analysis: C₃₂H₄₆N₄O₄ +0.75 H₂O

Calc: C, 68.12; H, 8.49; N, 9.93

Found: C, 68.22; H, 8.48; N, 10.17.

3) Compound 78

Analysis: C₃₂H₄₄N₄O₅

Calc. C, 68.06; H, 7.85; N, 9.92

Found: C, 67.84; H, 8,07; N, 9.62.

4) Compound 79

Analysis: C₃₃H₄₆N₄O₅ +H₂O

Calc: C, 66.42; H, 8.11; N, 9.39

Found: C, 66.73; H, 8.19; N, 9.23.

5) Compound 80

Analysis: C₃₅H₅₂N₄O₆

Calc: C, 67.28; H, 8.39; N, 8.97

Found: C, 67.08; H, 8.77; N, 8.41.

6) Compound 81

Analysis: C₃₃H₄₈N₄O₄ +H₂O

Calc: C, 68.01; H, 8.65; N, 9.61

Found: C, 68.42; H, 8.59; N, 9.17. 7) Compound 82

Analysis: C₃₆H₄₆N₄O₄ +0.5H₂O Calc: C, 71.14; H, 7.79; N, 9.21

Found: C, 71.41; H, 7.68; N, 9.10.

8) Compound 83

Analysis: C₃₂H₄₆N₄O₄ +1.2H₂O Calc: C, 67.15; H, 8.52; N, 9.79

Found: C, 67.21; H, 8.26; N, 9.47.

9) Compound 84

10) Compound 86

Analysis: C₃₅H₅₂N₄O₄

Calc: C, 70.91; H, 8.84; N, 9.45

Found: C, 70.37; H, 8.84; N, 8.77.

12) Compound 89

Analysis: C₃₁H₃₉N₅O₄ +0.7H₂O Calc: C, 66.71; H, 7.29; N, 12.54

Found: C, 66.91; H, 7.40; N, 12.14.

12) Compound 90

Analysis: C₃₃H₄₆N₄O₄ +0.8H₂O

Calc: C, 68.67; H, 8.31; N, 9.70

Found: C, 68.82; H, 8.11; N, 9.70.

13 ) Compound 91

Analysis: C₃₄H₄₈N₄O₄ + 0.3H₂O

Calc: C, 70.11; H, 8.41; N, 9.61

Found: C, 70.17; H, 8.64; N, 9.33. 14) Compound 92

Analysis: C₃₀H₄₂N₄O₄ + 1.2H₂O Calc: C, 66.14; H, 8.22; N, 10.29

Found: C, 66.18; H, 8.12; N, 10.31.

15) Compound 93

Analysis: C₃₂H₄₄N₄O₆ + 0.3H₂O Calc: C, 65.57; H, 7.67; N, 9.56

Found: C, 65.72; H, 7.50; N, 9.34.

16) Compound 94

Analysis: C₃₂H₄₄N₄O₄ + 0.3H₂O Calc: C, 70.04; H, 8.08; N, 10.21

Found: C, 70.34; H, 8.90; N, 8.93.

17) Compound 95

Analysis: C₃₃H₄₆N₄O₄ + .5H₂O Calc: C, 69.32; H, 8.28; N, 9.80

Found: C, 69.41; H, 8.25; N, 9.58.

18) Compound 99

Analysis: C₃₈H₅₄N₄O₄ + 1.5H₂O Calc: C, 69.37; H, 8.72; N, 8.51

Found: C, 69.48; H, 8.44; N, 8.36.

19) Compound 102

Analysis: C₄₀H₄₉N₅O₆ + 1.0 H₂O Calc: C, 67.30; H, 7.20; N, 9.81

Found: C, 67.50; H, 7.24; N, 9.53.

20) Compound 104

Analysis: C₃₂H₄₁N₅O₄ +0.75H₂O Calc: C, 67.04; H, 7.47; N, 12.21

Found: C, 67.16; H, 7.56; N, 11.95. 21) Compound 105

Analysis: C₃₂H₄₄N₄O₅ +0.5H₂O Calc: C, 66.99; H, 7.91; N, 9.77

Found: C, 67,00; H, 8.25; N, 9.50.

22) Compound 106

Analysis: C₃₂H₄₅N₅O₅ +0.8H₂O Calc: C, 64.69; H, 7.90; N, 11.78

Found: C, 64.93; H, 8.25; N, 11.12.

23) Compound 107

Analysis: C₃₁H₄₄N₃O₆S +0.3H₂O Calc: C, 61.42; H, 7.41; N, 9.24

Found: C, 61.43; H, 7.54; N, 9.05.

24) Compound 110

Analysis: C₃₅H₄₄N₄O₄

Calc: C, 71.89; H, 7.58; N, 9.58

Found: C, 71.65; H, 7.55; N, 9.34.

25) Compound 111

Analysis: C₃₄H₄₂N₄O₄ +0.5H₂O Calc: C, 70.44; H, 7.47, N, 9.66

Found: C, 70.82, H, 7.46; N, 9.20.

26) Compound 113

Analysis: C₃₄H₄₂N₄O₄ +0.3H₂O Calc: C, 70.88; H, 7.45; N, 9.72

Found: C, 71.12; H, 7.44; N, 9.32.

27) Compound 115

Analysis: C₃₄H₄₂N₄O₄ +0.7H₂O Calc: C, 69.99; H, 7.50; N, 9.60

Found: C, 70.14, H, 7.63; N, 9.25. 28) Compound 116

Analysis: C₃₄H₄₆N₄O₄ +1.4H₂O Calc: C, 68.06; H, 8.19; N, 9.33

Found: C, 68.40; H, 8.14; N, 8.87.

29) Compound 117

Analysis: C₄₀H₅₁N₅O₆ +0.6H₂O Calc: C, 67.79; H, 7.42; N, 9.88

Found: C, 67.81; H, 7.58; N, 9.76.

30) Compound 118

Analysis: C₃₃H₄₇N₅O₄ +0.7H₂O Calc: C, 67.14; H, 8.26; N, 11.86

Found: C, 67.54; H, 8.51; N, 11.28.

31) Compound 119

Analysis: C₃₅H₄₈N₄O₄ +1.25H₂O Calc: C, 68.76; H, 8.32; N, 9.16

Found: C, 69.07; H, 8.19; N, 8.75.

32) Compound 121

Analysis: C₃₄H₄₈N₄O₄ +1H₂O Calc: C, 68.66; H, 8.47; N, 9.42

Found: C, 69.02; H, 8.32; N, 9.06.

33) Compound 125

Analysis: C₃₇H₄₈N₄O₄ +0.5H₂O Calc: C, 71.47; H, 7.94; N, 9.01

Found: C, 71.65; H, 7.91; N, 8.73.

34) Compound 126

Analysis: C₄₁H₅₄N₄O₅ +2H₂O Calc: C, 68.83; H, 8.25; N, 7.64

Found: C, 69.03; H, 7.79; N, 7.50. 35) Compound 132

Analysis: C₃₅H₅₀N₄O₄

Calc: C, 71.15; H, 8.53; N, 9.48

Found: C, 70.90; H, 8.74; N, 9.12.

36) Compound 133

Analysis: C₄₀H₅₂N₄O₄ +0.9H₂O Calc: C, 71.80; H, 8.10; N, 8.37

Found: C, 71.86; H, 8.17; N, 8.18.

37) Compound 137

Analysis: C₃₇H₄₈N₄O₄ +0.8H₂O Calc. C, 70.85; H, 7.97; H, 8.93

Found: C, 71.01; H, 7.97; N, 8.54.

38) Compound 139

Analysis: C₃₉H₅₁N₅O₄ +0.8H₂O Calc: C, 70.09; H, 7.93; N, 10.48

Found: C, 70.18; H, 7.79; N, 10.42.

39) Compound 142

Analysis: C₃₈H₅₅N₅O₆

Calc: C, 67.33; H, 8.18; N, 10.33

Found: C, 67.02; H, 8.31; N, 9.89.

40) Compound 143

Analysis: C₃₄H₅₁N₅O₄

Calc: C, 68.77; H, 8.66; N, 11.80

Found: C, 68.57; H, 8.50; N, 11.53.

41) Compound 144

Analysis: C₃₈H₅₉N₅O₄ +0.4H4₂O Calc: C, 69.45; H, 9.17; N, 10.65

Found: C, 69.69; H, 9.02; N, 10 35. 42) Compound 145

Analysis: C₃₆H₄₇N₅O₄ +H₂O

Calc: C, 68.44; H, 7.82; N, 11.08

Found: C, 68.69; H, 7.74; N, 10.76.

43) Compound 148

Analysis: C₃₆H₄₇N₅O₄ +0.4H₂O Calc: C, 69.62; H, 7.56; N, 11.27

Found: C, 69.79; H, 7.70; N, 11.12.

44) Compound 149

Analysis: C₃₉H₅₀N₄O₄ +0.4H₂O Calc: C, 72.50; H, 7.93; N, 8.67

Found: C, 72.44; H, 7.99; N, 8.87.

45) Compound 150

Analysis: C₃₆H₄₇N₅O₄ +0.3H₂O Calc. C, 69.83; H, 7.74; N, 11.31

Found: C, 69.93; H, 7.65, N, 11.10.

46) Compound 151

Analysis: C₃₃H₄₇N₅O₄

Calc: C, 68.60; H, 8.20; N, 12.12

Found: C, 68.40; H, 8.16; N, 11.90.

47) Compound 245

Analysis: C₃₆H₄₄N₄O₄

Calc: C, 72.46; H, 7.43; N, 9.39

Found: C, 72.49; H, 7.49; N, 9.25.

48) Compound 246

Analysis: C₃₇H₄₆N₄O₄ +0.25H₂O Calc: C, 72.26; H, 7.61; N, 9.10

Found: C, 72.35; H, 7.83; N, 8.73. 49) Compound 154

Analysis: C₃₅H₄₈N₄O₄ +0.8H₂O Calc: C, 69.70; H, 8.29; N, 9.29

Found: C, 69.65; H, 8.27; N, 9.35.

50) Compound 158

Analysis: C₃₅H₄₈N₄O₄ +0.5H₂O Calc: C, 73.29; H, 7.65; N, 8.33

Found: C, 73.71, H, 7.75, N, 7.75.

51) Compound 159

Analysis: C₃₅H₄₈N₄O₄

Calc: C, 73.09; H, 7.66; N, 8.31

Found: C, 73.40; H, 7.75; N, 7.80.

52) Compound 160

Analysis: C₃₈H₄₈N₄O₄ +1.0H₂O Calc: C, 70.78; H, 8.12; N, 8.68

Found: C, 71.00; H, 8.05; N, 8.59.

53) Compound 161

Analysis: C₄₃H₅₂N₄O₄ +1H₂O Calc: C, 73.05; H, 7.70; N, 7.92

Found: C, 73.29; H, 7.95; N, 7.37.

54) Compound 166

Analysis: C₃₃H₄₆N₄O₄ +1.5H₂O Calc. C, 67.20; H, 8.37; N, 9.50

Found: C, 67.38; H, 7.98; N, 9.41.

55) Compound 171

Analysis: C₃₆H₅₂N₄O₆ +1.6H₂O Calc: C, 64.95; H, 8.36; N, 8.41

Found: C, 65.26; H, 8.15; N, 8.07. 56) Compound 177

Analysis: C₃₈H₄₇N₅O₄

Calc: C, 71.56; H, 7.43; N, 10.98

Found: C, 71.64; H, 7.62; N, 10.93.

57) Compound 178

Analysis: C₃₈H₅₀N₄O₄

Calc: C, 72.81; H, 8.04; N, 8.94

Found: C, 72.96; H, 8.17; N, 8.83.

58) Compound 179

Analysis: C₃₈H₄₇N₅O₄

Calc: C, 71.56; H, 7.43; N, 10.98

Found: C, 72.00; H, 7.55; N, 10.87.

59) Compound 180

Analysis: C₃₈H₄₇F₃N₄O₄

Calc: C, 67.04; H, 6.96; N, 8.23

Found: C, 67.02; H, 7.25; N, 8.23.

60) Compound 181

Analysis: C₃₈H₄₇F₃N₄O₄

Calc: C, 67.04; H, 6.96; N, 8.23

Found: C, 66.63; H, 6.98; N, 7.94.

61) Compound 182

Analysis: C₃₇H₄₇F N₄O_N

Calc: C, 70.45; H, 7.51; N, 8.88

Found: C, 70.28; H, 7.74; N, 8.82.

62) Compound 185

Analysis: C₃₄H₄₈N₄O₄

Calc: C, 70.80; H, 8.39; N, 9.71

Found: C, 70.44; H, 8.45; N, 9.51. 63) Compound 186

Analysis: C₃₇H₄₈N₄O₄ +0.8H₂O Calc: C, 70.85; H, 7.97; N, 8.93

Found: C, 71.12; H, 8.25; N, 8.45.

64) Compound 191

Analysis: C₄₂H₅₁N₅O₄ +.5CH₂C12 Calc: C, 69.78; H, 7.16; N, 9.58

Found: C, 69.75; H, 7.31; N, 9.68.

65) Compound 203

Analysis: C₃₇H₄₇N₄O₄ +1.1H₂O Calc: C, 68.31; H, 7.62; N, 8.61

Found: C, 68.32; H, 7.57; N, 8.53.

66) Compound 204

Analysis: C₃₇H₄₇CIN₄O₄

Calc: C, 68.66; H, 7.32; N, 8.66

Found: C, 68.32, H, 7.48; N, 8.42.

67) Compound 205

Analysis: C₃₈H₅₀N₄O₅ +0.7H₂O Calc: C, 69.63; H, 7.90; N, 8.54

Found: C, 69.72; N, 7.91; N, 8.54.

68) Compound 206

Analysis: C₃₉H₅₂N₄O₆ +0.8H₂O Calc: C, 68.15; H, 7.86; N, 8.15

Found: C, 68.01; H, 8.02; N, 8.15. EX AMPLE 12

A. [S-(R*,S*)]3,3-Diethyl-2-[4-[[[2-(4-hydroxy-1- piperidinyl)ethyl]amino]carbonyl]phenoxy]-N-[1-(4- methylphenyl)butyl]-4-oxo-1-azetidinecarboxamide

When 1-(2-aminoethyl)-4-benzyloxypiperidine is used in place of N,N,N'-trimethylethylene diamine in the procedure of Example l ib there is obtained [S-(R*,S*)] 3,3-diethyl-2-[4-[[[2-(4-benzyloxy-1-piperidinyl]ethyl]amino]carbonyl]phenoxy]-N-[1-(4-methylphenyl)-butyl]-4-oxo-1-azetidinecarboxamide.

B. [S-(R+,S*)]-3,3-diethyl-2-[4-[[[2-(4-hydroxy-1- piperidinyl)ethyl]amino]carbonyl]phenoxy]-N-[1-(4-methylphenyl)butyl]-4-oxo-1-azetidine-carboxamide

A solution of the amide from Step A above in 10 ml of glacial acetic acid containing 22 mg of 10% Pd/C is hydrogenated under 42 lb hydrogen pressure. When TLC indicate completion of the reaction, the mixture is filtered and concentrated in vacuo after the addition of 50 ml toluene. The residue is dissolved in ethyl acetate, washed with saturated sodium bicarbonate solution. The organic layer is dried with sodium sulfate and concentrated in vacuo. The residue is chromatographed on 15 g silica gel using 5% methanol in methylene chloride and yields 96 mg of [S-(R+,S*)]-3,3-diethyl-2-[4-[[[2-(4-hydroxy-1-piperidinyl)ethyl]amino]carbonyl]phenoxy]-N-[1-(4-methylphenyl)butyl]-4-oxo-1-azetidine-carboxamide.

Analysis: C₃₃H₄₆N₄O₅ +1.3H₂O

Calc: C, 65.83; H, 8.13; N, 9.30

Found: C, 66.10; H, 8.06; N, 8.91.

When 1-(2-aminoethyl)-4-benzyloxypiperidine is replaced in the procedure of Example 12 by the appropriate amines, the following compounds 123, 124, 129, 131 and 138 are obtained, for example: 1) Compound 129

Analysis: C₃₄H₄₈N₄O₅

Calc: C, 68.89; H, 8.16; N, 9.45

Found: C, 68.68; H, 8.18; N, 8.65.

2) Compound 131

Analysis: C₃₂H₄₄N₄O₅ +1H₂O

Calc: C, 65.92; H, 7.96; N, 9.61

Found: C, 66.07; H, 7.86; N, 9.45.

3) Compound 138

Analysis: C₃₃H₄₆N₄O₅ +0.5H₂O

Calc: C, 67.43; H, 8.06; N, 9.53

Found: C, 67.61; H, 8.06; N, 9.37.

Diamine Intermediates

The diamines used to prepare the amino amides described herein were commercially available or prepared according to the following routes

EXAMPLE 13

A. N-cyanomethylhomopiperazine

To a solution of 1.98 g homopiperazine in 50 ml acetone is added 4.25 g of powdered anhydrous sodium carbonate and 1.3 ml of chloroacetonitrite. After 24 hrs the reaction mixture is filtered and the filter cake washed with 100 ml acetone. The combined filtrates are concentrated in vacuo and the residue chromatographed on silica gel using methylene chloride as the eluent. The yield of N-cyanomethyl homopiperazine is 2.69 g. B. N-(2-aminoethyl)homopiperazine

To a suspension of 1.02 g lithium aluminum hydride in 50 ml of ether is slowly added a solution of 2.65 gm N-cyanomethyl homopiperazine in 25 ml ether. After the addition is complete the mixture is heated at reflux for 1 hour, then cooled to room temperature and quenched carefully with 1 ml water, 1 ml of 15% sodium hydroxide solution and 3 ml water. The mixture is filtered through sodium sulfate, the filter cake washed well with ether and the combined filtrates concentrated in vacuo to yield 2.60 g. N-(2-aminoethyl)homopiperazine.

EXAMPLE 14

N-(2-methylaminoethyl)homopiperazine

A. N-(2-formamidoethyl)homopiperazine

To a carefully prepared solution of 0.718 g of 60% sodium hydride dispension in 75 ml of absolute ethanol which has been cooled to 0°C is added 7.3 ml of ethyl formate. After 5 minutes there is added a solution of 2.55 gm of N-(2-aminoethyl)homopiperazine in 25 ml absolute ethanol. The mixture is stirred at room temperature overnight. Saturated sodium bicarbonate solution (15 ml) is then added and the reaction mixture is stirred with 150 ml ethyl acetate, filtered through MgSO₄ and the filtrate concentrated in vacuo. Chromatography of the residue on 150 gm silica gel using an eluent of methylene

chloride/methanol/conc. ammonium hydroxide (95/5/0.5) gives 2.78 g of N-(2-formamidoethyl)homopiperazine. B. N-(2-methylaminoethyl)homopiperazine

To a solution of 2.75 gm of N-(2-formamidoethyl)-homopiperazine in 20 ml of dry THF under N₂ is added carefully 60 ml of borane THF solution. After the addition is complete the reaction mixture is heated to reflux for 5 hours then stirred at room temperature overnight. The reaction mixture is quenched by the careful addition of 20 ml of 6 N HCl followed by refluxing for 1 hour. The reaction mixture is cooled, 50 ml of water added and solid KOH added carefully to alkaline pH. Extraction with ether gives the desired product N-(2-methylamino)homopiperazine.

EXAMPLE 15

A. N-benzyl-N,N'-dimethyl-N'-(2-phenylethyl)ethylenediamine

A mixture of 0.900 gm N-benzyl-N,N'-dimethylethyl-enediamine, 1.10 gm powdered sodium carbonate and 0.75 ml of 2- phenylethylbromide is refluxed for 5 hours. An additional 0.25 ml of bromide is added during this time. The reaction mixture is then cooled and filtered. The filterate is concentrated in vacuo and the residue chromatographed on silica gel using an eluent of CH₂Cl₂/CH₃OH/-NH₄OH (97/3/0.3) to yield 0.875 gm of N-benzyl-N,N'-dimethyl-N'-(2-phenylethyl)ethylenediamine.

B. N.N'-dimethyl-N-(2-phenylethyl)ethylenediamine

To a solution 0.870 gm N-benzyl-N-N'-dimethyl-N'-(2- phenylethyl)ethylenediamine in 10 ml ethanol and 5 ml acetic acid is added 0.18 gm Pd (OH)₂/C. The mixture is hydrogenated at 40 psi for 3.5 hours, then filtered and concentrated in vacuo. The residue is made akaline with 1N NaOH and extracted well with ethyl acetate (5 X 25 ml). The combined extracts are filtered through sodium sulfate and concentrated to yield N,N'-dimethyl-N-(2-phenylethyl)ethylenediamine.

EXAMPLE 16

A.

A solution of 1.28 gm 1-(2-amino-ethyl)piperdine and 1.07 gm pyridine-3-carboxaldehyde in 40 ml of toluene is heated to reflux under a Dean Stark trap. After 10 ml toluene distilled over the NMR of an aliquot indicated no aldehyde left. The reaction mixture was concentrated and the imine used directly in the next step.

B.

To a suspension of 0.380 gm of lithium aluminum hydride in 30 ml of dry THF which has been cooled to -10°C is added dropwise a solution of the above imine in 20 ml of dry THF. After about 1 hour the cold reaction mixture is quenched by the addition of 5 ml of 5 N NaOH, then diluted with 100 ml ether and 20 ml of water. The organic layer is separated, washed with brine, filtered thru sodium sulfate and concentrated to give 2.17 gm of 1-[2-(3-pyridylmethylamino)ethyl]-piperidine suitable for use in subsequent reactions. EXAMPLE 17

To 7.50 gm of N,N'-dimethylethylenediamine which has been cooled in an ice-ethanol bath is added portionwise over a 30 minute period 1.40 gm of 3-picolyl chloride. After stirring cold for 1 hour after the addition is completed, the reaction mixture is

concentrated in vacuo and the residue partitioned between 50 ml of ether and 10 ml of 5 N NaOH solution. The organic layer is separated and the aqueous layer extracted 2 times with 50 ml of ether. The combined organic extracts are dried through sodium sulfate and concentrated in vacuo. Chromatography on 150 gm silica gel using CH₂Cl₂/CH₃OH/NH₄OH (90/10/1) as eluent gives 0.930 g of N,N'-dimethyl-N-(3-pyridylmethyl)ethylenediamine.

EXAMPLE 18 Amino Acid→ diamine

A. To an ice cooled solution of 2.29 N-CBZ-D-Proline in 50 ml of CHCl₂ is added 1.35 gm 1-hydroxybenzotriazole hydrate followed by 2.06 gm of dicyclohexylcarbodumide. After 20 minutes, 0.85 ml of pyrrolidine is added and the reaction mixture stirred overnight after which time it is filtered and the filtrate concentrated in vacuo. The residue is partitioned between 100 ml ethyl acetate and 50 ml of 2 N hydrochloric acid. The organic layer is separated, washed with 50 ml of 1.0 N sodium hydroxide solution, dried through sodium sulfate and concentrated in vacuo. Chromatography on 150 gm of silica gel using ethylacetate in hexanes (30-100%) as eluent gives 2.04 gm of the desired pyrrolidine amide

B. To a solution of 1.519 gm of the amide (prepared in A) in 20 ml absolute ethanol is added 75 mg of 10% Pd on carbon catalyst. The mixture is hydrogenated at 40 psi for about an hour then filtrate and the filtrate concentrated to yield D-proline pyrrolidine amide.

C. To a suspension of 0.380 gm of lithium aluminum hydride in 15 ml of dry tetrahydrofuran is carefully added a solution of the D-proline amide (prepared in B above) in 10 ml tetrahydrofuran. The mixture is refluxed for 2 hrs then cooled and quenched with 2 ml of 2.5 N sodium hydroxide. The mixture is filtered through a pad of sodium sulfate and the filter cake washed with 2 × 50 ml of ether. The combined filtrates are concentrated in vacuo to yield 0.80 gm of desired 2-(1-pyrrolidinylmethyl)pyrrolidine.

EXAMPLE 19

[S-(R*,S*)]-2-[4-[[(4-Methyl)piperazin-1-yl]carbonyI] phenoxy]-((3,3-diethyl-N-[1-(methylphenyl)butyl]-4-oxo-1-azetidinecarboxamide

A solution of S-(R*,S*)]-4-(((3,3-diethyl-1- ((4-methylphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl chloride (3.8 mmol), prepared as in Example 11 A, in 50 ml of methylene chloride was cooled in an ice bath and a solution of 0.70 gm of N-methylpiperazine in 10 ml of methylene chloride was added over 5 min. The reaction was stirred for 1 hr and was then poured into a mixture of ice water and 10% potassium carbonate. The product was extracted with two portions of methylene chloride and each methylene chloride layer was washed with a portion of brine. The methylene chloride layers were combined, dried over sodium sulfate and evaporated. The residue was purified with flash chromatography using ethyl acetate, then 2% triethylamine/10% methanol/88% ethyl acetate to afford 2.1 gm of the title compound as a white solid.

Analysis: C₃₀H₄₂N₄O₄

Calc: C, 69.64; H, 7.92; N, 10.48

Found: C, 69.62; H, 8.23; N, 10.46.

EXAMPLE 20

[S-(R*,S*)]-2-[4-[[(4-Methyl)piperazin-1-yl]carbonyl] phenoxy]-((3,3-diethyl-N-[1-(3,4-methylenedioxyphenyl)butyl]-4-oxo-1-azetidine-carboxamide

When [S-(R*,S*)]-4-(((3,3-diethyl-1-((3,4-methylenedioxyphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl chloride (3.1 mmol), prepared as in Example 11 A, was reacted with N-methylpiperazine as in Example 19, there was obtained 1.75 gm of the title compound.

Analysis: C₃₁H₄₀N₄O₆

Calc: C, 65.94; H, 7.14; N, 9.92

Found: C, 65.80; H, 7.31; N, 10.05.

EXAMPLE 21

[S-(R*,S*)]-2-[4-[[(4-Hydroxyethyl)piperazin-1-yl] carbonyl]phenoxy]-((3,3-diethyl-N-[1-(methylphenyl) butyl]-4-oxo-1-azetidinecarboxamide

When [S-(R*,S*)]-4-(((3,3-diethyl-1-((4-methylphenyl)-butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl chloride (3.8 mmol), prepared as in Example 11 A, was reacted with N-(2-hydroxyethyl)piperazine (7.6 mmol) and diisopropylethylamine (3.8 mmol) as in Example 19, there was obtained 2.1 gm of the title compound.

Analysis: C₃₂H₄₄N₄O₅

Calc: C, 68.06; H, 7.85; N, 9.92

Found: C, 67.88; H, 7.87; N, 10.17. EXAMPLE 22

[S-(R*,S*)]-2-[4-[[(4-Hydroxyethyl)piperazin-1-yl] carbonyl]phenoxy]-((3,3-diethyl-N-[1-(3,4-methylenedioxyphenyl)butyl]-4-oxo-1-azetidinecarboxamide

When [S-(R*,S*)]-4-(((3,3-diethyl-1-((3,4-methylenedioxyphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl chloride (3.1 mmol), prepared as in Example 11 A, was reacted with N-(2-hydroxyethyl)piperazine (6.2 mmol) and diisopropylethylamine (3.1 mmol) as in Example 19, there was obtained 1.50 gm of the title compound.

Analysis: C₃₂H₄₂N₄O₇●1.5H₂O

Calc: C, 61.94; H, 6.89; N, 9.06

Found: C, 61.95; H, 6.92; N, 8.96.

EXAMPLE 23

[S-(R*,S*)]-2-[4-[[(4-Cyclopropyl)piperazin-1-yl] carbonyl]phenoxy]-((3,3-diethyl-N-[1-(4-methylphenyl)butyl]-4-oxo-1-azetidinecarboxamide

To a solution of [S-(R*,S*)]-4-(((3,3-diethyl-1-((4-methylphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl chloride (3.8 mmol), prepared as in Example 11 A, in 50 ml of methylene chlorine was added N-(cyclopropyl)piperazine dihydrochloride (5.7 mmol) and then a solution of diisopropylethylamine (15.8 mmol) in 10 ml of methylene chloride was added over 5 min with ice-bath cooling. The reaction was stirred for 1 hr at 0°C and then poured into ice water. The product was extracted with two portions of methylene chloride and each methylene chloride layer was washed with a portion of brine. The methylene chloride layers were combined, dried over sodium sulfate and ethyl acetate/50% hexanes, then 70% ethyl acetate/30% hexanes to afford 2.1 gm of the title compound as a white solid. Analysis: C₃₂H₄₂N₄O₄

Calc: C, 70.69; H, 7.91; N, 9.99

Found: C, 70.62; H, 8.04; N, 9.95.

EXAMPLE 24

[S-(R*,S*)]-2-[4-[[(4-Cyclopropyl)piperazin-1-yl] carbonyl]phenoxy]-((3,3-diethyl-N-[1-(3,4-methylenedioxyphenyl)butyl]-4-oxo-1-azetidinecarboxamide

When [S-(R*,S*)]-4-(((3,3-diethyl-1-((3,4-methylenedioxyphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl chloride (3.1 mmol), prepared as in Example 11 A, was reacted with N-(cyclopropyl)piperazine (4.6 mmol) and diisopropylethylamine (9.3 mmol) as in Example 23, there was obtained 1.80 gm of the title compound.

Analysis: C₃₂H₄₂N₄O₇

Calc: C, 67.10; H, 7.17; N, 9.49

Found: C, 67.03; H, 7.31; N, 9.47.

EXAMPLE 25

[S-(R*,S*)]-2-[4-[[(4-Piperazin-1-yl)carbonyl]phenoxy]-((3,3-diethyl-N-[1-(4-methylphenyl)butyl]-4-oxo-1-azetidinecarboxamide

Step A: [S-R*,S*)]-2-[4-[[(4-(t-Butoxycarbonyl))piperazin-1- yl]carbonyl]phenoxy]-((3,3-diethyl-N-[1-(4-methylphenyl)butyl]-4-oxo-1-azetidinecarboxamide

[S-(R*,S*)]-4-(((3,3-Diethyl-1-((4-methylphenyl)-butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl chloride (0.4 mmol), prepared as in Example 11 A, was reacted with N-(t-butoxycarbonyl)piperazine (0.6 mmol) and triethylamine (1.2 mmol) as in Example 23. The crude, title product so obtained was used directly in the following Step B. Step B: [S-R*,S*)]-2-[4-[(Piperazin-1-yl)carbonyl]phenoxy]- ((3,3-diethyl-N-[1-(4-methylphenyl)butyl-4-oxo-1- azetidine-carboxamide

The product from Step A was dissolved in 0.5 ml of anisole and 2 ml of cold TFA was added. The reaction was stirred at 0°C for 1 hr and was then diluted with methylene chloride and evaporated. The residue was taken up in methylene chloride, washed with 10% sodium carbonate and brine, dried over sodium sulfate and concentrated. The residue was purified by flash chromatography using 5, then 10% methanol/methylene chloride to afford 0.212 gm of title product.

Analysis: C₃₀H₄₀N₄O₅●1H₂O

Calc: C, 66.89; H, 7.85; N, 10.40

Found: C, 67.06; H, 7.55; N, 10.30.

EXAMPLE 26

[S-(R*,S*)]-2-[4-[[(4-Piperazin-1-yl)carbonyl]phenoxy]-((3,3-diethyl-N-[1-(3,4-methylenedioxyphenyl)butyl]-4-oxo-1-azetidinecarboxamide

Step A: [S-R*,S*)]-2-[4-[[(4-Benzyloxycarbonyl) Piperazin-1- yl]carbonyl]phenoxy]-((3,3-diethyl-N-[1-3,4-methylenedioxyphenyl)butyl]-4-oxo-1-azetidinecarboxamide

When [S-(R*,S*)]-4-(((3,3-diethyl-1-((3,4-methylenedioxyphenyl)butylamino)carbonyl)-4-oxo-2-azetidinyl)oxy)benzoyl chloride (0.41 mmol), prepared as in Example 11 A, was reacted with N-(benzyloxycarbonyl)piperazine (0.77 mmol) and diisopropylethylamine (1.6 mmol) as in Example 23, there was obtained 290 mg of the title compound.

Step B: [S-R*,S*)]-2-[4-[(Piperazin-1-yl)carbonyl]phenoxy]-((3,3- diethyl-N-[1-(3,4-methylenedioxyphenyl)butyl-4-oxo-1- azetidinecarboxamide

A solution of 250 mg of material from Example 26, Step A in 10 ml of ethanol was hydrogenated at 40 p.s.i. over 50 mg of 10% Pd/C for 16 hrs. The reaction was filtered and evaporated. The residue was purified by preparative TLC eluting with 2% TEA/10% methanol/-88% ethyl acetate to afford 150 mg of title product.

Analysis: C₃₀H₃₈N₄O₆●3H₂O

Calc: C, 59.59; H, 7.33; N, 9.29

Found: C, 59.66; H, 7.65; N, 9.61.

Claims

WHAT IS CLAIMED IS:

1. A pharmaceutical composition comprising a therapeutically effective, non-toxic amount of an (F)-actin shortening protein, a therapeutically effective amount of an elastase inhibitor and a pharmaceutically acceptable carrier.

2. A pharmaceutical composition comprising a therapeutically effective, non-toxic amount of the (F)-actin

shortening protein gelsolin and a therapeutically effective amount of a compound of Formula (I):

or a pharmaceutically acceptable salt thereof wherein:

R is C_1-6alkyl;

R¹ is C_1-6alkyl or C_1-6alkoxy-C_1-6alkyl;

M is

(1) hydrogen,

(2) C_{1 -6}alkyl,

(3) hydroxy C_1-6alkyl,

(4) halo C_1-6alkyl,

(5) C_2-6alkenyl, or

(6) C_1-6alkoxy-C_1-6alkyl;

Ra and Rb are each individually hydrogen;

R² and R³ are each independently

(1) hydrogen,

(2) C_{1 -6}alkyl, (3) halo, or

(4) C_{1 -6}alkoxy,

or R² and R³ are joined together to form a

methylenedioxy group or a furan ring,

R₄ is

wherein

Q is a covalent bond

Y is

R₉, R₁₀, R₁₁ and R₁₂ are each individually hydrogen or C_{1 -3}alkyl;

R₇ and R₈ are each individually

(a) hydrogen,

(b) C_{1 -6}alkyl,

(c) C_{1 -6}alkyloxy C_{2 -3}alkyl,

(d) hydroxy C_{2 -6}alkyl, or

n is 1, 2, 3, 4 or 5;

R₈ and R₉ are joined together to form a mono or di substituted saturated monocyclic ring of 6 to 7 atoms and having two hetero atoms which are the nitrogens to which R₈ and R₉ are attached; and the substituents are independently selected from hydrogen and C_{1 -3}alkyl.

3. A composition according to Claim 2 wherein

R is C_{1 -3}alkyl; R¹ is C_{1 -3}alkyl or C_{1 -3}alkoxy-C_{1 -3}alkyl;

M is

(1) hydrogen,

(2) C_{1 -3}alkyl,

(3) C_2-3 alkenyl, or

Ra and Rb are each hydrogen;

R² and R³ are each independently

(1) hydrogen,

(2) C_{1 -3}alkyl,

(3) C_{1 -3}alkoxy,

R² and R³ are joined together to form a

methylenedioxy group or a furan ring;

R₄ is

wherein

Q is a covalent bond;

^{Y is}

R₉, R₁₀, R₁₁ and R₁₂ are each individually hydrogen or

C_1-2alkyl;

R₇ and R₈ are each individually

(a) hydrogen,

(b) C_{1 -3}alkyl,

(c) C_1-3alkyloxy C_{2 -3}alkyl, n is 1, 2, 3 or 4; R₈ and R₉ are joined together to form a mono or di substituted saturated monocyclic ring of 6 to 7 atoms and having two hetero atoms which are the nitrogens to which R₈ and R₉ are attached; said rings selected from piperazinyl and homopiperazinyl; and the substituents are independently selected from hydrogen and C_{1 -3}alkyl.

4. A composition according to Claim 3 wherein

R is C_{1 -3} alkyl;

R₁ is C_{1 -3} alkyl;

M is

(a) C_{1 -2} alkyl, or

(b) C_{2 -3} alkenyl;

R² is

(a) hydrogen

(b) C_{1 -3} alkyl and

R³ is hydrogen, or

R² and R³ are joined together to form a methylenedioxy group or a furan ring;

R₇ and R₈ are each independently selected from

(a) hydrogen,

(b) C_{1 -2} alkyl,

(c) C_{1 -2} alkoxy C_{2 -3} alkyl,

R₈ and R₉ are joined together to form a mono or di substituted saturated monocyclic ring of 6 to 7 atoms and having two hetero atoms which are the nitrogens to which R₈ and R₉ are attached; said rings selected from piperazinyl and homopiperazinyl; and the substituents are independently selected from hydrogen and C_{1 -3}alkyl.

5. A composition according to Claim 4 wherein

R is methyl or ethyl;

R₁ is methyl or ethyl;

M is

(a) methyl a ethyl, or

(b) allyl; R² and R³ are each hydrogen, or

R² and R³ are joined together to form a methylenedioxy group or a furan ring;

n is 1 or 2;

R₈ and R₉ are joined together to form a mono or di substituted saturated monocyclic ring of 6 to 7 atoms and having two hetero atoms which are the nitrogens to which R₈ and R₉ are attached; said rings selected from piperazinyl and homopiperazinyl; and the substituents are independently selected from hydrogen and C_{1 -3}alkyl.

6. A pharmaceutical composition comprising a therapeutically effective, non-toxic amount of the (F)-actin

shortening protein gelsolin, a pharmaceutically acceptable carrier and a therapeutically effective amount of a compound of Formula (II):

wherein R is:

(1) -CH₃,

(2) 4-fluorophenyl,

(3) 3-chlorophenyl,

(4) phenyl,

(5) benzyl,

(6) H,

(7) i-Pr, (8) i-Bu,

(9) -CH₂CO₂Et,

(10) -CH₂CO₂H,

(11) Et,

(12) Pr,

(13) 2-pyrimidinyl,

(14) -CH₂CH₂OC(O)NHCH₃,

(15) cyclopropyl, or

(16) -CH₂CH₂OH.

7. A composition according to Claim 8 wherein the compound of Formula I is

[S-(R*,S*)]-2-[4-[[(4-methyl)piperazin-1-yl] carbonyljphenoxy]- 3,3-diethyl-N-[1-(3,4-methylenedioxyphenyl)butyl]-4-oxo-1- azetidinecarboxamide.

8. A method of treating lung disease in a patient in need of such treatment comprising:

administration to a patient in need of such treatment a composition according to Claim 2.

9. A method of treating lung disease in a patient in need of such treatment comprising:

administration to a patient in need of such treatment a composition according to Claim 3.

10. A method of treating lung disease in a patient in need of such treatment comprising:

administration to a patient in need of such treatment a composition according to Claim 1.

11. A method of treating lung disease in a patient in need of such treatment comprising: administration to a patient in need of such treatment a composition according to Claim 7.

12. A method of treating a patient with a lung disease, comprising:

administration to a patient in need of sputum viscosity reduction, a composition according to Claim 1 , wherein said amounts are effective to return the lung function of said patients to at least 60-75% of normal as measured by FEV₁.

13. A method of treating a patient with a lung disease, comprising:

administration to a patient in need of sputum viscosity reduction, a composition according to Claim 1, wherein said amounts are effective to return the lung function of said patients to at least 75-90% of normal as measured by FEV₁.