CA2196334A1 - Urokinase receptor ligands - Google Patents
Urokinase receptor ligandsInfo
- Publication number
- CA2196334A1 CA2196334A1 CA002196334A CA2196334A CA2196334A1 CA 2196334 A1 CA2196334 A1 CA 2196334A1 CA 002196334 A CA002196334 A CA 002196334A CA 2196334 A CA2196334 A CA 2196334A CA 2196334 A1 CA2196334 A1 CA 2196334A1
- Authority
- CA
- Canada
- Prior art keywords
- lower alkyl
- aryl
- halo
- compound
- cycloalkyl
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Abandoned
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Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K5/00—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof
- C07K5/04—Peptides containing up to four amino acids in a fully defined sequence; Derivatives thereof containing only normal peptide links
- C07K5/08—Tripeptides
- C07K5/0802—Tripeptides with the first amino acid being neutral
- C07K5/0804—Tripeptides with the first amino acid being neutral and aliphatic
- C07K5/0806—Tripeptides with the first amino acid being neutral and aliphatic the side chain containing 0 or 1 carbon atoms, i.e. Gly, Ala
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P27/00—Drugs for disorders of the senses
- A61P27/02—Ophthalmic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Abstract
Compounds of the invention inhibit urokinase plasminogen activator or uPAR, represented by general formula (I), where R10 is -CH(R9)XCH(R1)(R11) or a capping group, where X is NR12, CR12R15, O, S, SR12, or SR12R15; R1, R9, R11, R12, R15 are each H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-alkynyl, aryl-cycloalkyl, substituted with 0-3 halo, OH, NH2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, CN or NO2; R16 is -CH(R5)C(=O)NH2, H, lower alkyl, cycloalkyl, or lower alkenyl; R2 is aryl or aralkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl; R3 and R5 are each independently H
or lower alkyl; R4 is -CH2C(=O)NR13R14, where R13 is H, lower alkyl, phenyl or benzyl, and R14 is H, aryl, or aralkyl, where R6 and R7 are each independently H, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, and n and m are each independently an integer from 1 to 3; and pharmaceutically acceptable acid addition salts thereof.
or lower alkyl; R4 is -CH2C(=O)NR13R14, where R13 is H, lower alkyl, phenyl or benzyl, and R14 is H, aryl, or aralkyl, where R6 and R7 are each independently H, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, and n and m are each independently an integer from 1 to 3; and pharmaceutically acceptable acid addition salts thereof.
Description
W0 96/40747 2 1 9 6 3 3 4 ~ /U~ ,6~~
Urokinase Receptor Li ands Description Field of the Invention This invention relates to the fields of ligands of the urokinase . ' ~ ~, activator receptor, and methods for using and preparing the same.
o Background ofthe Invention Urokinase-type j ' O activator (uPA) is a - ' ' serine protease, having a catalytic "B" chain (amino acids 144-411), and an amino-teminal fragment ("ATF", aa 1- 143) consisting of a growth factor-like domain (4-43) and a kringle (aa 47-13~). The uPA kringle appears to bind heparin, but not fibrin, Iysine, o m -~ ' acid. The growth factor-like domain bears some similarity to the structure of epidermal growth factor (EGF), and is thus also referred to as an "EGF-like" domain. The single chain pro-uPA is activated by plasmin or other proteases, cleaving the chain into the two chan active fomm, which is linked together by a disulfide bond.
uPA binds to its specific cell surface receptor (uPAR). The binding interaction is 20 apparently mediated by the EGF-like domain (S.A. Rabbani et al., J Biol Chem (1992) 267: 14151 -~6). Cleavage of pro-uPA into active uPA is accelerated when pro-uYA and , ' O are receptor-bound. Thus, plasmin activates pro-uPA, which in tum activates more plasmin by cleaving I ' O This positive feedback cycle is apparently limited to the receptor-based proteolysis on the cell surface, since a large 2~ excess of protease inhibitors is found in plasma, including C~2 a , ' and PAI-l.
Plasmin can activate or degrade c.~t, a~,.,ll~lldl proteins such as fibrinogen, fibronectin, and zymogens, particularly of the matrix " r ot~ a~cs Plaslulllo activators thus can regulate ~lla~,~llula~ proteolysis, fibrin clot Iysis, tissue rt d"i~ ."~l cell migration, n ~ , and metastasis. Accordingly, there is great 30 interest in developing uPA inhibitors and uPA receptor antag--ni~ E. Appella et al., l Biol Chem (1987) ~.4437-40, detemlined that receptor binding activity is localized in , W096~40747 2 1 9 ~ 33~ ~"~ 1~
-2- :
the EGF-Gke dom~Zin, ~and that residues 12-32 appear to be critical for binditZg The critical domain a one ~uPAZ232) bound uPAR with an afrmity of 40 nM (about 100 fold lcss than intact ATF).
Recent studies have shown that the ~ "... of human turnor cell Gnes in vitro 5 corrddes with su face bound urokhZase, and that urokinase production itself is an prognostic indicdor in human breast cancer (W. Scblechte d al., Cancer CornmZ ~1990) _:173-79; H. Kobayashi et al., Br J Cancer (1993) 67:53744; J.A.
Foekens et ai., Cancer Res (199Z~ 52:G101-05~. It has a so been shown in both breast and colon cancer that urokinase is often madc by stromal cells (fibroblasts and U~ G ~)~
0 where~Zs the urokinaser~eceptor is found on tumor cells (C. Pyke et al., (~ Res (1993) 53:1911-lS; C.Pykeetal.,ArnJPath(19gl~138:1059-67). UPARhas- , y been identified as a monocyte activation antigen, MQ3, whose expression is induced in these n y celis upon activation ~.Y. Min et al., J Immunol (1992~ 148 3636-42~ as well as an activation antigen on human T 1~ , ~te,~ ~A I ;iykj~er et al., I
1~ munol~l994)152:5Q5-16~. Urokinasej 7~ activator knock-out mice(in which the uPA gcno is inactivated or dcleted throughout the body) have been dçveloped, and their ~ are deficiçnt in ,Atl~llul. I matriA d ,~r- in vitro (P.
Carmeliet et al., Fibrinolysis (1993~ 7 Suppl. I :27-28~. In addition, thesç mice show greatly rcduced smooth muscle cell l. c~ p. ,Lî~ Liuu aPer arterial wounding, 213 suggesting a possible role for uPA/uPAR in post-angioplasty restenosis.
The induction of urokinase and its receptor by agents known to be angiogenic in vivo, such as bFGF, vEGF, and TNFc, suggests a role for cell surface urokinase in h ~ -g~ (P. Mignatti et al., ~ç~Ql (I 991~ 113: 1193-202; L.E. Odekon et al., J
Cell Physiol (1992~ 258~3; M.J. Niedbala et ah, Blood ~1992) 79:678-87).
25 Although many factors are likely to be angiogen c m L~ O~- ~ conditions"li ,. n<l -0 of ~ I ".. 11 1 matrix by capillaly endothelial cells and release of matrix-bound pro-angiogenic factors by cell surface plasmin is likely a common sLe~p in these processes (D.
Weinstat-Saslo et al., ~ (1994) B:401-07). This is filrther supported by the ,1,~, ~ .ii.,., that several known anti l~ ~, substances reduce uPA expression (S.
Takano çt al., Cancer Res (1994) 54:26~4-60~. In vivo studies~ have shown that prevention of urokinase-receptor binding, by urokinase amtibodies or ~ u. .Il ~- iI ;. !I Yvith WO !16/40747 2 1 ~ 6 3 3 4 ~ ,, L S~
~ -3-inactive urokinase mutants, ~ reduces or eiiminates the metastatic potential of human prostate tumor ceiis in nude mice (C.W. Crowley et ai., Proc Natl Acad Sci USA
(1993) 90:5021-25; L. Ossowski et al., S~ll (1983) 35:611-19; L. Ossowski, J Cell Biol (1988) 107:2437-45). It has recentiy been shown in both in vitro and syngeneic in vivo 5 models that the protein uPAR antagonists are I . ~ (Min et al., Cancer Res (1996) 56:2428).
Aithough a primary role of uPAR is in the focusing of uPA dependent I ' ", activation to the cell surface, it aiso has other functions. For instance, uPAR
is involved in cell adhesion, functioning as a uPA dependent vitronectin receptor (Wei er al., I Biol Chem (1994) 269:32380-88). More recentiy, it has been shown that uPAR
interacts with integrins and is likely involved in cell shape changes and cell migration (Kindzelskii et al., I Immunol ( i 996) 156:297).
To date, oniy two smail molecules have been described which inhibit the uPA:uPAR interaction (suramin N. Behrendt et al., J Biol Chem (1993) 268:5985-89;
and8. " ,' ' ' -sulfonicacid: M.Plougetal.,E~ ' v(1994)33:8991-97). 1,'. '' i ~, these compounds are effective only at ~ ,lu...old. ~Jnc~ at Summary of the Invention We have now invented , ' which bind tightly to uPAR and are capab1e of 20 inhibitrng the uPA:uPAR interaction, and thus useful for treating disorders or diseases mediated by uPA and/or uPAR. The compounds have the generai structure:
R,oJ~ R16 R1~l,(X)~/
where Rlo is 1 ' 9 or a capping group, where X is NR~2, CR~2R~5, O, S, SRI2, or SRI2Rls; R~, R9, Ru, R~2 R~s are each i~ ly H, lower alkyl, lower25 alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-aikynyl, aryl-cycloalkyl, ' or substituted with 1-3 haio, OEL NH2, lower alkyl, halo-lower aii yl, lower .. . . . . . . . . . . _ . , . .. _ wo 96/40747 ~ 2 1 9 6 3 3 ~ Pcrlus~610g648 ~Nti2 alko y, lower ~llyl o~, lower alkylthio, CN or NO2; Rl,i is 5 , H, lower aikyl, cycioaikyl, or iower aikenyl; R~ is aryl or araikyl, ~ ' ' ' or substituted with 1-3 haio, OH, NH2, C~, NO2, lower aikyl, haio-iower aikyl, lower alko?y, lower aliyhmino, lower allylthio, or cycloalkyl, R3 and R5 arc each ' ', ~ ' ~ H or lower 5 alkyl;
IR1r5 --~N'R
RJ jS ~ ~:~ . where R,3 ;S ~ lower allyl, phenyl or ber~l, and R~ is H, aryi, ~(7~3)n ~ (R7)m arallyl, or especially , where R~s and R, are each ' ', ' l~, H, OH, NH2~ CN, NQ2, lower allyl, halo-lower alkyl, lower alkoxy, lower aikylarnino, lower aikylthio, or cycloalkyl, and n and m are each ' ' ',~, an integer from l to 3 10 inclusive; andplla~ acceptableacidadditionsaitsthereof.
Another aspect~of the invention is the method of treating tumor ~ by ' ' ' ~ a compormd ofthe invention to a wbject in need thereo~
Another aspect~of the invention is a ~ ,e~t.wl r. ~ .U. . comprising an effective amount of a mmpound ofthe invention and a ~Lca " '~ acceptable 1~ excipient.
;[)etaiied Description Definitions The terms "compound of the invention" and acompound of Formula I " refer to a compound of the formula: ~
~ - 5-R2 ~
R1~1,(X)~
where Rlo is 1 ~ 9 or a capping group, where X is NR~2, CRI2Rls, O, S, SR~2, or SRI2Rls; Rl, R9, R~l, Rl2 Rl5 are each ;.,~ p. ~ H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkeny'i, aryl-alkynyl, aryl-cycloalkyl, substituted s with 0-3 halo, OH, NH2, lower alkyl, haio-lower alkyl, lower alkoxy, lower alkylamino, lower alkyltbio, CN or NO2;
o \~NH2 R,6 is , H, lower aikyl, cycloalkyl, or lower alkenyl;
R2 is aryl or aralkyl, , - ,i ,~ . 4 or substituted with 1-3 halo, OH. NH2, CN, NO2, lower alkyl, halo-lower aikyl, lower alkoxy, lower alkylamino, lower alkylthio, or lo cycloalkyl; R3 and R5 are each ' ', ' 'y H or lower aikyl, iR13 ~N'R
R~ is , where Ru is H, lower a'ikyl, phenyl or benzyl, and R~ is ~J~(Rs)n [~(R7)m H, aryl, ara'ilcyl, or especially , where R6 and R7 are each 1 1.. 1.. lly H, OE~, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower aikylthio. or cycloalkyl, and n and m are each i I ~ ly an integer s from I to 3 inclusive, and ~,La~ acceptable acid addition salts thereo~
WO 96/40747 2 1 9 6 3 3 4 ~ ~"1 -~ ~
The terrn "alkyl" as used herein refers to saturated h~Lu~ ùn radicals containing from I to 30 carbon atoms, inclusive. A,kyl radicals may be straight,branched, or cyclic. :~emplary alkyl radicals irlclude ~pentyl, n-hexy~, n-octyl, n-dodecyl, 2-dodecyl, 4Octadecyl, 3,5~ k~yl t~ ~ yl, duryl~ and the like. The term5 "lower alkyr' as used~herein refers to straight, branched, and cyclic chain ~J~ u~ bù~
radicals having from I to 8 carbon atoms, such as methyl, ethyl, propyil isopropyl, ~1-butyl, s-butyl, t-bu~ n-pentyl, n-hexyl, cyclopentyl, cyclohexyl, 2~ u,yl,luy~ yl~
~,yl,lùy~...yl~ l, and the like. "Alkoxy" refers to radicals of the formula -OR, where R
is alkyl as defined aboYe: "lower alkoxy" refers to alkoxy radi; als wherein R is lower o aL1cyl. "Hydrûxy-lower alkyl" refers to r2dicals of the fûrmula HO-R-, where R ;s lower alkylene of I to 8 carbons, and may be stra;ght, branched, or :cyclic '~Iydroxy-lower alkoxy" refers to radicals of the formula HO-R-O-, where R is lower alkylene of I to 8 carbons, and may be s~r ight, branched, or cychc. "Lower alkoxy-lower alkyl" refers to ~roups ofthe formula R.~Rb-~ where R and R~ are each .: ' ~, ' 'y lower alkyl.
15 "Lower alkoxy-low~ alkoxy" refers to groups of the formula RO-RbO-, where R and R~
are each ', ' ~y lûwer alk~yl.
~ AIkenyl'7 refers to h) d~u.,~ ,., radicals of 2-20 carbon atoms having one or more double bonds. Alkenyl radica s may be straigltt, branched, or cyclic. Exempiary alkenyl radicais includc l-padenyl, 3-hexenyl, 1,4-octadienyl, 3,5 I;~.lh~ ,luL~ and the20 like. "Lower ar~cenyr' ~refers to all;enyl radicats hat~ing 2-8 carbon atDms The term Ualkynyl" refers to h~ll u~ uu radicals of 2-20 car~ûn atoms having one or more triple bonds. ALicynyl radicals may be straight, branched, or cyclic.
Exemplary alkynyl radicals include l-per,tynyl, 3-hexynyl, oct=a-2-yn-o-enyl, 3,5-d;"~Jt~,y~ L~ and the like. "Lower alkynyl" refers to alkynyt, radicals having 2-8 25 carbon atoms. ~ ~
The term "haloalkyl" refers to an aikyl radical substr~ted wi~h one or more halo&en atorns. Exemplary haloalkyl radic~s inciude l~inuu~ul~lllyl~ 2,2,2-trifiuoroethyl, 3C~ U~Y~ ' yl,2-bromo-3-~,luu~u~ ' ' yl,2,3-d;b~u.l,vbu~yl,and~helike.
"Aryr' refers to aromatic IIYdI UI,GI b~ having up to 14 carbon atoms~ preferably 3û phenyl, naphthyl, or b~nzhydryl. '~A ryl-lower alkyl" or "arall~yi" refers to radicais of the form Ar-R-, where Ar~ is aryl and R is lower aikyi. "Aryloxy" refers to radicals of the 21 ~6334 ~ -7-form Ar-O-, where Ar is aryl. "Aryloxy-lower alkyl" refers to radicals of the form ArO-R-, where Ar is aryl and R is lower alkyl. "Aryl-cycloalkyl" refers to a condensed ring radical having at least one aromatic ring, and at least one cycloalkyl ring, for example, 1-indanyl, S-indanyl, 9-duorenyl, 5,6,7,8-t ', J. u~ pLillyl, and the like.
The term "acyl" refers to a radical of the formula RCO-, in which R is H, alkyl as defmed above, phenyl, benzyl or naphthyl. Exemplary acyl groups include acetyl, propionyl, formyl, t-' y~bu..~l, benzoyl, and the like. "Lower acyl" refers to radicals wherein R is lower alkyl.
The term "halo" refers to a halogen radical, such as F, Cl, Br, or 1.
0 The term "capping group" refers to a small organic moiety commonly used for protecting amines and amides during synthesis. Exemplary capping groups include,without limitation, methyl, benzhydryl, 4,4'- ' ' yl,~,,lLh.~JIyl, and other acylating reagents (e.g., activated acids such as benzoic acids, benzoyl halides or anhydrides), and the like. In general, a capping group will preferably have a molecular weight less than about 500 g/mol, more preferably less than about 300 g/mol, and most preferably less than about 230 g/mol. Presently preferred capping groups include methyl, benzyl, phenyl, phenethyl, benzhydryl, and 4,4'-dimethoAyt~,.~h~d.yl.
The term "treatment" as used herein refers to reducing or alleviating symptoms in a subject, preventing symptoms from worsening or ~" uOI ~,.,..;.,g, inhibition or elimination 20 of the causative agent, or prevention of the infection or disorder in a subject who is free therefrom. Thus, for example, treatment may be reduction of or the prevention ofmetastasis in a patient having or susceptible to having a metastatic tumor.
The term "uPA- or uPAR-mediated disorder" refers to a disease state or malady which is caused or /~A~ I..II I by a biological activity of uPA or uPAR. The primary 25 biological activity exhibited is 1 ' O activation. Disorders mediated by I ' ~g activation include, without limitation, i~ lu~JIhlL~ ngi~-g.... ~ .g~., diabetic retinopathy, corneal ,~ , Kaposi's sarcoma, and the like), metastasis and invasion by tumor cells, and chronic i ~ (e.g., rheumatoid arthritis, ~ Jh.yb."l.a, and the like). EuGui"~l~icd ATF is also mitogenic for some tumor cells (e.g., SaOS-2 30 O~ ,ulllcl cells), which sometimes self-activate in an autocrine ~/O 96/40747 ~ 2 ~ 9 ~ 3 3 4 PCT~US96/09648 ~ - 8 -Accordingly, the huPAR antagonist of thc invention is effective ;A inhibiting the ,.uL~u. at;u.. of uPA-ac:tivated tumor cells.
The term "effe~d~e amountn refers to an amount of huPAR antagonist compound sufflcient to exhibit a d~etectable therapeutic effect. The therapeutic effect may include, 5 for example, without li~mitation, inhibiting the grow~h of undesired tissue or malignant cells, inhibiting ,, ~ a ~ lirniting tissue dama8e caused by chronic , amd the ~I;e. The precise effcctive aunount for a subject wiD depend upon the subject's size and h~alth the nature and severity of the condition to be treated, and the like. Thus, it is not possible to specify an exact effective amount in advance. However, o the effective amount for a given situation can be determined by routine based on the i ~ liUl~ provided herein.
The term "F ' '1~ acceptable" refers to ~ . ' and ~ o~:l ;. " .
which may be ' - ' to mammals without undue toxicity. Exemplary ~,h~u, '1~ acceptable salts include mineral acid salts such as hJd~, ' ' ;d~., 15 h, .' Ubl ~ ' I , r ~ sulfates, and the like; and the salts of organic acids such as acetates, IJlUIJ;UIla~, malonates, benzoates, and the like.
::
General M-~h-trlc, ' 1~ escription Compounds of the invention are easily synthesized by standard chemical methods.
2~ The presently-preferred~method of synthesis is the "~ .. ." technique described by P. Bartlett et al., WO9 111973~ h n ~ 5~ .J herein by reference. Briefly, an amine (generally bound to a solid phase~ is acylated by a reactant having a carbonyl group and a leaving group (and optwnally a side chain) to form an amide. This react;on is conducted under standard condhions for acylation of an amine, as described by Bartlett et al. The 25 acylating reagent is preferably in the form of an "activated" carbonyl, e.g., as an anhydride, acyl halide, carbonate, or the like. The leaving group is then displaced with a primary or secondary amine under conditions appropriate for S~2 ,', 'a t, as shown in the Scheme below: ;
WO 96140747 2 l 9 6 ~ ~ 4 PCTtU896/09648 g O O
L~X + H~N{~ ' ~NI~O
Rs Rs ~
~/ R4 O
Rs \ L J~X
\ +
~R~ 4 ~ Rs The cycles of acylation and S~,2 ~' r'- are repeated until a compound ofthe desired size is obtained. Either the tern inal amine or the terminal amide may be "capped"
with a suitable capping group, such as methyl or 4,4'-d;.. ,IIIOAYIJ~ dIYh for example 5 by reacting the compound with 4,4'-!" '' yb~...LIlyllyl alcohol under acidic conditions following cleavage from the synthesis resin.
The reactants employed in synthesis of the ~ G: ~1' are generally 'Iy available. Other reactants (e.g., less-common substituted amines) may be prepared by standard chemical means from amines that are ~ullll.~.,.~;dll~ available.
Compounds of the invention may be assayed for activity using standard protocols.For example, one may employ the protocol d~ LI aled in the Examples below to determine binding of C~J---r I of the invention to any desired receptor subtype (6.g., using different sources of tissue). Compounds which exhibit strong binding to receptors will exert either agonistic or (more usually) ~ ' ~ ' activity, which may be wo g6/40747 ~ ~ 2 1 9 6 3 3 4 Pcrlus96/og6~8 ~ o ~ --determined by means~:of .~y~,l " ~ ' tissue-based or in vivo assays known in the art.
Cl ~ ' within thc scope of the invention may easily be assayed for activity by standard receptor-binding aswys.
Cl , ' ofthe invention may be screcned for activity following any generaily 5 suitable aswy for urokinase activity or ir~lhibition. A ~ ly u~ful assay described in Goodson et al., Proc ~atl Acad Sci USA (19g4) 91:7129 ( ~",i~i herein by reference~. One may substitute fragments of urokinase for the ir~act molecule (e.g., one may use the EGF-likc binding domain alone, without the ~ P~tivc portion of uPA). In general, the:~l . ' should be tested against uPA receptors deriveci from o the species to be treated, as some specieD specificity is known to exist.
Compounds of:the invention are - ' t, i orally, topically, or by parenteral means, inciuding ~..l,....:-...., ~ and i~ - injection, , ' of sustained release depots, i"i...~ injection, intranasal - ' ~ , and thc like. lh~hen used to treat tumors, it may be ~,J~ ~ to apply the compound directly to the site, e.g.
t5 during surgery to rem~re the bulk of the tumor. A . ' ~ , . , ' of the invcntion antagonist may be - ' ~xi as a ~ u~ compr;sing the compound in ~ ' ' with a ~ "y acceptable excipient. Such may be aciueous solutions, emuisions, creams, ointments, Y~ , gels, iiposomal ! rl :~ and the like. Suitable excipients incl~lde water, saline~ Ringer's 20 solution, dextrose soh tion, and solutions of ethanol~ glucose7 sucrosc, dextran, mannose, mannitol, sorbitol, p.~ hyl~l~, glycol (PEG), phosphate, acetate, gelatin, collagen, Carbopol~9, vegetabie oils, and the like. One may ~ inc!ude suitable dti~.,D, stabilizers, . ' ' , ~ uiJ;~ls, and buffering agents, for example, BHA, BHT, citric acid,~ ascorbic ac;d, L~ , and the like. Cream or ointment bases 25 usefui in ~( ' ' inciude ianolin, S;lvadene~ ~Marion~, Aquaphor~9 ~DuLe T ~rAt~ri~Dc)~ and ~he like. Other topical r.,. ~Ul~l;OllD include aerosols, bandages, and other wound dressings. Aiternati~y, one may - l or ~ r the compound in a suitable polymer matrix or membrane, thus providing a sustained-release deliver,v device suitable for ,~' ' near the site to be treated locaily. Other devices indude ~o indv~elling catheters and devicesD such as the Aizeta9 minipump. Ophthalmic p. ~,~,.t. ali.JnD
may be formulated usi~g ~ available vehicles such as Sorbi-care~9 (Allergan), WO 96140747 2 1 9 6 3 3 4 r~ o. 61-N~d~Jn ~ (Merck, Sharp &: Dohme), Lacrilube~, and the iike, or may employ topicaf p~ Liu..s such as that described in US 5,124,155, ill~ lUI ~,d herein by reference. Further, one may provide a compound of the invention in solid form, especiafly as a Iyopfiiiized powder. Lyophilized r ~ typicafly contain stabilizing and 5 bulking agents, for example human serum albumin, sucrose, mannitol, and the like. A
thorough discussion of ph~,. . "~ acceptable excipients is available in Remington's r- Sciences (Mack Pub. Co.).
The amount of compound required to treat any particular disorder will of course vary depending upon the nature and severity of the disorder, the age and condition of the o subject, and other factors readify determined by one of ordinary skill in the art. The u~ i ' dosage may be determined by one of ordinary skill by following the methods set forth below in the examples. As a general guide, about û.0 1 mg/Kg to about 50 mglKg compound r ' ~ ~ cd i.v. ot ~ -- u- ~1 y is effective for inhibiting tissue damage due to chronic ~ n - For treating corneal A~g;~ ' ., the compound 15 may be ' ~ ~J locaffy in a gel or matrix at a _liùl~ of about 0.001 mg/Kg to about S mglKg.
Examples The examples presented below are provided as a further guide to the ~
20 of ordinary sfcilf in the art, and are not to be construed as limiting the invention in any way Example I
(Synthesis of Compounds of the Invention~
25 A.) Preparation of CHiR 5585 1.) Loading Elo.,.uacet;~, acid on Wang resin Rinfc resin (2.71 g, 1.98 mmole) with ~h~titllti~n 0.73 mmofe/g is swoffen with 15 mL
d;chfv-, ' (DCM) in a 50 mL reaction vessel and drained later. Blu~uac~ , acid (1.12g, 8 mmole) is mixed with IM DCC/NMP (8 mL, 8 mmole) and 10 mL DCM.
~o Dhl.~Lh,: r.~.hl;"e (58.5 mg, 0.48 mmole) is added into the resin. 18 mL ofactivated B-u...J~.~,etic acid/DCClNl~lDCM solution is then added into the reaction _ _ . . _ . _ . , .. . . . . .. . . .. _ . . _ .. . . _ _ vVo 96/40747 ~: ~ 21 9 6 3 3 4 PCT~US9h~
~2-vessel. The resin mixture is shaken for t~ min at room t~."~ and then drained and washed with 15 mL D~M 3 X3 15 mL DM F 2x and 15 mL IPA. The loaded resin (~ is dried under vacuo to ~vide IJ
2.) ~ (4.4'~ 'L )~1~ '~
Loaded resin t7~~ mg, 100 ~mole)(l) is sv~ilen with 2 mL DMSO in a 8 mL
reaction vessel, and then drained. Fmoc-protected N-(4~4'- " ' ,' ' ~ J.yl)-~j ' (5 mrnole~ is mixed with DMSO (1.907 mL3 to prepare a 2.5 mL solution of 2 M 2 - ' .~ ' - which is then aWed to the reaction vessd. The resin mixture is shaken at 45~C for 4 hr~ then drained and washed with 3 mL DMI~ 6X and 3 mL DCM
o 6X. to provide the loaded resin ~.
Urokinase Receptor Li ands Description Field of the Invention This invention relates to the fields of ligands of the urokinase . ' ~ ~, activator receptor, and methods for using and preparing the same.
o Background ofthe Invention Urokinase-type j ' O activator (uPA) is a - ' ' serine protease, having a catalytic "B" chain (amino acids 144-411), and an amino-teminal fragment ("ATF", aa 1- 143) consisting of a growth factor-like domain (4-43) and a kringle (aa 47-13~). The uPA kringle appears to bind heparin, but not fibrin, Iysine, o m -~ ' acid. The growth factor-like domain bears some similarity to the structure of epidermal growth factor (EGF), and is thus also referred to as an "EGF-like" domain. The single chain pro-uPA is activated by plasmin or other proteases, cleaving the chain into the two chan active fomm, which is linked together by a disulfide bond.
uPA binds to its specific cell surface receptor (uPAR). The binding interaction is 20 apparently mediated by the EGF-like domain (S.A. Rabbani et al., J Biol Chem (1992) 267: 14151 -~6). Cleavage of pro-uPA into active uPA is accelerated when pro-uYA and , ' O are receptor-bound. Thus, plasmin activates pro-uPA, which in tum activates more plasmin by cleaving I ' O This positive feedback cycle is apparently limited to the receptor-based proteolysis on the cell surface, since a large 2~ excess of protease inhibitors is found in plasma, including C~2 a , ' and PAI-l.
Plasmin can activate or degrade c.~t, a~,.,ll~lldl proteins such as fibrinogen, fibronectin, and zymogens, particularly of the matrix " r ot~ a~cs Plaslulllo activators thus can regulate ~lla~,~llula~ proteolysis, fibrin clot Iysis, tissue rt d"i~ ."~l cell migration, n ~ , and metastasis. Accordingly, there is great 30 interest in developing uPA inhibitors and uPA receptor antag--ni~ E. Appella et al., l Biol Chem (1987) ~.4437-40, detemlined that receptor binding activity is localized in , W096~40747 2 1 9 ~ 33~ ~"~ 1~
-2- :
the EGF-Gke dom~Zin, ~and that residues 12-32 appear to be critical for binditZg The critical domain a one ~uPAZ232) bound uPAR with an afrmity of 40 nM (about 100 fold lcss than intact ATF).
Recent studies have shown that the ~ "... of human turnor cell Gnes in vitro 5 corrddes with su face bound urokhZase, and that urokinase production itself is an prognostic indicdor in human breast cancer (W. Scblechte d al., Cancer CornmZ ~1990) _:173-79; H. Kobayashi et al., Br J Cancer (1993) 67:53744; J.A.
Foekens et ai., Cancer Res (199Z~ 52:G101-05~. It has a so been shown in both breast and colon cancer that urokinase is often madc by stromal cells (fibroblasts and U~ G ~)~
0 where~Zs the urokinaser~eceptor is found on tumor cells (C. Pyke et al., (~ Res (1993) 53:1911-lS; C.Pykeetal.,ArnJPath(19gl~138:1059-67). UPARhas- , y been identified as a monocyte activation antigen, MQ3, whose expression is induced in these n y celis upon activation ~.Y. Min et al., J Immunol (1992~ 148 3636-42~ as well as an activation antigen on human T 1~ , ~te,~ ~A I ;iykj~er et al., I
1~ munol~l994)152:5Q5-16~. Urokinasej 7~ activator knock-out mice(in which the uPA gcno is inactivated or dcleted throughout the body) have been dçveloped, and their ~ are deficiçnt in ,Atl~llul. I matriA d ,~r- in vitro (P.
Carmeliet et al., Fibrinolysis (1993~ 7 Suppl. I :27-28~. In addition, thesç mice show greatly rcduced smooth muscle cell l. c~ p. ,Lî~ Liuu aPer arterial wounding, 213 suggesting a possible role for uPA/uPAR in post-angioplasty restenosis.
The induction of urokinase and its receptor by agents known to be angiogenic in vivo, such as bFGF, vEGF, and TNFc, suggests a role for cell surface urokinase in h ~ -g~ (P. Mignatti et al., ~ç~Ql (I 991~ 113: 1193-202; L.E. Odekon et al., J
Cell Physiol (1992~ 258~3; M.J. Niedbala et ah, Blood ~1992) 79:678-87).
25 Although many factors are likely to be angiogen c m L~ O~- ~ conditions"li ,. n<l -0 of ~ I ".. 11 1 matrix by capillaly endothelial cells and release of matrix-bound pro-angiogenic factors by cell surface plasmin is likely a common sLe~p in these processes (D.
Weinstat-Saslo et al., ~ (1994) B:401-07). This is filrther supported by the ,1,~, ~ .ii.,., that several known anti l~ ~, substances reduce uPA expression (S.
Takano çt al., Cancer Res (1994) 54:26~4-60~. In vivo studies~ have shown that prevention of urokinase-receptor binding, by urokinase amtibodies or ~ u. .Il ~- iI ;. !I Yvith WO !16/40747 2 1 ~ 6 3 3 4 ~ ,, L S~
~ -3-inactive urokinase mutants, ~ reduces or eiiminates the metastatic potential of human prostate tumor ceiis in nude mice (C.W. Crowley et ai., Proc Natl Acad Sci USA
(1993) 90:5021-25; L. Ossowski et al., S~ll (1983) 35:611-19; L. Ossowski, J Cell Biol (1988) 107:2437-45). It has recentiy been shown in both in vitro and syngeneic in vivo 5 models that the protein uPAR antagonists are I . ~ (Min et al., Cancer Res (1996) 56:2428).
Aithough a primary role of uPAR is in the focusing of uPA dependent I ' ", activation to the cell surface, it aiso has other functions. For instance, uPAR
is involved in cell adhesion, functioning as a uPA dependent vitronectin receptor (Wei er al., I Biol Chem (1994) 269:32380-88). More recentiy, it has been shown that uPAR
interacts with integrins and is likely involved in cell shape changes and cell migration (Kindzelskii et al., I Immunol ( i 996) 156:297).
To date, oniy two smail molecules have been described which inhibit the uPA:uPAR interaction (suramin N. Behrendt et al., J Biol Chem (1993) 268:5985-89;
and8. " ,' ' ' -sulfonicacid: M.Plougetal.,E~ ' v(1994)33:8991-97). 1,'. '' i ~, these compounds are effective only at ~ ,lu...old. ~Jnc~ at Summary of the Invention We have now invented , ' which bind tightly to uPAR and are capab1e of 20 inhibitrng the uPA:uPAR interaction, and thus useful for treating disorders or diseases mediated by uPA and/or uPAR. The compounds have the generai structure:
R,oJ~ R16 R1~l,(X)~/
where Rlo is 1 ' 9 or a capping group, where X is NR~2, CR~2R~5, O, S, SRI2, or SRI2Rls; R~, R9, Ru, R~2 R~s are each i~ ly H, lower alkyl, lower25 alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-aikynyl, aryl-cycloalkyl, ' or substituted with 1-3 haio, OEL NH2, lower alkyl, halo-lower aii yl, lower .. . . . . . . . . . . _ . , . .. _ wo 96/40747 ~ 2 1 9 6 3 3 ~ Pcrlus~610g648 ~Nti2 alko y, lower ~llyl o~, lower alkylthio, CN or NO2; Rl,i is 5 , H, lower aikyl, cycioaikyl, or iower aikenyl; R~ is aryl or araikyl, ~ ' ' ' or substituted with 1-3 haio, OH, NH2, C~, NO2, lower aikyl, haio-iower aikyl, lower alko?y, lower aliyhmino, lower allylthio, or cycloalkyl, R3 and R5 arc each ' ', ~ ' ~ H or lower 5 alkyl;
IR1r5 --~N'R
RJ jS ~ ~:~ . where R,3 ;S ~ lower allyl, phenyl or ber~l, and R~ is H, aryi, ~(7~3)n ~ (R7)m arallyl, or especially , where R~s and R, are each ' ', ' l~, H, OH, NH2~ CN, NQ2, lower allyl, halo-lower alkyl, lower alkoxy, lower aikylarnino, lower aikylthio, or cycloalkyl, and n and m are each ' ' ',~, an integer from l to 3 10 inclusive; andplla~ acceptableacidadditionsaitsthereof.
Another aspect~of the invention is the method of treating tumor ~ by ' ' ' ~ a compormd ofthe invention to a wbject in need thereo~
Another aspect~of the invention is a ~ ,e~t.wl r. ~ .U. . comprising an effective amount of a mmpound ofthe invention and a ~Lca " '~ acceptable 1~ excipient.
;[)etaiied Description Definitions The terms "compound of the invention" and acompound of Formula I " refer to a compound of the formula: ~
~ - 5-R2 ~
R1~1,(X)~
where Rlo is 1 ~ 9 or a capping group, where X is NR~2, CRI2Rls, O, S, SR~2, or SRI2Rls; Rl, R9, R~l, Rl2 Rl5 are each ;.,~ p. ~ H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkeny'i, aryl-alkynyl, aryl-cycloalkyl, substituted s with 0-3 halo, OH, NH2, lower alkyl, haio-lower alkyl, lower alkoxy, lower alkylamino, lower alkyltbio, CN or NO2;
o \~NH2 R,6 is , H, lower aikyl, cycloalkyl, or lower alkenyl;
R2 is aryl or aralkyl, , - ,i ,~ . 4 or substituted with 1-3 halo, OH. NH2, CN, NO2, lower alkyl, halo-lower aikyl, lower alkoxy, lower alkylamino, lower alkylthio, or lo cycloalkyl; R3 and R5 are each ' ', ' 'y H or lower aikyl, iR13 ~N'R
R~ is , where Ru is H, lower a'ikyl, phenyl or benzyl, and R~ is ~J~(Rs)n [~(R7)m H, aryl, ara'ilcyl, or especially , where R6 and R7 are each 1 1.. 1.. lly H, OE~, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower aikylthio. or cycloalkyl, and n and m are each i I ~ ly an integer s from I to 3 inclusive, and ~,La~ acceptable acid addition salts thereo~
WO 96/40747 2 1 9 6 3 3 4 ~ ~"1 -~ ~
The terrn "alkyl" as used herein refers to saturated h~Lu~ ùn radicals containing from I to 30 carbon atoms, inclusive. A,kyl radicals may be straight,branched, or cyclic. :~emplary alkyl radicals irlclude ~pentyl, n-hexy~, n-octyl, n-dodecyl, 2-dodecyl, 4Octadecyl, 3,5~ k~yl t~ ~ yl, duryl~ and the like. The term5 "lower alkyr' as used~herein refers to straight, branched, and cyclic chain ~J~ u~ bù~
radicals having from I to 8 carbon atoms, such as methyl, ethyl, propyil isopropyl, ~1-butyl, s-butyl, t-bu~ n-pentyl, n-hexyl, cyclopentyl, cyclohexyl, 2~ u,yl,luy~ yl~
~,yl,lùy~...yl~ l, and the like. "Alkoxy" refers to radicals of the formula -OR, where R
is alkyl as defined aboYe: "lower alkoxy" refers to alkoxy radi; als wherein R is lower o aL1cyl. "Hydrûxy-lower alkyl" refers to r2dicals of the fûrmula HO-R-, where R ;s lower alkylene of I to 8 carbons, and may be stra;ght, branched, or :cyclic '~Iydroxy-lower alkoxy" refers to radicals of the formula HO-R-O-, where R is lower alkylene of I to 8 carbons, and may be s~r ight, branched, or cychc. "Lower alkoxy-lower alkyl" refers to ~roups ofthe formula R.~Rb-~ where R and R~ are each .: ' ~, ' 'y lower alkyl.
15 "Lower alkoxy-low~ alkoxy" refers to groups of the formula RO-RbO-, where R and R~
are each ', ' ~y lûwer alk~yl.
~ AIkenyl'7 refers to h) d~u.,~ ,., radicals of 2-20 carbon atoms having one or more double bonds. Alkenyl radica s may be straigltt, branched, or cyclic. Exempiary alkenyl radicais includc l-padenyl, 3-hexenyl, 1,4-octadienyl, 3,5 I;~.lh~ ,luL~ and the20 like. "Lower ar~cenyr' ~refers to all;enyl radicats hat~ing 2-8 carbon atDms The term Ualkynyl" refers to h~ll u~ uu radicals of 2-20 car~ûn atoms having one or more triple bonds. ALicynyl radicals may be straight, branched, or cyclic.
Exemplary alkynyl radicals include l-per,tynyl, 3-hexynyl, oct=a-2-yn-o-enyl, 3,5-d;"~Jt~,y~ L~ and the like. "Lower alkynyl" refers to alkynyt, radicals having 2-8 25 carbon atoms. ~ ~
The term "haloalkyl" refers to an aikyl radical substr~ted wi~h one or more halo&en atorns. Exemplary haloalkyl radic~s inciude l~inuu~ul~lllyl~ 2,2,2-trifiuoroethyl, 3C~ U~Y~ ' yl,2-bromo-3-~,luu~u~ ' ' yl,2,3-d;b~u.l,vbu~yl,and~helike.
"Aryr' refers to aromatic IIYdI UI,GI b~ having up to 14 carbon atoms~ preferably 3û phenyl, naphthyl, or b~nzhydryl. '~A ryl-lower alkyl" or "arall~yi" refers to radicais of the form Ar-R-, where Ar~ is aryl and R is lower aikyi. "Aryloxy" refers to radicals of the 21 ~6334 ~ -7-form Ar-O-, where Ar is aryl. "Aryloxy-lower alkyl" refers to radicals of the form ArO-R-, where Ar is aryl and R is lower alkyl. "Aryl-cycloalkyl" refers to a condensed ring radical having at least one aromatic ring, and at least one cycloalkyl ring, for example, 1-indanyl, S-indanyl, 9-duorenyl, 5,6,7,8-t ', J. u~ pLillyl, and the like.
The term "acyl" refers to a radical of the formula RCO-, in which R is H, alkyl as defmed above, phenyl, benzyl or naphthyl. Exemplary acyl groups include acetyl, propionyl, formyl, t-' y~bu..~l, benzoyl, and the like. "Lower acyl" refers to radicals wherein R is lower alkyl.
The term "halo" refers to a halogen radical, such as F, Cl, Br, or 1.
0 The term "capping group" refers to a small organic moiety commonly used for protecting amines and amides during synthesis. Exemplary capping groups include,without limitation, methyl, benzhydryl, 4,4'- ' ' yl,~,,lLh.~JIyl, and other acylating reagents (e.g., activated acids such as benzoic acids, benzoyl halides or anhydrides), and the like. In general, a capping group will preferably have a molecular weight less than about 500 g/mol, more preferably less than about 300 g/mol, and most preferably less than about 230 g/mol. Presently preferred capping groups include methyl, benzyl, phenyl, phenethyl, benzhydryl, and 4,4'-dimethoAyt~,.~h~d.yl.
The term "treatment" as used herein refers to reducing or alleviating symptoms in a subject, preventing symptoms from worsening or ~" uOI ~,.,..;.,g, inhibition or elimination 20 of the causative agent, or prevention of the infection or disorder in a subject who is free therefrom. Thus, for example, treatment may be reduction of or the prevention ofmetastasis in a patient having or susceptible to having a metastatic tumor.
The term "uPA- or uPAR-mediated disorder" refers to a disease state or malady which is caused or /~A~ I..II I by a biological activity of uPA or uPAR. The primary 25 biological activity exhibited is 1 ' O activation. Disorders mediated by I ' ~g activation include, without limitation, i~ lu~JIhlL~ ngi~-g.... ~ .g~., diabetic retinopathy, corneal ,~ , Kaposi's sarcoma, and the like), metastasis and invasion by tumor cells, and chronic i ~ (e.g., rheumatoid arthritis, ~ Jh.yb."l.a, and the like). EuGui"~l~icd ATF is also mitogenic for some tumor cells (e.g., SaOS-2 30 O~ ,ulllcl cells), which sometimes self-activate in an autocrine ~/O 96/40747 ~ 2 ~ 9 ~ 3 3 4 PCT~US96/09648 ~ - 8 -Accordingly, the huPAR antagonist of thc invention is effective ;A inhibiting the ,.uL~u. at;u.. of uPA-ac:tivated tumor cells.
The term "effe~d~e amountn refers to an amount of huPAR antagonist compound sufflcient to exhibit a d~etectable therapeutic effect. The therapeutic effect may include, 5 for example, without li~mitation, inhibiting the grow~h of undesired tissue or malignant cells, inhibiting ,, ~ a ~ lirniting tissue dama8e caused by chronic , amd the ~I;e. The precise effcctive aunount for a subject wiD depend upon the subject's size and h~alth the nature and severity of the condition to be treated, and the like. Thus, it is not possible to specify an exact effective amount in advance. However, o the effective amount for a given situation can be determined by routine based on the i ~ liUl~ provided herein.
The term "F ' '1~ acceptable" refers to ~ . ' and ~ o~:l ;. " .
which may be ' - ' to mammals without undue toxicity. Exemplary ~,h~u, '1~ acceptable salts include mineral acid salts such as hJd~, ' ' ;d~., 15 h, .' Ubl ~ ' I , r ~ sulfates, and the like; and the salts of organic acids such as acetates, IJlUIJ;UIla~, malonates, benzoates, and the like.
::
General M-~h-trlc, ' 1~ escription Compounds of the invention are easily synthesized by standard chemical methods.
2~ The presently-preferred~method of synthesis is the "~ .. ." technique described by P. Bartlett et al., WO9 111973~ h n ~ 5~ .J herein by reference. Briefly, an amine (generally bound to a solid phase~ is acylated by a reactant having a carbonyl group and a leaving group (and optwnally a side chain) to form an amide. This react;on is conducted under standard condhions for acylation of an amine, as described by Bartlett et al. The 25 acylating reagent is preferably in the form of an "activated" carbonyl, e.g., as an anhydride, acyl halide, carbonate, or the like. The leaving group is then displaced with a primary or secondary amine under conditions appropriate for S~2 ,', 'a t, as shown in the Scheme below: ;
WO 96140747 2 l 9 6 ~ ~ 4 PCTtU896/09648 g O O
L~X + H~N{~ ' ~NI~O
Rs Rs ~
~/ R4 O
Rs \ L J~X
\ +
~R~ 4 ~ Rs The cycles of acylation and S~,2 ~' r'- are repeated until a compound ofthe desired size is obtained. Either the tern inal amine or the terminal amide may be "capped"
with a suitable capping group, such as methyl or 4,4'-d;.. ,IIIOAYIJ~ dIYh for example 5 by reacting the compound with 4,4'-!" '' yb~...LIlyllyl alcohol under acidic conditions following cleavage from the synthesis resin.
The reactants employed in synthesis of the ~ G: ~1' are generally 'Iy available. Other reactants (e.g., less-common substituted amines) may be prepared by standard chemical means from amines that are ~ullll.~.,.~;dll~ available.
Compounds of the invention may be assayed for activity using standard protocols.For example, one may employ the protocol d~ LI aled in the Examples below to determine binding of C~J---r I of the invention to any desired receptor subtype (6.g., using different sources of tissue). Compounds which exhibit strong binding to receptors will exert either agonistic or (more usually) ~ ' ~ ' activity, which may be wo g6/40747 ~ ~ 2 1 9 6 3 3 4 Pcrlus96/og6~8 ~ o ~ --determined by means~:of .~y~,l " ~ ' tissue-based or in vivo assays known in the art.
Cl ~ ' within thc scope of the invention may easily be assayed for activity by standard receptor-binding aswys.
Cl , ' ofthe invention may be screcned for activity following any generaily 5 suitable aswy for urokinase activity or ir~lhibition. A ~ ly u~ful assay described in Goodson et al., Proc ~atl Acad Sci USA (19g4) 91:7129 ( ~",i~i herein by reference~. One may substitute fragments of urokinase for the ir~act molecule (e.g., one may use the EGF-likc binding domain alone, without the ~ P~tivc portion of uPA). In general, the:~l . ' should be tested against uPA receptors deriveci from o the species to be treated, as some specieD specificity is known to exist.
Compounds of:the invention are - ' t, i orally, topically, or by parenteral means, inciuding ~..l,....:-...., ~ and i~ - injection, , ' of sustained release depots, i"i...~ injection, intranasal - ' ~ , and thc like. lh~hen used to treat tumors, it may be ~,J~ ~ to apply the compound directly to the site, e.g.
t5 during surgery to rem~re the bulk of the tumor. A . ' ~ , . , ' of the invcntion antagonist may be - ' ~xi as a ~ u~ compr;sing the compound in ~ ' ' with a ~ "y acceptable excipient. Such may be aciueous solutions, emuisions, creams, ointments, Y~ , gels, iiposomal ! rl :~ and the like. Suitable excipients incl~lde water, saline~ Ringer's 20 solution, dextrose soh tion, and solutions of ethanol~ glucose7 sucrosc, dextran, mannose, mannitol, sorbitol, p.~ hyl~l~, glycol (PEG), phosphate, acetate, gelatin, collagen, Carbopol~9, vegetabie oils, and the like. One may ~ inc!ude suitable dti~.,D, stabilizers, . ' ' , ~ uiJ;~ls, and buffering agents, for example, BHA, BHT, citric acid,~ ascorbic ac;d, L~ , and the like. Cream or ointment bases 25 usefui in ~( ' ' inciude ianolin, S;lvadene~ ~Marion~, Aquaphor~9 ~DuLe T ~rAt~ri~Dc)~ and ~he like. Other topical r.,. ~Ul~l;OllD include aerosols, bandages, and other wound dressings. Aiternati~y, one may - l or ~ r the compound in a suitable polymer matrix or membrane, thus providing a sustained-release deliver,v device suitable for ,~' ' near the site to be treated locaily. Other devices indude ~o indv~elling catheters and devicesD such as the Aizeta9 minipump. Ophthalmic p. ~,~,.t. ali.JnD
may be formulated usi~g ~ available vehicles such as Sorbi-care~9 (Allergan), WO 96140747 2 1 9 6 3 3 4 r~ o. 61-N~d~Jn ~ (Merck, Sharp &: Dohme), Lacrilube~, and the iike, or may employ topicaf p~ Liu..s such as that described in US 5,124,155, ill~ lUI ~,d herein by reference. Further, one may provide a compound of the invention in solid form, especiafly as a Iyopfiiiized powder. Lyophilized r ~ typicafly contain stabilizing and 5 bulking agents, for example human serum albumin, sucrose, mannitol, and the like. A
thorough discussion of ph~,. . "~ acceptable excipients is available in Remington's r- Sciences (Mack Pub. Co.).
The amount of compound required to treat any particular disorder will of course vary depending upon the nature and severity of the disorder, the age and condition of the o subject, and other factors readify determined by one of ordinary skill in the art. The u~ i ' dosage may be determined by one of ordinary skill by following the methods set forth below in the examples. As a general guide, about û.0 1 mg/Kg to about 50 mglKg compound r ' ~ ~ cd i.v. ot ~ -- u- ~1 y is effective for inhibiting tissue damage due to chronic ~ n - For treating corneal A~g;~ ' ., the compound 15 may be ' ~ ~J locaffy in a gel or matrix at a _liùl~ of about 0.001 mg/Kg to about S mglKg.
Examples The examples presented below are provided as a further guide to the ~
20 of ordinary sfcilf in the art, and are not to be construed as limiting the invention in any way Example I
(Synthesis of Compounds of the Invention~
25 A.) Preparation of CHiR 5585 1.) Loading Elo.,.uacet;~, acid on Wang resin Rinfc resin (2.71 g, 1.98 mmole) with ~h~titllti~n 0.73 mmofe/g is swoffen with 15 mL
d;chfv-, ' (DCM) in a 50 mL reaction vessel and drained later. Blu~uac~ , acid (1.12g, 8 mmole) is mixed with IM DCC/NMP (8 mL, 8 mmole) and 10 mL DCM.
~o Dhl.~Lh,: r.~.hl;"e (58.5 mg, 0.48 mmole) is added into the resin. 18 mL ofactivated B-u...J~.~,etic acid/DCClNl~lDCM solution is then added into the reaction _ _ . . _ . _ . , .. . . . . .. . . .. _ . . _ .. . . _ _ vVo 96/40747 ~: ~ 21 9 6 3 3 4 PCT~US9h~
~2-vessel. The resin mixture is shaken for t~ min at room t~."~ and then drained and washed with 15 mL D~M 3 X3 15 mL DM F 2x and 15 mL IPA. The loaded resin (~ is dried under vacuo to ~vide IJ
2.) ~ (4.4'~ 'L )~1~ '~
Loaded resin t7~~ mg, 100 ~mole)(l) is sv~ilen with 2 mL DMSO in a 8 mL
reaction vessel, and then drained. Fmoc-protected N-(4~4'- " ' ,' ' ~ J.yl)-~j ' (5 mrnole~ is mixed with DMSO (1.907 mL3 to prepare a 2.5 mL solution of 2 M 2 - ' .~ ' - which is then aWed to the reaction vessd. The resin mixture is shaken at 45~C for 4 hr~ then drained and washed with 3 mL DMI~ 6X and 3 mL DCM
o 6X. to provide the loaded resin ~.
3.~ inp~ with Bl~ Acid ~BA-4~
The loaded resin ~2) is swollen ~qth 3 mL DCM in a 8 mL reaction vessel and then drained. BAA ~84 lli, 750 ,umole) is mixed w;th DIEA ~128 ul, ~50 ~mole) and DCM ~2.2 mL) to prep:are a 2.5 mL of 0.3 M BAAIDIEA/DC~ solution which is then added to the reaction=vessel. The resin mixture is shaken for 20 min at room lc~ lulr and then drained and washed with 3 mL DC~f. The resin sample is treated vith 2. 5 mL of 0.3 M BAAfDlEAl~C~f solution for 20 min again. It is driined and then washed with 3 mL DChif 6X and 3 mL DM~ 6X to provide the loaded resrn O-4.) Cml5tlinl~ ~ with 5 3 The loaded resin (_) u:lrwollc with 2 mL DMSO in a: 8 mL reaction vessel and then drained. 5-~ I (2.5 mmole) was dissolved in DMSO ~2.~ mL) to prepare a 2.5 mL of I M S a ~ ~ DMSO solution which is then added to the reaction vessel.
The resin mixture is shaken at 45~C fw 4 hr. It is th:en drained and washed with 3 mL
DMF 6X and 3 mL D~ 6X to provide the loaded resin (4).
The loaded resin ~2) is swollen ~qth 3 mL DCM in a 8 mL reaction vessel and then drained. BAA ~84 lli, 750 ,umole) is mixed w;th DIEA ~128 ul, ~50 ~mole) and DCM ~2.2 mL) to prep:are a 2.5 mL of 0.3 M BAAIDIEA/DC~ solution which is then added to the reaction=vessel. The resin mixture is shaken for 20 min at room lc~ lulr and then drained and washed with 3 mL DC~f. The resin sample is treated vith 2. 5 mL of 0.3 M BAAfDlEAl~C~f solution for 20 min again. It is driined and then washed with 3 mL DChif 6X and 3 mL DM~ 6X to provide the loaded resrn O-4.) Cml5tlinl~ ~ with 5 3 The loaded resin (_) u:lrwollc with 2 mL DMSO in a: 8 mL reaction vessel and then drained. 5-~ I (2.5 mmole) was dissolved in DMSO ~2.~ mL) to prepare a 2.5 mL of I M S a ~ ~ DMSO solution which is then added to the reaction vessel.
The resin mixture is shaken at 45~C fw 4 hr. It is th:en drained and washed with 3 mL
DMF 6X and 3 mL D~ 6X to provide the loaded resin (4).
5.) Acylating~rec~r~ with B~ u~ , Acid ~B~
BAA (84 ~11, 750 ~mde) is added to resin (4) as described aho~e in part 3.) tû provqde acylated resin (~ F~ the compound may be capped: at this point ~as a dimer~
by acylation with a carboxylic acid.
BAA (84 ~11, 750 ~mde) is added to resin (4) as described aho~e in part 3.) tû provqde acylated resin (~ F~ the compound may be capped: at this point ~as a dimer~
by acylation with a carboxylic acid.
6.) Couplin~ r~sin ~qth 1 h~J~u~y~Jh~l.,th The loaded resin (5) is swollen wqth 2 mL DMSO in a 8 rnL reaction vessel and then drained. ~ IIyl~u~ ' ,' (10 mmole3 is dissolved in DMSO (2.5 mL) to wo 96/40747 2 1 9 6 3 3 4 PCT/US96109648 ~ -13-prepare a 2.5 mL of Z M 4-h~dlu~-~, ' ' ,' 'DMSO solution which is then added to the reaction vessel. The resin mixture is shaken at 45~C for 4 hr. It is then drained and washed with 3 mL DMF 6X and 3 mL DCM 6X to provide the 10aded resin (~), which is dried under vacuo.
57.) Cleavin~ resin product The dried resin (~) is put in a 8 mL reaction vessel. 3 mL of 90% 1. illuu~ u..~li., acid/water is added into the reaction vessel. The resin is cleaved in TFA for 20 min at room t~ ,. alul ~ and then filtered into a 50 mL collection tube. All filtrate is altd to dryness under vacuo to give (1) (CH:IR 5585).
J~ O H~
HO ~
B.) Preparation of additionaH~
1.) C~R66g6 Compound CELR 6696 was prepared following steps I 4 of Part A above. The 15 loaded resin (g) was treated with phenylacetyl chloride in DCM/pyridine, then cleaved as set forth in step 7 to produce CHIR 6696 [~N~ IN~
~ CH
W0 96~40747 ~ 2 1 ~ 6 3 3 ~
2.) CHIR 10~:
C~IIR 10382 was prepared following part A above, ~ut . ' ~ ' ,' forN-~4,4'-' ' ~ ' instep2. 'rheresultingcompound vas clesved from the resin and capped with 4,4'~ hJ~I alcohol in 10%
H2SO~ldioxane to pro~ide CHIR 10382:
H O C~
Nl ~
3.) CHIR6?14: ;
CHIR 6714 ~ p~ cil in the same manner as ~HIR 10382 above~ but ammonia for N .. lllylKlyH ' ~A
: ~ OCH~
H O
~ NJI~N~
C.) Preparation of ~her compounds Similariy, proceetiing as~in part A.) above, the following ~u...~uu..d~ were made:
i~2 ~
wo g6/40747 2 1 9 6 3 3 4 PCT/USg610~648 ~ - 1 5 -CHIR# Rlo R2 R3 R, R, 5585 4 SJJ~u~yl ' ' ,1 5-indanyl H 414 -' Y~ JJ'Yl~ H
~ Syl ki~-5948 1 ' ' ~' ~ ' Jl ~-indanyl H 4,4'-!" ' yi~ J~yl- H
~,!~.~ ' ~ 5949 2,4,6-i h~ ' JA.~I' 5-indanyl H 4,4'-~' " yb~h~ ' yl- H
I,Lh~ yl Li~
5950 1 L~l~u~yi~ul~' - 5-indanyl H 4,4'-!'- " ylJw.L.,~ hyl- H
methyl 5951 iJW.L,~ ~ 'h~,'l 5-indanyl H 4,4-~' ' ylJw~hJJ~yl- H
5952 l~ul~ ' ' Jl 5-indanyl H 4,4'- i;u,~,,lluAyl,~,, LLyJlyl- H
~.ly ~ ., 5953 1 ' ~,' h~l 5-indanyl H 4,4'-~' " y~ Lh~Jlyl- H
" I ~
5954 4-hydlu~ylJi.c.. ~ yl- phenyl H 4,4'-dimethoxybenzhydryl- H
5955 1 LJI~U~ ' ',: 4-phenyl- H 4,4'-~" ' yb~.~h.~l~yl- H
' Jl phenyl "1~. ~ ~ ' 5956 1 i~Jlu~y~h ,l.,tllyl- phenethyl H 4,4'~i;u,~,,lluAyLI~ LllJJlyl- H
J ~,!~.- ' 5957 1 h~J~u~l' ' J cyclohexyl H 4,4'-~' ' y~ JJ~yl- H
5958 4-hydroxyphenethyl- 3-methyl- H 4,4'-d ' yb~,. LhJJ~yl- H
' jl enedioxy- ~ly~ ' phenyl 5959 1 h~J~u~yl' ''Jl 5-indanyl H 9-nuu~u.. J!~ ' ' H
- I
596û 1 h,.' u~ L~ Lllyl- 5-indanyl H 4-nl.~LlluAyl~ LylL'~. ' H
5961 1 hJI~u~yl ' ' J 5-indanyl H bWILL~1dIYILI.~I ' '- EI
5991 4-hydrûxyphenethyl- 5-indanyl Me 4,4'-dimethoxybenzhydryl- H
' Jl glycinamide 5992 1 h.~ ilU~.y, ' ' t~l 5-indanyl Me 4~4-J;I~I~1IIU~Yb~ JdIYI- H
il Ll ~ ~
599û 1-~1 h~ i~uA.~l ' 5-indanyl H 4,4'- i;ll ,Lhu~ylJ~,. Lh~dlyl- El ethylamino)ethyl ~lyl ~-5993 1-(1 hJ i~UA~' 5-indanyl H 4,4'- " ' yb~,.Lily i~yl- Me ethylamino)ethyl 5994 1 h~J~u7~y ' S; 5-indanyl Me 4,4' !' yl~.,. LLydryl- Me 5995 1-(4-hyJlu~yi' 5-indanyl H 44'-~' ' yi)~ J~yl- H
ethylamino)ethyl ~= ~
WO 96rr40747 ~ 2 1 9 6 3 3 ~ PCTIUS9~964X
~:
6696 benzyl ~ 5-indanyl H4,4' " ' ~ I,~I. ~,b yl- H
!r~
6697 phenethyl ~ 5-indanyl H4,4 ~ t ~ yl,~,~,JJ~yl H
"
6698 phenyl :~ S-ind nyl H4,4~ J,,,,,~ J . yl_ H
11509 ~rl~r~J.. J' ~ hyl 2-naphthyl H 4747-dh~ rl,w~J~llyl- H
11648 4 h,d~ " ' ~ ',1 2-naphthyl H 4,4'~ b~ JIlyi- H
Y
11649 q ~ yl 2-naphthyl H 4,4-,i ~ ~rt i yl- H
11650 2-ru,~ r ' _ 2-na~hthyl H 4~4'~ h.,d.yl- H
methyl ~ ~lY '' 11652 ~rlu~rJ.. ~ 1 3~4-di- H 4,4'-:'' ' yl~,.,Lh~d~yl- H
~1 nnethyl~ r phenyrl s 11653 1,~ ; ' J 2-naphthyl H 4,4'-d;.. ~lh~ d~yrl- H
~ ~ly.: ~ ' (Assay for uPA inhibitory Activity~t C~ , ' prepared as described in ExaTnple I above were screened in a human urokinase receptor radioligand ~ ., assay~ as described in Goodson et al., Proc Natl Ar~l Sci USA (1994) 91 :7129 (ill~,ul!~natcli herein by reference), except that the b~eled ligand used was an epitope ta~;ged version of the EGF-like domain of human urokinase, expressed and purified from ,~ ' ' yeast, ~The:activities observed are set forth in the Table below:
CEllR# ~IC~ M) % Inhibition C~ d~iOII tl~) 11509 ~'~0.033 ~ :
~.
11649 ~ 80 ~ 1 11650 80 ~ 1 11651 ~ 40 1165~ ~ 30 ~ I
116~3 =~ 85 :~ :1 10382 ~ 0.5 WO 9~40747 2 1 9 6 3 3 4 F~~ ,61_ 6714 10.0 5440 22.5 10 5585 0.2 5948 1.38 5949 2.6 5950 0.76 5 1 .4 5951 0.77 43.5 0.2 5952 0.7 32.8 0.2 5953 0.44 72.5 5954 1.8 5955 1.9 36.6 0.2 5956 ~10 24.1 2.5 5957 >10 19.3 0.2 5958 3.7 32.3 0.2 5959 48.7 10 5960 27.2 0.2 5961 0.93 5990 0.105 5991 31.8 0.2 5992 10.9 0.1 5993 0.12 27.2 0.1 5994 4.2 0.1 5995 20.5 0.1 6696 1.65 43.2 1.0 6697 0.2 80 1.0 6698 63.6 10
57.) Cleavin~ resin product The dried resin (~) is put in a 8 mL reaction vessel. 3 mL of 90% 1. illuu~ u..~li., acid/water is added into the reaction vessel. The resin is cleaved in TFA for 20 min at room t~ ,. alul ~ and then filtered into a 50 mL collection tube. All filtrate is altd to dryness under vacuo to give (1) (CH:IR 5585).
J~ O H~
HO ~
B.) Preparation of additionaH~
1.) C~R66g6 Compound CELR 6696 was prepared following steps I 4 of Part A above. The 15 loaded resin (g) was treated with phenylacetyl chloride in DCM/pyridine, then cleaved as set forth in step 7 to produce CHIR 6696 [~N~ IN~
~ CH
W0 96~40747 ~ 2 1 ~ 6 3 3 ~
2.) CHIR 10~:
C~IIR 10382 was prepared following part A above, ~ut . ' ~ ' ,' forN-~4,4'-' ' ~ ' instep2. 'rheresultingcompound vas clesved from the resin and capped with 4,4'~ hJ~I alcohol in 10%
H2SO~ldioxane to pro~ide CHIR 10382:
H O C~
Nl ~
3.) CHIR6?14: ;
CHIR 6714 ~ p~ cil in the same manner as ~HIR 10382 above~ but ammonia for N .. lllylKlyH ' ~A
: ~ OCH~
H O
~ NJI~N~
C.) Preparation of ~her compounds Similariy, proceetiing as~in part A.) above, the following ~u...~uu..d~ were made:
i~2 ~
wo g6/40747 2 1 9 6 3 3 4 PCT/USg610~648 ~ - 1 5 -CHIR# Rlo R2 R3 R, R, 5585 4 SJJ~u~yl ' ' ,1 5-indanyl H 414 -' Y~ JJ'Yl~ H
~ Syl ki~-5948 1 ' ' ~' ~ ' Jl ~-indanyl H 4,4'-!" ' yi~ J~yl- H
~,!~.~ ' ~ 5949 2,4,6-i h~ ' JA.~I' 5-indanyl H 4,4'-~' " yb~h~ ' yl- H
I,Lh~ yl Li~
5950 1 L~l~u~yi~ul~' - 5-indanyl H 4,4'-!'- " ylJw.L.,~ hyl- H
methyl 5951 iJW.L,~ ~ 'h~,'l 5-indanyl H 4,4-~' ' ylJw~hJJ~yl- H
5952 l~ul~ ' ' Jl 5-indanyl H 4,4'- i;u,~,,lluAyl,~,, LLyJlyl- H
~.ly ~ ., 5953 1 ' ~,' h~l 5-indanyl H 4,4'-~' " y~ Lh~Jlyl- H
" I ~
5954 4-hydlu~ylJi.c.. ~ yl- phenyl H 4,4'-dimethoxybenzhydryl- H
5955 1 LJI~U~ ' ',: 4-phenyl- H 4,4'-~" ' yb~.~h.~l~yl- H
' Jl phenyl "1~. ~ ~ ' 5956 1 i~Jlu~y~h ,l.,tllyl- phenethyl H 4,4'~i;u,~,,lluAyLI~ LllJJlyl- H
J ~,!~.- ' 5957 1 h~J~u~l' ' J cyclohexyl H 4,4'-~' ' y~ JJ~yl- H
5958 4-hydroxyphenethyl- 3-methyl- H 4,4'-d ' yb~,. LhJJ~yl- H
' jl enedioxy- ~ly~ ' phenyl 5959 1 h~J~u~yl' ''Jl 5-indanyl H 9-nuu~u.. J!~ ' ' H
- I
596û 1 h,.' u~ L~ Lllyl- 5-indanyl H 4-nl.~LlluAyl~ LylL'~. ' H
5961 1 hJI~u~yl ' ' J 5-indanyl H bWILL~1dIYILI.~I ' '- EI
5991 4-hydrûxyphenethyl- 5-indanyl Me 4,4'-dimethoxybenzhydryl- H
' Jl glycinamide 5992 1 h.~ ilU~.y, ' ' t~l 5-indanyl Me 4~4-J;I~I~1IIU~Yb~ JdIYI- H
il Ll ~ ~
599û 1-~1 h~ i~uA.~l ' 5-indanyl H 4,4'- i;ll ,Lhu~ylJ~,. Lh~dlyl- El ethylamino)ethyl ~lyl ~-5993 1-(1 hJ i~UA~' 5-indanyl H 4,4'- " ' yb~,.Lily i~yl- Me ethylamino)ethyl 5994 1 h~J~u7~y ' S; 5-indanyl Me 4,4' !' yl~.,. LLydryl- Me 5995 1-(4-hyJlu~yi' 5-indanyl H 44'-~' ' yi)~ J~yl- H
ethylamino)ethyl ~= ~
WO 96rr40747 ~ 2 1 9 6 3 3 ~ PCTIUS9~964X
~:
6696 benzyl ~ 5-indanyl H4,4' " ' ~ I,~I. ~,b yl- H
!r~
6697 phenethyl ~ 5-indanyl H4,4 ~ t ~ yl,~,~,JJ~yl H
"
6698 phenyl :~ S-ind nyl H4,4~ J,,,,,~ J . yl_ H
11509 ~rl~r~J.. J' ~ hyl 2-naphthyl H 4747-dh~ rl,w~J~llyl- H
11648 4 h,d~ " ' ~ ',1 2-naphthyl H 4,4'~ b~ JIlyi- H
Y
11649 q ~ yl 2-naphthyl H 4,4-,i ~ ~rt i yl- H
11650 2-ru,~ r ' _ 2-na~hthyl H 4~4'~ h.,d.yl- H
methyl ~ ~lY '' 11652 ~rlu~rJ.. ~ 1 3~4-di- H 4,4'-:'' ' yl~,.,Lh~d~yl- H
~1 nnethyl~ r phenyrl s 11653 1,~ ; ' J 2-naphthyl H 4,4'-d;.. ~lh~ d~yrl- H
~ ~ly.: ~ ' (Assay for uPA inhibitory Activity~t C~ , ' prepared as described in ExaTnple I above were screened in a human urokinase receptor radioligand ~ ., assay~ as described in Goodson et al., Proc Natl Ar~l Sci USA (1994) 91 :7129 (ill~,ul!~natcli herein by reference), except that the b~eled ligand used was an epitope ta~;ged version of the EGF-like domain of human urokinase, expressed and purified from ,~ ' ' yeast, ~The:activities observed are set forth in the Table below:
CEllR# ~IC~ M) % Inhibition C~ d~iOII tl~) 11509 ~'~0.033 ~ :
~.
11649 ~ 80 ~ 1 11650 80 ~ 1 11651 ~ 40 1165~ ~ 30 ~ I
116~3 =~ 85 :~ :1 10382 ~ 0.5 WO 9~40747 2 1 9 6 3 3 4 F~~ ,61_ 6714 10.0 5440 22.5 10 5585 0.2 5948 1.38 5949 2.6 5950 0.76 5 1 .4 5951 0.77 43.5 0.2 5952 0.7 32.8 0.2 5953 0.44 72.5 5954 1.8 5955 1.9 36.6 0.2 5956 ~10 24.1 2.5 5957 >10 19.3 0.2 5958 3.7 32.3 0.2 5959 48.7 10 5960 27.2 0.2 5961 0.93 5990 0.105 5991 31.8 0.2 5992 10.9 0.1 5993 0.12 27.2 0.1 5994 4.2 0.1 5995 20.5 0.1 6696 1.65 43.2 1.0 6697 0.2 80 1.0 6698 63.6 10
Claims (20)
1. A compound of tbe formula:
, where R10 is or a capping group, where X is NR12, CR12R15, O, S, SR12, or SR12R15, R1, R9, R11, R12 R15 are each independently H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-alkynyl, aryl-cycloalkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio. CN or NO2;
R16 is , H, lower alkyl, cycloalkyl, or lower alkenyl;
R2 is aryl or aralkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, R3 and R5 are each independently H or lower alkyl;
R4 is , where R13 is H, lower alkyl, phenyl or benzyl, and R14 is H, aryl, aralkyl, or , where R6 and R7 are each independently H, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, and n and m are each independently an integer from 1 to 3 inclusive;
and pharmaceutically acceptable acid addition salts thereof.
, where R10 is or a capping group, where X is NR12, CR12R15, O, S, SR12, or SR12R15, R1, R9, R11, R12 R15 are each independently H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-alkynyl, aryl-cycloalkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio. CN or NO2;
R16 is , H, lower alkyl, cycloalkyl, or lower alkenyl;
R2 is aryl or aralkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, R3 and R5 are each independently H or lower alkyl;
R4 is , where R13 is H, lower alkyl, phenyl or benzyl, and R14 is H, aryl, aralkyl, or , where R6 and R7 are each independently H, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, and n and m are each independently an integer from 1 to 3 inclusive;
and pharmaceutically acceptable acid addition salts thereof.
2. The compound of claim 1, wherein R4 is , and n and m are each 1, and R6 and R7 are each 4-methoxy, and R16 is , where R5 is H,
3. The compound of claim 2, wherein R2 is selected from the group consisting of 2-naphthyl and 5-indanyl.
4. The compound of claim 3, wherein R10 is , where R9, R11 and R12 are each H, and R1 is selected from the group consisting of , , and ~C~CH, where p is an integer fi om 0 to 6, q is an integer from 0 to 3, and R8 is OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl.
5. The compound of claim 4, wherein R3 is H.
6. The compound of claim 5, wherein R, is -C~H, and R2 is 2-naphthyl.
7. The compound of claim 5, wherein R1 is , q is 0, and R2 is 2-naphthyl.
8. The compound of claim 5, wherein R1 is , where p is 1, q is 1, R2 is 2-naphthyl, and R8 is OH.
9. The compound of claim 5, wherein R1 is where p is 1, q is 1, R2 is 2-naphthyl, and R8 is NH2.
10. The compound of claim 5, wherein R1 is where p is 0, q is 0, and R2 is 2-naphthyl.
11. The compound of claim 5, wherein R1 is where p is 1, q is 1, R2 is 5-indanyl, and R8 is OH.
12. The compound of claim 1, wherein R4 is , R3 and R5 are each H, R1 is , where p is 1, q is 1, R2 is 5-indanyl, and R8 is OH.
13. The compound of claim 1, wherein R4 is , and n and m are each 1, and R6 and R7 are each 4-methoxy, and R16 is H or lower alkyl.
14. The compound of claim 13, wherein R10 is benzyl, phenyl, phenethyl, 4-hydroxybenzyl, or 4-hydroxyphenethyl.
15. The compound of claim 14, wherein R3 is H, and R2 is 5-indanyl.
16. The compound of claim 15, wherein R10 is phenethyl.
17. The compound of claim 16, wherein R16 is methyl.
18. A composition comprising:
a compound of the formula where R10 is or a capping group, where X is NR12, CR12R15, O, S, SR12, or SR12R15; R1, R9, R11, R12, R15 are each independently H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-alkynyl, aryl-cycloalkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, CN or NO2;
R16 is , lower alkyl, cycloalkyl, or lower alkenyl;
R2 is aryl or aralkyl, unsubstituted or substituted with 1-3 halo, OH, NH12, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl;
R3 and R5 are each independently H or lower alkyl;
R4 is , where R13 is H, lower alkyl, phenyl or benzyl, and R14 is H, aryl, aralkyl, or , where R6 and R7 are each independently H, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, and n and m are each independently an integer from 1 to 3 inclusive, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.
a compound of the formula where R10 is or a capping group, where X is NR12, CR12R15, O, S, SR12, or SR12R15; R1, R9, R11, R12, R15 are each independently H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-alkynyl, aryl-cycloalkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, CN or NO2;
R16 is , lower alkyl, cycloalkyl, or lower alkenyl;
R2 is aryl or aralkyl, unsubstituted or substituted with 1-3 halo, OH, NH12, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl;
R3 and R5 are each independently H or lower alkyl;
R4 is , where R13 is H, lower alkyl, phenyl or benzyl, and R14 is H, aryl, aralkyl, or , where R6 and R7 are each independently H, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, and n and m are each independently an integer from 1 to 3 inclusive, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.
19. A method for treating a uPA- or uPAR-mediated disorder, comprising:
administering to a subject having a uPA- or uPAR-mediated disorder an effective amount of a compound of the formula , where R10 is or a capping group, where X is NR12, CR12R15, O, S, SR12, or SR12R15; R1, R9, R11, R12 R15 are each independently H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-alkynyl, aryl-cycloalkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, CN or NO2;
R16 is , H, lower alkyl, cycloalkyl, or lower alkenyl;
R2 is aryl or aralkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl;
R3 and R5 are each independently H or lower alkyl;
R4 is , where R13 is H, lower alkyl, phenyl or benzyl, and R14 is H, aryl, aralkyl, or , where R6 and R7 are each independently H, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, and n and m are each independently an integer from 1 to 3 inclusive, or a pharmaceutically acceptable acid addition salt thereof.
administering to a subject having a uPA- or uPAR-mediated disorder an effective amount of a compound of the formula , where R10 is or a capping group, where X is NR12, CR12R15, O, S, SR12, or SR12R15; R1, R9, R11, R12 R15 are each independently H, lower alkyl, lower alkenyl, lower alkynyl, aryl, aralkyl, aryl-alkenyl, aryl-alkynyl, aryl-cycloalkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, CN or NO2;
R16 is , H, lower alkyl, cycloalkyl, or lower alkenyl;
R2 is aryl or aralkyl, unsubstituted or substituted with 1-3 halo, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl;
R3 and R5 are each independently H or lower alkyl;
R4 is , where R13 is H, lower alkyl, phenyl or benzyl, and R14 is H, aryl, aralkyl, or , where R6 and R7 are each independently H, OH, NH2, CN, NO2, lower alkyl, halo-lower alkyl, lower alkoxy, lower alkylamino, lower alkylthio, or cycloalkyl, and n and m are each independently an integer from 1 to 3 inclusive, or a pharmaceutically acceptable acid addition salt thereof.
20. The method of claim 19, wherein said uPA- or uPAR-mediated disorder is tumor metastasis, tumor angiogenesis, restenosis, chronic inflammation, or corneal angiogenesis.
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US08/485,021 US5747458A (en) | 1995-06-07 | 1995-06-07 | Urokinase receptor ligands |
US08/485,021 | 1995-06-07 |
Publications (1)
Publication Number | Publication Date |
---|---|
CA2196334A1 true CA2196334A1 (en) | 1996-12-19 |
Family
ID=23926620
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CA002196334A Abandoned CA2196334A1 (en) | 1995-06-07 | 1996-06-06 | Urokinase receptor ligands |
Country Status (14)
Country | Link |
---|---|
US (2) | US5747458A (en) |
EP (1) | EP0777680B1 (en) |
JP (1) | JPH10503784A (en) |
AT (1) | ATE214711T1 (en) |
AU (1) | AU6164096A (en) |
BR (1) | BR9606429A (en) |
CA (1) | CA2196334A1 (en) |
DE (1) | DE69619940T2 (en) |
FI (1) | FI970446A (en) |
HU (1) | HUP9702220A3 (en) |
IL (1) | IL120101A0 (en) |
MX (1) | MX9700888A (en) |
NO (1) | NO970513D0 (en) |
WO (1) | WO1996040747A1 (en) |
Families Citing this family (10)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5519120A (en) * | 1989-04-07 | 1996-05-21 | Cancerforskningsfondet Af 1989 | Urokinase-type plasminogen activator receptor antibodies |
TR200000272T2 (en) * | 1997-07-31 | 2000-09-21 | F. Hoffmann -La Roche Ag | O-substituted hydrocycumaranone derivatives as antitumor and antimetastatic agents. |
AU3459999A (en) * | 1998-04-27 | 1999-11-16 | Warner-Lambert Company | Substituted diarylalkyl amides as calcium channel antagonists |
US6228985B1 (en) | 1998-05-21 | 2001-05-08 | Schering Corporation | Derivatives of aminobenzoic and aminobiphenylcarboxylic acids useful as anti-cancer agents |
ES2169690B1 (en) * | 2000-10-06 | 2004-03-16 | Diverdrugs Sl | QUARTERS OF N-ALQUILGLICINA CAPABLE OF PROTECTING NEURONS AGAINST EXCITOTOX AGGRESSIONS, AND COMPOSITIONS CONTAINING THEM. |
ES2169691B1 (en) * | 2000-10-11 | 2004-03-16 | Diverdrugs Sl | N-ALQUILGLYCIN QUARTERS ABLE TO BLOCK THE RESPONSE TO CHEMICAL SUBSTANCES, THERMAL STIMULES OR MEDIATORS OF THE INFLAMMATION OF NEURONAL RECEPTORS, AND COMPOSITIONS CONTAINING THEM. |
PL377856A1 (en) * | 2003-04-15 | 2006-02-20 | Merck Patent Gmbh | Identification of n-alkylglycine trimers for induction of apoptosis |
CN103270022B (en) | 2010-08-31 | 2015-08-19 | 巴勃罗·比咯斯拉达 | Neurotrophin receptor stimulant and the application as medicine thereof |
EP2647622B1 (en) * | 2010-12-03 | 2017-04-19 | Fujitsu Limited | Kinesin spindle protein inhibitors and application thereof |
IT202100023357A1 (en) | 2021-09-09 | 2023-03-09 | Cheirontech S R L | Peptides with anti-angiogenic activity |
Family Cites Families (7)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EP0138854B1 (en) * | 1983-03-08 | 1992-11-04 | Chiron Mimotopes Pty. Ltd. | Antigenically active amino acid sequences |
ATE81722T1 (en) * | 1984-07-24 | 1992-11-15 | Coselco Mimotopes Pty Ltd | PROCEDURE FOR DETERMINING MIMOTOPS. |
NZ215865A (en) * | 1985-04-22 | 1988-10-28 | Commw Serum Lab Commission | Method of determining the active site of a receptor-binding analogue |
IE66205B1 (en) * | 1990-06-14 | 1995-12-13 | Paul A Bartlett | Polypeptide analogs |
NZ256952A (en) * | 1992-09-24 | 1997-07-27 | Chiron Corp | Synthesis of n-substituted polyamides and pharmaceutical compositions |
US5536853A (en) * | 1994-04-11 | 1996-07-16 | Chiron Corporation | Opiate receptor ligands |
DE69532754T2 (en) * | 1994-04-26 | 2005-03-10 | Aventis Pharmaceuticals, Inc. | FACTOR XA INHIBITORS |
-
1995
- 1995-06-07 US US08/485,021 patent/US5747458A/en not_active Expired - Fee Related
-
1996
- 1996-06-06 DE DE69619940T patent/DE69619940T2/en not_active Expired - Fee Related
- 1996-06-06 AT AT96919248T patent/ATE214711T1/en not_active IP Right Cessation
- 1996-06-06 HU HU9702220A patent/HUP9702220A3/en unknown
- 1996-06-06 IL IL12010196A patent/IL120101A0/en unknown
- 1996-06-06 MX MX9700888A patent/MX9700888A/en unknown
- 1996-06-06 WO PCT/US1996/009648 patent/WO1996040747A1/en active IP Right Grant
- 1996-06-06 EP EP96919248A patent/EP0777680B1/en not_active Expired - Lifetime
- 1996-06-06 JP JP9501923A patent/JPH10503784A/en active Pending
- 1996-06-06 AU AU61640/96A patent/AU6164096A/en not_active Abandoned
- 1996-06-06 CA CA002196334A patent/CA2196334A1/en not_active Abandoned
- 1996-06-06 BR BR9606429A patent/BR9606429A/en not_active Application Discontinuation
- 1996-06-07 US US08/765,275 patent/US6121240A/en not_active Expired - Fee Related
-
1997
- 1997-02-03 FI FI970446A patent/FI970446A/en unknown
- 1997-02-05 NO NO970513A patent/NO970513D0/en unknown
Also Published As
Publication number | Publication date |
---|---|
US5747458A (en) | 1998-05-05 |
WO1996040747A1 (en) | 1996-12-19 |
DE69619940T2 (en) | 2002-12-19 |
HUP9702220A2 (en) | 1998-05-28 |
EP0777680A1 (en) | 1997-06-11 |
BR9606429A (en) | 1997-09-02 |
NO970513L (en) | 1997-02-05 |
ATE214711T1 (en) | 2002-04-15 |
US6121240A (en) | 2000-09-19 |
FI970446A (en) | 1997-03-26 |
DE69619940D1 (en) | 2002-04-25 |
FI970446A0 (en) | 1997-02-03 |
IL120101A0 (en) | 1997-04-15 |
JPH10503784A (en) | 1998-04-07 |
EP0777680B1 (en) | 2002-03-20 |
NO970513D0 (en) | 1997-02-05 |
MX9700888A (en) | 1997-04-30 |
HUP9702220A3 (en) | 1998-06-29 |
AU6164096A (en) | 1996-12-30 |
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Legal Events
Date | Code | Title | Description |
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FZDE | Discontinued |