CA2116653A1 - Peptides inhibiting il-1 beta release - Google Patents

Abstract

Di-, tri- and tetrapeptides in which the last .alpha.-amino acid is based on aspartic acid and attached to a residue A5 which is H; CF3; -Z1-Z2-Y2 wherein each of Z1 and Z2 independently is a direct bond or an .alpha.-amino acid residue and Y2 is NH2, C1-4 alkylamino, di-(C1-4 alkyl)amino or a heterocyclic radical attached by a nitrogen to Z2; -CH2-X1-Y3 wherein X1 is O or S and Y3 is heteroaryl; -CH2-Y3; substituted phenyl; ring substituted phenoxymethylene or phenylthiomethylene;
ring substituted pyridyloxymethylene; or a radical -CH2-X1-CO-Y4 wherein X1 is O or S and Y4 is trialkylmethyl or substituted phenyl or pyridyl, in free form or in salt form, have pharmacological activity, e.g. IL-1.beta. release inhibiting properties.

Description

W 0 93/O~t3~ 2 11 6 6 5 3 PCT/EP92/02~72 PEPTIDES INHIB~TING IL-1 8ETA RELEASE

The present invention relates to peptides having pharmaceutical - -~
utility, processes for their production, pharmac~utical compo-sitions comprising them and their use as pharmaceuticals. ~:
' ':, '.
More particularly the present invention provides:a compound of `~
formula I ~ .

R-[Al-A2ln-A3-A4-x-As (I) . .~ , .
-:
wherein : -~, R is hydrogen, an amino protecting group or optlonally ring substituted benzyloxy : ~:~
A1 is Val, Leu, Ala, Ile or trimethylsilyl-Ala : A2 is Phe or Tyr, n is 0 or 1, ~
:~ A3 is a direct bond, Val, Leu, Ala, Ile, trimethylsilyl-Ala or a :
divalent radical of formula (a) : ~ :

wherein ring A is optionally substituted by hydroxy or Cl - 4 alkoxy, ,' . '' W O 93!0913~ ~ l l 6 6 5 3 PCT/EP92/02472 A4 is a direct bond or a divalent radical of formula (b~
-N - CH-C0- .
( b Rl Y~

wherein -Rl is hydrogen or Cl_4alkyl, and Yl is the residue attaching to the ~-carbon atom of .`~-an a-aminO acid and optionally protected, :.:
-C~2-C~2-N(Cl 4alkyl)2, imidazol-2-yl-methyl, benzimidazol-2-yl-methyl, lH-1,2,4-triazol-3-yl~
methyl, pyrazol-3-y~l-methyI, indazol-3-yl-methyl or a radical:of formula ~c~ or ~d) `
R

~ ~R2 ~ -H~

(c)~ (d) : vherein each of R2~and R3,~independently, is~hydrogen,~
halogen:,.C1_~alky~1, C~3 or~trityl,~at mos~t one~
:~ of R2 ~and R3~belng~ ;and : : : each of R4 and R~s~:ind~pendently is~hydrogen,:
Cl_~alkyl,~hydroxy, Gl_4alkoxy,~CF3~,~phenyl or~
halogen, at most one ~f R4 and:R5~being :or A3 and A4 form together a radical of ~ormula:;(aa)~

~ -NH-en:- co-~- cn-co- ~ (aa Rl a Rl Y~
wherein Yl:~is~as defined above~and R1 and Rl,~form togethmr ; ~ -(C~z)~- wherein m is 2, 3, 4 or 5? and :.~
, : : ~ :.

....
; ~:, _ 3 ~ 100-7~21 ~,~

~ ., ., ~
1) X is a divalent radical of formula (e~

o O .-,.
~ ..

wherein R6 is ~ or Cl_4alkyl, and As is hydrogen; CP'3; a radi al -C~2-Xl-~3~herein Xl is 0 or S and~Y3 is heteroaryl;~ a r dical -C~2-Y3; or a ~:
radical of formulae (k): to (m) ' ~

-R : ~ `
R~o ~
-CE12-Xl-CO-Y4 ~ -C~-X~

wherein R7 .~.

Y4 iS ~rl-(C~ ~lkyl~_thyl or a residu-ring B is~ pyridyl, ; ~ R5 ring~C :is ;phenyl or pyridyl, each of R7 and R8 independently is Cl 4alkyl, ~Cl 4alkox~

~3U~'1T"~T7~ ~~
`~

CF3, halogen, nitro or cyano, and each of R9, Rl~ and Rll independently is nitro, cyano, CF3, carbamoyl, CO2Rl2, -CH=C~-CN or -C~C~CO2Rl2 wherein ;~
Rl 2 iS cl 6 alkY~

X being also a divalent radical of formula (e2) R
J 6 /OCl ~al'~Yl - N ~ (e ~ OCl 4~lkYl C02Cl 4dlkY~

when A5 is ~, or 2) X is a divalent radical o~ formula (f), .
R

C02H ~ ; ~

~ ` , and As is -Zl-Z2-Y2 wher~in each of Zl and Z2 independently : ~ ;
: ls a direct bond or an o,amino acid residu~ and Yz is NH2, Cl 4alkylamino, di-~Cl 4alkyl)amino or:a heterocy ~.
clic radical ac;tached by a nitrogen to~Z2;~or a r~adical of formulae (k~ to ~o) as defined above; or ORl3; or 14 15 erein Rl3 is Cl_l2alkyl optionally substituted by ~ or interrupted by O and each of Rl4 and Rls is~
independently hydrogen, Cl l2alkyI, C5 7cycloal~yl or benzyl, or - :: .

3) X is a divalent rad~cal of ormula (g) -R,. ;
I ~ ~-.`.`.
N ~ N~ (g) ' C02,'1 ~3U~T-~ T

W 0 93/0913~ 211 S 6 5 3 PCT/EP92/02472 and As is -Zl-Z2-Y2 as defined above, or 4) X is a divalent radical of formula (h) or (j) :'.' 'l6 ~ l6 0 ''`
- N ~ - N ~ C~ ~ C~

~02H CO~H ':~
( h ~

and A5 is a radical of formulae (k) to ~o). -C~2-Y3 or -CH2 -Xl -Y3 as defined above, , with th~ provisos tha~
only one of A3 and A4 can be a direct bond when n is 0, and each of ~3 and A4 ls other than a diFect bond when n is l. -and the physiologically-hydrolysable and -acceptable es~ers or ~ ; . ', amides thereof, in free form, in salt form or in rhe form of complexes.

Examples of protecting groups as R are e.g.:disclosed~i:n "Protec~'r~:
tive Groups in Orga~ic Synthesis"~T.;U.~Greene, Jb~iley~& Sons : '.
NY (19~1), 219-287, for:example acyl such as ace~tyl, methoxy~
succinyl, hydroxysuccinyl~or benzoyl optionall~substituted on ~ '', the phenyl ring with e.g~ p-methoxycarbonyl, p-meth~xy~or~ n,,~, p-'nitro; alkoxycarbonyl such as t-butyloxycarbonyl; arylme~ho- . .' xycarbonyl"such~as~9-1uorenylmet,hoxycarbonyl or ~enzyloxy;carbo-nyl opeionally substituted on the~ph~*yl~r~pg,wi;,th:p-methoxy, p-nitro, p-chloro or~m-phenyl; arylmethyl su~h as benzyl: aptio~
nally ring substituted with p-methoxy, p-nitro cr p-chloro;~or~
ary}sulfonyl such a~ phen~lmulfo~yl optionally ring substituted ,~ ., W O 93/0913~ 2 116 6 5 3 PC~/EP92/0247~

with p-methyl or p-methoxy, or naphthylsulfonyl optionally ring substituted with e.g. amino or di(Cl~alkyl)amino.

When R is ring substituted benzyioxy, it is preferably benzyloxy substituted with hydroxy or Cl_4alkoxy. Pr.eferably R is :
unsubstituted benzyloxy.

Halogen is preferably fluorine or chlorine. :~

:;
~hen A3 is a substituted radical of formula (a) it is pre~erably 1~ :
substituted by C1_~alkoxy, preferably in para to -C~0.

By -amino acid is meant a naturally occurring or eommercially available or non natural ~-amino acid or an optica~ isomer ~:
thereof. A non natural -amino acid is an a-amino acid which is not i.ncorporated into a protein under mRNA directi~n~ e.g. ~-Nal, a fluoro-a-amino acid~such as 1uoroalanine, trimethylsilyl-Ala ;~
or an a-amino acid such as : :

I`H ~ ;
H~ CH?.)n~
~ ;` I , : ; .. , :: ~ : : t~2N--CH-C~
: ,.:, wherein nl is an integer from 1 to 6 and n2 is an integer from~1 to 12.
. ' Protecting groups which may~be present in Yl are groups which :~
pro~ect the 0, S or N functionality in the ide chain:amino ~::
groups of an -acino a id. N-pro~ecting groups are e~.g. as ~`
disclosed above for R, or C3_salkyl such as isopropyl, fo~myl, a sugar residue such as 1-deoxy-fructosyl or ~-glucosyl(1-4)-~;"''"~
`~'' .

..

~ . . .

W O 93J0913~ 211~ 6 5 3 PCT/EP92/0247~

-'':'.:
deoxyfructosyl, dihydroxy-C3_6alkyl such as dihydroxypropyl, ~.
C5_7cycloalkyl such as cyclohexyl or tropinyl. O- and S-protecting groups for hydroxy and thiol functionalities are known and may be e.g. methyl, t;-butyl or benzyl. ~ :~

When Y2 is a heterocyclic radical, it may be e.g. a 5 or 6 ~
membered ring, e.g. piperidino or pyrrolidinyl. ;

Examples of heteroaryl as Y3 include e.g. 5-, 6- or 7-membered unsaturated h~terocyclic radicals, comprising at least one :~
nierogen and op~ionally further heteroatom9 ~uch~as N, O or S.
Preferably Y3 iS heteroaryl comprising from l to 4 nitrogen atoms, e.g. pyridyl, triazolyl, tetrazolyl, triazin-dionyl. ~:
. ~, ...
In ring B of radical (o), the nitrogen atom may be in o-, m- or .
para. When ring C in r~dical (k) is pyridyl, it may be 3-, 4- or 5-pyridyl.

Radicals (el), (e2), (:f), (g) and (j) are d~rived from Asp and comprise one asymetric:carbon atom and radical ~(h3~comprlses two asymetric arbon a~oms and accordingly:they lead to optical ;~
isomerism. It will be unders~ood~that ~the present inven~tion includes all individual isomeric forms;~and d1~astereoisomers as : ~.
well as mixtures5 e.g. racemates, unless otherwise stated. ~ ~ ~
,:
Radical of formula (el) attached to As which is hydrogen may ~ :
exist in both cyclic as well as in non-cyclic form e.g. as follows~
_ O OH ~ `
-NR6~11"H = ~\0 `-~

C02H ~1' :

...... ... .. ..

W O 9~/~913~ PCTIEP92/02472 .

It is to be understood that where tautomeric forms occur, the present invention embraces both lactol and oxo-carboxylic acid -~
forms, i.e. although compounds of formula I wherein X is a radical of formula (el) are defined for convenience by reference to the oxo-carboxylic acid form only, the invention is not to be understood as being in ~ny way limited by the particular ~::
nomenclature or graphic representation employed. Similar con-siderations apply in relation to starting materials exhibiting lactol/oxo-carboxylic acid tautQmerism as hereinafter described.

The same considerations also 2pply to radical of formul~ (h) which may exist in both linear and~cyclic form as follows:

' OH
NR6~ Ht/ O -NR6~<

C02H b~c/~rcr ~

O -~
: : , and to compou~ds of formula I comprisin~ a radical of formula (h)~ :.
and to the correepond~ng starting~materials.

~y the term ~Iphysio10gica1ly-hydrolysab1~ and -acceptab1e esters "
or amides" are me~nt est~rs and amides which~are hydrolysable ~ :
under physiological~conditions~to yield alcohols or amines:which :~
are themselves physiologically acceptabl~, i.e. ~which are~non-toxic at ~he desired dosage levels.

Such esters or amides are obtained by esterification or ~:
amidatio~9'respectively, of a compound of formu1a I wherein X is a radical bearing:a carboxy group.;Such esters include esters : :-with an aliphatic or alicyclic alcohol or polyol having l~to l2 carbo~ atoms. Such amides include amides with aliphatic amines, : ~ 1 e.g. C1_ 4 alkyl amiDe, cl _ 4 alkoxy-Cl_~alkyl amine such as ~

:

W O 93/0913~ 2 116 ~ 5 3 PCT/EP92/02~7~ ~

~-methoxy-ethyl amine, or aniline.

The compounds of formula I may exist e.g. in free form~ acid ad~
dition salt form or in the form of complexes thereof. Acid addit-ion salts may be formed with e.g. organic acids, polymeric acids ~-and inorganic acids. Such acid addition salt forms include e.g.
the hydrochlorides and acetates. Salt forms may also include those obtainable with the carboxylic group present in compounds of formula I, e.g. alkali metal salts such as sodium or potas-sium~ or substituted or unsubstituted ammonium salts. Complexes . . . ~
are e~g. formed from compounds of formula I on addition of inor- -ganic substances, e.g. inorganic salts or hydroxides such as Ca-and Zn-salts, and/or an addition of polymeric organic substances.

In the compounds of formula I, the follo~ing significances are preferred either individually or in any combination or sub- `-combination: `
~.:
1. R is an amino protecting group or benzyloxy, preferably an amino proteoting group. R is preferably benzyloxy when n is and A3 is a radical of formula ~a).

2. Each of A3 and A4 are other than a direct bond.
3. n is 0. ~ ~
4. A3 is a radical of formula (a) when n is O.

5. A3 is a direct bond, Val, Leu, Ala, Ile or tri~ethylslly-Ala. -~
.

6. Rl is hydrogen or methyl, preferably hydrogen.

7. Yl is the residue attaching to the a-carbon atom of an a-ami-no acid selected from Ala, Leu, ~is, Phe, Het9 Trp, trimethylsilyl-Ala and optionally side ehain protec~ed Arg~
Orn and Lys, or Yl is a radical of formula (c3.

W O 93/0913~ ~ 116 6 5 3 PCT/EP92/0247~

8. A3 and A~ ~ay also form together a radical of formula (aa) when n is 0.

9. m is 2 or 3.

10. X is a radical of formula (e1) and A5 is other than 11. X is a radical of formula (g)~

12. X is a radiral of formula (h) or (j).

13. R6 is hydrogen or methyl, preferably hydrogen.

14. Z~ in A5 is the residue of a natural ~-amino acid, preferably ..
of an aromatic/heterocycli~ -amino acid, par~icularly Pro.

15. Z2 in A5 is the residue~of a natural a-amino acid, preferably an aliphatic a-amino acid, part:icularly an al~pha~ic a-amin acid without further function~l group, mo~t preferab~y ~Jal.:
,~ ., 16. X is a radical of: formula (e1) optionally~esterified or :amidated and As is a radical of formula~(k),~ or (o).

17. X is a radical of formula ~el), (h) or (j) op~ionally este~
rified or amidated and A5 is -C~2-Xl-Y3, p:réferably -CH2-S-Y3, or -C~2-Y3.

18. X i5 a radical of formula (h) or (j) optionaIly esterified or amidated and A~ is (k).
'~' : :: ~ ' 19. X is a radic 1 of formula (j) optionally esteri~ied or amidated and A5 is a radical o~ formula (;m).
:'~, 20. Radical of formula (m~ is monosubstituted, pref~rably in ~116653 ~ ~

~ 100-7821 para, more preferably it is 4-nitrophenyl. ~:

21. All ~-amino acid residues present except X have the L
configuration. X has the D or L configuration.
~';'' 22. The asymetric carbon in (h) bearing the O~ group has the :.
configuration S~ Preferably the a~yme~rlc carbon in.(J) .:
bearing R6 ha~ th~ configuration R. ~'!`.
.' In a series of specific embodiments, the present invention also .,.
provides a compound of formula I wherein j n is 0, A3 is a direct bond, Yal, Leu, Ala, Ile or trime~ -thylsilyl-Ala, A4 is as defined above, or A3 and A4 form together ~.
a radical of ~ormula (aa) as defined above, and ;~
i) X is a radical of formula (e1) or (e2) as defined above and A~ ic ~, or : .
ii) ~ is a radical of formula ~el~ or (~) and A5 is a radical of ;~
formula (k~, (1) or (m) wher~in ~1 is 0, ;~
iii)X is a radical of formula (g~ and A5 is -Zl-Z2-Y2, or ~`
iv) X is a radical of formula (j) and ~5 iS a radical of~formuIa (k), (l~ or (m) wùerein Xl is 0. ~ ~ ~

The present invention als~ provides a proce~s ~or the production :~-of a compound of formula I, whi~h process comprises:

a) removing at least one protecting group from a compound of ~:~
formula I in protected form or adding a protecting group R at the N-ter~inal group of coapound of formula I;~or b3 converting one co~pound of formula I into another compound of formula I; or : ~.
~, c3 coupling together by an amide bond two peptide fragm~nts, :~

~ U ~ ~ -,7 W O 9~/~913~ 211~ 6 5 3 PCT/EP92/0247~t each of which contains at least one amino acid in protected or unprotected form and one peptide fragment containing a `
radical of formula (el~ to ~;) as defined above, the peptide fragments being such that a protect~d or unprotected peptide :-having the se~uence according to formula I above is obtained -~.
and, if necessary, removing the protecting group or groups ~ :
from a compound of formuIa I in protected form; or : ~
.,.
d) for ~he production of a compound of ormula I wherein~X is a radical of formula (e1) or (h) and As is a radical of formula (~), (l) or (o) or -CH2-Xl-Y3, reacting a compount:of formula ~-III . : ' : `
,'' '~ ' R~[Al~A23n~A3~A~~X'~c~2-z~ (III) . .

wherein R, Al to A4 and n are as defined above, X' is a radical of formula (el) or (h), cmd Z?~ iS a leaving group, ~`
: :.
e.g. halogen, ..
' ' ,'.
with a corresponding phenoly~thiophenol or ~Xl-pyridine;or an acid of formula HXl-CO-Y~ or a func~ional derivat~ive thereof , ~
or: ~Xl-Y3; or :

e) for the production of a compound of formula I ;

R~ l Al -A2 ] n ~A3 ~~4 ~N~ A5 ~ :

C02Rl 6 wherein R, A1 to As and n re as defined~:above and Rl6 is a C1_l2 aliphatic or alicyclic residue, oxid~zing~a ~ompound of formula V : :

" t . ~ ;j ~J " ~, ," i i, ~ - ' t ~,.

"`; ~t ~ t ;1~ ,s,~

W O 93/0913~ 2 116 6 ~ 3 PC~/EP~2~0247~

H 0~
R- [ Al -A2 ] r~ -A3 -A4 -~--A5 ~ .

~
'~
wherein R, Al to A5 9 n and Rl 6 are as defined above, `
-..; ~,.
and recovering a compound of formula I thus obtained in~free or salt form or in the form of a complex.

Processes (a) to (e) above may be carried out iD~accordance with `~
standard techniques known in the art.
" .
The removal of a protecting group in proc~ss step (a) may also ;~
include the removal of R o~ the N-terminal group of a compound of formula I. For example, whe~ R is benzyloxy carbonyl,~this group .~"
may be removed by hydrogenation in the presence of a catalyst, e.~g~ Pd.

In accordance with process~step (b) for example~, fo~r the production of a co~pound of formula I wherein X comprises a carbo~y group, a comp4und of:~ormuIa I wherein~X comprises an esterified ~r amidated:carboxy group~may be hydrolysed.:Such hydrolysis~may be effected:by treatment with:an~appropriate alkali or by acid hydrolysis,~for example in:t~e~presence of`
trifluoroacetic acid. : :~
~ - .
~urtùermore, in accordànce with process~ step (b), for:the production oE a compound of formula I wherein X comprises an ' ;:~
:es~terifie~ or amidated~carbo~y group?~a compound~of formula I.
~herein X comprises a carboxy group or an~esterified c~boxy group may be ~trans) esterified or amidated. Such ester~ formation o~
amida~ion may be:carried out using any of the: t~chniques ~nown;in ~:
the ar~, ~or example convertin~ the carbo~y group in a functional ,; ' `,'''',:
~ ,:

W O 93/0913~ 2 1 1 6 ~ 5 3 PCT/EP92/02~7 - 14 - ~
'''.'"'''`' reactive group, e.g. a corresponding carbonyl halide or anhydri-de, or using a compound of formula I wherein X is a radical of formula ~h) in the lactone form, and reacting such group with the selected alcohol or amine. ;:
, ~
In accordance with a further embodiment of process step (b)~ for ~.
the production of a compound of formula I wherein X is a radlcal ;;~
of formula (g), a compound of fo~mula I wherein X is a radical of formula (el) or (e23 and A5 iS ~ may be reacted with a compound ~
of formula II ~ ~-~2N-N~-CO~A5 (II) ~`
: '. .
wherein A5 is as defined above. This process may be earried out in analogy to the known techniques used for the preparation of semi carbazones, ,; :' . .; -A compound of formula I wherein X is a radical of~formula~(e may also be con~erted in acco~dance with ~nown techniques into a compound of formula I uherein X is~a radical of formula (e2) and vice versa.

Process step ~c)~may be carried ou~t by~the techniques~known~in ~ ~ -the ar~ of peptide chemistry. By peptide fragment comprising a radical of formula;(el) to (j~ is also meant the radical itself ~ ;
bearing a protecting group on the -NR6- moiety and an appropriate ending, e.g. As or C~2-Z~, on the other end.
'' `
Process step ~d) may conveniently be effected using a Dess-Martin reagent,~e.g. in~the presence o~ a base or~a halogen-precipita~
ting silver salt, or according to the Swern oxidation procedures.

Where desired, in these reactions, protecting~groups may be~used ~;
for functional groups which do not participate in the reaction.

:.

~ ' W O 93/~13~ 2 1 1 6 16 5 3 PCT/EP92tO247~ ~

These may be e.g. amino protecting groups, carboxy protecting groups, acetal groups etc. When the desired reaction is complete, ?'~
the protecting groups may then be removed.

Each of the above processes may be carried oue using starting materials in the form of one or other of the individual optical isomers or in the form of mixtures [relating to the asymetric carbons present in radicals of formulae (el) to (j) as X or in such a radical precursor~. Conveniently the starting materials `
are used as S- or R-enantiomers to produce a compound of formula ;
I wherein the asymetric carbon in radicsls~of fsrmulaè ~(el) to (j) has the S or R configuration, respectively.

The starting materials used in process steps (d) or ~e) may be prepared in analogy with process step (c~.
Insofar as the production of the starting materials is not particularly described, the compounds are known or may be prepared analogously to methods kno~1 and practiced in the art.

The following examples are illustrative of the invention. All temperatures are in ~C. ~
~ ,:
The following abbreviations are used~

THF = ~etrahydrofuran TFA = ~rifluoroacetic acid MeOH = methanol EtOAc = ethyl acetate DCC = dicyclohexylcarbodiimide `~
HOBT _ hydroxybenzotriazole `~
Z = benzyloxycarbonyl ~;
r.t. = room temperature Fmoc = 4-fluorenylmethoxycarbonyl a - amorph W O 93/09l3~ PCT/EP92/02~7~

E~AMPL~ 1. Z-Yal=Met-Asp~0~2-~ :
;
Z-Val-Met-Aspartic aldehyde dimethyl acetal-~-tert.butyl ester ~:
(1.17 g) is taken up in CH2Cl2 (150 ml), TFA (lS ml) and water (1 ml) added and s~irred for 2,5 hrs at r.t. The solvent is evaporated, toluene added twice and evaporated again. The ~ :
crystalline residue is dissolved in water (60 ml) upon heating, decanted from some undissolved material and crystallized at 5 , affording the first crop of the product: m.p. li7 . All mother liquors are collected and chromatographed (SiO2, acetone~hexane/- -EtOAc 60/40/0.5) to give a second crop of product, crystallizing , ~ .-upon trituration with water. . :

E~AMPL~ 2: Z-Val-~et-Asp(OH)-L Semicarbæzone ~:

Compound of Example 1 (0.27 g, 0.55 mmol) in MeaH (2 ml) is . :~
combined with semicarbazide, ~Cl (0.6 ml of 1 molar solution), then 5 drops of pyridine are added. The product crystallizes :
after 10 min. at r.t.: m.p. 233 - 235 . . .

EXAHPL~ 3: Z-Val ~e~-~sp(0~ semicarbazonyl-Pro-Val-N ~
. -: ~:
Compound of Example 1 (0.48 g, 1 mmol) is dissolved~in a mixture :~
of MeO~ (2 ml) water (0.5 ml) and 3 ~rops of pyridine. `~.
N-(hydrazinocarbonyl)-Pro-Val-N(CH3)2 ~0.3 g, 1 mmol) is dissolved in MeO~ (1 ml), water (2 ml) and 3 drops of 2N ~Cl.
Both solutions are combined and war~ed ~or 2 min. at 40 - 50 .
The product crystallizes upon cooling: m.p. 118 - 120 . ~-,:

By repeating ~he procedure of;Example 1, using the corresponding starting materials, following compounds may be prepared~

~xample 4 Z-Val-Phe-Asp~0~3 ~.,.

W 0 93/0913~ 2 1 1 ~ 6 5 3 PC~/EP92/02472 ~ ~
- l 7- :

~xample 5 Z-Val-His-Asp(OH)-H

By repeating the procedure of Example 2 or 3 respectively, using `
the corresponding starting materials, following compounds of formula IA

Z-A3-A4~ IA) \ C02H

wherein A3, A4 and A5 are as defined below, may~be prepared.
E~xample A3 A~ : ~s ~t . PO C ~6 Val Phe H a ~:
7 Val His H 150 ~ `;
8 Val Phe Pro-Val-N(CH3)2 126-129 9 Val His Pro-Val-N(C~3)z Ala-Tyr-Val Phe ~l 205:

~X~PLE 11: (3S)-3 (Z-valyl-phenylalanyl)-ami~o-5-(2t6-dimethy ~ ~ benzoylo~y)-4-oxo ~t:ano~:c~acid (35,4RS)-3-(Z-Val-Bis~a~ino-4-hydroxy-5-(2,6 dimethylbenzoyloxy) pentanoic a~id tert.-butyl es;~er (0.6 mmol) is t~reated:with Dess-Hartin reagent (O~.77 mmol) ln~C~2Cl2 (4 ml) for 45:~mln. and ;
filtered. Aceton and 0:.5 N NaOH are then~added and~aceton~is: : ::
e~aporated. The residual~crysta~line~material:is~washed~with~H2~0, aci~dified with 10 Z tartaric acid and extracted~with~EtOAc. The~
combined EtOAc extracts: a~fords after evaporation~the~tert. butyl:
ester of the pro~uct as yellow crystals. These are dissolved in .~.:
CH2Cl2 (~ ml)r TFA (4 ml) is:added:and the mixture is sti;rred at :
r.t. for 15 min. The mixture~is evaporated to~dryness~and chroma~ographed (sio2 ~ aceton/hexane~EtOAc 60/40/l)~affording:the~
product as slightly colored crystals.

,,~,..
^ ~. :,.
"`,;~
.
, - . , : : ~

2ll6653 W O 93/0913~ PCT~EP92/0247~ .

.
E~AMPL~ 12: (Z-valyl-alanyl)-~3R,4S)-3-amino-4-hydroxy-5- ~
.
(2,6-dichlorbenzoyloxy) .:
(Z-valyl-alanyl)-3R 9 4S)-3-amino-4,5-dihydroxy pentanoic acid ethyl ester and 4-dimethylaminopyridine are dissolved in pyridine and 2,6-dichlorobenzoyl chloride is added dropwise. The reaction mixture is stirred overnight at room temperature, then ice and ~ater are added and the mixture is extracted with AcOEt. The organic layer is ~ashed with water, then with NaGl solution, dried over Na2SO4, ~iltered, evaporated, and ~he crude product is :-chromatographed to give the title compound.

The (3R,4RS~ deri~ative of the title compound may be prepared as follows:

Z-Val-Ala-OH (0.26 mmol), DCC (53 mg, 0.26 mmol~ and ROBT~20 (39 mg, 0.26 mmol) are dissolved in T~F/DM~ (2 ml/2 ml) and stirred for 5 min., before (3S,4RS)-3-amin~-4-hydroxy-5(2,6-dimethylbenzoyloxy)-pen~anoic acid tert.butyl ester (87 mg, ;~
0.26 mmol) in T~F (2 ml) is added. The reaction mixture is :~
stirred overnight at r.t., evaporated and chro~atographed (SiO2, ~:~
EtOAc/MeO~/NH3 95/5/0.5) to afford the (3R,4RSj title co~pound.

E~HPL~ 13: ~-valyl-alanyl~-(3R)-3-a~i~o-4-oxo-5-~,6-dichloro-benz~ylo~y) pentanoic acid ethyl ester Dimethylsulfoxide in C~2Cl~ is added dropwise to a solution of : `
oxalyl chloride in C~2CI2 at -50~C. After 15 min- 9 a solution of compound of Examp~e 12 in C~2C12 is added dropwise and the reaction mix~ure stirred for 1 hour at -40C. Triethylamine is added and seirrin~ is continued for 3 hours at room tempera~ure.
Water is ~hen added and the reaceion mixture extraceed with C~2Cl2. The organic layer is washed with NaHCO3 solution and '~.:`''-' ';'' '~

W O 93/0913~ 2 11 6 6 ~ 3 PCT/EP92/02472 - 1 9 - ::

NaCl solution, dried over Na2S04, f;ltered and evaporated. The residue is chromatographed yielding the title compound. -~

By repeating the procedure of Examples 1-3 and 11 to 13, using :~.
the corresponding starting materials, following compounds of formula IB

Z-A3-A4-N~ ~ 0 Rx (IB) :~
~ C02R~

wherein A3, A4, Rx and Ry are as defined below~ may be prepared.

:;

~, .
'`"'`'.~' : .~
,.','.''`, ~, ~i, .. .

... ' ~.:
:
~ .

WO 93/0913: 2116 6 5 3 PCI`/EP92/0247 -2 ~

~ :,.. ~
3 ~ , :.
o ~ ....
o o ~ :
o C~ ~ _ .
_ o ~ ~ ,, ~ ~ ~ ~ ~ ~ .. `.
o? :~ ~ o o o N O o ~.;. .
N N N C: C C C N N
QJ C ~ C ~ ~ ~
~D D 1::D I I I 1 51 ~ - ~
~ I I QJ I ~7 ~ r~ ~ I I , 1:~ ~ 1 ~ o h~
CL ~ ) Z C~
I I O I I I I I

. '~
:~ 3~ 2 1~1; ~ ~"''`
~ '~

~t ~ 52 w c~

=o `:
: / ~ ~ , : : \: ~.-.-~ z: ~
~ ~ =~
: ~
~: _........ : ~ ~ : ;"
' E~
: ~: . ~ . .
u~ ~ t~ a~ cr o : : ''~
. ;'' r. ::
: :,.

WO 93/0913:~ _2 1 - PCI`/EP92/02472 ~ _ o C~
o ~ cr~
. . .
C~ o ~ ~ o o o ~ o o o C~
__ _ Cr~ ~ CO o~ :
~ h ~) ~
_- O ~ I I I I

~1 ~ ::
O C ' ~ :
N N N t~ ` ~
I¦ ~ :J C e ~ 3~
C~ N

"' .' ,' ,.:`',,`,.

~ U~ W U~ ~q ~7 '`'`''`''`;
~ ~C~

~
I ~
¢ E~
t~

~a ''', ~ in ~ ~ O ~ ,,, 6~3 WO 93/0913:` PCI`/EPg2/0247' . ~
- ~ .,.
.a) _ ~, . ~:.
~, o ~`
. . X
O ~ t ~ # * ~ -N a~: ~ : :
~ h _ O

t J N N N N N N N N O N N N N N N
e ~: c c cc: N C ~ C C
~ ~ ~ D D ~.D 5:~ D OU n ~ D D ~ ~
X I l I ~ I I I I I I ~ I I I I I I.:

I I I ~ I I I I I I I Is~

~ ~ o . ~ O~
: o N:C

¢ O d ¢ ~ 2 ~~a ~ m ~ J I ~ I ¢ ¢ . .
: : : ; ~: '~ .'~`:
: , : .. ~ .. ~, 1, ~ .
: : i.

¢ ~: ~ ~

c.~ ~ O o `~

:
: ~

WO 93/0~ 21~ 6 6 ~ 3 P~/EP~2/02472 ^,_ o ~
~ _ ~ V

o ~ ~
r~ 4~E
~ 1~ ':
_O

_l ~
Z:_ ~ '~ ~ ~ ':
N~ _~ N N N N
c O ~ c C:; D ~ / 8 ~ ~ v ~ e~ C`~ ~ ~ ;".,`.
..:;' '".

, ~ ~ v ~., V
~:; ~ ~ ~ r~ . . , C~ S~ V
, ,.,:, ,"

: ~`
~r ~ ~
'C , ::

,~
~=o ~

~: .:~ ¢ o=s~ ¢
.~. ' Ll 1~ ~ L~ Ll :
~

WO 93/0913~ 2116 6 5 3 PCI`/EP92/02~72 -~4-. . .
~ _ ~ ' o c~
~E: c, ~
c~ O ~' O ~
N Ci ~ ~ ~ ~
~ O
O ' ' ,` ~
. ~; ~) . ' ' .,~ .. , ,:
~ $
` O : ' ".- .

U~

a _ ~ 8 c I ~

o , .

~ C

r~
~ ~ ,~

.:

. .:
', W O 93/0913~ 2 1 1 6 6 S 3 PCT/EP92/0~472 2 ~-E2A~PL~ 58:
Cl Z-Elis-Asp-CH2-O CO--~ O >
Cl [a]20 = -20.3 c = 1 in MeO~

....
~XAMPLE 59: ~`
.,~, ., Cl Fmoc-Val~is-Asp-O-CO

Cl E~MPLB 60:
~, Cl C~ CH~-O ~ CO-~ls-Asp-C~-O-CO -[ a3 2 = -2I o 5 c = 1~ 07 HeO~

E~AHPL~ 6 Cl :

C6~s-C~2 ~ CO-~is-Asp-C~-O-CO~

O : Cl ,' . . .

W O 93/0913~ 2 ~16 fi 53 PCTtEP92/0247~
_ 2 6--E2AHPLE ~2: :
. .

(H3C~ N ~ Cl ¢ ~ SO2-Val-Ala-Asp-CH2-O-CO , ~ ~, Cl ; ~
By repeating the proeedure of Examples 1-3 and 12-13 and using : ~
the corresponding starting materials, following compounds of ::formula IC ~:

Z-Val-A~-N~ ~ 0 ~ IC -`
x 2 y '~

wherein A3, Rx and Ry are as flefined below, may be ;
prepared EXAMPLE - A~ Ry : R~ ~ ~

63 Ala ~ 2,6-diCl-benzoyl R, s : .
64 Ala C2H5 Cl :
~ -C0 _ ~ N ~ R : ~--y : ~.
Cl Ala ~.C4~9 2,6-diCl-benzoyl R,S
66 -. Ala -C2L4-OC~3 2,6-diCl-benzoyl R,S ~.

,, ~,0 93/0913~ G 6 5 3 PCT/EP92/02~72 ~ 7 EXA~PLE 67: (3RS)-3-(Z-valyl-phenylalanyl~amino-4-oxo-6-~p-nitrophenyl)-transhex~5-enoie acid tert~butyl ,:, (3RS)-3-(Z-Valyl-phenylalanyl)amino 4-oxo-pentanoic acid tert.butyl ester-5-diethylphosphonate (0.2 g~ 0.29 mM) is dissolv~d in THF (15 ml~ at 5C. NaB (25 mg, 0.59 mM) is added and the mixture stirred for 10 min. p-Nitrobenzaldehyde (0.18 g, 1.1 mM) in T~F (2 ml~ is added at O~C and the reaction mixture `stirred for 45 min. at this temperature. The reaction mixture is poored on 5 ~ tartaric acid, extracted with ethyl acetate, the organic phases dried over Na2SO4, evaporated and the product purified by chromatography yielding the title compound. ;

E~AMPL~ 68: Z-Val-Phe-~spartic aldehyde di~ethyl acetal-~-methyl .
ester Z Yal-Phe-Asp(O~ (0.8 g~ is dissolved~ in MeOH (25 ml) containing 8 % ~Cl, left at r.t. over night, evaporated to `~
dryness and the residual crystals washed with ether, providing the titl~ compound as white crystals. M.p.: 168 - 171 .

Starting materials may be prepared as follovs:

~AHPL~ 69: (3S3-3-(~luore~ylmetbo~ycarbonyl~a~ino-5-iodo-4-oxo ;
butanoic acid tert.butyl ester . ~ ~
Triethylamine (1.3 ml, 9.6 mmol), followed be ethylchloroformiate (0.92 ml, 9.6 mmol) are added to (3S)-3-(fluorenylmethoxycarbo- -;
nyl)amino-3-earboxy-butanoic acid tert~butyl ester (3.5 g, 8.5 mmol) in THF (60 ml) at - 10 . After 10 min., a solution of dia-zome~hane in~ether is added slowly, and the reaetion mixture stirred for 45 min. at O - 5 3- HCl (2N) in ether is added at 5 -10 until gas evolution has ceased. The reaction mixture is eva-porated to dryness, taken up in acetone (50 ml) NaI (4 g) added 21166~i~
W O 93/09l3~ PCT/EP92/02~7~ ~ .
- 2 S ~

and stirred for 1 hr at r.t. Ether (150 ml) is added, the reac-tion mixture filtered and evaporated. The residue is chroma~o-graphed (SiO2 ~ EtOAc/hexane), yielding the title compound a5 slightly yellow crystals.

EXANPL~ 70: ~3S)-3-(~luo enyl~ethoxycarbonyl)amino-5(2,6-. -,. ~
dimethylbenzo~lo~y)-4-oxo pentanoic acid tert.butyl :-~
~.,, ester . ;

3S-3-(Fluorenylmethoxycarbonyl)amino-5-iodo-4-oxo-butanoic acid tert,butyl ester. ~1 g, 1.8 mmol~, 2,6-dimethylbenzo~c acid~
(0.5 g, 3,3 mmol) and AgOAc (0.6 g9 3!6 mmol~ are dissolved in acetone t25 ml) and refluxed for 1 hr. After filtration and `
evaporation the crude product is chromatographed (SiO
ether/hexane 3/7) yielding ~he title compound.

~XAMPLE 71: t3S)-3-Fluo en~etho~ )onyl)a~i~o-5(4 nitroph~no~y)-4-o~o penta~?ic ~cid tert.butyl ester ~ .

i:
3S-3-(Fluorenylmethoxyca~bonyl)amino-5-iodo-4~oxo-butanoic açid tert.butyl ester (1 g, 1.8 mmol~, p-ritrophenol (0.5 g, 3.7 mmol~
and K2CO3 (0.38 g, 2.8 mmol):are xefluxed in acetone (~6 ml3~for 30 min., C~2C12 added:and the~organic phase washed with 2N ~:
Na~CO3. The combined organic phaszs ar:e dried,~ evap~rated and : ;~
chromatographed (SiO2,~ EtOAc/hexane 2/8):, yielding:the title compound as a yellow oil. ~ :
~ . ,, .
.

E~AHPL~ 72: (Z-val~l-ala~yl3-~3R,4S)-3-a~i~o-4,5-dihydro~y e l~jl e~el '~"'`' a~ Ethyl (3~4S)-3-benzylamino-4,5-(isopropylidenedioxy~
pentanoate (Y. Yamada, Tetrahedron Leters l983, 24, 3009) and ~ ~`
10% Pd/C in ethanol are shaken at r.~. for 30 m~ under B2. ~ ':
', , ~ ,.
`'',` ~

'' `'"

2 1 1 6 6 5 3 PCr/EP92/02~7? ~ :
2 g .

Filtration and evaporation of the reaction mixture give ethyl (3R,4S)-3-amino-4,5-(isopropylidenedioxy~pentanoate which is used without further purification.
: . '.
b) Z-Val-Ala-OH is dissolved in THF, HOBT~20 and DCC are added at ~ ;
5C. After stirring for 20 min. at 5C, diisopropylethylamine and ethyl (3R,4S)-3-amino-4,5-(isopropylidenedioxy)peneanoate in TH~ are added. Reaction mixture is stirred overnight at room temperature, filtered, evapored and chromatographed to afford (Z-valyl-alanyl)-~3R,4S)-3-aDino-4~5-(isopropyliden~
edioxy)pentanoic acid ethyl ester.

c) Compound 74 b) is dissolved in AcO~/R O (7S/25) and stirred at 40C for 4 hours. After evaporation, water is added and the mixture is extracted with AcOEt. The combi~ed extracts are washed with water, Na~CO3 solution~ NaCl solution, dried over Na2SO4, filtered and evaporated, yielding the title compound.
.;-.~,~.
The compounds of formula I their physiologically-hydrolysable and~
-acceptable esters and amides~and~their~pharmaceut~ically acceptable salts (hereinafter referred to as compounds of the invention) exhibit pharmaceutical activity and are, therefore, useful as pharmaceuticals.

In particular, the compounds of the invention inhibit IL~
secretion as indicated in the follo~ing in vitro test using T~P~
..
cells and in vivo test methods~

a) 900 ~l T~P-l cells (0.5 x 106 cells) together vith 100 U
y-interferonJO.g ml RPMI 1640 medium (containing 2 mM
L-glutamihe and~5 X heat-inactivated foetal calf serum) are pipetted into 24 vell culture plates. 100 ~l of the compound to be tested are ~hen added. After 3 hours at 37 ~ C in 5 X
C02/95 X air, 10 ~l~lipopolysaccharide 500 ~g/ml is added and ~ ,. .;, ',',, ,~.....

W ~ 93/0913~ ~ 6 PCT/EP~2/0247~ .

the incubation continued for a further 40 hours. Appropriate controls (with and without stimulus~ solvent~ ar also :
included. The media are then removed and clarified by centri- ~ .
fugation at 1000 g for 10 min. 1.0 ml digitonin 0.01 X is --added to the wells to lyse ~he c211s which are loosened by scraping with a rubber policeman and left at 4 C for :~
10 min. Lactate dehydrogenase measurements are then performed . immediately and the samples stored at - 20 ~ C until the other determinations can be made. The assays are: IL~
(medium and lysate), IL-6 (medium), TNF-a (medium), P~B2 -(medium and lysate3~ lactate dehydrogenase (LDH) and DNA
(lysates). IL-1~, IL-6 and TNF-~ assays are determine~ using :
commercially available ELISA kits (Cistron), P OE 2 is measured using a standard RIA and DNA fluorimetrically using nAPI. ;

In this test, the compounds of the invention selectively inhibit IL-l~ release in concen~rations from ~bout 0.01 to 100 ~M. In contrast IL-6, TNF-a, PG~2 and DNA levels remain substantially unaffected, and the compounds are:;non-toxic, since LD~ release is unchanged. It has for example been de~
termined that compounds of examples 40 and 50 have an IC50 : ~-value (concentration~of compound which i~hibits ~o 50% the release of IL-1 O of 1 and 0.1 ~M respectively. `~`
,.
b~ LPS-Fe~er ~. .
....
,~;' ".-A LPS-suspension (Sigma~ No. L-5Bû6~ lOO~g/Sml glucose solu-tion~kg s.c.) is injected in male Tuttlingen SD rats -~
(150-160g~. 2 hours later the body temperature is measured using a thermistor rectal probe connected to an ELLAB tele-thermometer. After 4 hours the test compound is ad~inistered ;~
p.o. 2 hours later (6 hrs ater LPS administration) the temperature is measured again. The temperature increment ~
shown by the untreated controls is taken as lOOX and that in : , ' ~
,~ .

',', `,'`.,~
- ~ . . .....

~ . ~

W O 93/0913~ 2 ~ 5 3 PCT/EP92/0247 the treated group is expressed as a percentage of ~his value.
The ED50 is the dose causing a 50Z inhibition of the tempera~
ture increase determined in the ~ontrol rats. In this test compounds of the invention inhibit the LPS-induced ~emperatu- :~
re increase when administered at a dosage in the range of from 0.001 to 0.1 mg/kg p.o. It has for example been deter-mined that compounds of examples 40 and 50 have each an ED50 value of 0.01 mg/kg p.o. and compound of example 51 an ED~o value of 0.05 mg/kg p.o.

c) Carrage~na~-Induced Pa~ ~de~a in the Rat : .
. . .

50FA male rats, 150-170g body weight, are used for each group. The test compound is administered orally as a :~
suspension in physiological saline/0.5X tragaeanth 1 hour prior to the carrageenan injection. Carrageenan (O.lml of a 1~ suspension in physiological saline~ is given:by subplantar injection into one hind paw. The swelling of the paw is measured by means of an antiphlogo-eter ~ccording to Kemper ~ :~
AmelmO A control readlng is taken immediately after ehe injection, and ~he swelling is measured after 3 and 5 hrs.
The mean value of the 3- and 5~hour r~ading is taken after deduction of the control reading, the values obtained from the treated animals are expressed as a percentage of the value obtained from non-treated controls. The ED50 is the dose causing a 50Z inhibition of the carrageenan-induced ~:~
s~elling after 3 hrs. In this test method compounds of the invention inhibit significantly the carrageenen-induced swelling when administered p~o. at a dosage in the range of from ~.02 to 5 mg/kg. It has for example been determined that ::
compound of example 40 and 50 have an EDso value of 0.2 and 1 mg/kg p.o. re8pectively.

'i"';

',':
'., W O 93/09135 2 11 6 6 53 PCT/EP92~02~7~ .

32 ~:

Compounds of the invention are therefore useful for the treatment of disorders with an aetiology associated ~ith or comprising excessive IL-l~ release, e.g. in a ~ide variety of inflammatory states and diseases, for example ~issus calcium depletion, ~-~
degenerative processes in bone and cartilage, e.g. rheumatoid arthritis and osteoarthritis, inflammatory bowel disease, irritable bouel disease, septic shock, psoriasis, asthma, adult ; -respiratory distress syndrome, diabetes type I, osteoporosis of various genesis including e.g. climacteric or post-menopausal osteoporosis as well as osteoporosi5 consequential to old age, immobilization or trauma, arterioscl~rosis and Alzheimer disease.

For the above uses the required dosage will of course vary depen~
ding on ~he mode of administration, the particular condition to be treated and the effect desired. In general however, satisfac~
tory re ults are achieved at daily dosage rates o~ from about ~:
0.001 to about lO0 mg/kg, pre~erably O~OOl to about 10:mg/kg ;~
animal body ueight. Suitable daily Idosage rat~s for larger mammals9 for example humans, are of the order of from about O.l :
mg to absut l g/da~, conveniently administered once, in divided dosages 2 to 4 ~/day, or in sustained release form. `:

Compounds of ~xamples 50 and 51 are preferred. ~ ;~

1,~ '',. j~' In accordance uith the foregoing the present invention also .
provides:

a~ A method for the treatment of disorders with an aetiology associated ~ith or comprising excessive IL-l~ release, e.g. ~
as fhdicated above in a subject in need thereof, ~hich method ~;`
comprises administering to said subject an eff~ctive amount of a comp~und of formula I 9 a physiologically-hydrolysable ~`
and -acceptable ester:or am;de thereof, or a pharmaceu~ically -~
acceptable sal~ thereof; s~

W O 93~09135 2 1 1 ~ 6 5 3 PCT/EP92/0247~ -- 33 - ~:

:' b) A compound of formuIa I, a physiologically-hydrolysable and -acceptable ester or amide thereof or a pharmaceuticalIy ~:
acceptable salt thereof, for use as a pharmaceutical, for example for use as an agent, e.g. in the method as discIosed above.

The compounds of the invention may be administered by any con- `~
ventional route, in particular nasally, enterally, preferably .:~
orally, e.g. in the for~ of tablets or~capsules, or parenterally e.g. in the form of:in~ectable solutions or suspensions or in a suppository form. Unit dosage for~s contain, for example from~
about 25 ~g to 500 mg of a compound of the invention.
r~
The compounds of the invention may be administered in free form ~ -or in pharmaceutically acceptable salt form. Such salts may be prepared in conventional manner and exhi~it the same orde~:of activity as the free compounds. :~
:
~: ~
~urthermore the present invention also provides~
c) A pharmaceutical~composition~comprising a compound:of formu~
la I~ a physiolog~cally-hyd~rolysable~and;-acceptable ester~;~or amide thereof or a:pharmaceutically aceeptable sal~t~thereof, : : ~.
as hereinbefore~defined, together~with a pharmaceutically~
acceptable diluent or ca~rier the~refor. Such~:compo:sltions m~ay~
be manufactured in convention~l:manner. They:may comprise up :~
to 99.9% by weight of~active :1ngredient.

; , .

' ''`~:
,.

", ~
:~

~: ~

Claims (10)

CLAIM

1. A compound of formula I

R-[A1-A2]n-A3-A4-X-A5 (I) wherein R is hydrogen, an amino protecting group or optionally ring substituted benzyloxy n is 0 or 1, A1 is Val, Leu, Ala, Ile or trimethylsilyl-Ala A2 is Phe or Tyr, A3 is a direct bond, Val, Leu, Ala, Ile, trimethylsilyl-Ala or a divalent radical of formula (a) (a) wherein ring A is optionally substituted by hydroxy or C1-4alkoxy, A4 is a direct bond or a divalent radical of formula (b) (b) wherein R1 is hydrogen or C1-4alkyl, and Y1 is the residue attaching to the .alpha.-carbon atom of an .alpha.-amino acid and optionally protected, -CH2-CH2-N(C1-4alkyl)2, imidazol-2-yl-methyl, benzimidazol-2-yl-methyl, 1H-1,2,4-triazol-3-yl-methyl, pyrazol-3-yl-methyl, indazol-3-yl-methyl or a radical of formula (c) or (d) or (c) (d) wherein each of R2 and R3, independently, is hydrogen, halogen, C1-4alkyl, CF3 or trityl, at most one of R2 and R3 being H, and each of R4 and R5 independently is hydrogen, C1-4alkyl, hydroxy, C1-4alkoxy, CF3, phenyl or halogen, at most one of R4 and R5 being H, or A3 and A4 form together a radical of formula (aa) (aa) wherein Y1 is as defined above and R1 and R1a form together -(CH2)m- wherein m is 2, 3, 4 or 5, and i) X is a divalent radical of formula (e1) (e1) wherein R6 is H or C1-4alkyl, and A5 is hydrogen; CF3; a radical -CH2-X1-Y3 wherein X1 is O
or S and Y3 is heteroaryl; a radical -CH2-Y3; or a radical of formulae (k) to (m) -CH2-X1-CO-Y4 (k) (l) or (m) (o) wherein Y4 is tri-(C1-4alkyl)methyl or a residue ring B is pyridyl, ring C is phenyl or pyridyl, each of R7 and R8: independently is C1-4alkyl, C1-4alkoxy, CF3; halogen, nitro or cyano, and each of R9, R10 and R11 independently is nitro, cyano, CF3, carbamoyl, CO2R12, -CH=CH-CN or: -CH=CHCO2R12 wherein R12 is C1-6alkyl, X being also a divalent radical of formula (e2) (e2) when A5 is H, or ii) X is a divalent radical of formula (f), (f) and A is -Z1-Z2-Y2 wherein each of Z1 and Z2 independently is a direct bond or an .alpha.-amino acid residue and Y2 is NH2, C1-4alkylamino, di-(C1-4alkyl)amino or a heterocyclic radical attached by a nitrogen to Z2; or a radical of formulae (k) to (o) as defined above; or OR13; or NR14R15 wherein R13 is C1-12alkyl optionally substituted by OH or interrupted by O and each of R14 and R15 is independently hydrogen C1-12alkyl, C5-7 cycloalkyl or benzyl, or iii)X is a divalent radical of formula (g) (g) and A5 is -Z1-Z2-Y2 as defined above, or iv) X is a divalent radical of formula (h) or (j) (h) (j) and A5 is a radical of formulae (k) to (o), -CH2-Y3 or -CH2-X1-Y3 as defined above, with the provisos that only one of A3 and A4 can be a direct bond when n is 0, and each of A3 and A4 is other than a direct bond when n is 1.

and the physiologically-hydrolysable and -acceptable esters or amides thereof, in free form, in salt form or in the form of complexes.

2. A compound of formula I according to claim 1, wherein n is 0, A3 is a direct bond, Val, Leu, Ala, Ile or trime-thylsilyl-Ala, A4 is as defined in claim 1, or A3 and A4 form together a radical of formula (aa) as defined in claim 1, and i) X is a radical of formula (e1) or (e2) as defined in claim 1 and A5 is H, or ii) X is a radical of formula (e1) or (f) and A5 is a radical of formula (k), (l) or (m) as defined in claim 1 wherein X1 is 0, iii)X is a radical of formula (g) and A5 is -Z1-Z2-Y2 as defined in claim 1, or iv) X is a radical of formula (j) and A5 is a radical of formula (k), (l) or (m) as defined in claim 1 wherein X1 is 0.

and the physiologically-hydrolysable and -acceptable esters or amides thereof, in free form, in salt form or in the form of complexes.

3. A compound of formula I according to claim 1 wherein n is 0 and each of A3 and A4 are other than a direct bond, and the physiologically-hydrolysable and -acceptable esters or amides thereof, in free form, in salt form or in the form of complexes.

4. A compound of formula I according to claim 1 or 3 wherein Y1 is the residue attaching to the .alpha.-carbon atom of an .alpha.-amino acid selected from Ala, Leu, His, Phe, Met, Trp, trimethyl-silyl-Ala and optionally side chain protected Arg, Orn and Lys, or a radical of formula (c), and the physiologically-hydrolysable and -acceptable esters or amides thereof, in free form, in salt form or in the form of complexes.

5. A compound of formula I according to claim 1, 3 or 4 wherein X is a radical of formula (e1), (h) or (j), As being other than H when X is (e1) and the physiologically-hydrolysable and -acceptable esters or amides thereof, in free form, in salt form or in the form of complexes.

6. (Benzyloxycarbonyl-valyl-alanyl)-3R-3-amino-4-oxo-5-(2,6-di chlorobenzoyloxy) pentanoic acid ethyl ester, (benzyloxy-carbonyl-valyl-alanyl)-3R,S-3-amino-4-oxo-5-(2,6-dichloro benzoyloxy) pentanoic acid ethyl ester, (benzyloxycarbo-nyl-valyl-alanyl)-3R-3-amino-4-oxo-5-(2,6-dichloro-pyridyl-4-carbonyloxy) pentanoic acid ethyl ester, (benzyloxycarbo-nyl-valyl-alanyl)-3R,S-3-amino-4-oxo-5-(2,6-dichlorobenzoyl oxy) pentanoic acid, (benzyloxycarbonyl-valyl-alanyl)-3R,S-3-amino-4-oxo-5-(2,6-dichlorobenzoyloxy) pentanoic acid isopropyl ester, (benzyloxycarbonyl-valyl-alanyl)-3R,S-3-amino-4-oxo-5-(2,6-dichlorobenzoyloxy) pentanoic acid t.-butyl ester, and (benzyloxycarbonyl-valyl-alanyl)-3S--3-amino-4-oxo-5-(2,6-dichlorobenzoyloxy) pentanoic acid in free form, in salt form or in the form of complexes.

7. A process for producing a compound of formula I and the phy-siologically-hydrolysable and -acceptable esters and amides thereof, as defined in claim 1, which process comprises a) removing at least one protecting group from a compound of formula I in protected form or adding a protecting group R
as defined in claim 1 at the N-terminal group of a compound of formula I; or b) converting one compound of formula I into another compound of formula I; or c) coupling together by an amide bond two peptide fragments, each of which contains at least one amino acid in protec-ted or unprotected form and one peptide fragment contai-ning a radical of formula (e1) to (j) as defined in claim 1, the peptide fragments being such that a protected or unprotected peptide having the sequence according to for-mula I above is obtained and, if necessary, removing the protecting group or groups from a compound of formula I in protected form; or d) for the production of a compound of formula I wherein X is a radical of formula (e1) or (h) and A5 is a radical of formula (k), (l) or (o) or -CH2-X1-Y3 as defined in claim 1, reacting a compound of formula III

R-[A1-A2]n-A3-A4-X'-CH2-Za (III) wherein R, A1 to A4 and n are as defined in claim 1, X' is a radical of formula (e1) or (h) as defined in claim 1, and Za is a leaving group, with a corresponding phenol, thiophenol or HX1-pyridine or an acid of formula HX1-CO-Y4 or a functional derivative thereof or HX1-Y3 wherein X1, Y3 and Y4 are as defined in claim 1; or e) for the production of a compound of formula I

(I) wherein R, A1 to A5 and n are as defined in claim 1 and R16 is a C1-12 aliphatic or alicyclic residue, oxidizing a compound of formula V

(V) wherein R, A1 to A5, n and R16 are as defined above, and recovering a compound of formula I or a physiologically-hydrolysable and -acceptable ester or amide thereof thus obtained in free or salt form or in the form of a complex.

8. A compound according to any one of claims 1 to 6 for use as a pharmaceutical.

9. A pharmaceutical composition comprising a compound of formula I as defined in claim 1 or a physiologically-hydrolysable and -acceptable ester or amide thereof, in free form or in phy-siologically acceptable salt form, together with a pharmaceu-tically acceptable diluent or carrier therefor.

10. A method for the treatment of disorders with an aetiology associated with or comprising excessive IL-1.beta. release, in a subject in need thereof, which method comprises administering to said subject an effective amount of a compound of formula I as defined in claim 1, a physiologically-hydrolysable and -acceptable ester or amide thereof, or a pharmaceutically acceptable salt thereof.