AMINO ACID AND PEPTIDE INHIBITORS OF HUMAN LEUCOCYTIC ELASTASE

Abstract: Potent and specific inhibitor compounds for elastase - type enzymes, especially for human leucocytic elastase (HLE), are provided by combining an amino acid or short peptide with a hydrocarbon-oxycarbonyl protecting group having at least 4 carbon atoms at the N-terminal of the amino acid or peptide, and a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms at the carbonyl group of the amino acid or peptide, the protecting group and the hydrophobic group being physiologically innocuous components, but providing the required inhibition. The inhibitor compounds can be expected to be useful in the treatment of elastase - type implicated diseases such as arthritis, tumor growth and emphysema.

Description: AMINO ACID AND PEPTIDE INHIBITORS OF HUMAN LEUCOCYTIC ELASTASE BACKGROUND OF THE INVENTION Field of the Invention: This invention provides potent and specific inhibitor compounds for elastase-type enzymes, especiall for human leucocytiσ elastase CHLEΪ , which enzyme is implicated in the growth of tumors, degradation of tissues in arthritis and in the destruction of lung tissues in emphysema, the molecular structure of the inhibitor compounds of the invention being designed to have an expected absence of undesirable side-effects in use. The invention also provides: processes for the prep¬ aration of said inhibitor compounds; pharmaceutical/veterinary compositions containing said inhibitor compounds; and methods for the clinical application of said inhibitor compounds and said pharmaceutical/veterinary compositions. Description of the Prior Art: There is evidence to implicate the neutral proteases . of human leucocytes (.polymorphonuclear leucocytes) in the degradation of cartilage in both rheumatoid and osteoarthritis. The chronic destruction of the elastic component of lung connective tissues by elastase-type enzymes, in particular by HLE and σathepsin G, is currently believed to result in the onset of chronic obstructive lung disease. These proteases are primarily inhibited by the major serum protease inhibitor - ~Ά. -_ proteinase inhibitor (c -PI) , which is also a normal constituent of bronchioalveolar lavage fluid (BAL) . -.- . , . . However, ,--PI is readily inactivated by oxidants such as those present in cigarette""smoke- or oxidative enzymes (i.e. myeloperoxidase) that normally ' function in phagocytic cells during inflammatory states.- " In addition, some persons 1. are genetically deficient in α, r-Pl with leyels of the inhibitor which are 25% of normal. Thus, persons with a compromised inhibitor screen are prime candidates for chronic obstructive lung disease. 5 There are many classes of compounds reported in the literature as inhibitors of HLE. Some of these compounds are the chloromethyl ketones, vide: J.C. Powers, B.F. Lupton, A.D. Harley, N. Nishino, R.J. Whitley, Biochem. and Biophys. Acta. 485, 156 (.1971 ) and K. Haveman & A. Janoff: "Neutral 10 proteases of Human Poly orphonuclear Leukocytes", p.221, Urban and Schwartzenberg, Baltimore (1977) ; the sulfonyl fluorides, vide: K. Haveman & A. Janoff: "Neutral proteases of Human Polymorphonuclear Leukocytes", p.221, Urban and Schwartzenberg, Baltimore C1977) ; the i idazole-N^carboxa- 15 mides, vide: W.C. Croutas, R.C. Budger, T.D. Ocain, D. Felter, J. Frankson, M. Theodorakis, Biochem. and Biophys. Research Commun. 1890 (1980)-; the azapeptides, vide: K. Haveman & A. Janoff, "Neutral proteases of Human Poly¬ morphonuclear Leukocytes", p.221, Urban and Schwartzenberg, 20 Baltimore (1977); cyclohexylamide, vide: CH. Hassall, W.H. Johnson, N.A, Roberts: Bio-Organic Chem. j3_, 299 (.1979) ; the adamantane sulphenyl peptides, vide: A.M.J. Blow, Biochem. J. 161, 13 (1977]; the cis-unsaturated fatty acids, vide: B.M. Ashe, M. Zimmerman, Biochem. and Biophys. Res. Commun. -5 7_5, 94 (1977); and the gold complexes such as gold thiomalate, vide: A. Baici, P. Salgam, K. Fehr, A. Boni, Biochem. Pharmac. -30_ 903 (1981) . The chloromethyl ketones, the sulfonyl fluorides, the imidazole-N-carboxamides, the aza peptides, the cis- 30 unsaturated fatty acids, and the adamantane sulphenyl peptides, all have reactive groups which make them inadvis¬ able, if not dangerous, to use. In general, it can be said that those compounds bearing highly reactive functional groups are difficult, if not impossible, to target onto ' 5 receptor sites, as they are likely to react with the many components of the body ayailable between the point of administration and the target receptor site. Oleiσ acid, a cis--insaturated carbσxylic acid, has been shown to be an acceptably good specific inhibitors of HLE, but not of porcine pancreatic elastase (PPE), trypsin, chymo- trypsin or cathepsin G, which has indicated to us that the principal difference between HLE and the other serine pro¬ teases could be due to the different hydrophobic character of a site near the active site, noting that HLE has not been fully sequenced, its 3-dimensional structure has not been determined, and its active site-stereochemistry is unknown. These proteases apparently prefer certain amino acids or short peptide sequences.as substrates, and previous workers in the field have sought to utilize this preference, either by using the synthetic substrate as a competitive inhibitor, or by functionalizing those substrates so as to cause them to react with the active site, thereby inactiv¬ ating the enzyme. SUMMARY OF THE INVENTION We have now conceived the idea of making an inhibitor for elastase-type enzymes especially for HLE, by combining a preferred amino acid or short peptide which on its own is a ubiquitous physiological component without appreciable toxicity, with a hydrocarbon-oxycarbonyl protecting group having at least 4 carbon atoms at the N-terminal of the amino acid or peptide, and a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms at the carbonyl group of the amino acid or peptide, the protecting group and the hydrophobic group being physiologically innocuous components, but providing the required inhibition. The advantage of this concept resides in that enzyme cleavage of the inhibitor would merely result in end products which are physiologically innocuous components. Thus, the present invention provides HLE and other elastase-type enzyme inhibitor compounds consisting of amino acid derivatives or peptide derivatives as defined by the general formula (I) and physiologically acceptable salts thereof, wherein: R, is a hydrocarbon-oxycarbonyl group with at least 4 carbon atoms, the hydrocarbon-σx oiety of which may be substituted or unsubstituted, for protecting the N-terminal of the amino 0 acid or peptide, which protecting group if cleaved in vivo, forms a physiologically innocuous compound; R_ and R, , which may be the same or different and may differ from unit to unit when n is greater than 1, are selected from hydrogen, alkyl of 1 to 10 carbon atoms, substituted alkyl 5 of 1 to 10 carbon atoms in which the substituents are those present in natural or non-natural amino acids, provided that they are physiologically innocuous, or R- and R- taken together with the adjacent nitrogen and carbon atoms may form a 5-membered ring with 4 carbon atoms, which ring may _0 optionally be substituted in which the substituents are those present in natural or non-natural amino acids provided that they are physiologically innocuous; R. is a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, which if cleaved in vivo, forms a physiologically innocuous .5 compound, said hydrophobic hydrocarbon chain being bonded direct to the carbonyl carbon or to the carbonyl carbon through an oxygen, sulphur or nitrogen hetero atom; and n is an integer 1 to 6. The group R, , which is a bulky, hydrophobic, apolar, amino-pro- 0 tecting group having steric influence on the elastase-type enzyme and materially contributing to the inhibitory effect of the 0 II present inhibitors , may be of the formula R ' - 0 - C - wherein R ' is s-elected from: (i) straightr-chain alkyl of 1 to _L0 carbon atoms, straight- chain alkenyl of 1 to 10 carbon atoms, straight-chain alkynyl of 1 to 10 carbon atoms, each of which is sub- stituted with one or -more of alkyl of 3 to 10 carbon atoms, alkenyl of 3 to 10 carbon atoms, and alkynyl of 3 to 10 carbo atoms to form a branched-r-chain; or substituted with one or more of cycloalkyl of 3 to 10 carbon atoms, cycloalkenyl of 3 to 10 carbon atoms, aryl of 6 to 10 carbon atoms, adamantyl, heterocyclic radicals such as pyrryl, f ryl, thiophenyl or pyrazolyl, any of which may be substituted, for example, with one ormore of halo, hydroxy, nitro, alkyl of 1 to 5 carbon atoms, or alkoxy of 1 to 5 carbon atoms; or (ii) branched ■ =-chain alkyl of 3 to 10 carbon atoms, branched- chain alkenyl of 3 to 10 carbon atoms, branched-chain alkynyl of 3 to 10 carbon atoms, cycloalkyl of 3 to 10 carbon atoms, cycloalkenyl of 3 to 10 carbon atoms, aryl of 6 to 10 carbon atoms, adamantyl, or heterocyclic radicals such as pyrryl, fur l, .thiophenyl or pyrazolyl, any of which may be substituted, for example, with one or more of halo, hydroxy, nitro, alkyl of .1 to 5 carbon atoms, or alkoxy of 1 to 5 carbon atoms, Thus, the amino-protecting group R-, may be: (a) a branched chai alkoxycarbonyl of 3 to 10 carbon atoms such as t-butyloxycarbon isopentyloxycarbonyl or isohexyloxycarbonyl; (b) an aryl substi uted straight-chain alkoxycarbonyl of 1 to 10 carbon atoms such as benzyloxycarbonyl; (c) a cycloalkyloxy carbonyl in which the cycloalkyl moiety has 3 to 10 carbon atoms, such as cyclohexyl- oxycarbonyl; (d) an aryloxycarbonyl in which the aryl moiety ha 6 to 10 carbon atoms, such as phenyloxycarbonyl ; or (e) a sub¬ stituted aryloxycarbonyl in which the aryl moiety has 6 to 10 carbon atoms, such as tolyloxycarbonyl or xylyloxycarbonyl . The groups R and R_ , which are constituent parts of natural or non-natural amino acids, may be the same or different and are selected from hydrogen, alkyl of 1 to 10 carbon atoms, for example, methyl, 'isopropyl or isobutyl; substituted alkyl of 1 to 10 carbon atoms in which the substituents are those present in natural or non-natural amino acids, for example, p-hydroxy phenylmethyl or 4-hydroxy-3 ,5-diiodobenzyl; or R and R-. taken together with the adjacent nitrogen and carbon atoms may form a 5-rmembered ring with 4 carbon atoms, for example, the 5-membered ring of proline, which 5*-membered ring may optionally be substituted in which the substituents are those present in natural or non-natural amino acids, for example, the 5-membered ring of either hydroxyproline or p-trifluoromethylproline. The group R, is a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, preferably 10 to 14 carbon atoms, more prefer¬ ably 12 carbon atoms, which, as indicated, if cleaved in vivo, forms a physiologically innocous compound, said hydro- phobic hydrocarbon chain being bonded direct to the carbonyl carbon or to the carbonyl carbon through an oxygen, sulphur or nitrogen hetero atom, preferably through nitrogen. For example, the group may be an alkyl, alkenyl, alkynyl, alkylamine, alkenylamine, alcohol or thiol, each of 8 to 16 carbon atoms, preferably 10 to 14 carbon atoms, more prefer¬ ably 12 carbon atoms. Moreover, the hydrophobic hydrocarbon chain group R. may be substituted, for example, with one or more ofhalo,hydroxy, nitro, alkyl of 1 to 5 carbon atoms, or alkoxy of 1 to 5 carbon atoms. Thus, the hydrophobic hydrocarbon chain group R may be octyl, nonyl, decyl, undecyl, dodecyl, tetradecyl, hexadecyl, amidyl octane, amidyl nonane, amidyl undecane, amidyl decane, amidyl dodecane, amidyl tetradeσane, or amidyl hexadecane. The present invention also provides pharmaceutical/ veterinary compositions comprising at least one amino acid derivative or peptide derivative of the general formula (I) above, or a physiologically acceptable salt thereof, in association with one or more non-toxic physiologically accept¬ able carriers. The present inyention further provides a method for inhibiting the growth of tumors or the degradation of tissues in arthritis or the destruction of lung tissues in emphysema, which comprises administering to an animal, including humans, suffering from any one of such conditions, an effective amount of at least one amino acid derivative or peptide derivative of the general formula Clϊ above, or a physiologically acceptable salt thereof, or a pharmaceutical/veterinary composition comprising at least one such derivative or salt. The amino acid derivatives or peptide derivatives defined by the general formula I above, and physiologically acceptable salts thereof, may be produced bymethods comprising: A: For the preparation of compounds when is i, as set out in the following reaction scheme A: (i) reacting an amino acid of formula II with a source of an R, protecting group such as di-t-butylcarbon- ate to form an amino acid of formula (III) so protected at its N-terminal; and (ii) reacting a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, with the carboxyl group of the amino acid of formula (.III) to form the desired compound of formula (.IV) . Reaction Scheme A 0 H - N - CH - C - OH (II) R- a source of an R, protecting group such as di-t-butylcarbonate O R- R- - 3 - a source of an R. hydrophobic bon chain of 8-16. carbon ch as dodecylamine R χ - N - CH - C -. R 4 (.IV) R 2 3 wherein R, , R 2 , R- and R, are as defined in formula (I) above. or the preparation of compounds when n is 2 to 6, s set out in the following reaction scheme B: (i) reacting an amino acid or peptide of formula (.V) with a source of a protecting group such as an alcohol, exemplified by methanol, to form an amino acid or peptide of formula (VI) protected at its carboxyl terminal; (ii) reacting an amino acid or peptide of formula (VII) with a source of an R-, protecting group, such as di-t-butylcarbonate, to form an amino acid or peptide of formula (VIII), so protected at its N- terminal; (iii) reacting the amino acid or peptide of formula (VI) with the amino acid or peptide of formula (VIII) to form a peptide of formula (IX) with both the N- ter inal and the carboxyl terminal of the peptide so protected; (iv) removing the protecting group from the carboxyl terminal of the peptide of formula (.IX) ; and (v) then reacting the peptide with a hydrophobic hydro¬ carbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired peptide of formula (.X) with the required R- ^ protecting group at the N-terminal of the peptide and the required hydro¬ phobic hydrocarbon chain at the carbonyl carbon of the peptide. Reaction Scheme B (V) a source of an R. protect¬ a source of a protect¬ ing group such as di-t- ing group (R) , such as butylσarbonate methanol -4- removal of the R protecting group, followed by reaction with a source of an R, hydro- phobic hydrocarbon chain of 8-16 carbon atoms such as dodecylamine wherein R 1( R., R 3 and R 4 are as defined in Formula (I) above; and a and b are integers of 1 to 5 , provided that a + b is no greater than 6. PREFERRED EMBODIMENTS OF THE INVENTION Preferred inhibitors defined by the general formula (I) above, are those wherein R-, is t-butyloxycarbonyl or benzyloxycarbonyl, more preferably, R, is t-butyloxycarbonyl; R_ and R-. are selected from hydrogen, methyl, isopropyl and isobutyl, or R_ and R_ taken together with the adjacent nitrogen and carbon atoms, form a 5-membered ring of proline, more preferably, 2 and R_ are selected from hydrogen, methyl and isopropyl; R. is a secondary amine with a hydrocarbon chain of 3 to 16 carbon atoms, more preferably, iO to 14 carbon atoms, most preferably, 12 carbon atoms; and n is 1 to 4, more preferably, 1 to 3; and physiologically acceptable salts thereof. Particularly preferred inhibitors defined by the general formula (I) above, are selected from the group: N-tert-butyloxycarbonyl-valylamidyl decane, N-tert-butyloxycarbonyl-valylamidyl dodecane, N-tert-butyloxycarbonyl-valyl-rvalylamidyl decane, N-tert-butyloxycarbonyl-valyl-valylamidyl dodecane, N-tert-butyloxycarbonyl-valyl-valyl-valylamidyl decane, N-tert-butyloxycarbonyl-valyl-valyl-valylamidyl dodecane, N-tert-butyloxycarbonyl-alanylamidyl decane, N-tert-butyloxycarbonyl-alanylamidyl dodecane, N-tert-but_loxycarbonyl-alanyl-alanylamidyl decane, N-tert-butyloxycarbonyl-alanyl-alanylamidyl dodecane, N-tert-butyloxycarbonyl-alanyl-alanyl-alanylamidyl decane, N-tert-butyloxycarbonyl-alanyl-alanyl-alanylamidyl dodecane, N-tert-butyloxycarbonyl-alanyl-prolyl-valylamidyl decane, N-tert-butyloxycarbonyl-alanyl-prolyl-valylamidyl dodecane, N-tert-butyloxycarbonyl-alanyl-alanyl-prolyl-yalylamidyl decane, N-tert-butyloxycarbonyl-alanyl-alanyl-prolyl-yalylamidyl dodecane, or a physiologically acceptable salt of any thereof. Possible salts of compounds of the general formula (I) are all the acid addition salts. The physiologically acceptable salts may be derived from inorganic or organic acids. Physiologically unacceptable salts, which may initially be obtained as process products, for example in the preparation of the compounds according to the invention on an industrial scale, are converted into physiologically acceptable salts by processes which are known to the skilled person. Examples of such suitable physiologically acceptable salts are water-soluble and water-insoluble acid addition salts, such as the hydrochloride, hydrobromide, hydroiodide, phosphate, nitrate, sulfate, acetate, citrate, gluconate, benzoate, butyrate, sulfosalicylate, maleate, laurate,. malate, fumarate, succinate, oxalate, tartrate, stearate, tosylate, mesylate and saliσylate. The preferred amino acid derivatives or peptide derivatives within the general formula (I) above, and phy¬ siologically acceptable salts thereof, may be produced by methods comprising: __ for the preparation of compounds when n is 1: (i) reacting an amino acid such as alanine or valine with a source of an R. protecting group such as di- t-butylcarbonate to form an amino acid so protected at its N-terminal, such as t-butyloxy- carbonyl alanine or t-butyloxycarbonyl valine; and (ii) reacting a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, with the carboxyl group of the amino acid of A (i) above to form the desired compound; 3: for the preparation of compounds when n is 2: (i) reacting an amino aciά such as valine with a source of a protecting group such as an alcohol, exempli- fied by ethanol, to form an amino acid protected at its carboxyl terminal, exemplified by valine methyl ester; (ii) reacting an amino acid such as valine with a source of anR. protecting group, such as di-t- butylcarbonate, to form an amino acid so protected at its N-terminal, exemplified by t-butyloxy¬ carbonyl valine; (iii) reacting the product of B (i) above with the product of B (ii) above to form a dipeptide with both the N-terminal and the carboxyl terminal of the dipeptide so protected; (iv) removing the protecting group from the carboxyl terminal of the dipeptide of B (iiil above; and (v) reacting the dipeptide of B (iv) above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired dipeptide with the required R, protecting group at the N-terminal of the dipeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the dipeptide; for the preparation of compounds when n is 3 : (i) reacting a dipeptide such as valylvaline with a source of a protecting group such as an alcohol, exemplified by methanol, to form a dipeptide protected at its carboxyl terminal, exemplified by valylvaline methyl ester; (ii) reacting an amino acid such as valine with a source of an R protecting group such as di-t-butylcarbonate, to form an amino acid so protected at its N-terminal, exemplified by t-butyloxycarbonyl valine; (iii) reacting the product of C (i) above with the product of C (.ii) above to form a tripeptide with both the N-terminal and the carboxyl terminal of the peptide so protected; (iv) removing the protecting group from the carboxyl terminal of the tripeptide of C (iii) above; and (v) reacting the tripeptide of C (iv) above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired tripeptide with the required R protecting group at the N-terminal of the tripeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tripeptide, or, alternatively: (vi) reacting an amino acid such as valine with a source of a protecting group such as an alcohol, exempli¬ fied by methanol, to form an amino acid protected at its carboxyl terminal , exemplified by valine methyl ester; (vii) reacting a dipeptide such as valylvaline with a source ofan R protecting group , such as di-t-butyl- σarbonate, to form a dipeptide so protected at its N-terminal, exemplified by t-butyloxycarbonyl- valylvaline; (viii) reacting the product of C (vi) above with the product of C (vii) above to form a tripeptide with both the N-terminal and the carboxyl terminal of the tri¬ peptide so protected; (ix) removing the protecting group from the carboxyl terminal of the tripeptide of C (viii) above; and (x) reacting the tripeptide of- (_ix)_ above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired tripeptide with the required R- protecting group at the N-terminal of the tripeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tripeptide, or the preparation of compounds when n is 4: (.i) reacting a tripeptide such as valylvalylvaline with a source of a protecting group such as an alcohol, exemplified by methanol, to form a tri¬ peptide protected at its carboxyl terminal, exemplified by valylvalylvaline methyl ester; (ii) reacting an amino acid such as valine with a source of an R, protecting group such as di-t- butylcarbonate, to form an amino acid so pro¬ tected at its N-terminal, exemplified by t-butyl¬ oxycarbonyl yaline; (iii) reacting the product of (.i) above with the product of D (.ii) above to form a tetrapeptide with both the N-terminal and the carboxyl terminal of the tetrapeptide so protected; (iv) removing the protecting group from the carboxyl terminal of the tetrapeptide of D (.iii) above; and (v) reacting the tetrapeptide of D Civ), above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired tetrapeptide with the required R, protecting group at the N-terminal of the tetrapeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tetrapeptide, or, alternatively: (vi) reacting an amino acid such as valine with a source of a protecting group such as an alcohol, exemplified by methanol, to form an amino acid protected at its carboxyl terminal, exemplifed by valine methyl ester; (vii) reacting a tripeptide such as valylvalylvaline with a source of an R, protecting group, such as di-t-butyl¬ carbonate, to form a tripeptide so- protected at. its N-terminal, exemplified by t-butylox carbony1- valylvalylvaline; (viii) reacting the product of D (vi). above with the product of _ (vii) above to form a tetrapeptide with both the N-terminal and the carboxyl terminal of the tetra¬ peptide so protected; (ix) removing the protecting group from the carboxyl terminal of the tetrapeptide of p_ (rviii) above; and (x) reacting the tetrapeptide of D (.ix). above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired tetrapeptide with the required R, protecting group at the N-terminal of the tetrapeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tetrapeptide, or, alternatively: (xi) reacting a dipeptide such as valylvaline with a source of a protecting group such as an alcohol, exemplified by methanol, to form a dipeptide protected at its carboxyl terminal, exemplified by valylvaline methyl ester ? (xii) reacting a dipeptide such as valylvaline with a source of an R. protecting group such as di-t- butylcarbonate, to form a dipeptide so protected at its N-terminal, exemplified by t-butyloxycar¬ bonyl valylvaline; - 1. - (xiii) reacting the product of D (xi). above with the product of D Ixii) above to form a tetrapeptide with both the N-terminal and the carboxyl terminal of the peptide so protected; (xiv) removing the protecting group from the carboxyl terminal of the tetrapeptide of D Cxiii) above; and (,xv) reacting the tetrapeptide of D (jxiv). above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired _tetrapeptide with the required R, protecting group at the N-terminal of the tetrapeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tetrapeptide. In carrying out the methods set out immediately above, the hydrophobic carbon chain of 8 to 16 carbon atoms is pre¬ ferably an alkylamine or alkenylamine of 8 to 16 carbon atoms. The compounds according to the invention are initially obtained either* as such or as their salts, depend¬ ing on the nature of the startingcompounds and depending on the reaction conditions. Moreover, salts are obtained by dissolving the free compounds in a suitable solvent, for example in a chlorinated hydrocarbon, such as ethylene chloride or chloroform, or a low-molecular aliphatic alcohol (ethanol or isopropanol) , which contains the desired acid or base, or to which the desired acid or base is subsequently added, if necessary in the precisely calculated stoichiometric amount. The salts are isolated by filtration, reprecipi- tation or precipitation or by evaporation of the solvent. Resulting salts can be converted into the free compounds by treatment with bases or acids, for example, with aqueous sodium bicarbonate or with dilute hydrochloric acid, and the compounds can in turn be converted into their salts. By this means, the compounds can be purified, or ' physiologically unacceptable salts can be converted into physiologically acceptable salts. PRACTICAL EMBODIMENTS OF THE INVENTION The following nσn-limitative practical examples are provided as illustrative of the preparation of the compounds of the invention: Example 1 Preparation of t-butyloxycarbonyl-L-valyl-L-valylamido-dodecane (a) Valine methyl ester hydrochloride CVal-OMe HCl) Valine (11.7g, 0.1 mole) was suspended in 100ml dry MeOH in a condenser to which was added dropwise thionyl chloride (15ml, 0.2 mole) . The solution was refluxed for 6 hrs and the solvent evaporated to yield a white solid in 98% yield (.13.lg) mp. 174-175°C. " 4. nmr (.DMSO-d.) δ: C-α-Val (1H, 3.9, q) , C-β-Val (1H, 2.4, m) , C-γ-Val (6H, 1.0, d) , OCH 3 (.3H, 3.9, s) . (b) t-Butyloxycarbonyl-L-valine (Boc-val) Valine (11.7g, 0.1 mole), was dissolved in 300ml of dioxane/ water (2:1) and 100ml 1M NaOH. The solution was cooled to 5 C and di-t-butylcarbonate (.24g, 0.11 mole) was added over a period of 10 min. The solution was then stirred at room temperature for 3hrs. The dioxane was evaporated and the aqueous solution was ice- cooled and mixed with 75ml ethyl acetate. The mixture was acidified to pH 2-3 with 0.5M KHS0 4 . The organic layer was separated and the aqueous layer was extracted with a further 2 x 60ml ethyl acetate. The combined organic layer was dried (Na 2 ≤0 4 ) and the solvent evaporated to yield a light yellow oil. The oil was left at 4°C to crystallize in 76% yield (16.5g) mp. 76-78°C. 1 H nmr (DMSO-d-) δ: C-α-Val (1H, 3.9, q) , C-β-Val (1H, 2.4, m) , C-γ-Val (.6K, 1.0, q) , (CH 3 )- (9H, 1.4, s) . - IB .- (c) t-Butyloxycarbonyl-L-valyl-L-valine-methyl ester (Boc- val-val-OMe) Boc-val (Ig, 4.6 mmole) was dissolved in 20ml dry THF and triethyl amine (0.7ml, 5.0 mmole) . The solution was cooled to -10 C with stirring before the addition of ethyl chloro- for ate (0.48ml, 5.0 mmole) . The solution was kept at -10 C for 20 min with continuous stirring. To this, solution was added, dropwise, a solution of Val-OMe (0.655g, 5.0 mmole) in 15ml THF, 2ml H 2 0 and triethyl amine (.0.7ml, 5.0 mmole) . The solution was then stirred for 15 hrs at room temperature. The solution was then placed in a separating funnel and 70ml ethyl acetate and 25ml H-O were added. The organic layer was collected and the aqueous layer was extracted with a further 2 x 40ml of ethyl acetate, The combined organic layers were then washed with 30ml of 1M HC1, H 2 0, sat.NaHCO, and H 2 0, dried (Na-SO and the solvent evaporated to yield a white solid, 85% yield (1.29g) mp. 132-134°C. 1 H nmr (DMSO-d g ) δ: C-α-Val (2H, 3.9, m) , C-β-Val (2H, 2.5, m) , C-γ-Val (12H, 0.9, d) , 0-CH 3 (.3H, 3,9, s). , (CH 3 ) 3 (.9H, 1,4, s) , NH-1 (1H, 7.0, broad) , NH-2 (1H, 8.2, broad) . (d) t-Butyloxycarbonyl-L-valyl-L-valine (.Boc^-val-val-OH) Boc-val-val-OMe (1.19g, 3.6 mmole) was dissolved in 30ml dioxane. With stirring at room temperature was added 10ml 1M NaOH, dropwise. The solution was then stirred for 5 hrs at room temperature. The solution was acidified with con¬ centrated HCl and extracted with 3 x 40ml of ethyl acetate, dried (Na 2 S0 4 ) and the solvent evaporated to yield a white solid. 95% yield (1.09g) mp. 144-147°C, X H nmr (DMSO-d-) δ: C-α-Val (2H, 4.0, m) , C-β-Val (2H, 2.4, m) , C-γ-Val (12H, 0.9, d) , (CH 3 ) 3 (.9H, 1.4, s) , COOH (lH, 11.2, broad) , NH-1 (1H, 6.9, broad) , NH-2 (1H, 7.8, broad) . (e) t-Buty oxycarbonyl-L-valyl-L-valylamido dodecane (.Boc- val-val-NH (CH 2 ) 11 CH 3 ) Boc-val-val-OH (0.3g, 0.95 mmole) was dissolved in 20ml dry THF and triethyl amine (.0.14ml, 1.0 mmole) . The solution was cooled to -10°C before ethyl chloroformate (.0.1ml, 1.0 mmole) was added. The solution was then stirred for 20 min at -10 C before a solution of dodecyl amine (0.1S5g, 1.0 mmole). dissolved in 20ml THF and triethylamine (,0.14ml, 1.0 mmole) was added dropwise. The solution was then stirred at room temperature for 15 hrs. The solvent was then evaporated to yield a white solid. This was dissolved in 75ml ethyl acetate and 25ml 5% NaHCO., . The organic layer was washed with 30ml H_0, 1M HCl and H 2 0, dried ( a 2 S0 4 l and the solvent evaporated to yield a white solid. Recrystallized ffirom ethyl acetate/hexane. 94% yield (.0.43g) mp.l08-110°C, 1. H nmr (DMSO-d-) δ : C-α-Val (.2H, 4.0, m). , C-8-Val (.2H, 2.5, m) , C-γ-Val (12H, 0.9, d) , ICH 3 ) 3 (,9H, 1.4, s) , C-l (.2H, 3.0, t) , CH 2 's (20H, 1.3, s) , C-T (.3H, 1,0, s) , NH-1 (.lH, 6.9, broad)., NH-2 (2H, 7.8, broad) , analysis: C-_H 53 N 3 0 4 requires C, 67.04; H, 11.04; N, 8.69; found: C, 67, 21; H, 11.22; N, 8.66. Example 2 Preparation of t-Butyloxycarbonyl-L-alanylamido dodecane (t-Boc- ala-NH(CH 2 ) ιL CH 3 ) T-Boσ-ala-OH(0.95g, 5 mmoles) , and dodecylamine (.2,3g, 15 mmoles) , was stirred at -5 C for 8 minutes (.activation time) in THF (12.5 mis) . After allowing the solution to warm to room temperature, the solvent was removed under vacuum. To the white crystalline solid was added ethyl acetate (,75mlsl and 5% NaHCO.- (25mls) . Following thorough shaking and then separation, the ethyl acetate layer was washed with water (.25mls) , IM.HCl(25mls) , and finally with water (25mls) . After drying over anhydrous Na 2 S0 4 f eva P oration -of the ethyl acetate yielded a white cry¬ stalline solid (1.25g, 70%) mp 67-68°C. Recrystallizing solvent: ethyl acetate/hexane (1:3), 'Hnmr(CDC1.-.) δ : C-α-Ala (1H, 4 J.q) , t-CH 3 (9H, 1.44s) C-1 (2H, 3.29) , CH 2 's (20H, 1.33, broad s) , C-T (3H, 0.9t), N-H(1H, 5.3d, 6.5 broad) , IR(KBr) (cm "1 ) N-Hstr (3330, 3350), Amide I (1600, 1695) , Amide II (.1525) , C-N (1260), C-Hstr (2900-3000), C-13nmr (CDCI3) ppm C-α-Ala (49.93), C-β-Ala (18.59) , CONH (172.64, 173.54) , C-0 (.155.47) C-1 (31.78), C-T (.13.97), CH 2 's (29.52) . Anal. Calc. C--H--N-Q- required C, 67.56, H, 11.06, N, 7.87; found C, 67.54, H, 11.40, N, 7.65. Example 3 Preparation of compounds with benzyloxy as the protecting group (a) Benzyloxycarbonyl-L-alanylamido dodecane (jZ-ala-NH(.CH-), , CiTJJ Z-ala-OH (l.lg, 5 mmoles) and dodecylamine (2.8g, 15 mmoles) , under the reaction conditions of Example 2, yielded a white crystalline solid (1.5g, 77%), mp 78-80°C. Recrystallizing solvent: ethyl acetate/hexane 'Hnmr(CDC1.,) δ : C-α-Ala (1H, 4.1 q) , C-β-Ala (3H, 1.43 d) C-1 (2H, 3.2 t) , CH- *s (20H, 1.38 broad s) , C-T (3H, 0.96 t) , CH- (2H, 5.1 s). , phenyl (H) (.5H, 7.3 s) , IR(KBr) (cm "1 ) N-Hstr (3300) , C-Hstr (.2900-3000) , Amide I (1655, 1700) Amide II (.1550) , C-N (1270) C-13nmr (CDCl-)ppm C-α-Ala (50.51) , C-β-Ala (18.77) , CONH (172.23, 171.85), C-0 (156.03), phenyl (.C) (128.47, 128.13, 127.91, 136.21), HN-C1 (66.86), CT (.14.05) CH 2 's (29.59) , (b) Benzyloxycarbonyl-L-valyl-L-alanylamido dodecane (,Z-val- ala-NH (CH-J-^CH- Z-val-ala-OH (0.6g, 3.7 mmoles) and dodecylamine (1.05g, 11 moles) , under the reaction conditions described in Example 2, yielded a white crystalline solid (0.35g, 34%) mp 46-48°C. _ 2 χ _ Recrystallizing solvent : ethyl acetate/hexane, 'Hnmr (CDC1 3 ) : C-α-Ala (IH, 4.1 g) , C-α-Val (IH, 4.7 m) , C-γ-Val (6H, 1.0 d, 0.98 d) , CH 2 -0 (2H, 5.07 s) , phenyl (K) (.5H, 7.24 s) , C-1 (2H, 3.2 t) , C-T (3H, 0.93 t) , CH- 's (.20H, 1.33 broad s) , IR(KBr) (cm "1 ) N-Hstr (.3300) , C-Hstr (.2900-3000) , Amide I (1645, 1695, 1750) , Amide II (1540) , C-N (1260) . (c) Benzyloxycarbonyl-L-valylamido dodecane (.Z-val-NH- (CH-)-,-, Z-val-OH (1.26 g, 5 mmoles) and dodecylamine (.2.8 g, 15 mmoles) , under the reaction conditions described in Example 2, yielded a white crystalline solid (1.36 g, 65%) , mp 90-92°C. Recry¬ stallizing solvent : ethyl acetate/hexane. 'Hnmr(.CDCI3) δ : C-α-Val (IH, 4.0 d) , C-β-Val (IH, 2.0m) , C-γ-Val (.6H, 0.98 d, 0.91 d) , CH--0 (2H, 5.0 s), phenyl (H) (,5H, 7.2 s) , C-1 (,2H, 3.16 t) , IR(KBr) Cc "1 ) N-Hstr (.3300) , C-Hstr (2900-2980), Amide I (1650, 1695) , Amide II (.1550) , C-N (.1255) C-13nmr (CDC_3)ppm C-α-Val (60.58) , C-β-Val (31.07) , C-γ-Val (.19.17, 17.98) , phenyl (C) (.128.39, 128.03, 127,31, 140.511, HN-Cl (66.83) , C-T (14.031, CONH (.171.21, 156,49. (d) Benzyloxycarbonyl-L-valyl-L-valylamido dodecane (.Z-val- Z-val-val-OH (0.91g, 2.6 mmoles) , and dodecylamine (1.4g, 7.8 mmoles) , under the reaction conditions described in Example 2, yielded a white crystalline solid (.0.89g, 70%) mp 134-136°C. Recrystallizing solvent : ethyl acetate/hexane 'Hnmr(CDCI3) δ : C-α-Val (IH, 4.0 m) , C-β-Val (,2H, 2.18m) , C-γ-Val (.12H, 0.98 d, 0.91 broad s) , CH--0 (2H, 4.96 s). , phenyl (H) C5H, 7.1s) , C-1 (2H, 3.1 t) , C-T (3H, 0.71 t) , IR(KBr) (cm "1 ) N-Hstr (2995) , C-Hstr (2900-2980) , Amide I (1640, 1695, 1725) , Amide II (1565) , C-N- (1260) . C-13nmr(.CDCl3) ppm C-α-Val(60.58 , 58.91) , C-β-Val (31.12, 31.90) , C-γ-Val (17.89, 18.07, 19.23, 19.35) , phenyl (C) (128.47, 128.08, 127,89, 132,26), CONH (172.06, 171.25, 171.19), C-0 156.69). , NH-C1 C66,951 C-T (14.071, CH 2 's (.29.62) . - Enzyme Inhibition Assays The specific and potent inhibition of HLE by the com¬ pounds defined by the general formula (II above, is seen from the data set out below, especially the data contained in Tables I to IV below, in which the assays were carried out in 0.1M HEPES buffer at 37° and pH = 7.5. The solution contained 0.5M NaCl and 15% dimethylsulfoxide. The total volume of the assay system was lOOOμl, which included 4 g of HLE protein. The molar concen- tration of the enzyme solution was calculated on the basis of 9.85. The activity of the enzyme was monitored using the synthe¬ tic substrate (O.lmM) N-succinyl-L-alanyl-L-alanyl-L-valyl-4- nitroanilide (SAAVN) , Liberation of 4-nitroaniline was monitored at 405nm. The sample cell contained inhibitor and enzyme in buffer, while the reference cell contained only inhibitor and no enzyme. The enzymatic reaction was initiated after 5 min, incubation at 37°C by the addition of substrate to both cells. Thus, as can be seen from Table I, a test compound consisting of an alkyl amide of six (.6) carbon atoms bonded to the carbonyl group of the amino acid valine with the protecting group t-butyloxycarbonyl bound to the N-terminal of valine, affords no inhibition of HLE at a concentration of O.lmM of test compound, similar results being obtained (hut not shown in Table I) when the number of carbon atoms in the alkyl amide chain exceeded sixteen (16) . On the other hand, the remaining compounds listed in Table I with eight (3) to fourteen (.141 carbon atoms in the alkyl amide bound to the carbonyl group of valine, show 100% inhibition of HLE at a concentration of 0.ImM of those compounds. TABLE I % INHIBITION OF HLE 0 . 05mM* 0 . OlmM* t-Boc-val-NH(CH-) -CH- 0 t-Boc-val-NH(CH-) CH- 100 73 0 t-Boc-val-NH(CH-) -CH, 100 89 53 - _ - Inhibitor 0 ,-LmM* 0 -, Q5mM* 0 . OlmM* t-Boc-val-rNH (CH- n H 3 JL O Q 86 50 t-Boc^-val-NH (CH 2 1 13 CH 3 100 88 4 0 t-Boc is tert ^ butyloxycarbonyl; Val is L-valyl; *concentration of inhibitor. As can be seen from Table II, substitution of t- butyloxycarbonyl as the protecting group at the N-terminal of the amino acid valine, by succinyl or methoxysuccinyl, which are not hydrophobic /apolar/bulky groups, results in compounds without any detected inhibitory effect on HLE at a concentration of O.lmM of test compound. TABLE II %INHIBITION OF HLE Inhibitor QMmM* 0.05mM* 0 ;OlmM* t-Boc-val-NH(CH 2 ) i:L CH 3 100 86 50 Sue-Va1-NH(CH-,) χ .^E- 0 MeO-Suc-val-NH(CH ' 2 ) ι;L CH 3 0 t-Boc is tert-bu.tyloxycarbonyl; sue is succinyl; MeO-suc is methoxysuccinyl; val is L-valyl; "Concentration of inhibitor. As can be seen from Table III, the optimum results are obtained when the amino acid is L-valine, although the inhibit¬ ory effect when L-alanine is substituted for L-valine is only marginally lower at a concentration of 0.1 mM of test compound. TABLE III %INHIBITION OF HLE Inhibitor 0.ImM* 0.05mM* 0.OlmM* t-Boc-ala-NH(CH-,)- CH-J 84 40 5 t-Boc-D-ala-NH(,CH 2 ^H ' . 54 27 13 Inhibitor O.lmM* 0.05mM* 0.OlmM* t-Boc-val-NH(CH 2 ) ιL CH 3 100 86 50 t-Boc-D-val-NH(CH-) ιL CH 3 82 42 15 t-Boc-leu-NH(CH 2 )nCH 3 24 t-Boc is tert-butyloxycarbonyl; ala is L-alanyl; D-ala is D-alanyl; val is L-valyl; D-val is D-valyl; leu is L-leucyl; * concentration of inhibitor. Further comparative data of the inhibition of HLE by the compounds of the present invention is set out in Table IV. TABLE IV % INHIBITION OF HLE (at 0.OlmM inhibitor concentration) t-Boc-val-NH(CH 2 ) g CH 3 53 t-Boc-val-NH(CH 2 )nCH 3 50 t-Boc-val-val-NH(CH 2 ) g CH 3 80 t-Boc_val-val-NH(CH-) ι:L CH 3 85 t-Boc-val-val-val-NH(CH 2 ) j _ 1 CH 3 97 t-Boc-ala-pro-val-NH(CH-,)nCH 3 57 t-Boc is tert-butyloxycarbonyl; val is L-valyl; ala is L-alanyl; pro is L-prolyl. As indicated in Table III, O.lmM t-butyloxycarbonyl- L-alanyl-NH(CH-), j CH- gave 84% inhibition of HLE. Comparative i tests with the same concentration of that inhibitor with the enzymes chymotrypsin and trypsin has been found to result in no inhibition of those two (2) enzymes, confirming the specifi¬ city of that inhibitor to HLE, Although variation of the protecting group at the N- terminal of the amino acid or peptide from tert-butyloxy carbonyl is envisaged, the greatest specificity for HLE inhibition in relation to other proteases is shown with t-butyloxycarbonyl as the protecting group. Thus, comparative tests in respect of .the percentage inhibition of each of HLE, PPE, chymotrypsin and trypsin with 0.05mM concentration of t-butyloxycarbonyl-L-alanyl NH(CH 2 )-,-, CH- has been found to show substantial inhibition of HLE and no detectable inhibition of any of PPE, chymotrypsin or trypsin. Moreover, whilst 0.05mM benzyloxy-L-valyl-NH (CH-,)-, , CH_ was found to give about half of the percentage inhibition of HLE obtained with t-butyloxycarbonyl-L-alanyl-NH (CH-) ,,CH- , the percentage inhibition of PPE and chymotrypsin with 0.05mM benzyloxy-L-valyl-NH (CH 2 ) 1 ιCH was nearly half that for HLE, although 0,05mM benzyloxy-L-yalyl-NH (CH-)^CH, was found to have no inhibitory effect on trypsin. Further comparative tests have indicated that the addition of small amounts of metal salts, especially copper salts, to compositions containing the compounds of the present invention, is beneficial as an adjuvant. Inhibitors in accordance with the present invention as set out above have been found not only to be very effective inhibitors of HLE but also to be: (a) substantially non-hydrolysed after incubation with human plasma for one hour at 37°C; and (b) relatively non-toxic, the LD 5Q values of the inhibitors being of the order greater than 3g/Kg of body weight in both mice and rats. Thus, the inhibitor compounds and salts of the invention can be expected to be useful in the treatment of HLE implicated diseases such as arthritis, tumor growth and emphysema. The possibility of inhaling a selected inhibitor compound or salt thereof as an aerosol in the treatment of emphysema, makes the exploitation of that area of use, attractive. The invention thus also relates to a method of treating animals suffering from any of the above-mentioned diseases. The method is characterized in that a thera- peutically active and physiologically acceptable amount of one or more of the inhibitors defined above is administered to the animal. The invention also relates to pharmaceutical/ veterinary compositions which contain one or more of the inhibitors defined by the general formula (I) and/or their physiologically acceptable salts. The pharmaceutical/veterinary compositions are pro¬ duced by processes which are known per se and with which the skilled person is familiar. The physiologically active com¬ pounds according to the invention are used either as such or, preferably, in combination with suitable pharmaceutical auxiliaries, in the form of tablets, dragees, capsules, suppositories, emulsions, suspensions or solutions. The skilled person is familiar with the auxiliaries which are suitable for the desired pharmaceutical formulations, Besides solvents, gelling agents, suppository bases, tableting auxiliaries and other excipients for active ingredients, it is also possible to use, for example, antioxidants, dispersing agents, emulsifiers, anti-foaming agents, flavor correctants, preservatives, -solubilizing agents and colorants. The optimum dosage and method of administration of the active compound required in each particular case can easily be determined by any skilled person. If the compounds according to the invention and/or their salts are to be used for treatment of the above- mentioned diseases, the pharmaceutical formulations, can also contain one or more physiologically active members of other groups of medicaments, such as steroidal and/or non-steroidal anti-inflamatory agents, immunosuppresants, sulfated glycosamines/glyσans and other sulfated carbo¬ hydrates, analgesics and antipyretics. In clinical use, the inhibitor compounds and salts of the present invention may be administered orally, rectally or by injection, for example, by trahsdermaL application for the treatment of arthritis, in the form of pharmaceutical prepara¬ tions comprising at least one of said compounds or salts thereof in association with a pharmaceutically acceptable carrier, which may be a solid or semi-solid or liquid diluent or capsule or aerosol applicator. Usually the active substance will constitute between 0.1 and 99% by weight of a solid/semi- solid/liquid preparation, more particularly, between 0.5 and 20% by weight for preparations intended for injection, and between 2 and 50% by weight for preparations suitable for oral administration. Dosage unit pharmaceutical preparations containing at least one compound or salt thereof in accordance with the invention for oral application, may be prepared by mixing the selected compound or salt with a solid pulverulent carrier such as lactose, saccharose, sorbitol, rαannitol, starches such as potato starch, corn starch or amylopectin, cellulose derivatives, or gelatine, and a lubricant such as magnesium stearate, calcium stearate, polyethylene glycol waxes, then compressed to form tablets. Coated tablets can be prepared by coating the tablets prepared as described above, with a concentrated sugar solution which may contain components such as gum arabic, gelatine, talcum, titanium dioxide, or the tablet can be coated with a lacquer dissolved in a readily volatile organic solvent or mixture of organic solvents. Soft gelatine capsules can be prepared by enclosing the selected compound or salt, mixed with a vegetable oil, in a soft gelatine shell. Hard gelatine capsules may contain the selected compound or salt in admixture with solid, pul- verulent carriers such as lactose, saccharose, sorbitol, mannitol, starches such as potato starch or corn starch or amylopectin, cellulose derivatives or gelatine. Dosage unit preparations for rectal application can be prepared in the form ' of suppositories comprising the active substance in admi__ture with a neutral fatty base, or gelatine rectal capsules comprising the active substance in admixture with vegetable oil or parafin oil. Liquid preparations for oral application can be in the form of syrups"or suspensions, such as solutions con- taining from about 0.2% to about 20% by weight of the selected compound, the balance being sugar and a mixture of ethanol, water, glycerol and propylene glycol. Solutions for parenteral application by injection can be prepared as an aqueous solution of the selected compound or the selected compounds preferably in a concen¬ tration of from about 0.5% to about 10% by weight. These solutions may also contain stabilizing agents and/or buffering agents and may conveniently be provided in various dosage unit ampoules. - _9 - Suitable transdermal daiϊy-dose administration of the selected compounds or salts in accordance with the invention can be 100-500 mg, preferably 200-300 mg, whilst weekly-dose administration can be in dosage of 25-2000 g every 1 to 3 weeks.

Claims: CLAIMS : 1. An amino acid derivative or peptide derivative defined by the general formula (I). : and physiologically acceptable salts thereof, wherein: 5 R is a hydrocarbon-oxycarbonyl group with at least 4 carbon atoms, the hydrocarbon-oxy moiety of which may be substituted or unsubstituted, for protecting the N-terminal of the amino acid or peptide, which protecting group if cleaved in vivo, forms a physiologically innocuous compound; 10 R_ and R„, which may be the same or different and may differ from unit to unit when n is greater than 1, are selected from hydrogen, alkyl of 1 to 10 carbon atoms, substituted alkyl of 1 to 10 carbon atoms in which the substituents are those present in natural or non-natural amino acids -5 provided that they are physiologically innocuous, or R ? and R 3 taken together with the adjacent nitrogen and carbon atoms may form a 5-membered ring with 4 carbon atoms, which ring may optionally be substituted in which the substituents are those present in natural or non- 0 natural amino acids provided that they are physiologically innocuous; R 4 is a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, which if cleaved in vivo, forms a physiologically innocuous compound, said hydrophobic hydrocarbon chain being bonded 5 direct to the carbonyl carbon or to the carbonyl carbon through an oxygen, sulphur or nitrogen hetero atom; and n is an integer 1 to 6. 2. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R 1 is a hydros carbon-oxycarbonyl group of the formula wherein R' is selected from: (i) straight-chain alkyl of 1 to 10 carbon atoms, straight- chain alkenyl of 1 to 10 carbon atoms, straight-chain alkynyl of 1 ' to 10 carbon atoms, each of which is sub¬ stituted with one or more of alkyl of 3 to 10 carbon atoms, alkenyl of 3 to 10 carbon atoms, and alkynyl of 3 to 10 carbon atoms to form a branched-chain; or is substituted with one or more of cycloalkyl of 3 to 10 carbon atoms, cycloalkenyl of 3 to 10 carbon atoms, aryl of 6 to 10 carbon atoms, adamantyl, or heterocyclic radicals, any of which may be substituted; or (ii) branched-chain alkyl of 3 to 10 carbon atoms, branched- chain alkenyl of 3 to 10 carbon atoms, branched-chain alkynyl of 3 to 10 carbon atoms, cycloalkyl of 3 to 10 carbon atoms, cycloalkenyl of 3 to 10 carbon atoms, aryl of 6 to 10 carbon atoms, adamantyl, or heterocyclic radicals, any of which may be substituted. 3. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R, is selected from: (a) a branched-chain alkoxycarbonyl of 3 to 10 carbon atoms; (b) an aryl substituted straight-chain alkoxycarbonyl of 1 to 10 carbon atoms; (c) a cycloalkyloxy carbonyl in which the cycloalkyl moiety has 3 to 10 carbon atoms; (d). an aryloxycarbonyl in which the aryl moiety has 6 to 10 carbon atoms; or (e) a substituted aryloxycarbonyl in which the aryl moiety has 6 to 10 carbon atoms. 4. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R, is selected from t- butyloxycarbonyl, isopentyloxycarbonyl, isohexyloxycarbonyl, benzyloxycarbonyl, cyclohexyloxycarbonyl, phenyloxycarbonyl, tolyloxycarbonyl or xylyloxycarbonyl. 5. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R-, is t-rbutyloxy- carbonyl or benzylσxycarbon l. 6. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R. is t-butyloxy¬ carbonyl, 7. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R. is a hydrophobic hydrocarbon chain of S to 16 carbon atoms bonded through nitrogen to the carbonyl carbon. 8. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R 4 is a secondary amine with a hydrocarbon chain of 1Q to 14 carbon atoms. 9. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R- and R 3 are independently selected from hydrogen, methyl, sopropyl and • isobutyl, or R- and R taken together with the adjacent nitrogen and carbon atoms form the 5-membered ring of proline, 10. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein n is 1 to 4, 11. An amino acid derivative or peptide derivative or salt thereof according to claim 1 wherein R, is t-butyloxy¬ carbonyl or benzyloxycarbonyl; R-, and R_ are independently selected from hydrogen, methyl, isopropyl and isobutyl, or R 2 and R- taken together with the adjacent nitrogen and carbon atoms form the 5-membered ring of proline; R. is a secondary amine with a hydrocarbon chain of 10 to 14 carbon atoms; and n is i to 4. 12. An amino acid derivative or peptide derivative or salt thereof according to claim 11wherein R, is t-butyloxy¬ carbonyl. - 2.3 - 13. An amino acid derivative or peptide derivative of claim 1 selected from the group: N-tert-butyloxycarbonyl-valylamidyl decane,. N-tert-butyloxycarbonyl-valylamidyl dodecane, N-tert-butyloxycarbonyl-valyl-valylamidyl decane, N-tert-butyloxyσarbonyl-valyl-valylamidyl dodecane, N-tert-butyloxycarbonyl-valyl-valyl-valylamidyl decane, N-tert-butyloxycarbonyl-valyl-valyl-valylamidyl dodecane, N-tert-butyloxycarbonyl-alanylamidyl decane, N-tert-butyloxycarbonyl-alanylamidyl dodecane, N-tert-butyloxycarbonyl-alanyl-alanylamidyl decane, N-tert-butyloxycarbonyl-alanyl-alanylamidyl dodecane, N-tert-butyloxycarbonyl-alanyl-alanyl-alanylamidyl decane, N-tert-butyloxycarbonyl-alanyl-alanyl-alanylamidyl dodecane, N-tert-butyloxycarbonyl-alanyl-prolyl-valylamidyl decane, N-tert-butyloxycarbonyl-alanyl-prolyl-valylamidyl dodecane, N-tert-butyloxycarbonyl-alanyl-alanyl-prolyl-valylamidyl decane, N-tert-butyloxycarbonyl-alanyl-alanyl-prolyl-valylamidyl dodecane, or a physiologically acceptable salt of any thereof. 14. An amino acid derivative or peptide derivative of claim 1 which is N-tert-butyloxycarbonylvalylamidyl decane, or N-tert-butyloxycarbonyl-valylamidyl dodecane, or a physiologi¬ cally acceptable salt of either thereof. « 15. An amino acid derivative or peptide derivative of claim 1 which is N-tert-butyloxycarbonyl-valyl-valylamidyl decane, or N-tert-butyloxycarbonyl-valyl-valylamidyl " odecane, or a physiologically acceptable salt of either thereof. 16. An amino acid derivative or peptide derivative of claim 1 which is N-tert-butyloxycarbonyl-valyl-valyl-valylamidyl decane, or N-tert-butyloxycarbonyl-valyl-valyl-valylamidyl dodecane, or a physiologically acceptable salt of either thereof. 17. An amino acid derivative or peptide derivative of claim 1 which is N-tert-butyloxycarbonyl-alanylamidyl decane, or N-tert-butyloxycarbonyl-alanylamidyl dodecane, or a physiologically acceptable salt of either thereof. 18. An amino acid derivative or peptide derivative of claim 1 which is N-tert-butyloxycarbonyl-alanyl-alanylamidyl decane, or N-tert-butyloxycarbonyl-alanyl-alanylamidyl dodecane, or a physiologically acceptable salt of either thereof. 19. An amino acid deriyatiye or peptide derivative of claim 1 which is N-tert-butyloxycarbonyl-alanyl-alanyl- alanylamidyl decane, or N-tert-butyloxycarbonyl-alanyl-alanyl- alanylamidyl dodecane, or a physiologically acceptable salt of either thereof. 20. An amino acid derivative or peptide derivative of claim 1 which is N-tert-butyloxycarbonyl-alanyl-prolyl- valylamidyl decane, or N-tert-butyloxycarbonylralanyl-prolyl- valylamidyl dodecane, or a physiologically acceptable salt of either thereof. 21. An amino acid derivative or peptide derivative of claim 1 which is N-tert-butyloxycarbonyl-alanyl-alanyl-prolyl- valylamidyl decane, or N-tert-butyloxycarbonyl-alanyl-alanyl- prolyl-valylamidyl dodecane, or a physiologically acceptable salt of either thereof. 22. A method of producing an amino acid derivative or peptide derivative defined by the general formula (II: and physiologically acceptable salts thereof, wherein; R, is a hydrocarbon-oxycarbonyl group with at least 4 carbon atoms, the hydrocarbon-oxy moiety of which may be substituted or unsubstituted, for protecting the N-terminal of the amino acid or peptide, which protecting group if cleaved in vivo, forms a physiologically innocuous compound; R- and R, , which may be the same or different and may differ from unit to unit when n is greater than 1, are selected from hydrogen, alkyl of 1 to 10 carbon atoms, substituted alkyl of 1 to 10 carbon atoms in which the substituents are those present in natural or non-natural amino acids provided that they are physiologically innocuous, or R-, and R- taken together with the adjacent nitrogen and carbon atoms may form a 5-membered ring with 4 carbon atoms, which ring may optionally be substituted in which the substituents are those present in natural or non-natural amino acids provided that they are physiologically innocuous; R. is a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, which if cleaved in vivo, forms a physiologically innocuous compound, said hydrophobic hydrocarbon chain being bonded direct to the carbonyl carbon or to the carbonyl carbon through an oxygen, sulphur or nitrogen hetero atom; and n is an integer 1 to 6, said method comprising: A: For the preparation of compounds when n is 1, as set out in the following reaction scheme A: (i) reacting an amino acid of formula (II). with a source of an R 1 protecting group such as di^t-butylcarbonate to form an amino acid of formula (.III) so protected at its N-terminal; and (ii) reacting a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, with the carboxyl group of the amino acid of formula (III) to form the desired compound of formula (LTV) , Reaction^ Scheme A 0 a. H - N - CH C - OH (ii; 1 a source of an R-, protecting group such as di-t-butylcarbonate O - N - CH - C - OH l (III) R, R. a source of an R. hydrophobic hydrocarbon chain of 8-16 carbon atoms such as dodecylamine O R, - N - CH c - R. (IV1 R ^ R-, wherein R. , R 2 , R 3 and 4 are as defined in formula (I) above; and B _ For the preparation of compounds when n is 2 to 6 , as set out in the following reaction scheme B: (i) reacting an amino acid or peptide of formula (V) with a source of a protecting group such as an alcohol, exemplified by methanol, to form an amino acid or peptide of formula (VI). protected at its carboxyl terminal; (ii) reacting an amino acid or peptide of formula (VII) with a source of an R-, protecting group, such as di-t-butyl-carbonate, to form an amino acid or peptide of formula (VIII). so protected at its N- terminal; (iii) reacting the amino acid or peptide of formula (VI) with the amino acid or peptide of formula (VIII) to form a peptide of formula (IX). with both the N- terminal and the carboxyl terminal of the peptide so protected; (iv) removing the protecting group from the carboxyl terminal of the peptide of formula (IX). ; and (v) then reacting the peptide with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired peptide of formula (X) with the required R, protecting group at the N-terminal of the peptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the peptide, Reaction- S'eheme B H (V) a source of an R, protect¬ a source of a protect¬ ing group such as di-t- ing group (R) , such as -if butylcarbonate methanol (VI) removal of the R protecting group, followed by reaction with a source of an 4 hydro¬ phobic hydrocarbon chain of 8-16 carbon atoms such as dodecylamine wherein R-, , R_ , R- and R 4 are as de ined in formula I above, and a and b are integers of 1 to 5 , provided that a+b is no greater than 6 , 23. A method of producing an amino acid derivative or peptide derivative defined by the general formula (IL: and physiologically acceptable salts thereof, wherein: R, is a hydrocarbon-oxycarbonyl group with at least 4 carbon atoms, the hydrocarbon-oxymoiety of which may be substituted or unsubstituted, for protecting the N-terminal of the amino acid or peptide, which protecting group if cleaved in vivo, forms a physiologically innocuous compound; R 2 and R- , which may be the same or different and may differ from unit to unit when n is greater than 1, are selected from hydrogen, alkyl of 1 to 10 carbon atoms, substituted alkyl of 1 to 10 carbon atoms in which the substituents are those present in natural or non-natural amino acids, provided that they are physiologically innocuous, or"R 2 and R, taken together with the adjacent nitrogen and carbon atoms may form a 5-membered ring with 4 carbon atoms, which ring may optionally be substituted in which the substituents are those present in natural or non- natural amino acids provided that they a-re physiologically innocuous; R. is a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, which if cleaved in vivo, forms a physiologically innocuous compound, said hydrophobic hydrocarbon chain being bonded direct to the carbonyl carbon or to the carbonyl carbon through an oxygen, sulphur or nitrogen hetero atom; and n is an integer 1 to 4, said method comprising: _ for the preparation of compounds when n is 1: (i) reacting an amino acid such as alanine or valine with a source of an R-, protecting group such as di-t-butylcarbonate to form an amino acid so protected at its N-terminal, such as t-butyloxy- carbonyl alanine or t-butyloxycarbonyl valine; and (ii) reacting a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, with the carboxyl group of the amino acid of A (i) above to form the desired compound; _____ for the preparation of compounds when n is 2: (i) reacting an amino acid such as valine with a source of a protecting group such as an alcohol, exempli- fied by methanol, to form an amino acid protected at its carboxyl terminal, exemplified by valine methyl ester; (ii) reacting an amino acid such as valine with a source of an R, protecting group, such as di-t-butyl¬ carbonate, to form an amino acid so protected at its N-terminal, exemplified by t-butyloxycarbonyl valine; (iii) reacting the product of B (ii) above with the product of B (iii above to form a dipeptide with both the N-terminal and the carboxyl terminal of the dipeptide so protected; (iv) removing the protecting group from the carboxyl terminal of the dipeptide of B (iii). above; and (v) reacting the dipeptide of B (iv) above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired dipeptide with the required R, protecting group at the N-terminal of the dipeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the dipeptide; C: for the preparation of compounds when n is 3 : (i) reacting a dipeptide such as valylvaline with a source of a protecting group such as an alcohol, exemplified by methanol, to form a dipeptide protected at its carboxyl terminal, exemplified by valylvaline methyl ester; (ϋ) reacting an amino acid such as valine with a source of an R, protecting group such as di-t-butylcarbon¬ ate, to form an amino acid so protected at its N- terminal, exemplified by t-butyloxycarbonyl valine; - 4 - (iii) reacting the product of C (i) aboye with the product of C (ii). above to form a tripeptide with both the N-terminal and the carboxyl terminal of the peptide so protected; (iv) removing the protecting group from the carboxyl terminal of the tripeptide of' C (iii). above; and (v) reacting the tripeptide of C (iv)l above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired tripeptide with the required R- protecting group at the N-terminal of the tripeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tripeptide, or, alternatively: (vi) reacting an amino acid such as valine with a source of a protecting group such as an alcohol, exemplified by methanol, to form an amino acid protected at its carboxyl terminal, exemplified by valine methyl ester; (vii) reacting a dipeptide such as valylvaline with a source of an R, protecting group, such as di-t- butylcarbonat , to form a dipeptide so protected at its N-terminal, exemplified by t-butyloxy- carbonyl-valylvaline; (viii) reacting the product of C (vi). above with the product of C (vii) above to form a tripeptide with both the N-terminal and the carboxyl terminal of the tripeptide so protected; (ix) removing the protecting group from the carboxyl terminal of the tripeptide of C (viii) above; and (x) reacting the tripeptide of C (ix). above with a Q5 hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired tripeptide with the required R, protecting group at the N-terminal of the tripeptide and the required hydrophobic hydrocarbon chain at the O carbonyl carbon of the tripeptide, D: for the preparation of compounds when n is 4: (i) reacting a tripeptide such as valylvalylvaline with a source of a protecting group such as an alcohol, exemplified by methanol, to form a tripeptide protected at its carboxyl terminal, _15 exemplified by valylvalylvaline methyl ester; (ii) reacting an amino acid such as valine with a source of an R, protecting group such as di-t- butyl-carbonate, to form an amino acid so protected at its N-terminal, exemplified by 20 t-butyloxycarbonyl valine; (iii) reacting the product of 13 (i) aboye with the product of D (ii) above to form a tetrapeptide with both the N-terminal and the carboxyl terminal of the tetrapeptide so protected; 15 (iv) removing the protecting group from the carboxyl terminal of the tetrapeptide of ( ii). above; and (v) reacting the tetrapeptide of D (iv). aboye with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired Q tetrapeptide with the required R-, protecting group at the N-terminal of the tetrapeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tetrapeptide, or, alternatively: (vi) reacting an amino acid such as yaline with a source of a protecting group such as an alcohol, ememplified by methanol, to form an amino acid protected at its carboxyl terminal, exemplified by valine methyl ester; (vii) reacting a tripeptide such as valylvalylvaline with a source of an R-, protecting group, such as di-t-butylcarbonate, to form a tripeptide so protected at its N-terminal, exemplified by t- butyloxycarbonyl-valylvalylvaline; (viii) reacting the product of D (vii above with the product of D (vii) above to form a tetrapeptide with both the N-terminal and the carboxyl terminal of the tetrapeptide so protected; (ix) removing the protecting group from the carboxyl terminal of the tetrapeptide of D (viii). above; and (x) reacting the tetrapeptide of D_ (ix). above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired _» tetrapeptide with the required R. protecting group at the N-terminal of the tetrapeptide and the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tetrapeptide, or, alternatively: (xi) reacting a dipeptide such as valylvaline with a source of a protecting group such as an alcohol, exemplified by methanol, to form a dipeptide protected at its carboxyl terminal, exemplified by valylvaline methyl ester; (xii) reacting a dipeptide such as valylvaline with a source of an R, protecting group such as di-t- butylcarbonate, to form a dipeptide so protected at its N-terminal, exemplified by t-butylσxy- carbonyl valylvaline; (xiii) reacting the product of D (xiϊ above with the 70 product of D (xii). above to form a tetrapeptide with both the N-terminal and the carboxyl terminal of the peptide so protected; (xiv) removing the protecting group from the carboxyl terminal of the tetrapeptide of D (xiii) above; and 75 (xv) reacting the tetrapeptide of D (xiv). above with a hydrophobic hydrocarbon chain of 8 to 16 carbon atoms, such as dodecylamine, to form the desired tetrapeptide with the required R-, protecting group at the N-terminal of the tetrapeptide and 30 the required hydrophobic hydrocarbon chain at the carbonyl carbon of the tetrapeptide. 24. The method of claim 23 or 23, wherein the hydrophobic hydrocarbon chain of 8 to 16 carbon atoms is an alkylamine or alkenylamine of 8 to 16 carbon atoms. 25. A pharmaceutical or veterinary composition comprising at least one amino acid derivative or peptide derivative or physiologically acceptable salt thereof according to claim 1 in association with one or more non-toxic acceptable carriers. 26. A pharmaceutical or veterinary composition of claim 25 in which said derivative or salt is formulated into tablet, capsule, pill, powder, granule, suppository, bougie, or other pharmaceutically acceptable form. 2 ~ l . A pharmaceutical or veterinary composition of claim 25 in dosage unit form. 28. A pharmaceutical or veterinary composition of claim 25, which includes an adjuvant amount of a apetal salt. 29. A pharmaceutical or -veterinary composition of claim 28 wherein the metal salt is a copper salt. 30. The use of an effective amount of at least one amino acid derivative or peptide derivative or physiologi¬ cally acceptable salt thereof according to claim 1, or a composition according to claim 25, as an inhibitor for elastase - type enzymes. 31. A method for inhibiting the growth of tumors or the degradation of tissues in a . rthritis or the destruction of lung tissues in emphysema, which comprises administering to an animal suffering from any one of such conditions, an effective amount of at least one amino acid derivative or peptide derivative or physiologically acceptable salt thereof according to claim 1, or a composition according to claim 25.

Inventors: TERNAI BELA, TERNAI, BELA
Applicants: UNIV TROBE, LA TROBE UNIVERSITY
Application year: 1985
Main classification: C07C125
IPC codes: A61K38, C07C67, C07C239, C07C271, C07K1, C07K5, C07K7, C07K14, C12N9