DE2712781A1 - POLYPEPTIDE DERIVATIVES AND PHARMACEUTICAL PREPARATIONS CONTAINING THEM - Google Patents

Description

PATENT LAWYERS 97i? 7fi1

Dlpl.-Infl. P. WIRTH Dr. V. SCH MIED-KOWARZIK Dipl.-Ing. G. DANNENBERG ■ Dr. P. WEINHOLD Dr. D. GUDEL

335024 SIEGFHIEDSTHASSi: 8

TELEPHONE: (089) _{Q0QQ M} q _{NCHEN 4q}

P.P.A. 109-107 Wd / Sh

State of Michigan
Lansing, Michigan
United States

Polypeptide derivatives and pharmaceutical preparations containing them .

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The present invention relates to compositions for treating schizophrenia. In particular, it relates to a method of treating schizophrenia using polypeptides, λχ. -N-acylated and / or carbon-containing esters of polypeptides. Finally, it also relates to polypeptides with 3 to about 8 c * -amino acid molecule parts and the following characteristic structure:

0 0 0

-NH-CH-C-NH-CH-C-NH-CH-Cf I.
CHO- CH

^{1 /} \ / 'n ^ ^CH 3 CHv CH-r CHv (C / 5H- »)

especially the N-acylated ones, which have the T-V-L structure Polypeptides and the corresponding amides, e.g. the N-acylthreonylvalylleucinamides.

Schizophrenia is the most serious of all mental illnesses that the psychiatrist must deal with. The incidence rate is about 1 to 1000, the predisposition frequency about 1 to 100. schizophrenia is the cause of 25% to 50 i "of all admissions in state mental hospitals or psychiatric wards of general hospitals (see Kolb, Modern Clinical Psychiatry, pages 311-312 , 1973).

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'40 *

In most patients, this disease sets in slowly and unobtrusively a; however, acute onset can be observed in a small percentage of patients. The disease is characterized by disturbances in the emotional behavior with dullness and inappropriate language; by Impairment of the mind, such as the inability to form a goal, fragmentation, disorders of association, Blocking, neologism etc .; by withdrawing from the environment and constantly brooding; by the loss of the ability Experiencing pleasure, and most commonly from cognitive disorders that manifest as hallucinations and prejudices manifest.

Unfortunately, schizophrenia tends to begin in their final years of growth in adolescents, or in young adults, and Even the use of anti-psychotic drugs cannot prevent the sufferer any longer or his entire life is less disturbed.

There is much evidence that this disease is a genetically determined condition; but there is no doubt that environmental influences also play a role, especially experiences in early childhood play an essential role. However, there seems to be no doubt that there is a basic metabolic disorder, which is further described below.

Unfortunately, it cannot be proven whether animals also suffer from schizophrenia suffer because you don't talk to an animal

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and in this way can determine if it is schizophrenic. The macaques of the Wisconsin Primate Laboratory, raised without a mother and therefore emotionally neglected, show behavioral disorders (fearfulness, aggressive behavior towards one another, the inability to function sexually) similar to Behavior of schizophrenic patients come very close (Harlow, "The Development of Patterns of Affections," Thomas William Salmon Lectures, New York Academy of Medicine, New York, December 1960). The most severely damaged monkeys have been examined and show a disorder in a cK-2 globulin of their blood described below which is similar to the disorder characteristic of schizophrenics (Beckett, Frohmann et al, "Schizophrenic-like Mechanisms in Monkeys", The American Journal of Psychiatry, 119: 835-842, 1963). The electroencephalogram of these monkeys also showed similar results, especially in the septum of the brain Disorders as already described as typical of schizophrenic patients (Heath, "Electroencephalographic Studies in Isolation-Raised Monkeys with Behavioral Impairment "., Diseases of the Nervous System, 33: 157-163 * 1972).

Animals have therefore mainly been used to detect the presence of abnormal metabolic substances in the blood of schizophrenics To prove the patient. If you give hats an abnormal blood protein in an attempt in which they are rewarded with food for climbing a rope, their performance becomes considerable slowed down (Bergen et al, "Further Experiments with Plasma Proteins from Schizophrenics," Serological Fractions in Schizophrenia, edited by R. E. Heath, p. 67, Paul B. Hoeber,

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Inc., Medical Book Department of Harper & Roy /, New York, 1965).

There is certain evidence that extracts from the limbic system ("limbic system") of the brain of deceased patients produce a simulated schizophrenic response when injected into normal subjects (Garey, "Focal Electroencephalographic Changes Induced by Anti-Septal Antibodies ", Biological Psychiatry, 8: 75-88, '974). The most conclusive Investigations consisted in isolating the crf> from the blood of schizophrenic patients -2-globulin was injected in microliter quantities into the ventricles of rats that had electrodes had been implanted in the median forebrain bundle. Rats with such implanted electrodes reward themselves by pressing a button as often as they can; all other activities are neglected. If you now inject microscopic amounts of the isolated oC-2 globulin into the ventricle, there is a reduction of keystrokes to be observed, which does not occur when taking a comparable substance from the blood of a control person injected. Some of the rats injected with the isolated protein from the blood of schizophrenic patients linger in it a position above the key as if to press it, however, fluctuate and are unable to continue moving; this corresponds to a schizophrenic motor disorder. The release of keypresses is interpreted to mean that the Animal - similar to the schizophrenic patient - has lost the ability to experience pleasure (Caldwell et al, "The Effects

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of the S-Protein on Intracranial Self-Stimulation in the Rat ", Biological Psychiatry, 8: 235-244, 1974).

For many years there has been intensive research into the biochemical causes of schizophrenia and the possibilities of this disease to be treated psychopharmacologically. It was found that a protein with a high molecular weight estimated at about 263,000, made up of about 80 y £ of lipoids, is more active in schizophrenics than in normal people; this protein is all the more active, the more the state of the Schizophrenics worsened. This protein was classified as an oC -2 globulin and called an S protein; please refer Prohman, C.E., Latham, L.K., Beckett, P.G.S., and Gottlieb, J.3., "Biochemical Studies of a Serum Factor in Schizophrenia", Molecular Basis of Some Aspects of Mental Activity, Volume 2, edited by 0. Walaas, pp. 241-255, Academic Press, New York (1967); and Prohman, CE., "Studies on the Plasma Factors in Schizophrenia, "Mind as a Tissue, edited by C. Rupp, pp. 181-195, Hoeber Med. Div., Harper & Row, New York. If this S protein is given to rats, for example, it blocks the pleasure of pleasure stimuli in these rats without to noticeably impair their avoidance reflexes.

Attempts to reduce the activity of the S protein in schizophrenics result in extracts from the brain, among other things e.g. obtained from cattle, from the pineal glands of cattle etc.,

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to find a substance with which to concentrate or Can regulate the activity of the 5-protein. In "Control of the Plasma Factor in Schizophrenia," Recent Advances in Biological Psychiatry, Volume 7, pages 45 to 51, 1964, is described by Prohman et al that an isolated protein from the tissue of animals counteracts the activity of the S protein. This protein is known as the anti-S protein and is found in both human as well as animal tissue. In schizophrenics the S-protein shows an Ot-HeIix-arrangement, while

/ ("random") it in normal people predominantly as randomly arranged Chain or in ß-helix form. The manufacture of therapeutic Quantities of the anti-S protein is extremely difficult and expensive, and its extraction for general medicinal purposes Application is impracticable. In addition, in order to obtain a few milligrams of the anti-S protein from obstacles, you need about 100,000 head of cattle. Thus, the naturally occurring anti-S protein is apparently sufficient at most to treat a few Patients and the cost would be prohibitive. more publishments about this research are: C.E. Prohman, R.E. Arthur, H.S. Yoon and J.S. Gottlieb, "Distribution and Mechanism of Action of the Anti-S-Protein in Human Brain ", Biological Psychiatry, Volume 7, No. 1, pages 53-61, 1973; and CR. Harmison and C.E. Prohman, "Conformational Variation in a Human Plasma Lipoprotein ", Biochemistry, Vol. 11, No. 26, pp. 4485-4493, 1972.

Furthermore, it was found that the extracted anti-S protein is an antigen. Schizophrenia can be downright

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bluffing to be suppressed when looking at a patient the anti-S protein administered for the first time, but the anti-S protein is quickly inactivated by the formation of antibodies; with subsequent administration of the anti-S protein, this can be inactivated occur so quickly that practically no effect of the treatment is noticeable. An attempt was therefore made to use the To break down anti-S protein into fragments that exhibit acceptable activity in the treatment of schizophrenia without being considered Antigen to act.

According to the present invention, digestion of the Anti-S proteins with e.g. pepsin and trypsin to produce small peptides with a molecular weight of less than about 1000 a fraction containing tripeptide obtained which reduces the undesired activity of the S-protein. The tripeptide causes that the S-protein present in oC-helix form in a arbitrarily arranged chain is converted. This tripeptide was named threonyl-valyl-leucine. Try this Synthesizing tripeptide has resulted in a practically pure, i.e. crystalline, form of the tripeptide. It's effective compared to the S-protein of schizophrenics, i.e. it leads to the conversion of the <K -helix form of the S-protein into the arbitrary one arranged shape. However, if the tripeptide is used in experiments to counteract the effect of the S protein in rats, thus it turns out that its effectiveness lasts only a short time; To effectively treat schizophrenia, one would need the tripeptide therefore be administered at short intervals.

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It has also been found that TVL peptides and their derivatives can be used to alleviate schizophrenic symptoms in mammals exhibiting such symptoms. These polypeptide materials contain the residues of at least three C 1-4 amino acids and have the TVL structure listed above. The polypeptide can be an amino acid, ie one terminal amino acid molecule part in acid form and the other terminal amino acid molecule part in crt -amino form (-NHp). However, polypeptide materials are preferably used to eliminate schizophrenic symptoms which have at least one of the amino acid end groups in derivative form, in particular an acyl-substituted <f> -amino end group or a carboxylic acid end group in the form of an amide or ester. Both end groups are preferably substituted in this case, and the acylated amides are particularly preferred. In addition, the polypeptide materials can also be otherwise substituted, and the pharmaceutically usable, non-toxic derivatives, for example the acid salts or basic salts, the polypeptides and their derivatives can also be used according to the invention for the treatment of schizophrenic symptoms.

The T-V-L-containing polypeptides according to the invention are for treatment schizophrenic symptoms because they obviously cause the "in the inner system, e.g. the brain, of the oQ -Helix-Porm of the S-protein present in schizophrenic patients merges into the arbitrarily arranged form. Possess the polypeptides or acylated polypeptides and their esters and amides following structure:

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¹¹ ο 0 0 0

H-fR ^a -C-hr (A) + -NH-CH-C-NH-CK-C-NH-CH-C- (B) -R ^Y
Pt,,

CH

CHOH CH

CH ₅ CH ₅ CH _{5 x} _ ^,

CH,

In this formula, R stands for a saturated aliphatic
Hydrocarbon group such as an alkylene group having 1 to about 20 carbon atoms, preferably a lower alkylene group having e.g. 1 to about 4 carbon atoms; R is a group -0 (R ^e ) in which R ^{e is} hydrogen or a saturated aliphaticche
Hydrocarbon group, for example lkylgruppe an _A, with 1 is to about 20 carbon atoms, preferably a lower alkyl group having for example 1 to about 4 carbon atoms, or an aryl or aralkyl group having 1 to about 4 rings, preferably having from 6 to about 24 carbon atoms, or R ^ is -NR ⁰ R, where R ^c and R ^{d can have the} same or different meanings and represent hydrogen or a lower alkyl group with, for example, 1 to

CD
are about 6 carbon atoms, or R and R are lower

Alkylene groups and, together with the nitrogen atom to which they are attached, form a cyclic structure; A and B are each a Monoiminoacylrest, preferably an egg -Monoiminoacylrest, having 2 to about 10 carbon atoms, and B is hydrolyzable from the (L) component of the essential TVL structure so that the carboxyl function of the (L) component in the patient respectively. \ 7irt is available; ρ is 0 to 2;
t is 0 to about 5 >> x is 0 to about 5; and t plus χ together are also 0 to about 5. If t stands for a number of 2 or more,

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so the individual radicals A can be the same or different, so that (A) + can be, for example, phenylalanylprolyl. In the same way, the individual radicals B can also be the same or different if χ stands for 2 or more. In a preferred group of compounds -R stands for -IiR ⁰ R or at least one of the symbols p, t or χ is a number other than 0. The (T) component in the structural formulas above and in the structural formulas listed below can be incorporated into any in diastereoisomeric form, e.g. as threonyl or allothreonyl, whereby the stereochemistry at the ß-carbon is reversed. Preferably the (T) component is threonyl. The (L) component in the structural formulas above and below can be either the group

-K-CH-C- (commonly referred to as leucyl), which are

is preferred, or in another form, e.g. as group 0

ti

-N-CH-C- as in isoleucyl.

CH-CH,
I ³

CH ₀ -CH

The present invention also encompasses those compounds in which any substituent, e.g. Monoiminoacyl residue on the carboxyl group of the (L) components in the essential T-V-L structure can be hydrolyzed. So that the desired activity is guaranteed, is the substituent

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Usually hydrolyzable under the conditions prevailing in the host for which it is intended. Suitable such hydrolyzable substituent which hydrolyze in the presence of red blood cells at an incubation temperature that of 7 / irtes corresponds to approximately body temperature, for example at about 35 ° to 40 ⁰ C. A simple method for determining whether a substituent hydrolyeierbar or not, consists in that the compound, which has the essential TVL structure and a substituent on the (L) part of the molecule, is incubated in a liver medium at about 37.degree.

The peptide moieties in the compounds according to the invention can have optical activity. Different parts of the peptide molecule can have a form designated as L- or D-. The monoiminoacyl radicals A of the formula II and the essential T-V-L part of the compounds can be in the form L- or D- or as a mixture of these forms, e.g. as a racemate. Since peptides of the form D-, which are substituents, are often attached. hydrolyze the carboxyl functions of the peptides only with difficulty leave in order to maintain the carboxylic acid function, the monoiminoacyl radicals B of the formula II expediently have none optical activity or, if they are optically active, are in the form L-. Compounds are preferred in which the (L) component of the essential T-V-L structure has the form D-.

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Da3 tripeptide described here threonyl-valyl-leucine or
its derivatives, for example the corresponding amides, can be obtained in relatively high purity and be crystalline, for example
in a purity of at least about 90 ° or 95 °, preferably practically 100 °.

Some compounds according to formula II are intermediate products for the production of materials of this formula, which can then be used effectively for the treatment of schizophrenia. For example, a compound in which ρ stands for the number 0 and R stands for hydrogen , an intermediate for the corresponding
Be material in which ρ is 1 and / or R is another
Has significance as hydrogen. Particularly suitable for the treatment of schizophrenia are polypeptides in which ρ stands for a number from 0 to 2, as well as polypeptides which contain at least three peptide or amino acid molecule parts and in which t stands for 0 or more, regardless of whether they are Amino acid or derivative. Polypeptides in which at least one of the symbols p, t or χ stands for a number other than 0, show
a longer duration of activity than the tripeptide TVL in amino acid form.

Preferred acylated polypeptides which can be used as active ingredients or as Intermediate products that can be used for such active ingredients have the following structural formula:

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¹¹¹ O 0 0 0 0 0

HfR ^a -C> fKH-CH-cV NH-CH-C-NH-CH-C-NH-CH-CfNH-CH-C * -R ^{b p} I ₁ ^x IIII ₁

R * CHOH .CH CH ₀ R ¹

CH, CH, CH,, CH

5 y ⁵ / \

CH ₅ CH ₅

Il

In this formula the group fNH-CH-C ·) · is a monoiniinoacyl-

rest and R stands, for example, for hydrogen, a lower alkyl group

having for example 1 to about 4 carbon atoms or a group -CH-OR.

2 3

in which H and R each represent hydrogen, a lower alkyl group with e.g. 1 to about 4 carbon atoms or an aryl or aralkyl groups having 1 to about 4 rings, preferably 6 to about 24 carbon atoms. Such groups can

a b may be substituted, such as in the case of tyrosyl. R, R, p, t and χ have the same meanings as in formula II.

In the compounds represented by formula III, R is part of an oC-amino acid or peptide molecule part or residue, such as in the case of threonyl or allothreonyl, where R is a C-hydroxyethyl group; Leucyl, where R is a 2-methylpropyl group is; Valyl, where R is an isopropyl group; Seryl, where R is a hydroxymethyl group; Isoleucyl, where R is a Is 1-methylpropyl group; Glycyl, where R is hydrogen; or

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₁ 271278]

c <-alanyl, where R is a methyl group; Arginyl, where Ii

NH
is a group C-NH (CH ₂ ), -; Tyrosyl, where R is a

NH ₂

Group is HO-T ^ ^ V-CH ₂ -; and the like. V / enn the R encompassing

Group contains an o <-hydroxy group, it can be converted into a lower Alkoxy or aryloxy group are converted, preferably into a benzyloxy or t-butyloxy group. This can be done during the Synthesis take place in order to protect the hydroxyl function. the Peptide-LIolekülteile, i.e. Ivlonoiminoacylreste attached to the essential T-V-L structure can also be prolyl, arginyl or the like, '' 'they can also be a dicarboxylic acid group be, e.g. a residue of glutamic or aspartic acid.

The group R ^{a of} the compounds described herein can contain up to about 20 carbon atoms; however, it is expediently an alkylene group with up to about 4 carbon atoms.

Q Il

The N-acyl group H {-R -Cf can therefore, for example, be an acetyl, propanoyl, butinoyl, pentanoyl, hexanoyl, heptanoyl, octanoyl, nonanoyl, decanoyl, undecanoyl, dodecanoyl, tridecanoyl, Tetradecanoyl, pentadecanoyl, palmitoyl, heptadecanoyl, stearyl or the like. The ester group R can be methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl, decyl, undecyl, dodecyl, tridecyl, tetradecyl, pentadecyl, Heptadecyl, octadecyl, phenyl, benzyl,

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tertiary butoxycarbonyl, tertiary butyl, dihydro-epi-hydroxy androsteronyl group or the like. Preferably they are

/ one N-acyl and the ester group normal and ^{therefore have 7} straight lines term carbon-to-carbon arrangement. The connection of the above formula or its precursor intermediates contain substituent groups that do not interfere with the desired chemical or pharmacological activity of the material.

Intermediate products from which the active compounds according to the invention can be prepared are, for example, compounds of following formula

^IV 0 0 0

D- (A) _t -NH-CH-C-NH-CH-C-NH-CH-C- (B) _x -E

CHOIR ^{4 .CH}

I / \

CH, CH, CH,

J 5 5

In this formula, t, z, A and B have the meanings given above; D is H or a group protecting the oC -amino group; E is -OH, a terminal group protecting the carboxylic acid group, a metal complex of e.g. zinc, copper, nickel, cobalt, iron, magnesium or aluminum, preferably a metal hydroxide complex, or an acid or base addition salt; R is H, an aliphatic, preferably saturated, hydrocarbon group, e.g., an alkyl group having 1 to about 20 carbon atoms, such as a lower alkyl group having e.g. 1 to 4 carbon atoms, or an araliphatic group or aryl group

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with 1 to about 4 rings, preferably with 6 to about 24 carbon atoms, an acyl group, including the alkanoyl or aralkanoyl groups with 1 to about 20 carbon atoms, preferably a lower acyclic acyl group having 1 to about 4 carbon atoms, a tetrahydropyranyl group, a monovalent one Metal such as an alkali metal such as sodium, or the like. A metal such as zinc, copper, nickel, cobalt, iron, magnesium or aluminum can complex with functions of the polypeptide, e.g., with a carbonyl function.

As protective groups for the (^ -amino groups can be used are: (1) acyl or thioacyl groups having preferably 2 to about 21 carbon atoms, e.g., lower aliphatic acyl groups having 2 to about 7 carbon atoms such as acetyl groups, etc .; lower araliphatic acyl groups having 8 to about 15 carbon atoms such as phthalyl, naphthoyl, benzoyl groups, etc .; Trifluoroacetj4-, Chloroacetyl, £ T-chlorobutyryl, toluenesulfonyl, benzoyleulfonyl, Nitrophenylsulfenyl, tritylsulfenyl, o-nitrophenoxyacetyl groups and the like; (2) protective aromatic urethane groups such as benzyloxycarbonyl and substituted benzyloxycarbonyl groups, e.g. p-chlorobenzyloxycarbonyl, p-nitrobenzyloxycarbonyl, p-bromobenzyloxycarbonyl, p-methoxybenzyloxycarbonyl groups; (3) protective aliphatic urethane groups, such as tert-butyloxycarbonyl, diieopropylmethoxycarbonyl, isopropyloxycarbonyl, Ethoxycarbonyl, allyloxycarbonyl groups; (4) protective cycloalkyl urethane groups such as cyclopentyloxycaibonyl, Cyclohexyloxycarbonyl groups; (5) protective thiourethane groups such as phenylthiocarbonyl groups; (6) protective

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Alkyl and aralkyl groups such as triphenylmethyl (trityl) and Benzyl groups; and (7) trialkyisilane groups, e.g., hmethylailane groups.

The following can be used to protect the carboxylic acid end groups: (1) -OG, where G can be, for example, an aliphatic moiety with preferably 1 to about 20 carbon atoms or an araliphatic moiety with 1 to about 4 rings, preferably 6 to about 24 carbon atoms, and an ester terminated with a carbon-containing radical provides, or acyl groups, for example lower acyclic acyl groups with preferably 1 to about 6 carbon atoms; (2) an amide-yielding puncture, _{e.g. -NH 2} , aminoaliphatic or aminoaraliphatic groups, e.g. monocyclic araliphatic groups, with 1 to about 10 carbon atoms, so that the amine can be primary, secondary or tertiary, e.g. -NR ⁰ R, where R ^c and R have the meanings given above; or (3) an anchoring agent such as the resin carrier -NH-CH-CgH ^ -, the resin carrier -0-CH ₂ -C ₆ H.-,

^C 6 ^H 5

chloromethylated resin, hydroxymethyl resin, Merrifield resin and The like. The aliphatic or araliphatic groups mentioned are preferably hydrocarbyl groups.

The polypeptides according to the invention, for example the compounds of Formula II, can be used to alleviate the symptoms of schizophrenia in humans or mammals, and it it is believed that this is achieved by reducing the activity of the S protein, i.e. by reducing the occurrence of

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least of oC helix porms of the S protein in warm-blooded organisms. These active compounds can relate to the symptoms and manifest forms of schizophrenia, such as anxiety and tension, Manic-depressive psychoses etc. counteract when administered to a living being in a pharmacological manner will. The active ingredients according to the invention can be mixed with a pharmaceutical carrier, e.g., in the digestive tract; parenteral administration is preferred. The active substance can be in solid or liquid form and with a pharmaceutically acceptable carrier as a pesticide, solution, suspension, Emulsion or the like. Be combined. Parenteral administration can be, for example, subcutaneous, intraperitoneal, intravenous, intraarterially, intramuscularly or on similar V / eioe. For oral administration, one active ingredient and one pharmaceutical useful carrier e.g. processed into pills, dragees, tablets, capsules or liquid suspensions. Suitable carriers are solids such as lactose, magnesium stearate, calcium stearate, Starch, terra alba, dicalcium phosphate, sucrose, talc, stearic acid, gelatin, agar-agar, pectin, acacia or the like, and liquids, such as sesame oil, olive oil, water and the like. A coating of, for example, cellulose acetate phthalate can also be used. the compositions according to the invention can contain preservatives, Stabilizers, wetting agents, emulsifying agents, buffers or salts and the like. Contain. The active compounds of the invention or compositions containing these compounds can be used together with other physiologically active materials and / or drugs such as buffering agents, antacids, sedatives,

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Sedatives, anesthetics, hormones, and the like, may be administered, or they may include such materials.

The active compounds can be used in different dosages, depending on the compound chosen The severity of the condition to be treated, the mode of administration, the desired effect on the respective host, e.g. a mammal, and other factors. So the amount of active compound can vary but must be sufficient to make the schizophrenic Significantly reduce symptoms in mammals. Generally the dosage will be at least about 0.01 mg per kg Body weight of the living being to be treated per day (mg / kg / day) up to about 2 mg / kg / day or more, preferably at about 0.1 up to 1 mg / kg / day. Treatment can consist of taking the selected daily dose all at once or in partial amounts administered at regular intervals, e.g., about 2 to 4 doses of about 0.05 to 0.2 mg / kg within a day. the active compound can also be entered as a drug with long-term effect, in which the active ingredient over a longer period of time Period is released in the body. The duration of effectiveness can be extended by converting the polypeptides into organic, predominantly polymeric compounds are incorporated, e.g. in gelatin, polyphloretin phosphate or polyglutamic acid.

The compounds according to the invention, i.e. both those for treatment Compounds suitable for schizophrenic symptoms, as well as their precursor intermediates, can be synthesized from individual

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Nn amino acids or from polypeptides v / ground that provide the desired T-V-L series peptides. In one type of production, amino acid components are in the desired Sequence converted to this peptide structure, e.g. the area function of a first amino acid with the carboxylic acid function a second amino acid is condensed into a dipeptide structure, which is then further formed into the desired structure reacted with 3 to about 8 amino acid units or molecule parts can be. With it the desired combination of amino acid molecule parts Is obtained, it is generally convenient to block the sites of the amino acid that lead to a unwanted side reactions, e.g. the carboxylic acid function of a first amino acid reaction partner an amine function of a molecule of a first or second Amino acid. The blocking can be done in a known manner. Particularly suitable blocking groups for the amine function of the amino acids are the protective Λ-amino groups, e.g. phthalyl, carbobenzyloxy or t-butyloxycarbonyl groups, because they can easily be bound to the amino groups without leading to a racemate formation of the amino acid are among the Reaction conditions are relatively inert and can be easily separated from the amino group without damaging the amino acid. These protecting groups can be used as acid halide, e.g. chloride, Eater or acid anhydride, e.g., mixed acid anhydride, can be reacted with alkyl formate, such as a lower alkyl formate, of the blocking group. The reaction can take place at room temperature in an inert, liquid organic solvent, e.g.

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Benzene. If desired, higher or lower temperatures, e.g. 3. about 0 ° to 50 ⁰ C, can be worked. The carboxylic acid function of the amino acid can be blocked by reaction with the amino group of the amino acid which is in front of it in the peptide series, or - if the acid is at the beginning of the series - with another suitable blocking group.

So that the amino acid synthesis product can easily be obtained after each reaction stage, it can be useful that the at the beginning or at the end of the amino acid moiety is a resin ester or resin amide. Particularly suitable resins are, for example, benzhydrilamine resin, chloromethyl resin, Merrifield resin and hydroxymethyl resin. The manufacture of benzhydrilamine resin is made by Rivallile et al in HeIv. 54, 2772 (1971) and the preparation of a hydroxymethyl resin by Bodanszky et al in Cnem. Ind. (London) 58, 1597-98 (1968)

The blocking groups must be present in the reagent and under the reaction conditions used to prepare the peptide be stable. Blocking groups for the terminal carboxylic acid group must be stable under the conditions that occur during the removal of the protective </, amino group in each stage of the synthesis to rule. The blocking group should retain its protective properties under the synthesis conditions, i.e. not to be cleaved, and the reaction conditions to be applied to the blocking group upon completion

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To remove the synthesis, the peptide chain must not change in an undesirable .'eise. The blocking groups can be removed by known methods depending on the type of blocking group in question, e.g. in presence of trifluoroacetic acid, by hydrogenolysis with e.g. hydrogen and a catalyst such as platinum, palladium or the like, with hydrogen bromide in glacial acetic acid or trifluoroacetic acid or with anhydrous hydrofluoric acid. The medium for the Hydrogenolysis is preferably practically anhydrous.

The reaction between an existing peptide unit and a subsequent amino acid can be carried out in the solid phase. In the production of resin esters or amides, the resin carrier and amino acid can be intimately mixed in an inert organic solvent such as tetrahydrofuran, dioxane, dimethylformamide, benzene, ethanol or the like. The reaction occurs at room temperature, but, if appropriate, higher or lower temperatures can be applied, for example, between about 0 ° to 8O ⁰ C. In general, the reaction will be allowed to proceed substantially to completion in order to obtain the maximum yield of peptide in the desired order. A particularly complete reaction is achieved by employing substantial excesses of amino acid reactants, for example at least about 1.5 to about 200 times the stoichiometric amount or more, and operating with long reaction times, for example at least about 1 hour and preferably at least about 5 hours to 500 hours or longer. To what extent the

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Reaction has progressed, can through the Kaiser color reaction tea to be determined. In this test, a positive reaction indicates that the amino acid has unsubstituted sites. The reaction can also be terminated by Dorman titration be determined.

Suitable binding processes for producing the polypeptide chain according to the invention are already known. In general, the amino acid and / or peptide fragments are bound to one another by reacting, for example, an amino acid or a peptide having a protected C ^ amino group and a terminal carboxyl group with an amino acid or a peptide in which a free cC amino group and a protected carboxyl group is present; or an amino acid or a peptide with an active oC -amino group and a protected carboxylic acid end group is reacted with an amino acid or a peptide with a free carboxylic acid end group and a protected <A -amino group eie converts into an acid azide, anhydride or imidazolide or into an activated ester such as cyanoethyl ester, thiophenyl ester, p-nitrothiophenyl ester, thiocreeyl, p-methanesulfonylphenyl, p-nitrophenyl, 2,4-dinitrophenyl, 2,4, 5- or 2,4,6-trichlorophenyl, pentachlorophenyl, N-hydroxysuccinimide, N-hydroxyphthalimide, 8-hydroxyquinoline, N-hydroxypiperidine ester, or by reaction with a carbodiimide (optionally with the addition of N-hydroxysuccinimide) or Ν, Ν'-carbonyldiimidazole or an isoxazolium salt, see Woodward reagent; the amino group

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can e.g. be activated by reaction with phosphite. Frequently For example, the carbodiimide process, the 'tfeygand-Wuensch process, is used (Carbodiimide in the presence of N-hydroxysuccinimide), the azide process, a process using activated esters, the anhydride process and the Merrifield process and methods using N-carboxy anhydrides or N-thiocarboxy anhydrides.

It is expedient to work with the carbodiimide process; as a coupling agent, for example, N-ethyl-N '- (^ - diaethylaminopropylcarbodiimide) or a dihydrocarbylcarbodiinide, z.3. having a dialkyl group of 1 to about 20 carbon atoms such as dicyclohexylcarbodiimide can be used. The carbodiimide can be placed in an inert solvent such as methylene chloride to facilitate its handling. Most of the time, the molar ratio of carbodiimide to the lowest concentration of amino acid reactants is about 0.9: 1 to 10: 1 Amount, for example at least 1 ecm to about 100 ecm © which is more per gram of amino acid, used. The reaction takes place at room temperature, but can also be carried out at higher or lower temperatures, for example between about -3O ⁰ C and 50 ⁰ C «

The blocking of the amino acid or peptide fragment can for further reaction in any way and by means of known

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Proceedings are repealed. Appropriately be Blocking agents to amino groups, e.g. acyl blocking groups, removed in a two-step process in which in a hydrogen halide such as hydrogen chloride is used in both stages. The product thus obtained is an acid addition salt, E.g. a hydrogen chloride salt, which with a base such as triethylamine, pyridine, sodium or Kaliuir.hydroxyd, Sodium or potassium carbonate or the like, neutralizes earth can. The hydrogen halide is used by passing it through a liquid medium such as dioxane or ethylene chloride, lets rise. A rosin ester can be converted to an unblocked peptide by adding hydrogen halide, such as e.g. hydrogen bromide, passed in gaseous form through a medium containing the peptide arzester and trifluoroacetic acid.

'. As stated earlier, the polypeptide that makes the T-V-L bond has, suitably modified to a compound that is effective in the treatment of schizophrenic symptoms for a long time is as the amino acid tripeptide; the modified polypeptide can then be converted to a jflL-N-acylated polypeptide. Known acylation processes can be used for this purpose; A process that does not impair the optical activity of the araic acid moieties may be particularly desirable. the oC-amino group of the polypeptide can be carried out in a known manner Reaction with the corresponding acid halide or an acid anhydride or the like. Acylated. The carboxylic acid or Kster-] Ondgruppen can be esterified by means of known processes or be transesterified to produce a peptide having a

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has greater effectiveness. The esterification can take place by reacting the polypeptide, which has a non-blocked, terminal carboxylic acid function, with the corresponding alcohol. Amides with the characteristic TVL bond of the compositions according to the invention can also be produced by known processes. For example, the corresponding acid or the acid halide, for example the chloride, of the peptide can be reacted with ammonia or a primary or secondary amine. Ammonia or amine are usually used in at least the stoichiometric amount that is required for complete conversion with the peptide structure. The reaction can be carried out in an inert solvent, such as tetrahydrofuran, and conveniently at a temperature of about 10 to 50 ^° C. or more. In general, it can be said that the higher the molecular weight of the acyl and ester groups on the polypeptide, the slower and longer the effect of a polypeptide in the treatment of schizophrenic symptoms.

Suitable derivatives of the compounds according to the invention and intermediates are the pharmaceutically acceptable salts of the polypeptides, including the acid and base addition salts. The acid addition salts are obtained by reacting the polypeptide with an organic or inorganic acid, e.g. with hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid, Acetic acid, maleic acid, monoic acid, citric acid, malic acid, ascorbic acid, benzoic acid and the like. The compounds

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with the T-V-L bond can also be present as metal complexes, which were made by mixing the polypeptides with the moderately soluble Salt, hydroxide or oxide of a metal in contact

/ b? w. Brought cations. Suitable metals / are e.g. cobalt, copper, calcium, Iron, zinc, magnesium, sodium, potassium and ammonium. Likewise nickel and aluminum can be used. For example, you can make a metal complex by adding the polypeptide and a moderately soluble metal salt, hydroxide or oxide in one aqueous medium, or by adding an alkaline medium to an aqueous solution of the polypeptide and a practically insoluble one Metal salt gives and forms an insoluble polypeptide-metal hydroxide complex. An insoluble polypeptide-metal salt complex can also be prepared in situ by adding an aqueous alkaline medium with the polypeptide and charged with a metal salt.

The following examples illustrate the present invention. Unless otherwise stated, all peptide molecular parts are the show an optical activity, present in the form L-.

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Example 1: Preparation of leucine benzyl ester p-toluenesulfonate In a 200 ccm three-necked flask fitted with a Dean-Stark trap, reflux condenser (with attached drying tube) and mechanical lifting device, 15 g (0.0789 UoI) of p- Toliolsulfonic acid monohydrate heated under reflux until one equivalent of water had been released. Then 10 g (0.07634 mol) of leucine and 17.0 g (0.15 mol) of benzyl alcohol were added. The mixture was refluxed for 4 hours and then allowed to cool to room temperature. The solvents were removed in vacuo and a white solid residue was obtained. The white plague was washed with anhydrous ether and recrystallized from ethanol / ether, 28.2 g of leucine benzyl ester p-toluenesulfonate being obtained in three fractions; Yield = 94 i °; F = 157-158 ⁰ C (with decomposition).

Example 2: Preparation of Benzyl-y-oC-t-butyloxycarbonylvalylleucinate

About 13.2 grams (0.0336 moles) of benzyl leucinate p-toluenesulfonate was treated with sodium bicarbonate; the benzylleucinate thus obtained mirde obtained by methylene chloride extraction. In a 200 ccm three-necked flask equipped with inlet and outlet adapters for nitrogen gas and a magnetic stirrer, 6.25 g (0 _f 34 WoI) of dicyclohexylcarbodiimide were dissolved in 30 ecm of freshly distilled, dry CHpCIp, and then the benzyl leucinate was added. The mixture was kept under a Stickotoffatmophäre and cooled with the aid of dry ice and carbon tetrachloride to about -30 ⁰ C. Well were

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7 »45 g (0.0336 mol) of N-cK-t-butyloxycarbonylvaline, dissolved in 50 ecm of dry CHpGIP», were added dropwise to the mixture. The mixture was stored for 2 days at -10 ^0C. The white precipitate which had formed during this time was filtered off and the methylene chloride layer was washed successively with 25% aqueous acetic acid, aqueous sodium bicarbonate solution, water and a saturated aqueous sodium chloride solution. The organic phase was then dried over anhydrous MgSOi and the solvent removed in vacuo. The residue was taken up in ethyl acetate, filtered and concentrated again in vacuo. The white pesticide obtained in this way was recrystallized from ethanol / hexane and yielded 9 »17 g of benzyl No ^ -t-butyloxycarbonylvalylleucinate in two fractions. Yield = 65 *; P = 90-91 ⁰ C

Example 3: Preparation of Benzyl-N-c <-t-butyloxycarbonyl-O-benzylthreonylvalylleucinate

In a 100 cc three-necked flask equipped with inlet and outlet adapters for gas and a magnetic stirrer, 4.08 g (0.00972 mol) of benzyl-No <-t-butyloxycarbonylvalyl leucinate were dissolved in 50 ecm of glacial acetic acid. The mixture was ^{cooled to 5 °} C., whereupon hydrogen chloride gas was bubbled through the mixture for 30 minutes. The mixture was then dried in vacuo. The white, powdery benzylvalylleucine hydrochloride obtained in this way was washed several times with anhydrous ether. Then, the powder in 30 cc of dry CH ₂ Cl ₂ was suspended and cooled to -3O ⁰ C, with 1.38 cc (0.010 mol) of triethylamine and treated stirred for 30 minutes to a solution.

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The triethylamine solution thus obtained was added to 1.94 g (0.010 mol) of dicyclohexylcarbodiimide, dissolved in 30 ecm of freshly distilled, dry CHpClp ». This solution was kept under a nitrogen atmosphere, and then 3.0 g (0.00972 mol) of N-ot-t-butyloxycarbonyl-O-benzylthreonine, dissolved in 30 ecm of dry CHpClp, were added dropwise. The reaction mixture was allowed to stand for two days at -1O ⁰ C. The white precipitate was then filtered off and the methylene chloride layer was washed successively with water, 25% aqueous acetic acid, aqueous sodium bicarbonate solution, water and saturated aqueous salt solution. The organic layer was dried over anhydrous MgSO4 and the solvent removed in vacuo. The residue was taken up in ethyl acetate, filtered and concentrated in vacuo to give 3.56 g of benzyl N-oi-t-butyloxycarbonyl-O-benzylthreonylvalyl leucinate in the form of a white powder. Yield = 60 i '\ F = 115-119 ° C.

Example 4: Preparation of Benzyl-O-benzylthreonylvalylleucinate hydrochloride

Hydrogen chloride gas was slowly passed through a ^{stirred solution, cooled to 5 °} C., of 4.1 g (6.71 mol) of benzyl-N-ot-t-butyloxycarbonyl-O-benzylthreonylvalylleucinate in 50 ecm of glacial acetic acid; the treatment lasted 50 minutes. Then the solvent was removed in vacuo. The residue was washed with ether and recrystallized from ethanol / ether and yielded 3.14 g of benzyl-O-benzylthreonylvalylleucinate hydrochloride. Yield = $ 86; P = 200-202.5 ° C.

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Example 5 For the preparation of threonylvalyl leucine Ea, 1 g of benzyl-O-benzylthreonylvalylleucinate hydrochloride was neutralized to give benzyl-O-benzylthreonylvalylleucinate. A hydrogenolysis mixture was then prepared containing the benzyl O-benzyl threonyl valyl leucinate, 0.5 g of 10% palladium on charcoal per 0.01 mole of the material to be hydrogenated, and absolute ethanol. 1.1 molar equivalents of acetic acid, based on the benzyl ester, were added to this mixture. The mixture was hydrogenated for 4 hours. The catalyst was then filtered off and the solvents removed in vacuo. The residue was washed with anhydrous ether and recrystallized from ethanol / ether to give 544 mg of the free peptide threonylvalylleucine; Yield = $ 85>

Example 6; Preparation of Benzyl-N-ot-acetyl-O-benzylthreonylvalylleucinate

Following the procedure of Example 5, benzyl-O-benzylthreonylvalylleucinate was prepared from 1 g of benzyl-O-benzylthreonylvalylleucinate hydrochloride and dissolved in acetic acid, whereupon 2 equivalents of acetic anhydride, based on the benzyl ester, were added. The mixture was stirred overnight. The solvents were then removed in vacuo and the residue was recrystallized from methanol / ether. 806 mg of benzyl-Hc ^ -acetyl-O-benzylthreonylvalylleucinate were obtained; Yield> 85%.

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Example 7? Preparation of No (-acet.ylthreonylvalylleucinamide 1 g of benzyl-N- O ^ -acetyl-O-benzylthreonylvalylleucinate was dissolved in ethanol. Ammonia was bubbled through the solution for 8 hours. After stirring for 48 hours at room temperature, The solvents were removed in vacuo, the residue was washed with ether and yielded 795 mg (99% yield) of No (-acetyl-O-bonzylthreonylvaluucinamide. 1 g of the Nc ^ -acetyl-O-benzylthreonylvaiylleucinamide was then subjected to hydrogenolysis in absolute ethanol , whereby 705 mg of N- oC-acetylthreonylvalylleucinamide were obtained; yield = 90 #.

Example 8; Preparation of Threonylvalylleucine by Merrifield Synthesis

A solution of 5.3 g (21.2 millimoles) of t-butyloxycarbonyi-L-leucine, 2.5 ecm (18 millimoles) of triethylamine in 2-methy! Tetrahydrofuran was prepared and mixed with 10 g of Merrifield resin (a 2 # Divinylbenzene crosslinked polystyrene), which provided 21.2 millimoles of chlorine, was added. The analysis indicated that the Merrifield resin contained approximately 1.06 millimoles of chlorine per gram of resin. The resin was dried prior to use with methanol, distilled tfasser, washed ethanol and methylene chloride and then at 100 ⁰ C in vacuo. Also before use, the 2-methyltetrahydrofuran was distilled over a sodium dispersion and the triethylamine first over phenyl isocyanate and then over a sodium dispersion. The mixture was refluxed for about 70 hours to remove the blocked amino acids with the resin

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to esterify. After the esterification, the resin was washed with tetrahydrofuran, ethanol, glacial acetic acid, ethanol, distilled water, ethanol and methylene chloride and ^{dried in vacuo at 100 °} C. for 3 hours. About 21.5 g to 22 g of t-butyloxycarbonyl-L-leucine resin ester were obtained.

The t-butyloxycarbonyl-L-leucine resin ester was dissolved in 200 ecm of methylene chloride suspended and stirred while very pure nitrogen was bubbled through the suspension. The t-3utyloxycarbonyl protecting group was removed by

400 ecm of a mixture of equal parts by volume of trifluoroacetic acid and added methylene chloride. The resin was washed and with 400 ecm of a 10% strength by volume triethylainin solution in chloroform neutralized. Then the resin was treated with chloroform and methylene chloride washed. About 4.82 grams of t-butyloxycarbonyl-L-valine would be incorporated into the resin and then given an equivalent amount, i.e. about 4.57 g (24 millimoles) of dicyclohexylcarbodiimide, and the coupling reaction was allowed to proceed for at least 16 hours. The t-butyloxycarbonyl-L-valyl-L-leucyl resin ester thus obtained was now washed with methylene chloride, practically anhydrous ethanol, glacial acetic acid, ethanol and methylene chloride to the as Remove by-product obtained dicyclohexylurea. A Kaiser color reaction was used to check whether the reaction had ended.

In practically the same way, the protecting group, the t-butyloxycarbonyl group, was removed. In the no longer blocking

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- Υί-

th resin esters were 500 g of t-butyloxycarbonyl-O-benzyl-L-threonine and then an equivalent amount of dicyclohexylcarbodiimide (3.6 g, 18.4 millimoles) and the coupling reaction was allowed to take place Progress 4 hours. The rosin ester thus obtained was washed and over in the manner described above Dried overnight at room temperature in vacuo over phosphorus pentoxide. About 22 g to 23 g of t-butyloxycarbonyl-O-benzyl-L-threonyl-L-valyl-L-leucyl resin ester were obtained obtain.

The tripeptide was split off from the resin by the dried t-butyloxycarbonyl-O-benzylthreonylvalylleucine resin ester suspended in 100 ecm of 100 # trifluoroacetic acid, through which anhydrous hydrogen bromide was bubbled. In this way became the ester linkage and the protecting group split off the O-benzyl group on the threonine, whereby the soluble hydrobromide and trifluoroacetate salts of threonylvalylleucine in trifluoroacetic acid.

The trifluoroacetic acid solution was then dried by evaporation and the residue is dissolved in 100 ecm of a mixture of equal parts by volume of methanol and distilled water. The mixture was dried under reduced pressure and the Rücketand dissolved in 100 ecm ethanol and dried under reduced Pressure won. The dried material was used in 10

/Lot

Dissolved up to 20 ecm of a minimal amount of 50% glacial acetic acid and filtered. Solid sodium carbonate or sodium bicarbonate was slowly added until a precipitate formed. The finished one

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Tripeptide product was obtained by recrystallized from ethanol / water, or by dissolved nan the product into a large excess of distilled water, the solution concentrated by Abdeetillieren part of the water and at refrigerator temperature, that is about 5 ⁰ C, crystallize left . About 657 mg of L-threonyl-L-valyl-L-leucine were obtained. Yield = 75 #; F = 240-242 ⁰ C (with decomposition).

Example 9: Preparation of L-threonyl-L-valyl-D-leucine The procedure of Example 8 was repeated except that D-leucine was used in place of L-leucine and L-threonyl-L-valyl-D-leucine was obtained. The product was a white crystalline mixture had a decomposition point of about 240 ⁰ C (under charring).

Example 10: Preparation of N-oC-acetylthreonylvalylleucine Following the procedure of Example 8, t-butyloxycarbonyl-O-benzylthreonylvalylleucine resin ester was prepared. The protective t-butyloxycarbonyl group was removed in practically the same manner as the groups on the leucyl resin ester and valyl leucyl resin ester of Example 8, and O-benzylthreonylvalylleucine resin ester was obtained. About 5.0 g of the dried O-benzylthreonylvalylleucine resin ester were acetylated by first washing the resin ester with dimethylformamide and then reacting it with 100 com acetylating agent, which consists of 44 parts by volume of dimethylformamide, 5 parts by volume of acetic anhydride and 1 part by volume Triethylamine consisted; the reaction took 20 to 50 minutes, and the resin was then with dimethylformamide and with

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Washed methylene chloride and dried in vacuo at room temperature overnight.

The N-acetyl peptide was cleaved from the resin according to the procedure of Example 8. The trifluoroacetic acid solution was then dried by evaporation under reduced pressure. The residue was washed with a mixture of equal parts by volume of methanol and water, dried by evaporation under reduced pressure, washed with ethanol and dried again under reduced pressure. The product was recrystallized from ethanol / ether, and 400 mg of N- <k -acetylthreonylvalylleucine were obtained. Yield = 55 Jt; P = 210-214 ° C (with decomposition).

Example 11: Merrifield synthesis of the hydrochloride 3 of glycylthreonylvalylleucine

10 g of the O-benzylthreonylvalylleucine resin ester prepared according to Example 10 were coupled with t-butyloxycarbonylglycine by adding the resin ester to 100 ecm of methylene chloride suspended, 1.94 g (10 millimoles) of t-butyloxycarbonylglycine and an equimolar amount of dicyclohexylcarbodiimide, i.e. 2.06 g (10 millimoles), added and condense the mixture for 4 hours

haree'ster Hess. The ■ t-butyloxycarbonyl-glycyl-O-benga.threonylvalylleucine ^ · was cleaved from the resin in the manner described in Example 8. After washing with methanol, distilled water and ethanol and adding sodium carbonate until the pH was 4.5, the product was dried under reduced pressure.

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The glycylthreonylvalylleucine product was converted to the corresponding hydrochloride by dissolving ea in 100 ecm of a mixture of 3 parts by volume of ethanol per part by volume of ether, through which anhydrous hydrogen chloride gas was bubbled for about 10 minutes. The product was filtered, dried overnight at room temperature in a vacuum dryer to provide 600 mg of the hydrochloride of glycylthreonylvalylleucine. Yield = 60 Ji; P = 225-226 ⁰ C (with decomposition).

Example 12; Merrifield synthesis of N-cA-acetyltyrosylthreonylyaly! Leucine

7.4 g of the 0-benzylthreonylvalylleucine resin ester prepared according to Example 10 were obtained placed in a reaction flask and suspended in 75 ecm of methylene chloride. In this suspension Then 3.735 g (10 millimoles) of t-butyloxycarbonyl-O-benzyltyrosine were added and an equivalent amount of dicyclohexylcarbodiimide (2.06 g, 10 millimoles) is added, followed by the coupling reaction 4 hours was carried out. Agitation was carried out by blowing very pure nitrogen through the suspension. The resin was washed with methylene chloride, ethanol, acetic acid, ethanol and methylene chloride to remove the dicyclohexylurea by-product to remove, and then dried overnight at room temperature in vacuo over phosphorus pentoxide. The weight of the dried Product, the O-benzyltyrosyl-0-benzylthreonylvalylleucine resin ester, typically increased to 7.50 to 7.55 g.

The N-acetylation was carried out in the same way as in Example 10 for the synthesis of N-O ^ -acetylthreonylvalylcucine

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was described, but 7.5 ecm acetic anhydride, 2.5 ecm triethylamine and 50 ecm dimethylformamide were used. After the N-acetylation, the resin was washed with dimethylformamide and then with methylene chloride and dried overnight Dried in vacuo at room temperature.

Cleavage of the peptide from the Merrifield resin was carried out according to the procedure of Example 8; However, in order to prevent a possible electrophilic aromatic substitution of the tyrosine ring by bromine, the hydrogen bromide was first passed through a mixture containing 2 g of resorcinol per 100 ecm of trifluoroacetic acid to remove all traces of B ^. As a further precautionary assumption against electrophilic substitution, 20 ecm of anisole were added directly to the vessel in which the cleavage was carried out. The N-ot-acetyltyrosylthreonylvalylleucine obtained as product was recrystallized from ethanol / ether and yielded 900 mg of the tetrapeptide; Yield = 59 $>', P = 234.5-235 ° C (with decomposition). Amino acid analysis of the product found 0.93 tyrosine, 1.00 threonine, 1.00 valine and 1.00 leucine.

Example 13: Merrifield Synthesis of Threonylvalylisoleucine A solution of 5.3 g (21.2 millimoles) of t-butyloxycarbonyl-L-isoleucine and 2.5 ecm (18 millimoles) of triethylamine in 2-methyltetrahydrofuran was prepared and mixed with 20 g of Merrifield- Hars shifted. Before use, the resin was washed with methanol, distilled water, ethanol, and methylene chloride and added to

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10O ⁰ C dried in vacuo; the 2-methyltetrahydrofuran was distilled over a sodium dispersion and the triethylamine was distilled first over phenyl isocyanate and then over a sodium dispersion. The mixture was refluxed for about 72 hours to esterify the blocked amino acids with the resin. After the esterification, the resin with tetrahydrofuran, ethanol, glacial acetic acid, ethanol, distilled water, and methylene chloride was washed and dried for 3 hours at 10O ⁰ C in vacuo. About 21.5 to 22 g of t-butyloxycarbonyl-L-isoleucine resin ester were obtained. The coupling of t-butyloxycarbonylvaline with the t-butyloxycarbonylisoleucine resin ester was carried out practically in the same way as the coupling of t-butyloxycarbony1-L-valine with the t-butyloxycarbonyl-L-isoleucine resin ester according to Example 8. Because of the even greater steric hindrance, the However, the reaction lasts for 24 hours. The resin was then washed in the usual manner to remove the by-products, and the completion of the reaction was checked by the Kaiser color reaction test. The addition of t-butyloxycarbonylthreonine to produce t-butyloxycarbonylthreonylvalylisoleucine resin ester, the cleavage of the peptides from the resin to obtain t-butyloxycarbonylthreonylvalylisoleucine and the subsequent isolation and purification of the product were also carried out according to the method of Example 8. The tripeptide threonylvalylisoleucine was obtained in an amount of about 600 mg; Yield = 68 36.

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Example 14: Merrifield Synthesis of Threonylvalylleucinylarinine

Following the procedure of Example 8, 6.78 grams (21.2 millimoles) of t-butyloxycarbonylnitro (guanidinyl) -L-arginine as an amino acid was esterified with 20 grams of the Merrifield resin. The esterification was carried out by treating the mixture heated about 72 hours under Hückflusa and Harzester according to Example 8 was washed and dried _v; about 22 to 23 g of t-butyloxycarbonylnitroarginine resin ester were obtained. About 5 g of t-butyloxycarbonyl-L-leucine (24 millimoles) was coupled to the nitroarginine resin ester by adding an equivalent amount of dicyclohexylcarbodiimide (4.6 g, 24 uillimole) and allowing the reaction to run for 16 hours. After the rosin ester was washed, it was coupled according to the procedure of Example 8 with t-butyloxycarbonyl-L-valine and t-butyloxycarbonyl-O-benzyl-L-threonine. There was thus obtained about 24 g of t-butyloxycarbonyl-O-benzyl-L-threonyl-L-valyl-L-leucylarginine resin ester.

Since the nitro group shows good stability for hydrogen bromide, the protected peptide resin ester was cleaved with anhydrous hydrogen fluoride in order to remove the nitro group at the same time. After treatment with a suitable ion exchange resin, the eluate was concentrated to an oil and then isoelectric precipitated by adding solid sodium carbonate. The L-threonyl-L-valyl-L-leucyl-L-arginine obtained as product was recrystallized from an ethanol-water solution and yielded about 900 mg of the tetrapeptide.

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Peptides with the characteristic T-V-L structure can also prepared by reaction in the solid phase with a chloromethylated resin, i.e. a polystyrene crosslinked with divinylbenzene will. In these methods of attaching peptide parts of the molecule for example, a t-butyloxycarbonyl leucine resin ester in dioxane suspended and freed from the blockages using 4N hydrochloric acid. The hydrochloride obtained in this way is with Triethylamine neutralized and washed. In order to couple the additional amino acid parts of the molecule, a coupling agent such as dicyclohexylcarbodiimide is used. However, this process is not as economical as other synthesis processes, e.g. the Merrifield synthesis.

Example 15: Threonylvalylleucinnethylester were added crude Threonylvalylleucin in a 5 cc Heagenzglas and dried overnight at 100 ⁰ C in vacuum over phosphorus pentoxide a desiccant 0.066 g (0.2 mmol). The dried tripeptide was dissolved in 0.54 ecm anhydrous methanol. A previously prepared mixture of dimethoxypropane and hydrogen chloride (reagent quality) in a weight ratio of 542: 831 was added in an amount of 0.63 ecm into the reaction mixture. Then the test tube was tightly closed. The mixture was mixed thoroughly and allowed to stand overnight to form the threonylvalylleucine methyl ester. The solvents were removed by evaporation under a stream of nitrogen, and then the ester was dried overnight at 10O ⁰ C in vacuum with the aid of a phosphoric anhydride.

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Example 16; IJ-O-Di ac e ty lthreonylvaly 11 eucinme thy lest er The product of Example 15 was dissolved in 0.38 ecm of pyridine. As the acylating agent was 0.1 cc (2 millimoles) of acetic anhydride was added and the reaction mixture allowed to stand for 20 minutes at 37 ⁰ C in a sealed Iieagenzglas. The solvent was removed from the aeylierten product by evaporation under a stream of dry nitrogen, after which the product overnight at 100 ⁰ C under vacuum over phosphoric anhydride Einein-drying agent was dried.

The aylated NO-diacetylthreonylvalylleucine methyl ester can be purified by dissolving the crude product in methanol and subjecting the sample to high pressure liquid 3 chromatography. A »yaters modular liquid chromatograph is used for this, which consists of a column 3» 05 m in length and 9.5 mm in diameter filled with “Phenylporasil-B”; methanol is used as the eluting agent. The elution is monitored for changes in the index of refraction and for UV absorption at 225 nm; the flow rate of the methanol eluent is 1 cc / min. The largest maximum with a retention volume of about 100 ecm is collected as a purified fraction. The solvent from this fraction is removed by evaporation under a stream of dry nitrogen, whereupon ^{drying is carried out in vacuo at 100 °} C. and with a phosphoric anhydride drying agent overnight.

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The following examples illustrate the isolation and recovery of the threonylvalylleucine tripeptide from animal sources.

Example 17: Extraction and Isolation of Threonylvalylleucine Containing hindbrain polypeptide

The hypothalami of several hind brains were excised and placed in a buffer solution - hereinafter referred to as tris-citrate buffer denoted - homogenized, which was prepared in the following way:

It became a 0.498 molar aqueous solution of the tris-citrate buffer prepared by adding 9.58 g of citric acid and 49 »9 g Tris (hydroxymethyl) aminomethane in 1 liter of distilled water gave. The pH of the solution thus obtained was adjusted to 8.65 with the aid of a 0.1 molar aqueous solution of sodium hydroxide.

The hypothalami were homogenized in 4 parts by volume (ocm) of Tris-citrate buffer per part by weight (g) of the hypothalami. The homogenized mixture was centrifuged for 20 minutes with 15,000 V / Hin at 4 ⁰ C, and the supernatant fluid containing the extracted anti-S-protein Paktor together with numerous impurities, was collected.

4 kg of starch hydrolyzate were then washed once with distilled water and twice with the tris-citrate buffer. The wet

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Thickness became 123 cm long, 60 cm wide and 5 cm thick block processed. The two ends of the block were held together by filter paper to remove excess buffer could run out of the block. After the liquid had drained off, the block was only about a thickness 1.5 cm.

Across the width of the block, starting at a point 43.5 cm from the end of the block, a 2.5 ca-wide depression was dug. The block was brought to a temperature of 4 ^° C. Then the supernatant liquid obtained by centrifugation (about 35 to 40 ecm in total), which contained the anti-S-protein factor, was mixed with 10 drops of bromophenol blue and then with the starch dug out of the well. This mixture was poured into the 2.5 cm wide well of the block.

Now the block was subjected to an electrophoretic current of 750 volts and 50 milliamps with the cathode attached to the end of the block 43.5 cm from the well was removed, and the anode on the opposite end. The stream was continuously passed through the block until the blue line running across the width of the block (caused by the bromophenol blue) is 42 cm had to relocate the anode. This took about 18 hours. During this time a bright red line (running through the Hemoglobin in the hypothalami extract entotand) about 5 cm from the depression away towards the cathode.

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The after completion of the electrophoresis from the red and blue Lines of bounded area were divided into 10 strips, each strip running the width of the block and approximately Was 5 cm wide. The material within these strips has been separately and completely excavated and each about 10 minutes in 12 ecm of the tris-citrate buffer. Each of the 10 fractions thus obtained was according to the tryptophan uptake method - described in Biological Psychiatry, Volume 7, No. 1, page 53 (1973) - rated that one sign for the amount of Anti-S-Protein Paktora present. The two or three active fractions were retained. Number the strips on the starch block from 1 to 10 and if the numbering begins with the strip next to the blue line, the fractions with the greatest activity will be of the anti-S-protein factor generally from the strips 2, 9 and 10 received.

The procedure described above was repeated until 20 fractions had been obtained which had a relatively high activity of anti-S protein factor. These 20 factions were combined and evaporated through dialysis tubing until their original volume was 100 to 160 ecm had been reduced to about 40 to 60 ecm.

For chromatography of the concentrated solution of the crude anti-S-protein factor the following column was prepared:

* The slurry was then passed through a Buchner funnel under vacuum filtered.

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DEAE cellulose column

800 g of diethylaminoethyl cellulose ("Selectacel", Brown Company, Berlin, New Hampshire) washed twice with 19 l each of an aqueous 0.19 N solution of sodium hydroxide. Afterwards the filler was filled with 19 liters of 0.19 N hydrochloric acid each time washed. Finally it was still with 0.005 molar phosphate buffer washed until it was free of chloride (generally 25 v / ash). Then the filler material was converted into a 110 ecm given high glass column, which had an inner diameter of 2.5 cm.

The concentrated, crude anti-S-protein factor solution was added to the DEAE column and the column was filled with a Two-piston gradient elution system elutes at the piston A 1000 ecm of an aqueous, 0.005 molar cation phosphate buffer solution (pH value 7.4) and flask B 1000 ecm of an aqueous, 0.04 molar cation phosphate buffer solution (pH value 4.3) contained, supplemented by 405.5 g of sodium chloride per 50 l of buffer solution. The eluate was collected in 15 cc fractions; all in all there were about 120 parliamentary groups. Each fraction was determined for de3 activity using the tryptophan uptake method Anti-S protein factor was examined, and the 10 fractions with the strongest activity were pooled and lyophilized concentrated to about 10% of their original volume.

The entire procedure described above was repeated until such an amount of concentrate was collected that

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the anti-S protein factor contained that total protein content - determined by the Lowry method - was about 200 mg ". This requires about 300 to 400 ecm of concentrate, i.e. about 400 to 1000 g of bovine hypothalami and thus about 160 cattle.

The concentrate containing about 200 mg of protein was dialyzed in distilled water for 24 hours. 4.25 ecm 0.1 molar trypsin and 2.1 ecm 1 molar CaClp were added to each 17 ecm of concentrate. The pH of the mixture was adjusted to 7.8 with 1 molar NaOH, and then the hybrid was ^{incubated at 37 °} C. for 2 hours in a water bath shaker. After the incubation, the solution was filtered through a Diaflo membrane with a molecular weight of 10,000 (Amicon UM-10), and 4.25 ecm 0.1 molar pepsin solution was added to each 17 ecm piltrate. The pH was adjusted to 1.5 with 1 molar HCl, and the mixture was again ^{incubated at 37 °} C. for two hours. After incubation, the pH was increased to 7.8 with 1 molar HaOH, and the solution was again filtered through a Diaflo membrane with a molecular weight of 1000 (Amicon UM-2).

Flash evaporation reduced the volume of the filtrate to about 50 ecm lowered. The resulting mixture was centrifuged and the supernatant liquid was collected. The pH of the supernatant Liquid was adjusted to less than 2.2 by adding concentrated hydrochloric acid. The acidified one

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• Λ.

Liquid was diluted with distilled water to a volume of 75 ecm. Then it became the anti-S-Proteln factor containing solution is chromatographed in the following column:

First column filled with sulfonated resin "Aminex 50-WX2" (Bio Rad Company), a hydrogen ion form of the "Dowex" cation exchange resin with a particle size of 200-325 mesh, was used as the filler material. The resin is a sulfonated polystyrene that is nominally crosslinked with 2 $> pivinylbenzene. This filler material was washed with 0.1N hydrochloric acid in a Buchner funnel until the washing liquid was colorless. It was then washed with distilled water until the washing water showed a pH of 5, and then with about 3 parts by volume of an aqueous 2.0 N sodium hydroxide solution. After washing again with distilled water until the washing water had a pH value of 5, the filler material was placed in a beaker and slurried for 2 to 3 hours at 35 ^° C. in twice the volume of aqueous 1N sodium hydroxide solution. The resulting slurry was filtered through a Buchner funnel and the filter cake was washed with distilled water until the washing water showed a pH of 5. The filler material was then slurried in 2 volumes of aqueous 0.2 molar sodium citrate solution (pH 3.1) and the slurry was poured into a 150 cm high glass column which had an internal diameter of 2 cm.

0.044-0.074 mm

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The acidified aqueous solution of the crude anti-S-protein factor was poured onto this column, and then the column with it eluted on a two-piston gradient elution system; Pistons A contained 2000 ecm of an aqueous 0.2 iaolar solution of sodium citrate (pH value 3.1), and flask B contained 2000 ecm of acetate citrate buffer (pH 9.1) made from 31 g citric acid and 402 g of sodium acetate in 4 l of deionized water, The column would be under a nitrogen pressure of 0.7 kg / cm held. The eluate was collected in 10 cc units; all in all about 120 fractions were obtained.

These fractions were numbered in the order in which they were removed from the column. Each of the factions was with The activity of the anti-S-protein factor was examined with the help of the tryptophan uptake procedure. A graphic representation was prepared with the number of fractions along the x-axis and the activity of the fractions along the y-axis was entered. Such a graphical representation shows at least one activity maximum, in general between the 20th and 30th fraction, and often other maxima can also be found, e.g. one between 40 and 50, one between 70 and 80, one between 90 and 100 and one between 110 and 120. The fractions which give rise to these maxima, e.g. about 2 to 3 fractions from the group between 20 and 30 are combined for further treatment. Any combination was then treated separately in the following way:

7098A0 / 0890

Trennun ^ n-Chronato ^ raphie

The combined fractions that have strong activity The anti-S protein factor was concentrated to about 5 to 10 ecm by flash evaporation on a rotary evaporator. The concentrate thus obtained was subjected to "curtain" electrophoresis on a "curtain" electrophoresis device Beckman-Spinco model CP exposed. An electrolyte solution was prepared by adding 22.4 ecm of an aqueous 0.5 molar Solution of KHpPO- with 259.2 ecm of an aqueous 0.5 molar Solution of Na ^ HPOj mixed, the mixture with distilled Water made up to 20 1 and the pH of the resulting solution adjusted to 8.0 by adding either phosphoric acid or sodium hydroxide was added. It was powered by direct current (50 milliamps, 950 V) worked. 5 liters of the electrolyte solution were used for each batch. The fractionated by electrophoresis Solution of the anti-S-protein factor plus impurities was obtained in 32 fractions of approximately 15 ecm each from the electrophoresis device collected. Each fraction was determined by the tryptophan uptake method for the activity of the anti-S protein factor examined. The fractions with the greatest activity were retained and added. Usually this is the case around the 8th to 12th fractions that are removed from the device.

The combined fractions showing the greatest activity were flashed on a rotary evaporator

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' St

concentrated to a volume of about 1 to 5 ecm. The concentrated Solution, which now only contained about 5 mg of polypeptide material, was made with the aid of the following molecular sieve columns separated:

Two glass columns were connected in series, each of which was 200 μm high and had an internal diameter of 0.81 cm. Both columns were filled with molecular sieve material ("Sephadex G-15 fine", Pharmacia Pine Chemicals, Piscataway, New Jersey) filled. This material is a finely divided dextran with a particle size of 40 to 120iA.

The solution containing the anti-S protein factor was passed through the columns described above, 0.02 molar Acetic acid was used as the solvent. The eluate was collected in 2 cc fractions; a total of about 120 Factions received. Each of these fractions was determined by the tryptophan uptake method for the activity of the anti-S protein pactors investigated and also by UV absorption (wavelength 220 nanometers) for the total polypeptide content. A sufficient amount of the fractions showing the strongest anti-S-protein factor activity was added together to ao that the total polypeptide content is about 60 / µg to 1.5 mg fraud. The solution obtained in this way was evaporated to a volume of about 1 to 3 ecm by flash evaporation on a rotary evaporator concentrated.

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The concentrated solution of the Anti-S-Protein-Paktor3 plus impurities was then subjected to ion exchange chromatography in the following column:

Second column filled with sulfonated resin

The glass column had a height of 200 cm and an internal diameter of 0.81 cm. It was filled with the above-described cation exchange resin ("Aminex 50-V / X2"). The filling was placed in the column and brought into equilibrium with an O ^ -n sodium citrate buffer solution (pE value 3.0); then the system was heated to a temperature of 35 ⁰ C.

The solution to be chromatographed was added to the column and then the column with a two-piston gradient elution system eluted; Flask A contained 1 l of a 0.2 N sodium citrate buffer solution (pH - '. Yert 3.0), and flask B contained 250 ecm of a buffer solution prepared in the following manner had been; A solution of 14.6 g citric acid, 20.6 g sodium acetate, 3.1 ecm glacial acetic acid and 800 ecm distilled Produced water, brought to a pH value of 4.0 by adding concentrated hydrochloric acid and then with distilled Water made up to 1 1.

The eluate from the column was in 5 ccm fractions in a Amount of about 4 to 5 fractions collected per hour. A total of about 130 to 150 such fractions were obtained. Every Fraction was assayed for anti-S protein factor activity by the tryptophan uptake method. The parliamentary group

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respectively the fractions that showed the strongest activity were relatively pure solutions of the anti-S-protein factor, i.e. they were free from most of the other peptide material, tissue cells etc. that accompany the paktor when using the triscitrate buffer solution extracted from the bovine hypothalamus. The greatest concentration of anti-S-protein factor is common to be found in factions 80 to 86. The anino analysis of this Fractions revealed that the factor is a polypeptide that consists of a combination of some of the following amino acids: Glutamic acid, threonine, valine, leucine, phenylalanine, tyrosine, aspartic acid, serine and glycine. The statistical evaluation These amino acid analysis data and the study of the enzymes suggest that the anti-S-protein factor is the Contains tripeptide L-threonyl-L-valyl-L-leucine.

Procedure ^

Another possibility to separate the combined fractions from the fractionation described above treat and purify is reverse phase separation chromatography. A Waters Associates Model 660 Solvent Programmer and a "Water Associates Model 6000 Solvent Delivery System" and an approximately 1.8 to 3 m high column filled with Phenyl Porasil B. with a diameter of 9 »5 mm (also available from Waters Associates). The column is in a constant temperature vessel made from styrofoam blocks there is a constant temperature bath (Haake Type FE). the

709840/0890

Samples are injected with a universal injector (Water Associates Model U6K; 1 / ul to 10 ecm) introduced into the column, and the Effluent from the column is collected with a fraction collector (Scientific Manufacturing Industries 1205). Of the Outflow from the column is monitored with a UV spectrophotometer (Beckman Model 25) that includes a microcell element (Water Associates LC-25) and a differential refractometer (Waters Associates R-401).

The pooled fractions obtained from the column filled with sulfonated resin were dried, in each case 5 to 10 ecm of solvent dissolved and introduced into the column. The eluate from the column was examined for refractive index and absorption at 225 nm. As soon as maxima occurred, the Interrupted flow from the column and examined a sample for UV absorption at 350 to 210 nm. It became factions 5 ecm each were collected, and homogeneous maxima were combined. The combined fraction thus obtained was dried by amino acid analysis and the tryptophan uptake method the maxima with inhibition activity were determined.

The fraction containing the L-threonyl-L-valyl-L-leucine, could be N-acylated according to the procedures of Examples 10, 15 and 16 and / or esterified.

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As already stated, a © C-2 globulin (c <-Helix-S protein) was isolated from the plasma of schizophrenic patients in the course of schizophrenia research, which shows a different activity than a similar <* -2-globulin ( randomly arranged S-protein) from the plasma of normal humans; see above, Prohman, Recent Advances in Biological Psychiatry. If this K- Helix-S protein, isolated from schizophrenic patients, is administered to rats, for example, it leads to clear psychological reactions in these rats. In the publication cited above; by Caldwell, Biological Psychiatry, it is described that self-stimulation of the rats to generate feelings of pleasure clearly decreased when the rats were administered the (λ In this way, the effectiveness of the compounds according to the invention in the treatment of schizophrenic symptoms in humans can be demonstrated Show inhibition of tryptophan uptake; As noted in Example 17, the tryptophan uptake method can be used to isolate to contribute ng of peptide practices that are active against the <X-Helix-S protein.

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Example 18

The Vex-search of intracranial seibat stimulation in rats, described by Caldwell, Biological Psychiatry, pp. 237-239, was repeated in the following manner. By observing the rats into which electrodes had been implanted in the median forebrain bundle, it was determined how often the animals stimulated each other by pressing the button within a certain period of time. Then the o <-helix-S protein was administered "intercisternally" to the rats, and the frequency of keypresses decreased to about 77 Ά the original number. The rats were injected intramuscularly with various polypeptides in an amount of 0.2 mg / kg body weight to determine whether these polypeptides could reverse the effects of the σ <-helix-S protein. In addition, the tryptophan uptake test was carried out with the individual polypeptides. The inhibition of tryptophan uptake was determined after 10 minutes. The results of these tests are summarized in the table below.

Administered peptide Inhibition
d.Tryptophan-
record, # Another increase
of the buttons
press fo Threonylvalylleucine 19.8 85 N-acetylthreonylvalylleucine 20.2 100 Grlycylthreonylvalylleucine 24.3 100 Phenylalanylprolylthreonyl-
valylleucylprolylphenyl-
alanin 39.5 100 N-acetylthreonylvalylleucine-
aiaid 56.5 100 Threonylvalylleucinamide 21.6 90 Threonylvalylisoleucine 5.6 41

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The following polypeptides showed inhibition of tryptophan on would take after 10 minutes determined.

Inhibition of peptide administered tryptophan on would take j "

N - (/ »-acetylthreonylvalyl-
leucine methyl ester 24.2 Ν-Λ-acetyltyrosylthreonyl-
valylleucine 21.3 Prolylthreonylvalylleucyl-
proly! glycine 44.2

For the following polypeptides, the inhibition was dex ' Tryptophan uptake determined after 10, 15 and 30 minutes.

Inhibition of tryptophan uptake in - / » after

Administered peptide 10 min. 15 min. 30 min.

D-Alanyl-L-threonyl-L-valyl-

L-leucine 15 16 0

L-threonyl-L-valyl-D-leucine 12 18 21

With the above. In the experiment, amino acid molecule parts in D-form were used to inhibit tryptophan uptake. With D-Alanyl-L-threonyl-L-valyl-L-leucine show results obtained that the inhibition of tryptophan uptake can decrease after a certain time. The duration of the polypeptide's effectiveness with T-V-L structure was however improved when the Leucine part of the molecule was present in D-form.

For comparison purposes, the experiment described above was repeated with the following polypeptides. The inhibition of the Tryptophilus susceptibility was determined after 10 minutes.

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Inhibition
d.Tryptophan-
micrograph, io Another impetus
of the buttons
pressing on io 0 3 0 toxic 0 toxic 0 toxic 2 not examined

Administered peptide

Glycyl glycyl glycine

O-Benzylthreonylbalylleucinamide

N-cC-acetyl-O-benzylthreonylvalylleucinainide

Benzyl-O-benzylthreonylvalylleucate hydrochloride

L-threonyl-L-valyl-L-leucyl-D-alanine

As from the one performed with L-threonyl-L-valyl-L-leucyl-D-alanine Experimentally evident, the peptide shows no substantial tryptophan uptake when the (L) -component of the essential T-V-L structure has a substituent which is relatively difficult to hydrolyze from a carboxylic acid function of an amino acid leaves.

Using the procedure described, N-acetyltyrosylthreonylvalylleucine was also produced and prolylthreonylvalylleucylprolylglycine to determine their effect on rats, which the o (. -Helix-S protein had been administered. It was found that both polypeptide materials led to a renewed increase in key presses in the rats. Another effective one Polypeptide that can be used to treat schizophrenic symptoms is e.g. threonylvalylleucylarginine.

709840/0890

Claims (1)

Patent claims: 1. / Compound of the following formula as well as their pharmaceutical usable salts: 0 0 0 (A) ₊ -KH-CH-C-NH-CH-C-Nh-CH-C- (B) -R ^b * II ι ^x CHOH .CH I CH, CH, OH, νCoH? where in this formula A stands for an eC -monoiminoacyl radical having 2 to about 10 carbon atoms; B represents a hydrolyzable <X monoiminoacyl radical having 2 to about 10 carbon atoms; R ^{a is} an alkylene group having 1 to about 20 carbon atoms; R is -0R ^e or -NR ⁰ R, where R is hydrogen, an alkyl group with 1 to about 20 carbon atoms or an aryl or aralkyl group with 6 to cd is about 24 carbon atoms and R and R have the same or different meanings and stand for hydrogen or a lower alkyl group, or R ^c and R are lower alkylene groups and together with the nitrogen atom form to which they are attached a cyclic structure; ρ is 0 to t and χ each stand for a number from 0 to 5 and the sum of t plus χ - is also a number from 0 to 5. 709840/0890 ORIGINAL INSPECTED 2. A compound according to claim 1, characterized in that the part of the molecule -NH-CH-C- is derived from threonyl and the CHOH CH ₃ Rest B, if it is optically active, is in ! Configuration J. Compound according to claim 2, characterized in that the part of the molecule -NH-CH-C- is derived from leucyl. I 4. Compound according to claims 1-3 »characterized in that A and B for the same or different peptide Kolekülteile from the group of prolyl, threonyl, AlIothreonyl, leucyl, isoleucyl, glycyl, seryl, arginyl , Phenylalanyl, glutamyl, asparagyl and alanyl groups. 5 · Connection according to claims 1-3 »characterized in that O H (H C) is an acetyl group. 6. Compound according to claim 1-3, characterized in that R stands for a methyloxy group or a group -NH ₂ or -OH. 7. A compound according to claims 1-3, characterized in that O the moiety -NH-CH-C- is a D-leucyl group. (C ₂ H ) ^ ^{3 2 5} ^ -CH ₃ 709840/0890 8. L-threonyl-L-valyl-D-leucine 9. Phenylalanylprolylthreonylvalylleucylprolylphenylalanine 10. N-acetylthreonylvalylleucinamide 11. A compound according to claims 1-3, characterized in that 0 0 the part of the molecule -NH-CH-C-NH-CH-C-NH-CH-C- a ι i CHOH CH I / \ CH, CH, CH, Is L-threonyl-L-valyl-L-leucine residue . 7098AO / 0890 12. Pharmaceutical composition for de-scaling schizophrenics Symptoms in living things suffering from schizophrenia consisting of a pharmaceutically acceptable one A carrier and an amount of a compound according to claims 1-11 sufficient to relieve schizophrenic symptoms or a pharmaceutically acceptable salt of this compound) 709840/0890 Compound of the following formula or pharmaceutically useful one Salts of this compound: 0 0 0 D- (A) + - ra-CH-C-NH-CH-C-NH-CH-C- (B) -E i I i CHOIR ⁴ CH I CH-z CH-I CIw (Cr> H'7 ^{5 3 5 2 3} ^ CH ₅ where in this formula A stands for a c <monoiminoacyl radical having 2 to about 10 carbon atoms; B represents a hydrolyzable monoiminoacyl radical having 2 to about 10 carbon atoms; D is -H or a protecting group for the c < -amino group; E for -OH, a protecting group for a carboxylic acid end group, a metal hydroxide complex or 4th
is an acid addition salt; R is H, an alkyl group having 1 to 20 carbon atoms, an acyl group having 1 to about 20 carbon atoms, a tetrahydropyranyl group, or a monovalent metal; t and χ each stand for a number from 0 to 5 and the sum of t plus χ is also a number from 0 to 5. 14. A compound according to claim 13 »characterized in that η
the part of the molecule -NH-CH-C- is derived from threonyl and the CHOIR ⁴
CH ₅ Remainder B, if it is optically active, is in the L configuration. 7098A0 / 089Q - Ürr- 15. A compound according to claim 13 »characterized in that Il the part of the molecule -NlI-CH-C- is derived from Lcucyl. I,
⁵ 16. A compound according to claim 13-15, characterized in that DACs A and B represents identical or different peptide rMolekül- parts from the group consisting of prolyl, threonyl, 0-Benzylthreonyl-, Allothreonyl-, leucyl, isoleucyl, glycyl -, seryl, arginyl, phenylalanyl, glutamyl, asparagyl, and alanyl groups. 17. A compound according to claim I3-I6, characterized in that .. that D is a t-butyloxycarbonyl group. 18. Compound according to claim 13-17, characterized in that E is a benzyloxy group. 19. A compound according to claim 13-18, characterized in that E is a benzyl group. 20. A compound according to claim 13-19, characterized in that the moiety -NH-CH-C- is a D-leucyl group. I 709840/0890 21. A compound according to claim 13-20, characterized in that sr. 0 0 0 the part of the molecule -NH-CH-C-NH-CH-C-NH-CH-C- I I CHO CH CH, CH, CH, a CH, CH, L-threonyl-L-valy1-L-leuc inre st is. 709840/0890