WO2018161976A1 - Tetrapeptide propylene oxide derivative, preparation method therfor, and use thereof - Google Patents

Abstract

Disclosed in the present invention are a tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof, as well as a preparation method therefor, and the use thereof. The structure of the tetrapeptide propylene oxide derivative or the pharmaceutically acceptable salt thereof is shown in formula I. The compound of the present invention can selectively inhibit the chymotrypsin-like activity of 20S proteasome, and is also expected to display anti-inflammatory properties and inhibition of cell proliferation.

Description

Tetrapeptide propylene oxide derivative, preparation method and use thereof Technical field

The invention belongs to the field of drug synthesis, and in particular relates to a novel class of tetrapeptide propylene oxide derivatives and their use in pharmacodynamics.

Background technique

Currently, malignant tumors are still one of the major diseases that threaten people's lives. Although cancer treatment has made great progress, it has not yet been able to treat cancer radically. Although the anti-cancer drugs currently on the market have certain curative effects, they are mostly cytotoxic drugs with serious toxic and side effects. Therefore, how to study targeted new anticancer drugs from effective tumor targets has become a top priority for medical workers.

The Ubiquitin-Proteasome Pathway (UPP) regulates the levels of proteins involved in cell cycle control, and this pathway has important relationships with cancer, cardiovascular and cerebrovascular diseases, and the pathogenesis of neurodegenerative diseases. . The use of some potent inhibitors to inhibit this pathway from over-degrading important proteins will provide new insights into the treatment of these diseases.

Carfilzomib is a new generation of highly selective irreversible proteasome blockers for the treatment of multiple myeloma (MM) with at least 2 drugs (including bortezomib and immunomodulators). It was approved for marketing on July 20, 2012 by the US Food and Drug Administration (FDA). A number of clinical studies have confirmed that carfilzomib alone or in combination with other drugs have a strong anti-MM effect, low toxicity, especially low incidence of peripheral neuropathy, good tolerance and high safety.

Summary of the invention

OBJECT OF THE INVENTION In view of the above technical problems, an object of the present invention is to provide an epoxyketone compound having a novel structure and having a function of inhibiting proteasome. As a 20S proteasome inhibitor, they can block tumor cell proliferation and induce tumor cell apoptosis, which can be used for the treatment and prevention of various diseases such as malignant tumors in humans and animals.

Another object of the present invention is to provide a process for the preparation of the above compounds.

Still another object of the present invention is to provide an application of the above compound for the preparation of an antitumor drug.

Technical Solution: The present invention discloses a tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof, which has the structure shown in Formula I,

among them:

R ₁ is a substituted or unsubstituted C _1-10 alkyl group, a C _3-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group;

R ₂ is a substituted or unsubstituted C _1-10 alkyl group, a C _1-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group;

R ₃ is a substituted or unsubstituted C _1-10 alkyl group, a C _3-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group;

R ₄ is a substituted or unsubstituted C _1-10 alkyl group, a C _3-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group;

Z is selected from the following fragments:

P is hydrogen or a substituted or unsubstituted C _1-10 alkyl group, a C _1-10 alkoxy group, a phenyl group, a naphthyl group, a tetrahydronaphthyl group, a heterocycloalkyl group or a heterocyclic aryl group.

Preferably, when R ₁ , R ₂ , R ₃ and R ₄ are a substituent group, the substituent is a C _1-4 alkyl group, a C _1-4 alkoxy group, a cyano group, a hydroxyl group, a decyl group, an amino group. , a substituted amino group, or a halogen; P group is a substituent, wherein the substituent is C _1-4 alkyl, C _1-4 alkoxy, halogen or C _1-4 haloalkyl.

Preferably, the heterocycloalkyl group has 3, 4, 5, 6 or 7 ring-forming atoms; the aryl group has 4, 5, 6, 7, 8, 9 or 10 ring-forming atoms; The aryl group has 4, 5, 6, 7, 8, 9 or 10 ring-forming atoms.

R ₁ is preferably a C _1-10 alkyl group, a C _1-6 cycloalkyl group, a heterocycloalkyl group or a heterocyclic group, a phenyl group, a naphthyl group, a fluorenyl group, a thiazolyl group, a thienyl group, a benzothienyl group. a hetero atom-containing aromatic group such as an imidazolyl group, or optionally substituted by a C _1-4 alkyl group, a C _1-4 alkoxy group, a cyano group, a nitro group, a hydroxyl group, a decyl group, an amino group or a halogen; _{1 is} more preferably a C _1-10 alkyl group, a phenyl group, a naphthyl group, an anthracenyl group, a thiazolyl group, a thienyl group, a benzothienyl group, an imidazolyl group, or an alkyl group optionally C _1-4 , C. Alkoxy, cyano, nitro, hydroxy, decyl, amino, substituted amino or halogen substituted with from _{1 to 4} ; R _{1 is} most preferably fluorenyl, thiazolyl, thienyl, benzothienyl, imidazolyl, or It is optionally substituted by a C _1-4 alkyl group, a C _1-4 alkoxy group, a nitro group or a halogen.

R ₂ is preferably a C _1-10 alkyl group, a C _3-6 cycloalkyl group, a heterocycloalkyl group or a heterocyclic group, a phenyl group, a naphthyl group or a fluorenyl group, a thiazolyl group, a benzothiazolyl group or the like. An aromatic group of an atom, or optionally substituted by a C _1-4 alkyl group, a C _1-4 alkoxy group, a cyano group, a nitro group, a hydroxyl group, a decyl group, an amino group, a substituted amino group or a halogen; R _{2 is} more preferably Is a C _1-10 alkyl group, a C _{3 -6} cycloalkyl group, a phenyl group, a naphthyl group, a fluorenyl group, a thiazolyl group, a benzothienyl group, an imidazolyl group, or an alkyl group optionally substituted by C _1-4 . group, C alkoxy, cyano, nitro, hydroxy, mercapto, amino or halogen substituted _{1 to 4;} R ₂ and most preferably _{1 to 10} C alkyl group, a cycloalkyl group of C _{3 ~ 6,} indole A group, a thiazolyl group, a thienyl group, a benzothienyl group, an imidazolyl group, or optionally substituted with a C _1-4 alkyl group, a C _1-4 alkoxy group, a nitro group or a halogen.

R ₃ is preferably a C _1-10 alkyl group, a C _1-6 cycloalkyl group, a heterocycloalkyl group or a heterocyclic group, a phenyl group, a naphthyl group, a fluorenyl group, a thiazolyl group, a thienyl group, a benzothienyl group. a hetero atom-containing aromatic group such as an imidazolyl group, or optionally a C _1-4 alkyl group, a C _1-4 alkoxy group, a cyano group, a nitro group, a hydroxyl group, a decyl group, an amino group, a substituted amino group or a halogen. Substituted; R _{3 is} more preferably C _1-10 alkyl, phenyl, naphthyl, anthracenyl, thiazolyl, thienyl, benzothienyl, imidazolyl, or optionally C _1-4 alkane a group of C _{1 to 4} alkoxy, cyano, nitro, hydroxy, decyl, amino or halogen; R _{3 is} most preferably fluorenyl, thiazolyl, thienyl, benzothienyl, imidazolyl, or It is optionally substituted by a C _1-4 alkyl group, a C _1-4 alkoxy group, a nitro group or a halogen.

R ₄ is preferably a C _1-10 alkyl group, a C _1-6 cycloalkyl group, a heterocycloalkyl group or a heterocyclic group, a phenyl group, a naphthyl group, a fluorenyl group, a thiazolyl group, a thienyl group, a benzothienyl group. a hetero atom-containing aromatic group such as an imidazolyl group, or optionally a C _1-4 alkyl group, a C _1-4 alkoxy group, a cyano group, a nitro group, a hydroxyl group, a decyl group, an amino group, a substituted amino group or a halogen. Substituted; R _{4 is} more preferably C _1-10 alkyl, phenyl, naphthyl, anthryl, thiazolyl, thienyl, benzothienyl, imidazolyl, or optionally C _1-4 alkane Substituted, C _1-4 alkoxy, cyano, nitro, hydroxy, decyl, amino or halogen; R _{4 is} most preferably C _1-10 alkyl, phenyl, thiazolyl, or optionally A C _1-4 alkyl group, a C _1-4 alkoxy group, a nitro group or a halogen is substituted.

Preferably, Z is selected from the following fragments:

P is hydrogen or a C _1-10 alkanoyl group, a C _1-10 alkoxy group, an aryl group or a heterocyclic group (such as a hetero atom containing N, S or O), or optionally a C _1-4 alkane a group, a C _1-4 alkoxy group, a halogen or a C _1-4 halogenated alkyl group; P is preferably hydrogen, morpholinyl, isoxazolyl, phenyl, naphthyl, tetrahydronaphthyl, n-propyl Oxy or isopropoxy.

The "optionally substituted" in the R ₁ , R ₂ , R ₃ , R ₄ and P groups of the present invention means that the groups of R ₁ , R ₂ , R ₃ , R ₄ and P may be substituted by these groups. It is also possible not to be substituted by these groups, that is, not limited to the case of being substituted by the groups recited, but also by the case where these groups are not substituted. This expression is such that "R ₁ is a substituted or unsubstituted C _1-10 alkyl group, a C _1-6 cycloalkyl or heterocycloalkyl group, a phenyl group, a naphthyl group or a fluorenyl group, wherein the substituent is The C _{1 to 4} alkyl group, the C _1-4 alkoxy group, the cyano group, the hydroxy group, the decyl group, the amino group, the substituted amino group or the halogen are expressed in the same manner, but the substitution or substitution is not limited to C _{1 .} An alkyl group of _～10 , but extended to all such groups, including substituted or unsubstituted C _3-6 cycloalkyl or heterocycloalkyl, substituted or unsubstituted benzyl, substituted or unsubstituted A naphthyl group, a substituted or unsubstituted fluorenyl group or the like, wherein the substituent is a C _1-4 alkyl group, a C _1-4 alkoxy group, a cyano group, a hydroxyl group, a decyl group, an amino group or a halogen.

The "R ₁ is a substituted or unsubstituted C _1-10 alkyl group, a C _3-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group" means, for example, R ₁ is a substitution. When a C _1-10 alkyl group is substituted with a certain group of the alkyl group, and R _{1 is an} unsubstituted C _1-10 alkyl group, the alkyl group is not substituted by another group. Replaced by the regiment.

The term "alkyl" is used to mean a saturated hydrocarbon group, the C _1-10 alkyl group means a saturated hydrocarbon group having 1 to 10 carbon atoms, and the C _1-4 alkyl group means a saturated hydrocarbon group having 1 to 4 carbon atoms. .

The term "cycloalkyl" refers to a non-aromatic carbocyclic group, including cyclized alkyl groups. Cycloalkyl groups can include bicyclic or polycyclic systems. Examples of the cycloalkyl group include a cyclopropyl group, a cyclobutyl group, a cyclopentyl group, a cyclohexyl group, a cycloheptyl group, and a C _3-6 cycloalkyl group means a cycloalkyl group having 3 to 6 carbon atoms.

The term "benzyl" refers to benzyl, and substituted benzyl means that at least one hydrogen atom on the phenyl ring of the benzyl group is substituted by a non-hydrogen moiety, and the substituent of the benzyl group may be halogen, -CN, -OH,- SH, -NH ₂ , a straight or branched alkyl group of 1 to 6 carbons, a substituted linear or branched alkyl group of 1 to 6 carbons.

The term "heterocycloalkyl" refers to a non-aromatic heterocarbocyclyl group, including cyclized alkyl groups, wherein one or more ring-forming carbon atoms are replaced by a hetero atom such as an O, N or S atom. The heterocycloalkyl group of the invention preferably has 3, 4, 5, 6 or 7 ring-forming atoms.

The term "heterocyclic aryl" refers to a cyclic aromatic group containing a hetero atom O, N or S, such as furan, thiophene, benzothiophene, pyrrole, thiazole, oxazole, imidazole, pyridine, pyridazine, pyrimidine, pyridyl Oxazine, quinoline, isoquinoline, indole, benzofuran, anthracene, acridine and the like. The heterocyclic aryl group of the invention preferably has 4, 5, 6, 7, 8, 9 or 10 ring-forming atoms.

"Alkoxy" means an -O-alkyl group which generally has from 1 to 10 carbon atoms. Examples of the alkoxy group include a methoxy group, an ethoxy group, a propoxy group (e.g., n-propoxy group and isopropoxy group), a t-butoxy group and the like.

"Aryl" means an aromatic carbocyclic group including monocyclic or polycyclic aromatic hydrocarbons such as phenyl, naphthyl, anthracenyl, phenanthryl and the like. The aryl group of the invention preferably has 4, 5, 6, 7, 8, 9 or 10 ring-forming atoms.

The "aryloxy group" means an -O-aryl group, and the concept of the aryl group is as described above, and the most preferable example of the aryloxy group is a phenoxy group. "Halogen" includes fluorine, chlorine, bromine and iodine.

The amino acid (synthesis raw material) substituted with the R ₁ , R ₂ , R ₃ and R ₄ groups in the compound of the present invention may be a racemate or may be optically active, and R ₁ , R ₂ and R ₃ in the present invention. The amino acid substituted with the R ₄ group is preferably in the S configuration.

Preferred compounds of the invention are:

The invention also discloses a preparation method of the tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof, and the total preparation route is as follows:

In the reaction formula, each group P, R ₁ , R ₂ , R ₃ , R ₄ , Z is as defined above, and the formula (I-1), (I-2) is reacted under the action of a condensing agent to obtain a formula ( I-3), formula (I-3) is produced under the action of trifluoroacetic acid (I-4). Formula (I-4), (I-5) produces formula (I-6) under the action of a condensing agent, and formula (I-6) produces (I-7) under the action of trifluoroacetic acid, wherein formula (I-7) And the carboxylic acid substituted with the P group is reacted under the action of the peptide condensing agent to form the formula (I-8), which forms (I) under the action of LiOH and water.

The preparation of the compounds of the invention is detailed below:

P, R ₁ , R ₂ , R ₃ , R ₄ , Z are as defined above.

The preparation method of the compound (I) includes the following steps:

1) an amino acid of the formula (I-1) and an amino acid methyl ester of the formula (I-2) are obtained by a condensing agent to give a compound of the formula (I-3);

2) After dissolving the compound of the formula (I-3) in DCM, trifluoroacetic acid is added to react to form a compound of the formula (I-4).

3) A compound of the formula (I-4) and an amino acid of the formula (I-5) are condensed under the action of a condensing agent to form a compound of the formula (I-6).

4) reacting a compound of the formula (I-6) with trifluoroacetic acid to obtain a compound of the formula (I-7);

5) A compound of the formula (I-7) and a compound (I-8) are condensed by a condensing agent to form a compound of the formula (I-9).

6) A compound of the formula (I-9) is subjected to a saponification reaction to obtain a compound of the structure (I).

Finally, compounds (I) and (II) are reacted in the presence of a certain condensing agent to form (III). The condensing agent used is 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (abbreviated as EDC.HCl) or benzotriazol-1-yl-oxy hexafluorophosphate Tripyrrolidinylphosphorus is abbreviated as (PYBOP), 1-hydroxybenzotriazole (abbreviated as HOBt).

The present invention also provides the use of the tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof for the preparation of a medicament for treating inflammation, cancer, hyperproliferative diseases or immune-related diseases.

Further, there is provided a use of a tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof for modifying various antigenic peptides produced by a proteasome in an organism.

The use of enzyme inhibitors has a number of biological effects of proteasome inhibition. It has been reported that at the cellular level, after treatment of cells with various proteasome inhibitors, accumulation of polyubiquitinated proteins, changes in cell morphology, and apoptosis occur. Inhibition of proteasomes has also been suggested as a possible anti-tumor therapeutic strategy. Epoxomicin was first identified in the screening of anti-tumor compounds, confirming that the proteasome is an anti-tumor chemotherapeutic drug target. Therefore, these compounds are useful for treating cancer. Inhibition of the proteasome is also linked to inhibition of NF-κB activation and stabilization of p53 levels. Thus, the compounds of the invention are also useful for inhibiting NF-κB activation and stabilizing p53 levels in cell culture. Since NF-κB is a key regulator of inflammation, it is an attractive target for anti-inflammatory therapeutic interventions. Thus, the compounds of the invention are useful in the treatment of chronic inflammation-related diseases including, but not limited to, COPD, psoriasis, bronchitis, emphysema, and cystic fibrosis.

The compounds of the present disclosure are useful for treating conditions directly mediated by proteolytic function of the proteasome (e.g., muscle wasting) or indirectly mediated by proteasome processed proteins (e.g., NF-[kappa]B). Proteasomes are involved in the rapid elimination and post-translational processing of proteins (eg, enzymes) involved in cell regulation (eg, cell cycle, gene transcription and metabolic pathways), intercellular communication, and immune responses (eg, antigen presentation). Specific examples set forth below include: ?-amyloid and regulatory proteins such as cyclin, TGF-? and the transcription factor NF-?B.

Other embodiments of the invention relate to cachexia and muscular atrophy. The proteasome degrades many proteins in mature reticulocytes and growing fibroblasts. In cells lacking insulin or serum, the rate of proteolysis is almost doubled. Inhibition of the proteasome reduces proteolysis, thereby reducing muscle protein loss and kidney or liver nitrogen load. The inhibitor of the present invention can be used for the treatment of diseases such as cancer, chronic infectious diseases, fever, muscle abortion (atrophy) and denervation, nerve damage, fasting, acidosis-related renal failure, diabetes and liver failure. See, for example, U.S. Patent 5,340,736 to Goldberg. Thus, embodiments of the invention include methods for reducing the rate of muscle protein degradation of cells; reducing the rate of intracellular protein degradation; reducing the rate of p53 protein degradation by cells; and inhibiting the growth of p53-associated cancer. All of the above methods involve contacting a cell (in vivo or in vitro, such as a patient's muscle) with an effective amount of a compound of the invention (e.g., a pharmaceutical composition).

Another protein that is processed by the proteasome is NF-κB, a member of the Rel family of proteins. The transcriptional activators of the Rel family can be divided into two groups. The first group required proteolytic processing, including p50 (NF-κB1, 105 kDa) and p52 (NF-κ2, 100 kDa). The second group does not require proteolytic processing, including p65 (RelA, Rel (c-Rel) and RelB). Both homodimers and heterodimers can be formed by members of the Rel family; for example, NF-κB is a p50-p65 heterodimer. After phosphorylation and ubiquitination of IκB and p105, the two proteins are degraded and processed, respectively, to produce active NF-κB, which is transported from the cytoplasm to the nucleus. Ubiquitinated p105 is also processed by purified proteasome (Palombella et al, Cell (1994) 78:773-785). Active NF-κB forms a stereospecific enhancer complex with other transcriptional activators and, for example, HMG I(Y), which induces selective expression of specific genes.

NF-κB regulates genes involved in immune, inflammatory, and mitotic events. For example, an immunoglobulin light chain kappa gene, an IL-2 receptor alpha chain gene, a class I major histocompatibility complex gene, and encoding, for example, IL-2, IL-6, granulocyte colony-stimulating factor, and IFN-β. NF-κB is required for expression of many cytokine genes (Palombella et al, Cell (1994) 78:773-785). Some embodiments of the invention include methods of affecting the expression levels of IL-2, MHC-I, IL-6, TNFα, IFN-β, or any of the other aforementioned proteins, each method comprising administering to the patient an effective amount of a compound of the present disclosure. Complexes comprising p50 are rapid mediators of acute inflammatory and immune responses (Thanos, D. and Maniatis, T., Cell (1995) 80: 529-532).

NF-κB is also involved in the expression of cell adhesion genes encoding E-selectin, P-selectin, ICAM and VCAM-1 (Collins, T., Lab. Invest. (1993) 68: 499-508). One embodiment of the invention is a method of inhibiting cell adhesion (e.g., E-selectin, P-selectin, ICAM or VCAM-1 mediated cell adhesion), the method comprising reacting a cell with an effective amount of a compound of the invention ( Or a pharmaceutical composition) is contacted, or an effective amount of a compound (or pharmaceutical composition) of the invention is administered to a patient.

Intracellular proteolysis produces small peptides for presentation to T lymphocytes, thereby inducing a class I MHC-mediated immune response. The immune system screens autologous cells that have been infected with the virus or have undergone cancer transformation. One embodiment is a method of inhibiting antigen presentation by a cell comprising contacting a cell with a compound of the invention. The compounds of the invention may be used to treat immune-related diseases such as allergies, asthma, organ/tissue rejection (graft versus host disease) and autoimmune diseases including, but not limited to, lupus, rheumatoid arthritis, psoriasis , multiple sclerosis and inflammatory bowel disease (eg ulcerative colitis and Crohn's disease). Thus, another embodiment is a method of inhibiting a patient's immune system (e.g., inhibiting transplant rejection, allergy, autoimmune disease, and asthma), the method comprising administering to the patient an effective amount of a compound of the invention.

Yet another embodiment is a method of altering a library of antigenic peptides produced by proteasomes or other multi-catalytically active Ntn. For example, if the PGPH activity of the 20S proteasome is selectively inhibited, the antigenic peptide group produced by the proteasome and presented to the cell surface by MHC molecules is not identical to the chymotrypsin-like sample without any enzyme inhibition or, for example, the proteasome. The activity is selectively inhibited by either of the two cases of the resulting and presented antigenic peptide group.

Certain proteasome inhibitors block the degradation and processing of ubiquitinated NF-κB in vitro and in vivo. Proteasome inhibitors also block IκB-α degradation and NF-κB activation (Palombella et al, Cell (1994) 78: 773-785; Traenckner et al, EMBO J. (1994) 13: 5433-5441). One embodiment of the invention is a method of inhibiting degradation of IκB-α, the method comprising contacting a cell with a compound of the invention. Another embodiment is a method of reducing the cellular content of NF-κB in a cell, muscle, organ or patient, the method comprising contacting a cell, muscle, organ or patient with a compound of the invention.

Other eukaryotic transcription factors that require proteolytic processing include the universal transcription factor TFIIA, the herpes simplex virus VP16 accessory protein (host cytokine), the viral-induced IFN-regulatory factor 2 protein, and the sterol regulatory element binding protein 1 that binds to the membrane.

Other embodiments of the invention are methods of affecting the cyclin-dependent eukaryotic cell cycle, the method comprising contacting a cell (in vitro or in vivo) with a compound of the invention. Cyclin is involved in cell cycle regulation. Proteasomes are involved in the degradation of cyclins. Examples of cyclins include mitotic cyclins, G1 cyclins, and cyclin B. Degradation of cyclins causes cells to exit one cell cycle phase (eg, mitosis) and enter another phase (eg, divide). All cyclins are thought to associate with the p34.sup.cdc2 protein kinase or related kinase. The proteolytic targeting signal is localized to amino acid 42-RAALGNISEN-50 (degradation box). There is evidence that cyclins are converted to a form that is susceptible to disruption by ubiquitin ligase, or that cyclin-specific ligase is activated during mitosis (Ciechanover, A., Cell, (1994) 79: 13-21) . Inhibition of the proteasome inhibits cyclin degradation, thereby inhibiting cell proliferation in, for example, cyclin-associated cancers (Kumatori et al, Proc. Natl. Acad. Sci. USA (1990) 87: 7071-7075). One embodiment of the invention is a method of treating a proliferative disorder (e.g., cancer, psoriasis or restenosis) in a patient, the method comprising administering to the patient an effective amount of a compound of the invention. The invention also encompasses a method of treating cyclin-associated inflammation in a patient, the method comprising administering to the patient a therapeutically effective amount of a compound of the invention.

Further embodiments are methods of affecting proteasome-dependent modulation of oncoproteins and methods of treating or inhibiting cancer growth, each method comprising contacting a cell (in vivo, such as a patient, or in vitro) with a compound of the invention. HPV-16 and HPV-18-derived E6 proteins stimulate ATP- and ubiquitin-dependent conjugation and degradation of p53 in crude reticulocyte lysates. It has been confirmed that the recessive oncogene p53 accumulates at a non-permitted temperature in a cell line having a mutant heat labile E1. High levels of p53 may cause apoptosis. Examples of proto-oncogene proteins that are degraded by the ubiquitin system include c-Mos, c-Fos, and c-Jun. One embodiment is a method of treating apoptosis in p53-associated cells, the method comprising administering to the patient an effective amount of a compound of the invention.

Finally, the compounds of the invention may also be used as diagnostic reagents (e.g., in diagnostic kits or clinical laboratories) for screening proteins (e.g., enzymes, transcription factors) processed by Ntn hydrolases (including proteasomes). The compounds of the invention are also useful as research reagents for specifically binding to the X/MB 1 subunit or alpha chain and inhibiting the proteolytic activity associated therewith. For example, the activity of other subunits of the proteasome (and its specific inhibitor) can be determined.

Most cellular proteins undergo proteolytic processing during maturation or activation. The enzyme inhibitors disclosed herein can be used to determine whether a cell, development or physiological process or output is modulated by the proteolytic activity of a particular Ntn hydrolase. One such method comprises obtaining an organism, an intact cell preparation or a cell extract; contacting the organism, cell preparation or cell extract with a compound of the invention; contacting the organism, cell preparation of the compound of the invention Or the cell extract signals and then monitors the process or output. The high selectivity of the compounds of the invention allows for the rapid and accurate elimination or effect of Ntn (e.g., the 20S proteasome) in specific cellular, developmental or physiological processes.

The present invention also provides a pharmaceutical composition comprising the tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof and a pharmaceutically acceptable carrier.

Administration

Depending on the disease to be treated and the age, health and weight of the patient, the compounds prepared according to the methods described herein can be administered in a variety of different forms, as is well known in the art. For example, when the compounds are intended for oral administration, they may be formulated as tablets, capsules, granules, powders or syrups; or for parenteral administration, they may be formulated as injections (intravenous, intramuscular) Or subcutaneous), infusion preparation or suppository. When administered by the ocular mucosal route, they can be formulated as eye drops or eye ointments. These preparations can be prepared by a conventional method, and if necessary, the active ingredient can be combined with any conventional additives or excipients (for example, a binder, a disintegrating agent, a lubricant, a flavoring agent, a solubilizing agent, a suspending agent, an emulsifier, a coating) Mixtures, cyclodextrins and/or buffers). Although the dosage will depend on the condition, age and weight of the patient, the nature and severity of the condition to be treated or prevented, the route of administration and the form of the drug, in general, the recommended daily dose of the compound of the invention for an adult patient is 0.01 mg. -2000 mg, which can be administered as a single dose or as multiple divided doses. The amount of active ingredient prepared in a single dosage form in admixture with the carrier is usually the amount of the compound which produces a therapeutic effect.

The precise administration time and/or composition dose to achieve optimal therapeutic effect will depend on the activity, pharmacokinetics and bioavailability of the particular compound, the physiological condition of the patient (including age, etc.) in terms of the therapeutic effect on a particular patient. Gender, type and stage of disease, general physical condition, response to a specific dose, and type of drug), route of administration, and the like. In any event, the above criteria can be used as a basis for precise adjustment therapy, such as determining the optimal dosing time and/or dosage to be administered, which requires only routine experimentation, including monitoring the patient and adjusting the dosage and/or administration time.

The term "pharmaceutically acceptable" as used herein means that the ligands, materials, compositions and/or dosage forms are within reasonable medical judgment and are suitable for contact with human tissues and animal tissues without excessive toxicity, irritation, Allergies or other problems or complications with a reasonable benefit/risk ratio.

The term "pharmaceutically acceptable carrier" as used herein refers to a pharmaceutically acceptable starting material, ingredient or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material. All carriers must be "acceptable", ie compatible with the other formulation ingredients of the formulation, and not deleterious to the patient. Some examples of pharmaceutically acceptable carriers include: (1) sugars such as lactose, glucose and sucrose; (2) starches such as corn starch, potato starch and substituted or unsubstituted beta cyclodextrin; (3) fibers And its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; Excipients such as waxes for cocoa butter and suppositories; (9) oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols such as propylene glycol; (11) Polyhydric alcohols such as glycerin, sorbitol, mannitol and polyethylene glycol; (12) esters such as ethyl oleate and ethyl laurate; (13) agar; (14) buffers such as hydrogen Magnesium oxide and aluminum hydroxide; (15) alginic acid; (16) pyrogen-free water; (17) isotonic saline; (18) Ringer's solution; (19) ethanol; (20) phosphate buffer Solution; (21) other non-toxic compatible substances used in pharmaceutical preparations. In certain embodiments, the pharmaceutical compositions of the invention are non-pyrogenic, i.e., do not cause significant elevation of body temperature upon administration to a patient.

The term "pharmaceutically acceptable salts" refers to relatively non-toxic inorganic acid addition salts and organic acid addition salts of the inhibitors. These salts can be prepared in situ during the final isolation and purification of the inhibitor, or the free base form of the purification inhibitor can be reacted with a suitable organic or inorganic acid alone, and the salt thus formed can be isolated. Representative salts include hydrobromide, hydrochloride, sulfate, hydrogen sulfate, phosphate, nitrate, acetate, valerate, oleate, palmitate, stearate, lauric acid Salt, benzoate, lactate, phosphate, tosylate, citrate, maleate, fumarate, succinate, tartrate, naphthylate, methane An acid salt, a glucoheptonate, a lactobionate, a lauryl sulfonate, an amino acid salt, and the like. (See, for example, Berge et al. (1977) "Pharmaceutical Salts", J. Pharm. Sci. 66: 1-19).

In other instances, the inhibitors useful in the methods of the invention may comprise one or more acidic functional groups and, therefore, are capable of forming pharmaceutically acceptable salts with pharmaceutically acceptable bases. In these instances, the term "pharmaceutically acceptable salts" refers to relatively non-toxic inorganic base addition salts and organic base addition salts of the inhibitors. These salts can likewise be prepared in situ during the final isolation and purification of the inhibitor, or the free acid form of the purification inhibitor alone with a suitable base (eg a hydroxide, carbonate or hydrogen carbonate of a pharmaceutically acceptable metal cation). Salt), ammonia or pharmaceutically acceptable organic primary, secondary or tertiary amines. Representative alkali metal or alkaline earth metal salts include lithium salts, sodium salts, potassium salts, calcium salts, magnesium salts, and aluminum salts, and the like. Representative organic amines useful for the formation of base addition salts include ethylamine, diethylamine, ethylenediamine, ethanolamine, diethanolamine, piperazine, and the like (see, for example, Berge et al., supra).

Wetting agents, emulsifiers and lubricants (such as sodium lauryl sulfate and magnesium stearate) as well as coloring agents, release agents, coating agents, sweeteners, flavoring agents, flavoring agents, and preservatives may also be added to the composition. Agents and antioxidants.

Examples of pharmaceutically acceptable antioxidants include: (1) water-soluble antioxidants such as ascorbic acid, cysteine hydrochloride, sodium hydrogen sulfate, sodium metabisulfite, sodium sulfite, etc.; (2) oil-soluble antioxidants, for example Vitamin C ester of palmitic acid, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha-tocopherol, etc.; (3) metal chelating agents such as citric acid, ethylenediamine four Acetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and the like.

Formulations suitable for oral administration may be capsules, cachets, pills, tablets, lozenges (using a flavored base, usually with sucrose and gum arabic or tragacanth), powders, granules, or aqueous Or a solution or suspension in a non-aqueous liquid, either an oil-in-water or water-in-oil liquid emulsion, or an elixir or syrup, or a soft lozenge (using an inert base such as gelatin and glycerin, or Sucrose and gum arabic) and/or mouthwashes and the like, all of which contain a predetermined amount of an inhibitor as an active ingredient. The composition can also be administered as a bolus, granule or paste.

In an oral solid dosage form (capsules, tablets, pills, lozenges, powders, granules, etc.), the active ingredient is mixed with one or more pharmaceutically acceptable carriers, such as sodium citrate or dicalcium phosphate and/or Any carrier: (1) a filler or extender such as starch, cyclodextrin, lactose, sucrose, glucose, mannitol and/or silicic acid; (2) a binder such as carboxymethylcellulose, alginic acid Salt, gelatin, polyvinylpyrrolidone, sucrose and/or gum arabic; (3) humectants such as glycerin; (4) disintegrants such as agar, calcium carbonate, potato starch, tapioca starch, alginic acid, certain silicic acid a salt and sodium carbonate; (5) a dissolution delaying agent such as paraffin; (6) an absorption enhancer such as a quaternary ammonium compound; (7) a wetting agent such as acetyl alcohol and glyceryl monostearate; (8) an adsorbent, For example, kaolin and bentonite; (9) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, and mixtures thereof; (10) colorants. In the case of capsules, tablets and pills, the pharmaceutical compositions may also contain buffering agents. Solid compositions of a similar type may also be employed as fillers in soft and hard-filled gelatin capsules, using such excipients as lactose or milk sugar as well as high molecular weight polyethylene glycols.

Tablets may be prepared by compression or molding, optionally with one or more accessory ingredients. The compressed tablet may be a binder (for example, gelatin or hydroxypropylmethylcellulose), a lubricant, an inert diluent, a preservative, a disintegrant (for example, sodium starch glycolate or croscarmellose sodium). , surfactant or dispersant preparation. Molded tablets can be prepared by molding a powdered inhibitor mixture moistened with an inert liquid diluent in a suitable machine.

Tablets and other solid dosage forms (e.g., lozenges, capsules, pills, and granules) can optionally be scored or prepared to have coatings and shells such as enteric coatings and other coatings known in the pharmaceutical art. They may also be formulated for sustained or controlled release of the active ingredient using, for example, hydroxypropyl methylcellulose in varying proportions, other polymeric matrices, liposomes and/or microspheres to provide the desired release rate. They can be sterilized, for example, by filtration through a bacterial filter, or by incorporating a sterilizing agent in the form of a sterile solid, which can be dissolved in sterile water or some other sterile injectable medium before use. These compositions may also optionally contain opacifying agents, which may be compositions which release the active ingredient only, or preferentially, in certain parts of the gastrointestinal tract, and optionally in a delayed release mode. Examples of embedding compositions that can be used include polymers and waxes. The active ingredient may also be in the form of microcapsules, optionally with one or more of the above-mentioned excipients.

Oral liquid dosage forms include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active ingredient, the liquid dosage form may contain inert diluents conventional in the art such as water or other solvents, solubilizers and emulsifiers such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzoic acid Benzyl ester, propylene glycol, 1,3-butanediol, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil and sesame oil), glycerin, tetrahydrofuranol, polyethylene glycol, sorbus Fatty acid esters of polysaccharides and mixtures thereof.

Besides the inert diluent, the oral compositions may contain adjuvants such as wetting agents, emulsifying agents, suspending agents, sweetening agents, flavoring agents, coloring agents, flavoring agents and preservatives.

In addition to the active ingredient, the suspension may contain suspending agents, for example, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar , tragacanth and their mixtures.

The rectal or vaginal preparation may be a suppository, which may be prepared by mixing one or more inhibitors with one or more non-irritating suitable excipients or carriers including, for example, cocoa Oils, polyethylene glycols, suppository waxes or salicylates, which are solid at room temperature and liquid at body temperature, and therefore will melt in the rectum or vaginal cavity, releasing the active agent.

Formulations suitable for vaginal administration also include pessaries, tampons, creams, gels, pastes, foams or sprays, including suitable carriers known in the art.

Dosage forms for topical or transdermal administration of inhibitors include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches, and inhalants. The active ingredient may be admixed under sterile conditions with apharmaceutically acceptable carrier and any such preservative, buffer, or propellant.

Ointments, pastes, creams and gels may contain, in addition to inhibitors, excipients such as animal and vegetable oils, oils, waxes, paraffin, starch, tragacanth, cellulose derivatives, poly Ethylene glycol, silicone, bentonite, silicic acid, talc, zinc oxide or a mixture thereof.

Powders and sprays can contain, in addition to inhibitors, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicate, polyamide powder or mixtures of these materials. Sprays can also contain conventional propellants such as chlorofluorocarbons and volatile unsubstituted hydrocarbons such as butane and propane.

The inhibitor can also be administered by aerosol. This can be accomplished by preparing an aqueous aerosol, liposomal formulation or solid particles containing the composition. Non-aqueous (e.g., fluorocarbon propellant) suspensions can be used. Sonic nebulizers are preferred because they minimize shear forces that can cause degradation of the compound.

In general, aqueous aerosols are prepared by formulating an aqueous solution or suspension of the drug with conventionalpharmaceutically acceptable carriers and carriers. The carrier and stabilizer vary depending on the requirements of the particular composition, but typically include nonionic surfactants (Tween, Pluronic, sorbitan ester, lecithin, Cremophor), pharmaceutically acceptable solubilizing agents (eg, polyethylene glycol). ), harmless proteins (such as serum albumin), oleic acid, amino acids (such as glycine), buffers, salts, sugars or sugar alcohols. Aerosols are usually prepared using an isotonic solution.

Transdermal patches have more advantages in controlling the administration of body inhibitors. Such dosage forms can be prepared by dissolving or dispersing the drug in a suitable medium. Absorption enhancers can also be used to increase the flux of the inhibitor across the skin. Such migration rates can be controlled by rate-regulating membranes or by dispersing the inhibitor into a polymer matrix or gel.

A pharmaceutical composition of the invention suitable for parenteral administration comprises one or more inhibitors and one or more pharmaceutically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions, or prior to use Sterile powders which can be reconstituted into sterile injectable solutions or dispersions may contain an antioxidant, a buffer, a bacteriostatic agent, a solute, a suspending agent or a thickening agent which renders the formulation isotonic with the blood of the intended recipient.

Examples of suitable aqueous and nonaqueous vehicles which may be employed in the pharmaceutical compositions of the present invention include water, ethanol, polyols (e.g., glycerol, propylene glycol, polyethylene glycol, and the like), suitable mixtures thereof, vegetable oils (e.g., olive oil), and An organic ester for injection, such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of a coating material such as lecithin, by the maintenance of the desired particle size of the dispersion, and by the use of surfactants.

These compositions may also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. The addition of various antibacterial and antifungal agents prevents the action of microorganisms such as parabens, chlorobutanol, phenol, sorbic acid and the like. A tonicity adjusting agent such as sugar, sodium chloride or the like may also be required in the composition. In addition, the absorption of the injectable pharmaceutical preparation can be prolonged by the addition of agents which delay absorption, such as aluminum monostearate and gelatin.

In some cases, in order to prolong the effect of the drug, it is necessary to slow the rate of absorption of the drug by subcutaneous or intramuscular injection. For example, absorption of a parenterally administered drug is delayed by dissolving or suspending the drug in an oil vehicle.

Injection depot formulations are prepared by forming microencapsule matrices of the inhibitor in a biodegradable polymer such as polylactide-polyglycolide. The release rate of the drug can be adjusted depending on the ratio of drug to polymer and the specific polymer properties used. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by encapsulating the drug in liposomes or microemulsions which are compatible with body tissues.

The pharmaceutical preparations can be administered orally, parenterally, topically or rectally. Of course, it is administered in a dosage form suitable for various administration routes. For example, they are administered in the form of tablets or capsules, administered by injection, inhalation, eye wash, ointment, suppository, infusion solution; topically with a lotion or ointment; and administered rectally with a suppository. Oral administration is preferred.

The term "parenteral administration" as used herein refers to a mode of administration other than enteral and topical administration, generally referred to as injection and infusion, including but not limited to intravenous, intramuscular, intraarterial, intrathecal, Intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subepidermal, intra-articular, subcapsular, subarachnoid, intraspinal and intrasternal injections.

The terms "systemic administration" and "peripheral administration" as used herein mean that the ligand, drug or other substance is not directly administered into the central nervous system such that they enter the body of the patient and thus undergo metabolic or other similar processes, such as subcutaneous administration.

These inhibitors can be administered to humans or other animals for therapeutic purposes, and any suitable route of administration can be employed, including oral, nasal (eg, by spray), rectal, intravaginal, parenteral, intracisternal, and topical (eg, Use powders, ointments or drops, including buccal and sublingual administration).

Regardless of which route of administration is selected, the inhibitors of the invention (which may be used in their appropriate hydrated form) and/or the pharmaceutical compositions of the invention may be formulated as pharmaceutically acceptable dosage forms by conventional methods known in the art.

The actual dosage level of the active ingredient of the pharmaceutical compositions of the present invention can be varied to provide an effective amount of the active ingredient to achieve the desired therapeutic response without poisoning the patient, for the particular patient, composition, and mode of administration.

The concentration of a compound of the invention in a pharmaceutically acceptable mixture will vary depending on a variety of factors, including the dosage of the compound administered, the pharmacokinetic profile of the compound employed, and the route of administration. In general, the compositions of the present invention can be provided as aqueous solutions containing from about 0.1% to about 10% w/v of the compound of the invention for parenteral administration. A typical dose is from about 0.01 mg/kg body weight to about 50 mg/kg body weight per day, administered in 1-4 divided doses. The same or different compounds of the invention may be included in each sub-dose. The dosage administered will be an effective dose, and the effective dosage will depend on a variety of factors, including the overall condition of the patient, the formulation of the selected compound, and the route of administration.

Another aspect of the invention provides a combination therapy wherein one or more additional therapeutic agents are administered with a proteasome inhibitor of the invention. Such combination therapies can be achieved by administering the components simultaneously, sequentially or separately.

In certain embodiments, a compound of the invention is administered in combination with one or more other proteasome inhibitors.

In certain embodiments, the compounds of the invention are administered in combination with a chemotherapeutic agent. Suitable chemotherapeutic agents may include natural products such as the vinca alkaloids (i.e., vinblastine, vincristine, and vinorelbine), paclitaxel, epidipodophyllotoxin (i.e., etoposide, teniposide). , antibiotics (dactinomycin (actinomycin D), daunorubicin, doxorubicin and idarubicin), anthracyclines, mitoxantrone, bleomycin, pucamycin (Glucosin) and mitomycin, enzyme (L-asparaginase, which systematically metabolizes L-asparagine, removes cells that cannot synthesize their own asparagine); antiplatelet agents; antiproliferative/antibiotics Mitotic alkylating agents such as nitrogen mustards (nitrogen mustard, cyclophosphamide and its analogues, melphalan, chlorambucil), aziridines and methyl melamines (hexamethylene melamine and thiophene) , sulfonate alkyl ester (Bai'an), nitrosourea (carmustine (BCNU) and its analogues, chain star), trazenes-dacarbazinine (DTIC); Anti-proliferative/anti-mitotic antimetabolites such as folic acid analogs (methotrexate), pyrimidine analogs (fluorouracil, fluorouridine and cytarabine), purine analogs Related inhibitors (mercaptopurine, thioguanine, pentastatin and 2-chlorodeoxyadenosine); aromatase inhibitors (anastrozole, exemestane and letrozole); platinum coordination complexes ( Cisplatin, carboplatin, procarbazine, hydroxyurea, mitoxantrone, aminoglutethimide; histone deacetylase (HDAC) inhibitors (triccobacterin, sodium butyrate, apicidan, suberoylanilide) Suberoyl anilide hydroamic acid; hormones (ie estrogens) and hormonal agonists, such as luteinizing hormone releasing hormone (LHRH) agonists (goserelin, leuprolide and triptoride) forest). Other chemotherapeutic agents may include nitrogen mustard, camptothecin, ifosfamide, tamoxifen, raloxifene, gemcitabine, noviben or any analog or derivative of the foregoing.

In certain embodiments, the compounds of the invention are administered in combination with a cytokine. Cytokines include, but are not limited to, interferon-γ, -α and -β, interleukin 1-8, 10 and 12-granulocyte monocyte colony-stimulating factor (GM-CSF), TNF-α and -β, TGF -β.

In certain embodiments, the compounds of the invention are administered in combination with a steroid. Suitable steroids include, but are not limited to, 21-acetoxypregnenolone, amlodipine, alpha progesterone, ansinide, beclomethasone, betamethasone, budesonide, prednisone, clobetasol, Clodroxolone, clonpredil, corticosterone, cortisone, cortisol, difluxate, dexamethasone, deshydroxymetasone, dexamethasone, diflurazon, difluorocoron, two Fluoric acid ester (difuprednate), glycyrrhetinic acid, fluconate, flucline, flumethasone, flunisolide, fluocinolone, fluocinolone acetonide, flubutylbutyrate, fluconazole, fluorometholone, Flueperone acetate, flurbidine acetate, fluprednisolone, hydrofluoric acid, fluticasone propionate, formoterol, Hasinide, halobetasol propionate, halomethasone, hydrocortisone, chloride Pano ethyl carbonate, equine ketone, metformin, methylprednisolone, methylprednisolone, mometasone furoate, palmidone, prednisolone, prednisolone, prednisolone 25-two Ethylaminoacetate, prednisolone sodium phosphate, prednisone, prednisolone, prednisolone, rimexolone, teicosone, triamcinolone, ko Desonide, benzene, triamcinolone acetonide, triamcinolone hexacetonide, and salts and / or derivatives thereof.

In certain embodiments, the compounds of the invention are administered in combination with an immunotherapeutic. Suitable immunotherapeutic agents include, but are not limited to, MDR modulators ( verapamil, valspordar, bicepoda, tariquidar, laniquidar), cyclosporin, thalidomide, and monoclonal antibodies. Monoclonal antibodies can be naked monoclonal antibodies or conjugated monoclonal antibodies such as rituximab, tocilizumab, alemtuzumab, epratuzumab, temimumab, gemtuzin Anti-ozomibine (gemtuzumab ozogamicin), bevacizumab, cetuximab, erlotinib and trastuzumab.

detailed description

Synthesis of the first part of the compound

The preparation of the compounds of the invention can be carried out as follows:

1. Preparation of Compound (I)

1. Preparation of Compound (I-3):

After compound (I-1), HOBt was dissolved in anhydrous DCM and stirred at -5 ° C for 10 min, EDICI was added at this temperature, stirred for 15-20 min, and then compound (I-2) was added and stirred for 15-20 min, then added. The DIPEA was stirred for 20 minutes and moved to room temperature for reaction. After the completion of the reaction, the mixture was poured into water and extracted with DCM. EtOAc (EtOAc m.

2. Preparation of compound (I-4):

The compound (I-3) was dissolved in anhydrous DCM, and TFA was slowly added dropwise at -5 ° C, stirred for 0.5 hours, and then stirred at room temperature for 3 hours and then detected. After completion of the reaction, the reaction mixture was concentrated to give a brown-yellow oil.

3. Preparation of compound (I-6):

After compound (I-5), HOBt was dissolved in anhydrous DCM and stirred at -5 ° C for 10 min, EDICI was added at this temperature, stirred for 15-20 min, and then compound (I-4) was added and stirred for 15-20 min, then added. The DIPEA was stirred for 20 minutes and moved to room temperature for reaction. After the completion of the reaction, the mixture was poured into water and extracted with EtOAc. EtOAc (EtOAc)

4. Preparation of Compound (I-7):

The compound (I-6) was dissolved in anhydrous DCM, and TFA was slowly added dropwise at -5 ° C, stirred for 0.5 hour, and then stirred at room temperature for 3 hours and then detected. After completion of the reaction, the reaction mixture was concentrated to give a brown-yellow oil.

5. Preparation of compound (I-9):

The P group-substituted carboxylic acid (I-8), HOBt was dissolved in anhydrous DCM and stirred at -5 ° C for 10 min. At this temperature, EDICI was added, stirred for 15-20 min, and then compound (I-7) was added and stirred. After -20 min, DIPEA was added and stirred for 20 minutes, and moved to room temperature for reaction. After the completion of the reaction, the mixture was poured into water and extracted with EtOAc. EtOAc (EtOAc)

Preparation of Compound (I):

The compound (I-9) was dissolved in MeOH/H ₂ O, and a LiOH aqueous solution was added dropwise at 0 ° C, stirred for 2 hours, and allowed to react to room temperature for a certain period of time. Water was added and the pH was adjusted to 6-7 with hydrochloric acid, and extracted with ethyl acetate. The organic phase was washed with brine, dried over anhydrous sodium sulfate and evaporated

2. Preparation of Compound (II)

1. Preparation of Compound (II-2):

The compound (II-1) and HOBt were dissolved in DCM, and EDC.HCl was added. After stirring at -5 ° C for 15 min, dimethylhydroxylamine hydrochloride was added. After 15 min, DIPEA was added, and the reaction was carried out at low temperature for 25 min. After the reaction at room temperature, DCM was used. extraction, the organic phase washed with 1N HCl, 5% NaHCO ₃ wash, brine, dried over anhydrous sodium sulfate, the solvent was distilled off to give compound (II-2).

2. Preparation of compound (II-3):

The compound (II-2) was dissolved in tetrahydrofuran, and ethylmagnesium bromide was added dropwise at -20 ° C, and the mixture was added dropwise to room temperature. After completion, the reaction was quenched by dropwise addition of 1 N diluted hydrochloric acid, and extracted with ethyl acetate. The organic layer was dried and concentrated to give the compound (II-3).

3. Preparation of compound (II-4):

The compound (II-3) was dissolved in tetrahydrofuran, piperidine acetate and piperidine were added, and paraformaldehyde was added in portions, and after refluxing for 3 hours, an appropriate amount of water was added, and then extracted with ethyl acetate, and 1 N diluted hydrochloric acid and saturated brine were respectively used. After washing, the organic phase is dried and concentrated to give the compound (II-4).

4. Preparation of compound (II-5):

The compound (II-4) is dissolved in toluene, and aluminum isopropoxide and isopropanol are added, and the reaction is carried out at 50 ° C. After the reaction, the mixture is extracted with water and ethyl acetate, washed with 1N diluted hydrochloric acid and brine, and then dried and concentrated. Compound (II-5).

5. Preparation of compound (II-6):

The compound (II-5) was dissolved in DCM, then vanadium acetylacetonate was added, and the mixture was cooled to 0 ° C under ice-cooling, and t-butanol was slowly added dropwise. After the completion of the reaction, an appropriate amount of water is added, and the mixture is extracted with dichloromethane, washed with saturated sodium thiosulfate and saturated brine, and then dried and concentrated to afford compound (II-6).

6. Preparation of compound (II-7):

The compound (II-6) is dissolved in dimethyl sulfoxide, diisopropylethylamine is added, and pyridine sulfur trioxide is added in portions under ice-cooling, and the reaction is carried out to room temperature until the reaction is complete. The extract was washed with 1 N of dilute hydrochloric acid and saturated brine, and the organic phase was dried and concentrated to give Compound (II-7).

7. Preparation of compound (II):

The compound (II-7) was dissolved in anhydrous DCM, and TFA was slowly added dropwise at -5 ° C, stirred for 0.5 hours, and then stirred at room temperature for 3 hours and then detected. After completion of the reaction, the reaction mixture was concentrated to give a brown-yellow oil.

3. Preparation of Compound (III)

Preparation of Compound (III):

After dissolving compound (I), HOBt in anhydrous DCM and stirring at -5 ° C for 10 min, add EDIHCl at this temperature, stir for 15-20 min, then add compound (II) and stir for 15-20 min, then add DIPEA and stir for 20 min. Move to room temperature for reaction. After completion of the reaction, the mixture was poured into water and extracted with EtOAc. EtOAc (EtOAc)

The preparation of the compounds of the invention is described below by the synthesis of specific compounds:

First, the preparation of acid fragments:

Take the preparation of N-2-morpholinylacetyl-L-thiazole alanyl-L-leucyl-L-phenylalanine as an example:

Compound 1 (3.22 g, 13.91 mmol), HOBt (2.82 g, 20.87 mmol) was dissolved in anhydrous DCM (100 mL). After stirring at -5 °C for 10 min, then HCl HCl (4.0 g, 20. After stirring for 15-20 min, compound 2 (3.0 g, 13.91 mmol) was added and stirred for 15-20 min, then DIPEA (10 mL, 62.60 mmol) was added and stirred for 20 min. After completion of the reaction, poured into ice water, extracted with DCM, and the combined organic phases were with 0.4N HCl, 5% NaCO ₃ sodium hydrogencarbonate, saturated brine, dried over anhydrous sodium sulfate, the solvent was evaporated to give Compound 3.

Compound 3 (6.79 g, 17.29 mmol) was dissolved in anhydrous DCM (100 mL). EtOAc (1············· After completion of the reaction, the reaction mixture was concentrated to give a brown brown oil.

Compound 5 (5 g, 18.4 mmol), HOBt (3.73 g, 27.6 mmol) was dissolved in anhydrous DCM (100 mL). After stirring at -5 °C for 10 min, then EtOAc (5.32 g, 27.6 mmol) After -20 min, compound 4 (7.48 g, 18.4 mmol) was added and stirred for 15-20 min, then DIPEA (20 mL, 124.2 mmol) was added and stirred for 20 min. After completion of the reaction, poured into ice water, extracted with DCM, and the combined organic phases were with 0.4N HCl, 5% NaCO ₃ sodium hydrogencarbonate, saturated brine, dried over anhydrous sodium sulfate, the solvent was evaporated, to give compound 6.

Compound 6 (6.6 g, 12.1 mmol) was dissolved in anhydrous DCM (100 mL), and TFA (10 mL) was slowly added dropwise at -5 ° C, stirred for 0.5 hour, and then stirred at room temperature for 3 hours. After completion of the reaction, the reaction mixture was concentrated to give a brown brown oil.

Compound 8 (2.67 g, 18.4 mmol), HOBt (3.74 g, 27.6 mmol) was dissolved in anhydrous DCM (100 mL). After stirring at -5 °C for 10 min, then HCl (5.32 g, 27.6 mmol) was stirred at this temperature. After 15-20 min, compound 7 (7.48 g, 18.4 mmol) was added and stirred for 15-20 min, then DIPEA (12.16 mL, 73.6 mmol) was added and stirred for 20 min. After completion of the reaction, poured into ice water, extracted with DCM, and the combined organic phases were with 0.4N HCl, 5% NaCO ₃ sodium hydrogencarbonate, saturated brine, dried over anhydrous sodium sulfate, the solvent was evaporated to give compound 9.

Compound 9 (9.6g, 16.7mmol) was dissolved in _{MeOH / H 2 O (200mL /} 50mL), was added dropwise at 0 ℃ _{LiOH · H 2 O (1.01g,} 23.38mmol) (10mL) solution of H ₂ O, After stirring for 2 hours, the mixture was allowed to react to room temperature for a certain period of time. Water was added and the pH was adjusted to 6-7 with hydrochloric acid, and extracted with ethyl acetate. The organic phase was washed with brine, dried over anhydrous sodium sulfate Rate 89%, mp: 177-178; ¹ H NMR (400 MHz, CDCl ₃ ) δ 0.81 - 0.73 (m, 3H), 0.85 (t, J = 10.2 Hz, 3H), 1.25 (d, J = 4.5 Hz) , 2H), 1.60–1.49 (m, J=7.1 Hz, 1H), 2.19 (s, 8H), 2.70 (s, 2H), 3.27 (s, 2H), 3.79 (d, J = 12.5 Hz, 2H) , 4.33 (s, 1H), 4.67 (s, 1H), 4.79 (s, 1H), 7.13 (s, 1H), 7.26 - 7.15 (m, 4H), 7.27 (s, 1H), 7.41 (s, 1H) , 8.49 (s, 1H), 8.65 (s, 1H), 8.71 (d, J = 4.9 Hz, 1H); MS (ESI) m/z: 674.0 [M+H] ⁺ .

The synthesis method of all the acid fragment compounds in the present invention is similar to that of 10.

The specific compounds synthesized and their names are as follows.

Second, the preparation of amine fragments:

Compound (S)-2-Amino-4-methyl-1-((R)-2-methyloxiran-2-yl)pentan-1-one 2,2,2-trifluoroacetic acid The preparation of salt (38) is an example:

Compound 31 (10.0 g, 43.23 mmol), HOBt (5.8 g, 43.23 mmol) was dissolved in DCM (100 mL), EDC.HCl (11.65 g, 64.85 mmol) was added, and stirred at -5 ° C for 15 min, then dimethylamine salt was added. Acid salt (4.21 g, 43.23 mmol), after 15 min, DIPEA (13.97 g, 108.08 mmol) was added, and the reaction was carried out at low temperature for 25 min. After completion of the reaction at room temperature, it was extracted with DCM, washed with 1N HCl, 5% NaHCO _? It was washed with water, dried over anhydrous sodium sulfate and evaporated.

N-tert-butoxycarbonyl-L-leucine-N'-methoxy-N'-carboxamide 32 (0.5 mol) was weighed into a reaction flask, dissolved in 500 mL of tetrahydrofuran, and dropped at -20 ° C. Ethyl magnesium bromide (2.0 M, 750 mL) was added and the mixture was taken to room temperature overnight. The reaction was quenched by dropwise addition of 1N EtOAc (EtOAc)EtOAc.

Weigh 33 (0.4 mol), dissolve in 400 mL of tetrahydrofuran, add piperidine acetate (1.5 mol) and piperidine (1.0 mol) and paraformaldehyde (2.0 mol), reflux for 3 h, then add paraformaldehyde (2.0 mol The reaction was completed by TLC. After adding an appropriate amount of water, the mixture was extracted with ethyl acetate, and washed with 1N diluted hydrochloric acid and brine, and then dried and concentrated to give 34.

Weigh aluminum azide (0.3 mol) and isopropanol (3 mol), add 200 mL of toluene and 34 (0.3 mol), dissolve in 100 mL of toluene, add dropwise to the reaction system at room temperature, drop, react at 50 ° C, TLC The reaction was detected to be complete. After adding an appropriate amount of water, the mixture was extracted with ethyl acetate. The mixture was washed with 1N diluted hydrochloric acid and brine, and then dried and concentrated.

Weigh 35 (0.2 mol), add 200 mL of dichloromethane to dissolve, then add vanadium acetylacetonate (0.04 mol), under nitrogen protection, cool to 0 ° C in an ice bath, slowly add dropwise t-butanol, and stir vigorously overnight. The TLC was used to detect the disappearance of the starting material. After adding an appropriate amount of water, the mixture was extracted with methylene chloride, washed with saturated sodium thiosulfate and saturated brine, and then dried and concentrated.

36 (0.12 mol) was dissolved in 100 mL of dimethyl sulfoxide, diisopropylethylamine (0.24 mol) was added, and pyridine sulfur trioxide (0.24 mol) was added portionwise in an ice bath, and the mixture was allowed to react at room temperature, and the reaction was analyzed by TLC. After completion, the mixture was extracted with ethyl acetate, and washed with 1N diluted hydrochloric acid and brine, and then dried and evaporated.

Compound 37 (1.0 g, 3.69 mmol) was dissolved in anhydrous DCM (10 mL). EtOAc (3 mL) was slowly added dropwise at -5 ° C, stirred for 0.5 hour, then warmed to room temperature and stirred for 3 hours. After completion of the reaction, the reaction mixture was concentrated to give a brown brown oil.

The 39 to 42 synthesis method and the 38 synthesis method are similar in the present invention.

The specific compounds synthesized and their names are as follows.

3. Preparation of a compound of formula (III)

For example, the preparation of N-2-morpholinyl-L-thiazolepropionyl-L-leucyl-L-phenylpropionyl-L-leucyl-methyloxirane (43):

Compound 10 (7 g, 10.55 mmol), HOBt (21.3 g, 15.83 mmol) was dissolved in DCM, EHCCI (3 g, 15.83 mmol) was added, and stirred at -5 ° C for 15 min, then 39 (2.8 g, 10.55 mmol), after 15 min was added DIPEA (4mL, 4.75mmol), the reaction at low temperature 25min, after reaction at room temperature, extracted with DCM, the organic phase washed 1N HCl, 5% NaHCO ₃ wash, brine, dried over anhydrous sodium sulfate, the solvent was evaporated to give compound 43, yield 30%, mp: 108-109 ° C; ¹ H NMR (400 MHz, CDCl ₃ ) δ 0.80 (-CH ₃ , ddd, J = 15.3, 9.0, 4.9 Hz, 6H), 0.90 (-CH _{3 )} , dt, J=8.5, 4.2 Hz, 6H), 1.25 (-CH ₃ , s, 3H), 1.28 (-CH, d, 1H), 1.33 (-CH, d, J = 4.1 Hz, 1H), 1.50 (-CH ₂ , d, J = 3.9 Hz, 2H), 1.64 (-CH ₂ , s, 2H), 2.48 (-CH ₃ , t, 3H), 2.87 (-CH ₂ , d, J = 5.1 Hz, 1H), 3.05–2.97 (-CH ₂ , m, 3H), 3.26–3.07 (-CH ₂ , m, 2H), 3.37–3.27 (-CH ₂ , m, 2H), 3.72 (-CH ₂ , t, 4H), 4.31 - 4.20 (-CH, m, 1H), 4.74 - 4.49 (-CH, m, 3H), 6.42 (-CONH, d, J = 7.1 Hz, 1H), 6.57 (-CONH, d, J = 8.1 Hz, 1H), 7.12 (-Ph, d, J = 1.9 Hz, 1H), 7.15 (-Ph, s, 1H), 7.17 (-Ph, d, J = 1.4 Hz, 1H), 7.25 - 7.19 (-Ph,m,3H), 7.27 (-Ph,d,J=3.1Hz,1H), 8.36(-CONH,d,J=6.4Hz,1H),8.78(-CONH,d,J=2.0Hz , 1H) _{^{13 C NMR (101MHz, CDCl 3}} ) δ16.63,21.20,21.54,22.89,23.29,24.65,24.97,29.61,32.59,36.93,39.88,40.32,49.98,52.19,52.25,52.75,53.74,53.8858.96,61.70 , 66.84, 116.18, 126.71, 128.39, 129.12, 136.96, 152.36, 153.42, 170.66, 170.73, 170.88, 171.78, 207.87; MS (ESI) m/z: 714.0 [M+H] ⁺ , 736.1 [M+Na] ⁺ ;HRMS calcd for C ₃₆ H ₅₂ N ₆ Na O ₇ S, [M+Na] ⁺ 735.35504, found 735.35104.

The synthesis of all compounds in the present invention is similar to that of 43.

The specific compounds synthesized and their names are as follows.

The second part inhibits the determination of proteasome activity

I. Proteasome inhibitory activity

The present invention utilizes a fluorescent polypeptide substrate Suc-Leu-Leu-Val-Tyr-AMC (abbreviated as Suc-LLVY-AMC, Suc represents succinyl group, AMC stands for 7-amide-4-methylcoumarin) to determine proteasome Chymotrypsin-like enzyme activity.

The proteasome used in the present invention is the human erythrocyte 20S proteasome, and the enzyme, fluorescent substrate and test buffer are all purchased from Enzo. The experimental system was 16 μL, in which the substrate was 8 μL, the proteasome 4 μL (0.8 ng), the final concentration was 50 μM, the drug (inhibitor) 4 μL, and the final concentration was 2×10 ^-6 M to 4.88×10 ^-10 M, the last concentration. It is 0M, and the actual concentration is 8×10 ^-6 M～1.95×10 ^-9 M, and the last concentration is 0M. The specific experimental process is as follows:

1, drug configuration:

The drug was weighed and dissolved in DMSO to a concentration of 10 ^-2 M. Pipette 2 μL to 98 μL of DMSO to obtain 2 × 10 ^-4 M, then take 8 μL from the 2 × 10 ^-4 M concentration of the drug and add 198 μL of H ₂ O to obtain 8 × 10 ^-6 M, using the same method. A drug having a concentration of 2 × 10 ^-6 M, 5 × 10 ^-7 M, 1.25 × 10 ^-7 M, 3.12 × 10 ^-8 M, 7.8 × 10 ^-9 M, 1.95 × 10 ^-9 M was obtained, and the last concentration was 0 M. For no medication.

2. Substrate preparation:

25 mg of the fluorescent polypeptide substrate was dissolved in 654 μL of DMSO to obtain a 50 mM stock solution, which was stored at -20 ° C, diluted 500-fold when used, and 8 μL was added to each sample so that the final substrate concentration in the reaction system was 50 μM.

3. Preparation of reaction system:

The 20S proteasome was diluted from 2 ng/μL into a concentration of 8 ng/μL with a buffer solution, added to a 384-well fluorescent plate, 4 μL was added to each well, and 4 μL of the sample to be tested was added to each well. The drug Carfilzomib was a positive control drug and was reacted at 37 ° C for 15 min. After the completion of the reaction, 8 μL of the fluorescent substrate was added to each well, and the reaction was carried out at 37 ° C for 1 hour in the dark, and the fluorescence value was measured using a 360 nm/460 nm fluorescent plate reader (BMG LABTECH POLARstar OPTIMA Microplate Reader).

4, data processing

It was calculated after deduction of background fluorescence resulting product at different concentrations of the drug, using GraphPad Prism software to calculate IC ₅₀ drug concentration proteasome inhibition.

Compound number	IC 50 (nM)	Compound number	IC 50 (nM)	Compound number	IC 50 (nM)
43	22.30	57	>2000	71	44.31
44	354.17	58	24.68	72	18.02
45	23.29	59	17.76	73	12.88
46	34.41	60	23.43	74	29.46
47	40.89	61	14.95	75	>2000
48	26.70	62	9.73	76	20.61
49	30.97	63	17.86	77	>2000
50	812.16	64	15.54	78	NA
51	51.52	65	20.15	79	61.76
52	30.60	66	12.35	80	164.71
53	52.79	67	27.37	Carfilzomib	11.42
54	62.41	68	NA
55	11.56	69	21.45
56	17.24	70	16.31

Second, cell line inhibitory activity

The detection solution used in the present invention is a single-solution cell proliferation assay kit from Promega; the cells used are U266, RPMI8226. The experimental system is 110uL, which contains 90μL of cell suspension, 10μL of detection solution, 10μL of drug (inhibitor), and the final concentration is 4.54×10 ^-8 M～1.77×10 ^-9 M. The last concentration is 0M, the actual concentration is It is 5 × 10 ^-7 M to 1.95 × 10 ^-8 M, and the last concentration is 0M. The specific experimental process is as follows:

1, drug configuration:

The drug was accurately weighed and dissolved in DMSO to 10 ^-2 M. Pipette 1 μL to 199 μL DMSO to obtain 5×10 ^-5 M, then pipette 3.3 μL and 326.7 μL serum-free RPMI1640 medium from 5×10 ^-5 M concentration to obtain 5×10 ^-7 M, 1.5 Diluted by a gradient to obtain 3.3 × 10 ^-7 M, 2.2 × 10 ^-7 M, 1.48 × 10 ^-7 M, 9.87 × 10 ^-8 M, 6.58 × 10 ^-8 M, 4.38 × 10 ^-8 M, 2.92 × 10 ^-8 M, 1.95 × 10 ^-8 M concentration of the drug, the last concentration of 0M is not added.

2. Cell suspension configuration:

After the cells were separately counted, the dilution configuration U266 was 1 × 10 ⁴ /well, and the RPMI 8226 was 1 × 10 ⁴ /well.

3. Preparation of reaction system:

90 μL of cell suspension was added to each well of 96-well fluorescence plate for 24 h; then 10 μL of the sample to be tested was added to each well, and the commercially available drug Carfilzomib was used as a positive control for 24 h. After the reaction was completed, 10 μL was added to each well. The solution was incubated for 2-3 h and the absorbance was measured by a 490 nm fluorescence microplate reader (BMG LABTECH POLARstar OPTIMA Microplate Reader).

4, data processing

The absorbance of the product obtained under different concentrations of the drug after subtraction of the background was calculated, and the IC ₅₀ concentration (nM) of the drug against cytotoxicity was calculated using GraphPad Prism software.

The results of some of the compounds are as follows:

Numbering	RPMI8226	U266B1	Numbering	RPMI8226	U266B1
43	27.59	24.46	70	54.37	45.38
55	42.22	50.88	71	87.44	80.98
56	25.52	36.94	72	77.70	56.87
59	46.95	45.48	73	23.26	16.45
60	35.28	33.1	74	52.90	55.95
61	14.69	23.18	Carfilzomib	39.09	35.72
67	64.91	50.81

Third, the toxicity of compounds

In the present invention, the compounds 43 and 73 were administered at a dose of 15 mg/kg in the tail vein, and 5 ICR rats in each group did not have serious side effects such as death, and the body weight decreased slightly after administration, but the body weight recovered after the 10th day. While the control drug Carfilzomib was administered at a dose of 10 mg/kg, all of the five ICR mice died after one administration, and thus the toxicity of the compound of the present invention was significantly lower than that of the control compound.

The specific experimental process is as follows:

1, drug preparation

0.2110 g of citric acid monohydrate was weighed into a beaker, and 100 ml of physiological water was weighed into a beaker, dissolved, transferred to a container, and placed in a 10 Mm citric acid buffer solution.

Weigh 4mg of the test substance into the beaker, measure 4ml of 10Mm citric acid buffer into the beaker, adjust the pH to 2-3, and ultrasonically dissolve the test substance to obtain 1mg/ml of the test solution. .

4 mg of Carfilzomib powder was weighed into a mortar, ground with a small amount of 10% sulfobutyl-β-cyclodextrin, and added to 4 ml with 10 Mm of citric acid buffer to obtain a 1 mg/ml Carfilzomib test solution.

2. Dosing regimen

Germline: ICR mouse sex: male age: 6-8 weeks; health level: SPF level

Source / Origin: Shanghai Xipuer - Bikai Experimental Animal Co., Ltd.

3. Monitoring indicators:

The body weight of the mice was measured daily from the date of administration, and the breathing, activity, coat color, and death were observed.

4, data processing:

The body weight of the mice was counted and correspondingly calculated by SPSS 19.0, and the differences between the groups were analyzed.

Fourth, solubility experiment

The compounds 43 and 73 and Carfilzomib of the present invention were each dissolved in a pH 2.0 phosphate buffer solution, a pH 4.0 acetate buffer solution, a pH 6.8 phosphate buffer solution, and pure water to form a saturated solution. The saturation solubility was quantitatively determined by HPLC, and the results are shown in the following table. Experimental data showed that compounds 43 and 73 were significantly more soluble in the solvent and water at different pH than the positive drug Carfilzomib.

The therapeutic dose of the compound designed in the present invention can be determined depending on the mode of administration, the purpose of the treatment, the health of the patient, and the prescription of the physician. The concentration and proportion of the compounds contemplated by the present invention in a combination drug will vary depending on a number of factors, including the route of administration, the dosage administered, and the chemical nature (e.g., hydrophobicity). For example, the compounds contemplated by the present invention can be provided in aqueous physiological buffers containing from about 0.1 to 10% w/v of the compound for parenteral administration. Some conventional dosage ranges range from about 1 [mu]g/kg to 1 g/kg per day. In a particular embodiment, the dosage ranges from about 10 [mu]g/kg body weight to 100 mg/kg body weight per day. The dosage will vary depending on the route of administration, the state of health of the patient, the type and degree of progression of the disease or disorder, the relative biological potency of the compound, and the formulation of the excipient. The effective dose can be derived from the dose response curve of the in vitro or animal model test system.

Claims (10)

1. a tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof, which has the structure shown in Formula I,

   

   among them:

   R ₁ is a substituted or unsubstituted C _1-10 alkyl group, a C _3-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group;

   R ₂ is a substituted or unsubstituted C _1-10 alkyl group, a C _1-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group;

   R ₃ is a substituted or unsubstituted C _1-10 alkyl group, a C _3-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group;

   R ₄ is a substituted or unsubstituted C _1-10 alkyl group, a C _3-6 cycloalkyl group, a heterocycloalkyl group, an aryl group or a heterocyclic aryl group;

   Z is selected from the following fragments:

   

   P is hydrogen or a substituted or unsubstituted C _1-10 alkyl group, a C _1-10 alkoxy group, an aryl group, a heterocycloalkyl group or a heterocyclic aryl group.
2. The tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof according to claim 1, wherein when R ₁ , R ₂ , R ₃ and R ₄ are a substituent group, the substituent is C _{1 ～4} alkyl, C _1-4 alkoxy, cyano, hydroxy, decyl, amino, substituted amino or halogen; when P is a substituent, the substituent is C _1-4 alkyl, C _{1 ～ 4} alkoxy group, halogen or C _1-4 halogenated alkyl group.
3. The tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof according to claim 1, wherein the heterocycloalkyl group has 3, 4, 5, 6 or 7 ring-forming atoms; There are 4, 5, 6, 7, 8, 9 or 10 ring-forming atoms; the heterocyclic aryl has 4, 5, 6, 7, 8, 9 or 10 ring-forming atoms.
4. The tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof according to claim 1, wherein:

   R ₁ is a C _1-10 alkyl group, phenyl, naphthyl, anthryl, thiazolyl, thienyl, benzothienyl, imidazolyl, or optionally C _1-4 alkyl, C _{1 ～4} alkoxy, cyano, nitro, hydroxy, decyl, amino or halogen substituted;

   R ₂ is a C _1-10 alkyl group, a C _1-6 cycloalkyl group, a phenyl group, a naphthyl group, an anthracenyl group, a thiazolyl group, a benzothienyl group, an imidazolyl group, or optionally a C _1-4 Alkenyl, C _1-4 alkoxy, cyano, nitro, hydroxy, decyl, amino or halogen substituted;

   R ₃ is C _1-10 alkyl, phenyl, naphthyl, anthracenyl, thiazolyl, thienyl, benzothienyl, imidazolyl, or optionally C _1-4 alkyl, C _{1 ～4} alkoxy, cyano, nitro, hydroxy, decyl, amino or halogen substituted;

   R ₄ is a C _1-10 alkyl group, phenyl, naphthyl, anthryl, thiazolyl, thienyl, benzothienyl, imidazolyl, or optionally C _1-4 alkyl, C _{1 ～4} alkoxy, cyano, nitro, hydroxy, decyl, amino or halogen substituted;

   P is hydrogen, morpholinyl, isoxazolyl, phenyl, naphthyl, tetrahydronaphthyl, n-propoxy or isopropoxy, or optionally C _1-4 alkyl, C _{1 ～4} alkoxy group, halogen or C _1-4 halogenated alkyl group.
5. A method for producing a tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4, which is characterized in that it is synthesized as follows:

   

   Formula IV is the tetrapeptide propylene oxide derivative of the present invention or a pharmaceutically acceptable salt thereof.
6. A pharmaceutical composition comprising the tetrapeptide propylene oxide derivative of any one of claims 1 to 4, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
7. Use of the tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4 for the preparation of a medicament for inhibiting proteasome.
8. Use of a tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4 for the preparation of a medicament for the treatment of inflammation, cancer or hyperproliferative diseases.
9. Use of the tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4 for the preparation of a medicament for the treatment of an immune-related disease.
10. Use of a tetrapeptide propylene oxide derivative or a pharmaceutically acceptable salt thereof according to any one of claims 1 to 4 for the preparation of a medicament for modifying various antigenic peptides produced by a proteasome in an organism.