BRPI0919711B1 - Pharmaceutical compositions with attenuated release of phenolic opioids and the use of said compositions in the treatment of pain. - Google Patents

Abstract

PHARMACEUTICAL COMPOSITIONS WITH ATTENUATED RELEASE OF PHENOLIC OPIOIDS, USES OF SAID COMPOSITIONS, AND METHOD OF REDUCING THE DRUG ABUSE POTENTIAL OF A COMPOSITION CONTAINING A PHENOLIC OPIOID PRODRUG. The present invention relates to pharmaceutical compositions and their methods of use, wherein the pharmaceutical compositions comprise a phenolic opioid prodrug, which provides enzymatically controlled release of a phenolic opioid, and an enzyme inhibitor that interacts with the enzyme(s) mediating the enzymatically controlled release of the phenolic opioid from the prodrug, so as to attenuate the enzymatic cleavage of the prodrug.

Description

Cross-reference to Related Request

[001] This application claims the benefit of priority, under 35 U.S.C. *119(e), to U.S. Provisional Patent Application No. 61/106,400, filed October 17, 2008.

Introduction

[002] The present invention relates to phenolic opioids that are susceptible to creating dependence. Consequently, it is necessary to control access to these drugs. Controlling access to drugs is costly to administer and can lead to denial of treatment for patients who are unable to present themselves for dosing. For example, patients suffering from acute pain may be denied treatment with an opioid unless they have been admitted to a hospital.

[003] International patent application publication number WO 2007/140272 describes certain prodrugs that confer controlled release of phenolic opioids. The prodrugs are resistant to misuse, being stable in the presence of household chemicals such as vinegar or yeast, and require enzymatic activation in the intestines to initiate the release of the phenolic opioid. The prodrugs are believed to release the phenolic opioid via an enzyme-activated cyclization release mechanism. Thus, it is believed that enzyme-induced cleavage of an amide bond confers a nucleophilic nitrogen atom, which then undergoes a cyclization release reaction.

[004] The prodrugs described in WO 2007/140272 resist the release of the phenolic opioid when subjected to conditions commonly used by those intending to abuse the drug, but release the phenolic opioid when administered orally. This provides substantial protection against dependence. However, there are situations in which oral consumption of this prodrug can potentially lead to excessive exposure to the phenolic opioid, either through excessive abuse or accidental use.

Summary

[005] The present description provides pharmaceutical compositions and their methods of use, wherein the pharmaceutical compositions comprise a phenolic opioid prodrug, which provides enzymatically controlled release of a phenolic opioid, and an enzyme inhibitor that interacts with the enzyme(s) that mediate the enzymatically controlled release of the phenolic opioid from the prodrug, so as to attenuate the enzymatic cleavage of the prodrug.

[006] Consequently, according to one aspect, embodiments of the invention include pharmaceutical compositions comprising a trypsin inhibitor and a compound of general formula (I):X-C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5)(I)

[007] or a pharmaceutically acceptable salt thereof, wherein:

[008] X represents a residue of a phenolic opioid, in which the hydrogen atom of the phenolic hydroxyl group is replaced by a covalent bond to form -C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5);

[009] R1 represents a (1-4C)alkyl group;

[0010] R2 and R3, each independently, represent a hydrogen atom or an (1-4C)alkyl group;

[0011] n represents 2 or 3;

[0012] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[0013] R5 represents a hydrogen atom, an N-acyl group or a residue of an amino acid, a dipeptide or an N-acyl derivative of an amino acid or dipeptide.

[0014] The embodiments provide a pharmaceutical composition comprising a trypsin inhibitor and a compound of general formula (II): X-C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5)(II)

[0015] or pharmaceutically acceptable salt thereof, wherein:

[0016] X represents a residue of a phenolic opioid, in which the hydrogen atom of the phenolic hydroxyl group is replaced by a covalent bond to form -C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5);

[0017] R1 is selected from alkyl, substituted alkyl, arylalkyl, substituted arylalkyl, aryl and substituted aryl;

[0018] each R2 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl and aminoacyl;

[0019] each R3 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl and aminoacyl;

[0020] or R2 and R3, together with the carbon to which they are attached, form a substituted cycloalkyl and cycloalkyl group, or two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, form a substituted cycloalkyl or cycloalkyl group;

[0021] n represents an integer from 2 to 4;

[0022] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[0023] R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or dipeptide.

[0024] The embodiments provide a pharmaceutical composition comprising a trypsin inhibitor and a compound of general formula (III): X-C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5)(III)

[0025] or pharmaceutically acceptable salt thereof, wherein:

[0026] X represents a residue of a phenolic opioid, in which the hydrogen atom of the phenolic hydroxyl group is replaced by a covalent bond to form -C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5);

[0027] R1 represents a (1-4C)alkyl group;

[0028] R2 and R3, each independently, represent a hydrogen atom or an (1-4C)alkyl group;

[0029] n represents 2 or 3;

[0030] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[0031] R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or dipeptide.

[0032] The embodiments provide a pharmaceutical composition comprising a trypsin inhibitor and a compound of general formula (IV):

[0033] or pharmaceutically acceptable salt thereof, wherein:

[0034] Ra is hydrogen or hydroxyl;

[0035] Rb is oxo (=O) or hydroxyl;

[0036] the dashed line is a double link or a single link;

[0037] R1 represents a (1-4C)alkyl group;

[0038] R2 and R3, each independently, represent a hydrogen atom or a (1-4C)alkyl group;

[0039] n represents 2 or 3;

[0040] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[0041] R5 represents a hydrogen atom, an N-acyl group or a residue of an amino acid, a dipeptide or an N-acyl derivative of an amino acid or dipeptide.

[0042] The embodiments provide a pharmaceutical composition comprising a trypsin inhibitor and a compound of general formula (V):

[0043] or a pharmaceutically acceptable salt thereof, wherein:

[0044] Ra is hydrogen or hydroxyl;

[0045] Rb is oxo (=O) or hydroxyl;

[0046] the dashed line is a double link or a single link;

[0047] R1 is selected from alkyl, substituted alkyl, arylalkyl, substituted arylalkyl, aryl and substituted aryl;

[0048] each R2 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl and aminoacyl;

[0049] each R3 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl and aminoacyl;

[0050] or R2 and R3, together with the carbon to which they are attached, form a substituted cycloalkyl and cycloalkyl group, or two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, form a substituted cycloalkyl or cycloalkyl group;

[0051] n represents an integer from 2 to 4;

[0052] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[0053] R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or dipeptide.

[0054] The embodiments provide a pharmaceutical composition comprising a trypsin inhibitor and a compound of general formula (VI):

[0055] or a pharmaceutically acceptable salt thereof, wherein:

[0056] Ra is hydrogen or hydroxyl;

[0057] Rb is oxo (=O) or hydroxyl;

[0058] the dashed line is a double link or a single link;

[0059] R1 represents a (1-4C)alkyl group;

[0060] R2 and R3, each independently, represent a hydrogen atom or a (1-4C)alkyl group;

[0061] n represents 2 or 3;

[0062] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[0063] R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or dipeptide.

Brief description of the figures

[0064] Figure 1 is a graph comparing mean blood concentrations over time of hydromorphone (HM) after PO administration to rats of Compound 1 alone and Compound 1 with varying amounts of Glycine max (soybean) trypsin inhibitor (SBTI).

[0065] Figure 2 is a graph comparing mean plasma concentrations over time of hydromorphone (HM) after PO administration to rats of Compound 1 alone, Compound 1 with ovalbumin (OVA), and Compound 1 with ovalbumin and SBTI.

[0066] Figure 3 is a graph comparing individual blood concentrations over time of hydromorphone (HM) after PO administration to rats of Compound 1 alone and Compound 1 with Bowman-Birk trypsin-chymotrypsin inhibitor (BBSI).

[0067] Figure 4 is a graph comparing mean plasma concentrations over the release time of hydromorphone (HM) after PO administration to rats of Compound 2 alone and Compound 2 with SBTI.

[0068] Figure 5 is a graph comparing mean plasma concentrations over the release time of hydromorphone (HM) after PO administration to rats of Compound 3 alone and Compound 3 with SBTI.

[0069] Figure 6 is a graph comparing mean plasma concentrations over the release time of hydromorphone (HM) after PO administration to rats of Compound 4 alone and Compound 4 with SBTI.

[0070] Figures 7A and 7B are graphs indicating the results of exposure to a certain combination of Compound 4 and trypsin, in the absence of any trypsin inhibitor or in the presence of SBTI, Compound 107, Compound 108, or Compound 109. Figure 7A illustrates the disappearance of Compound 4, and Figure 7B illustrates the appearance of hydromorphone, over time under these conditions.

[0071] Figure 8 is a graph comparing mean plasma concentrations over time of hydromorphone (HM) release after PO administration to rats of Compound 3 alone and Compound 3 with Compound 101.

[0072] Figure 9 is a graph comparing mean plasma concentrations over the release time of hydromorphone (HM) after PO administration to rats of Compound 4 alone and Compound 4 with Compound 101.

Definitions

[0073] The following terms have the following meanings, unless otherwise indicated. Any undefined terms have their meanings recognized in the field.

[0074] As used herein, the term "alkyl," by itself or as part of another substituent, refers to a saturated monovalent hydrocarbon radical with a branched or linear chain derived by the removal of a hydrogen atom from a single carbon atom of a parent alkane. Typical alkyl groups include, but are not limited to, methyl; ethyl, propyls, such as propan-1-yl or propan-2-yl, and butyls, such as butan-1-yl, butan-2-yl, 2-methyl-propan-1-yl or 2-methyl-propan-2-yl. In some embodiments, an alkyl group comprises from 1 to 20 carbon atoms. In other embodiments, an alkyl group comprises from 1 to 10 carbon atoms. In still other embodiments, an alkyl group comprises from 1 to 6 carbon atoms, as well as from 1 to 4 carbon atoms.

[0075] "Alkenyl", by itself or as part of another substituent, refers to an unsaturated alkyl radical with a branched, linear or cyclic chain having at least one carbon-carbon double bond derived by the removal of a hydrogen atom from a single carbon atom of a parent alkene. The group may be in the cis or trans conformation around the double bond(s). Typical alkenyl groups include but are not limited to ethenyl; propenyls, such as prop-1-en-1-yl, prop-1-en-2-yl, prop-2-en-1-yl (allyl), prop-2-en-2-yl, cycloprop-1-en-1-yl; cycloprop-2-en-1-yl; butenyls, such as but-1-en-1-yl, but-1-en-2-yl, 2-methyl-prop-1-en-1-yl, but-2-en-1-yl, but-2-en-1-yl, but-2-en-2-yl, buta-1,3-dien-1-yl, buta-1,3-dien-2-yl, cyclobut-1-en-1-yl, cyclo-but-1-en-3-yl, cyclobuta-1,3-dien-1-yl, etc., and the like.

[0076] "Alkynyl", by itself or as part of another substituent, refers to an unsaturated alkyl radical with a branched, linear or cyclic chain having at least one carbon-carbon triple bond derived by the removal of a hydrogen atom from a single carbon atom of a parent alkyne. Typical alkynyl groups include but are not limited to ethinyl; propynyls, such as prop-1-yn-1-yl, prop-2-yn-1-yl, etc.; butynyls, such as but-1-yn-1-yl, but-1-yn-3-yl, but-3-yn-1-yl, etc., and the like.

[0077] "Acyl", by itself or as part of another substituent, refers to a -C(O)R30 radical, wherein R30 is hydrogen, alkyl, cycloalkyl, cycloheteroalkyl, aryl, arylalkyl, heteroalkyl, heteroaryl, heteroarylalkyl, as defined herein. Representative examples include but are not limited to formyl, acetyl, cyclohexylcarbonyl, cyclohexylmethylcarbonyl, benzoyl, benzylcarbonyl, piperonyl and the like. Substituted acyl refers to substituted versions of acyl and include, for example but not limited to, succinyl and malonyl.

[0078] The term "aminoacyl" and "amide" refers to the group -C(O)NR21R22, wherein R21 and R22 are independently selected from the group consisting of hydrogen, alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, aryl, substituted aryl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, heteroaryl, substituted heteroaryl, heterocyclic and substituted heterocyclic, and wherein R21 and R22 are optionally joined to the nitrogen attached to them, forming a heterocyclic or substituted heterocyclic group, and wherein alkyl, substituted alkyl, alkenyl, substituted alkenyl, alkynyl, substituted alkynyl, cycloalkyl, substituted cycloalkyl, cycloalkenyl, substituted cycloalkenyl, aryl, substituted aryl, heteroaryl, substituted heteroaryl, heterocyclic, and substituted heterocyclic are as defined herein.

[0079] "Alkoxy", by itself or as part of another substituent, refers to a radical -OR31 wherein R31 represents an alkyl or cycloalkyl group, as defined herein. Representative examples include but are not limited to methoxy, ethoxy, propoxy, butoxy, cyclohexyloxy and the like.

[0080] "Alkoxycarbonyl", by itself or as part of another substituent, refers to a radical -C(O)OR31 wherein R31 represents an alkyl or cycloalkyl group, as defined herein. Representative examples include, but are not limited to, methoxycarbonyl, ethoxycarbonyl, propoxycarbonyl, butoxycarbonyl, cyclohexyloxycarbonyl and the like.

[0081] "Aryl", by itself or as part of another substituent, refers to a monovalent aromatic hydrocarbon radical derived by removing a hydrogen atom from a single carbon atom of a parent aromatic ring system. Typical aryl groups include, but are not limited to, groups derived from aceantrylene, acenaphtylene, acephenantrylene, anthracene, azulene, benzene, chrysene, coronene, fluoranthene, fluorene, hexacene, hexaphene, hexalene, as-indacene, s-indacene, indane, indene, naphthalene, octacene, octaphene, octalene, ovalene, penta-2,4-diene, pentacene, pentalene, pentaphene, perylene, phenalene, phenanthrene, picene, pleiadene, pyrene, pyranthrene, rubicene, triphenylene, trinaphthalene, and the like. In some embodiments, an aryl group comprises from 6 to 20 carbon atoms. In other embodiments, an aryl group comprises from 6 to 12 carbon atoms. Examples of aryl groups are phenyl and naphthyl.

[0082] "Arylalkyl", by itself or as part of another substituent, refers to an acyclic alkyl radical in which one of the hydrogen atoms bonded to a carbon atom, typically a terminal or sp3 carbon atom, is substituted by an aryl group. Typical arylalkyl groups include but are not limited to benzyl, 2-phenylethyl-1-yl, naphthylmethyl, 2-naphthylet-1-yl, naphthobenzyl, 2-naphthophenylethyl-1-yl and the like. In some embodiments, an arylalkyl group is (C7-C30) arylalkyl, for example, the alkyl portion of the arylalkyl group is (C1-C10) and the aryl portion is (C6-C20). In other embodiments, an arylalkyl group is (C7-C20) arylalkyl, for example, the alkyl portion of the arylalkyl group is (C1-C8) and the aryl portion is (C6-C12).

[0083] Compounds can be identified by their chemical structure and/or chemical name. The compounds described herein may contain one or more chiral centers and/or double bonds and, consequently, may exist in the form of stereoisomers, such as isomers in double bonds (i.e., geometric isomers), enantiomers, or diastereomers. Accordingly, all possible enantiomers and stereoisomers of the compounds, including the stereoisomerically pure form (e.g., geometrically pure, enantiomerically pure, or diastereomerically pure) and enantiomeric and stereoisomeric mixtures are included in the description of the compounds herein. Enantiomeric and stereoisomeric mixtures can be resolved into their component enantiomers or stereoisomers using separation techniques or chiral synthesis techniques well-known to the skilled. The compounds may also exist in various tautomeric forms, including the enol form, the keto form, and mixtures thereof. Accordingly, the chemical structures represented here encompass all possible tautomeric forms of the illustrated compounds. The compounds described also include isotopically labeled compounds, in which one or more atoms have an atomic mass different from the atomic mass conventionally found in nature. Examples of isotopes that may be incorporated into the compounds described here include, but are not limited to, 2H, 3H, 11C, 13C, 14C, 15N, 18O, 17O, etc. The compounds may exist in non-solvated as well as solvated forms, including hydrated forms. Certain compounds may exist in multiple crystalline or amorphous forms. In general, all physical forms are equivalent for the uses contemplated herein and are intended to fall within the scope of this description.

[0084] "Cycloalkyl", by itself or as part of another substituent, refers to a saturated cyclic alkyl radical. Typical cycloalkyl groups include, but are not limited to, groups derived from cyclopropane, cyclobutane, cyclopentane, cyclohexane, and the like. In some embodiments, the cycloalkyl group is (C3-C10) cycloalkyl. In other embodiments, the cycloalkyl group is (C3-C7) cycloalkyl.

[0085] "Cycloheteroalkyl", by itself or as part of another substituent, refers to a saturated cyclic alkyl radical in which one or more carbon atoms (and any associated hydrogen atoms) are independently substituted by an identical or different heteroatom. Typical heteroatoms to substitute the carbon atom(s) include but are not limited to N, P, O, S, Si, etc. Typical cycloheteroalkyl groups include but are not limited to groups derived from epoxides, azirines, thiranes, imidazolidine, morpholine, piperazine, piperidine, pyrazolidine, pyrrolidine, quinuclidine and the like.

[0086] "Heteroalkyl, heteroalkenyl and heteroalkynyl", by themselves or as part of another substituent, refer to alkyl, alkenyl and alkynyl groups, respectively, in which one or more of the carbon atoms (and any associated hydrogen atoms) are independently substituted by identical or different heteroatomic groups. Typical heteroatomic groups that may be included in these groups include, but are not limited to, -O-, -S-, -O-O-, -S-S-, -O-S-, -NR37R38-, =N-N=, -N=N-, -N=N-NR3+R40, -PR41-, -P(O)2-, -POR42-, -O-P(O)2-, -SO-, -SO2-, -SnR43R44- and the like, wherein R37, R38, R3+, R40, R41, R42, R43 and R44 are independently hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, arylalkyl, substituted arylalkyl, cycloalkyl, substituted cycloalkyl, cycloheteroalkyl, substituted cycloheteroalkyl, heteroalkyl, substituted heteroalkyl, heteroaryl, substituted heteroaryl. heteroarylalkyl or substituted heteroarylalkyl.

[0087] "Heteroaryl", by itself or as part of another substituent, refers to a monovalent heteroaromatic radical derived by removing a hydrogen atom from a single atom of a parent heteroaromatic ring system. Typical heteroaryl groups include but are not limited to groups derived from acridine, arsindole, carbazole, β-carboline, chromane, chromene, cinoline, furan, imidazole, indazole, indole, indoline, indolizine, isobenzofuran, isochromene, isoindole, isoindoline, isoquinoline, isothiazole, isoxazole, naphthyridine, oxadiazole, oxazole, perimidine, phenanthridine, phenanthroline, phenazine, phthalazine, pteridine, purine, pyran, pyrazine, pyrazole, pyridazine, pyridine, pyrimidine, pyrrole, pyrrolizine, quinazoline, quinoline, quinolizine, quinoxaline, tetrazole, thiadiazole, thiazole, thiophene, triazole, xanthene and the like. In some embodiments, the heteroaryl group is a heteroaryl group with 5-20 members. In other embodiments, the heteroaryl group is a heteroaryl group with 5-10 members. In still other embodiments, heteroaryl groups are derivatives of thiophene, pyrrole, benzothiophene, benzofuran, indole, pyridine, quinoline, imidazole, oxazole, and pyrazine.

[0088] "Heteroarylalkyl", by itself or as part of another substituent, refers to an acyclic alkyl radical in which one of the hydrogen atoms bonded to a carbon atom, typically a terminal or sp3 carbon atom, is substituted by a heteroaryl group. In some embodiments, the heteroarylalkyl group is a 6-30 membered heteroarylalkyl group, for example, the alkyl portion of the heteroarylalkyl group has 1-10 members and the heteroaryl portion is a 5-20 membered heteroaryl group. In other embodiments, the heteroarylalkyl group is a 6-20 membered heteroarylalkyl group, for example, the alkyl portion of the heteroarylalkyl group has 1-8 members and the heteroaryl portion is a 5-12 membered heteroaryl group.

[0089] "Opioid" refers to a chemical substance that exerts its pharmacological action by interacting with opioid receptors. "Phenolic opioid" refers to a subset of opioids that contain a phenol group. Examples of phenolic opioids are provided below.

[0090] "Parent Aromatic Ring System", by itself or as part of another substituent, refers to an unsaturated cyclic or polycyclic ring system with a conjugated π electron system. Specifically included in the definition of "parent aromatic ring system" are fused ring systems in which one or more of the rings are aromatic and one or more of the rings are saturated or unsaturated, such as, for example, fluorene, indane, indene, phenolene, etc. Typical parent aromatic ring systems include, but are not limited to, aceantrylene, acenaphtylene, acephenantrylene, anthracene, azulene, benzene, chrysene, coronene, fluoranthene, fluorene, hexacene, hexaphene, hexalene, as-indacene, s-indacene, indane, indene, naphthalene, octacene, octaphene, octalene, ovalene, penta-2,4-diene, pentacene, pentalene, pentaphene, perylene, phenalene, phenanthrene, picene, pleiadene, pyrene, pyranthrene, rubicene, triphenylene, trinaphthalene, and the like.

[0091] "Paternal Heteroaromatic Ring System", by itself or as part of another substituent, refers to a paternal aromatic ring system in which one or more carbon atoms (and any associated hydrogen atoms) are independently substituted with identical or different heteroatoms. Typical heteroatoms to substitute the carbon atoms include but are not limited to N, P, O, S, Si, etc. Specifically included in the definition of "paternal heteroaromatic ring systems" are fused ring systems in which one or more of the rings are aromatic and one or more of the rings are saturated or unsaturated, such as, for example, arsindole, benzodioxane, benzofuran, chromane, chromene, indole, indoline, xanthene, etc. Typical paternal heteroaromatic ring systems include but are not limited to arsindole, carbazole, □-carboline, chromane, chromene, cynoline, furan, imidazole, indazole, indole, indoline, indolizine, isobenzofuran, isochromene, isoindole, isoindoline, isoquinoline, isothiazole, isoxazole, naphthyridine, oxadiazole, oxazole, perimidine, phenanthridine, phenanthroline, phenazine, phthalazine, pteridine, purine, pyran, pyrazine, pyrazole, pyridazine, pyridine, pyrimidine, pyrrole, pyrrolizine, quinazoline, quinoline, quinolizine, quinoxaline, tetrazole, thiadiazole, thiazole, thiophene, triazole, xanthene and the like.

[0092] "Pharmaceutical composition" refers to at least one compound and a pharmaceutically acceptable vehicle with which the compound is administered to a patient.

[0093] "Pharmaceutically acceptable salt" refers to a salt of a compound that possesses the desired pharmacological activity of the parent compound. These salts include: (1) acid addition salts, formed with inorganic acids, such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like; or formed with organic acids, such as acetic acid, propionic acid, hexanoic acid, cyclopentanopropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl)benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, 4-methylbicyclo[2.2.2]-oct-2-ene-1-carboxylic acid, glucoheptonic acid, acid 3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid, laurylsulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid and the like; or (2) salts formed when an acidic proton present in the parent compound is replaced by a metal ion, for example, an alkali metal ion, an alkaline earth ion, or an aluminum ion; or coordinated with an organic base, such as ethanolamine, diethanolamine, triethanolamine, N-methylglucamine and the like.

[0094] The term "solvate," as used herein, refers to a complex or aggregate formed by one or more molecules of a solute, that is, a compound of the embodiments, or a pharmaceutically acceptable salt thereof, and one or more molecules of a solvent. These solvates are typically crystalline solids having a substantially fixed molar ratio of solute and solvent. Representative solvents include, by way of example, water, methanol, ethanol, isopropanol, acetic acid, and the like. When the solvent is water, the solvate formed is a hydrate.

[0095] "Pharmaceutically acceptable vehicle" refers to a diluent, adjuvant, excipient, or carrier with which or in which a compound is administered.

[0096] "Patient" includes humans, and also other mammals, such as livestock, zoo animals and companion animals, such as a cat, dog or horse.

[0097] "Prevent" or "prevention" or "prophylaxis" refers to a reduction in the risk of a condition, such as pain, occurring.

[0098] "Prodrug" refers to a derivative of an active agent that requires a transformation within the body to release the active agent. It is common, but not necessary, for prodrugs to be pharmacologically inactive until they are converted into the active agent.

[0099] "Carrier moiety" refers to a form of protecting group that, when used to mask a functional group in an active agent, converts the active agent into a prodrug. Typically, the carrier moiety will be linked to the drug via linkage(s) that is/are cleaved by enzymatic or non-enzymatic means in vivo.

[00100] "Protecting group" refers to a group of atoms that, when attached to a reactive functional group in a molecule, masks, reduces, or prevents the reactivity of the functional group. Examples of protecting groups can be found in Green et al., "Protective Groups in Organic Chemistry," (Wiley, 2nd edition 1991), and Harrison et al., "Compendium of Synthetic Organic Methods," Volumes 1-8 (John Wiley and Sons, 1971-1996). Representative amino protecting groups include, but are not limited to, formyl, acetyl, trifluoroacetyl, benzyl, benzyloxycarbonyl ("CBZ"), tert-butoxycarbonyl ("Boc"), trimethylsilyl ("TMS"), 2-trimethylsilylethanesulfonyl ("SES"), trityl and substituted trityl, allyloxycarbonyl, 9-fluorenylmethyloxycarbonyl ("FMOC"), nitroveratryloxycarbonyl ("NVOC"), and the like. Representative hydroxy protecting groups include, but are not limited to, those in which the hydroxyl group is acylated or alkylated, such as benzyl and trityl ethers, as well as alkyl ethers, tetrahydropyranyl ethers, trialkylsilyl ethers, and allyl ethers.

[00101] "Replaced" refers to a group in which one or more hydrogen atoms are independently replaced by identical or different substitutes^). Typical substituents include, but are not limited to, alkylenedioxy (such as methylenedioxy), -M, -R60, -O-, =O, -OR60, -SR60, -S-, =S, -NR60R61, =NR60, -CF3, -CN, -OCN, -SCN, -NO, -NO2, =N2, -N3, -S(O)2O-, -S(O)2OH, -S(O)2R60, -OS(O)2O-, -OS(O)2R60, -P(O)(O-)2, -P(O)(OR60)(O-), -OP(O)(OR60)(OR61), -C(O)R60, -C(S)R60, -C(O)OR60, -C(O)NR60R61, -C(O)O-, -C(S)OR60, -NR62C(O)NR60R61, -NR62C(S)NR60R61, -NR62C(NR63)NR60R61 and -C(NR62)NR60R61, wherein M is a halogen; R60, R61, R62 and R63 are independently hydrogen, alkyl, substituted alkyl, alkoxy, substituted alkoxy, cycloalkyl, substituted cycloalkyl, cycloheteroalkyl, substituted cycloheteroalkyl, aryl, substituted aryl, heteroaryl or substituted heteroaryl, or optionally R60 and R61, together with the nitrogen atom to which they are attached, form a cycloheteroalkyl or substituted cycloheteroalkyl ring.

[00102] "Treating" or "treatment" of any condition, such as pain, refers, in certain modalities, to the improvement of the condition (i.e., halting or reducing the development of the condition). In certain modalities, "treating" or "treatment" refers to the improvement of at least one physical parameter, which may or may not be discernible by the patient. In certain modalities, "treating" or "treatment" refers to the inhibition of the condition, either physically (e.g., stabilization of a discernible symptom), physiologically (e.g., stabilization of a physical parameter), or both. In certain modalities, "treating" or "treatment" refers to delaying the onset of the condition.

[00103] "Therapeutically effective amount" means the amount of a compound that, when administered to a patient to prevent or treat a condition, such as pain, is sufficient to effect that treatment. The "therapeutically effective amount" will vary depending on the compound, the condition and its severity, and the patient's age, weight, etc.

Detailed Description

[00104] Before the present invention is further described, it should be understood that this invention is not limited to the particular embodiments described, as these may obviously vary. It should also be understood that the terminology used herein is intended only to describe particular embodiments and is not intended to be limiting, as the scope of the present invention will be limited only by the appended claims.

[00105] It should be noted that, as used here and in the accompanying claims, the singular forms "a", "an" and "the" include plural referents, unless the context clearly dictates otherwise. It should also be noted that claims may be drafted in such a way as to exclude any optional element. As such, this statement is intended to serve as a precedent for the use of exclusive terminology, such as "only", "just" and the like, related to the recitation of claim elements, or the use of a "negative" limitation.

[00106] It should be understood that, as used herein, the term "an" entity refers to one or more of that entity. For example, a compound refers to one or more compounds. As such, the terms "an," "a," "one or more," and "at least one" may be used interchangeably. Similarly, the terms "comprising," "including," and "possessing" may be used interchangeably.

[00107] The publications discussed herein are provided only by their disclosure prior to the filing date of this application. Nothing presented herein should be construed as an admission that the present invention is not entitled to anticipate such publications by virtue of prior invention. Additionally, the publication dates provided may differ from the actual publication dates and may need to be independently verified.

[00108] Unless otherwise defined, all technical and scientific terms used herein have the same meaning usually understood by a professional in the field to which this invention pertains. Although any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention, preferred methods and materials will now be described. All publications mentioned herein are incorporated herein by reference to disclose and describe the methods and/or materials in connection with which the publications are cited.

[00109] Unless otherwise noted, the methods and techniques of the present embodiments are generally implemented in accordance with conventional methods well-known in the field and as described in various general and more specific references that are cited and discussed throughout the present specification. See, for example, Loudon, "Organic Chemistry", Fourth Edition, New York: Oxford University Press, 2002, pages 360-361, 1084-1085; Smith and March, "March's Advanced Organic Chemistry: Reactions, Mechanisms, and Structure", Fifth Edition, Wiley-Interscience, 2001.

[00110] The nomenclature used here to designate the compounds in question is illustrated in the Examples presented here. Where possible, this nomenclature has generally been derived using the commercially available AutoNom software (MDL, San Leandro, Calif.).

[00111] It has now been discovered that attenuated release of a phenolic opioid can be achieved by administering a soy-derived trypsin inhibitor in combination with a particular prodrug described in WO 2007/140272.

Representative Modalities

[00112] This description provides pharmaceutical compositions, and their methods of use, wherein the pharmaceutical compositions comprise a phenolic opioid prodrug, which provides enzymatically controlled release of a phenolic opioid, and an enzyme inhibitor, which interacts with the enzyme(s) mediating the enzymatically controlled release of the phenolic opioid from the prodrug, so as to attenuate the enzymatic cleavage of the prodrug. The description provides pharmaceutical compositions comprising a trypsin inhibitor and a phenolic opioid prodrug containing a trypsin unstable moiety which, when cleaved, facilitates the release of the phenolic opioid. Examples of phenolic opioid prodrugs and trypsin inhibitors are described below.

Phenolic opioid prodrugs

[00113] According to certain embodiments, a phenolic opioid prodrug is provided that provides enzymatically controlled release of a phenolic opioid. The phenolic opioid prodrug is a corresponding compound in which the phenolic hydrogen atom has been replaced by a leaving spacer group containing a nitrogen nucleophile that is protected with a portion suitable for enzymatic cleavage, wherein the configuration of the leaving spacer group and the nitrogen nucleophile is such that, by enzymatic cleavage of the portion suitable for cleavage, the nitrogen nucleophile is able to form a cyclic urea, releasing the compound from the leaving spacer group so as to provide a phenolic opioid.

[00114] The enzyme capable of cleaving the portion suitable for enzymatic cleaving may be a peptidase - in which the portion suitable for enzymatic cleaving is linked to the nucleophilic nitrogen through an amide bond (for example, a peptide: -NHCO-). In some embodiments, the enzyme is a protein digestive enzyme.

Formulas I-VI

[00115] As shown here, Formula I describes compounds of Formula II, wherein R1 is an (1-4C)alkyl group; R2 and R3 each independently represent a hydrogen atom or an (1-4C)alkyl group, and R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid, or a dipeptide.

[00116] Formula III describes compounds of Formula II, wherein R1 is an (1-4C)alkyl group; R2 and R3, each independently, represent a hydrogen atom or an (1-4C)alkyl group, and R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid, or a dipeptide.

[00117] Formula IV describes compounds of Formula I, in which "X" is structurally substituted with certain phenolic opioids.

[00118] As also shown here, Formula IV describes compounds of Formula V, wherein R1 is an (1-4C)alkyl group; R2 and R3, each independently, represent a hydrogen atom or an (1-4C)alkyl group, and R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or a dipeptide.

[00119] Formula VI describes compounds of Formula V, wherein R1 is an (1-4C)alkyl group; R2 and R3 each independently represent a hydrogen atom or an (1-4C)alkyl group, and R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid, or a dipeptide.

[00120] For Formulas l-lll, X represents a residue of a phenolic opioid, in which the hydrogen atom of the phenolic hydroxyl group is replaced by a covalent bond, forming -C(O)-NR1- (C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5).

[00121] As described above, "opioid" refers to a chemical substance that exerts its pharmacological action through interaction with opioid receptors. "Phenolic opioid" refers to a subset of opioids that contain a phenol group. For example, phenolic opioids include but are not limited to buprenorphine, dihydroetorphine, diprenorphine, etorphine, hydromorphone, levorphanol, morphine (and metabolites thereof), nalmefene, naloxone, N-methylnaloxone, naltrexone, N-methylnaltrexone, oxymorphone, oripavine, ketobemidone, dezocine, pentazocine, phenazocine, butorphanol, nalbuphine, meptazinol, O-desmethyltramadol, tapentadol, nalorphine. The structures of the aforementioned phenolic opioids are shown below:

[00122] In certain embodiments, the phenolic opioid is oxymorphone, hydromorphone, or morphine. Formulas I - VI are now described in more detail below. Formula I

[00123] The compounds of formula (I) correspond to compounds described in WO 2007/140272, in which the nucleophilic nitrogen atom is linked to an L-arginine or L-lysine residue.

[00124] Examples of values for the phenolic opioid as provided in X are oxymorphone, hydromorphone, and morphine.

[00125] Examples of values for R1 are methyl and ethyl groups.

[00126] Examples of values for each of R2 and R3 are hydrogen atoms. An example of a value for n is 2.

[00127] In one embodiment, R4 represents - CH2CH2CH2NH(C=NH)NH2.

[00128] An amino acid can be a naturally occurring amino acid. It will be appreciated that naturally occurring amino acids usually have the L configuration.

[00129] With reference to R5, examples of particular values are:

[00130] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group;

[00131] for an amino acid: alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine or valine, and

[00132] for a dipeptide: a combination of any two amino acids selected independently from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine and valine.

[00133] Examples of particular values for R5 are:

[00134] a hydrogen atom;

[00135] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group, and

[00136] for a residue of an amino acid, a dipeptide, or an N-acyl derivative of an amino acid or dipeptide: glycinyl or N-acetylglycinyl.

[00137] In one embodiment, R5 represents N-acetyl, glycinyl or N-acetylglycinyl, such as N-acetyl.

[00138] An example of the group represented by -C(O)-CH(R4)-NH(R5) is N-acetylarginyl.

[00139] In a particular embodiment, the compound of Formula (I) is hydromorphone 3-(N-methyl-N-(2-N'-acetylarginylamino)) ethylcarbamate, or a pharmaceutically acceptable salt thereof. This compound is described in Example 3 of WO 2007/140272. Formula II

[00140] The embodiments provide a compound of general formula (II): X-C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5)(II)

[00141] or pharmaceutically acceptable salt thereof, wherein:

[00142] X represents a residue of a phenolic opioid, in which the hydrogen atom of the phenolic hydroxyl group is replaced by a covalent bond, forming -C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5);

[00143] R1 is selected from alkyl, substituted alkyl, arylalkyl, substituted arylalkyl, aryl and substituted aryl;

[00144] each R2 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl and aminoacyl;

[00145] each R3 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl and aminoacyl;

[00146] or R2 and R3, together with the carbon to which they are attached, form a cycloalkyl or substituted cycloalkyl group, or two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, form a cycloalkyl or substituted cycloalkyl group;

[00147] n represents an integer from 2 to 4;

[00148] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[00149] R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or dipeptide.

[00150] In formula II, examples of values for the phenolic opioid as provided in X are oxymorphone, hydromorphone, and morphine.

[00151] In formula II, R1 can be selected from alkyl, substituted alkyl, aryl alkyl, substituted aryl alkyl, aryl, and substituted aryl. In certain cases, R1 is (1-6C)alkyl. In other cases, R1 is (1-4C)alkyl. In other cases, R1 is methyl or ethyl. In other cases, R1 is methyl. In some cases, R1 is ethyl.

[00152] In certain cases, R1 is a substituted alkyl group. In certain cases, R1 is an alkyl group substituted with a carboxyl group or a carboxyl ester. In certain cases, R1 is -(CH2)5-COOH, -(CH2)5-COOCH3, or -(CH2)5-COOCH2CH3.

[00153] In certain cases, in formula II, R1 is an arylalkyl or substituted arylalkyl. In certain cases, in formula II, R1 is an arylalkyl. In certain cases, R1 is a substituted arylalkyl. In certain cases, R1 is a carboxyl-substituted arylalkyl group or a carboxyl ester. In certain cases, R1 is -(CH2)q(C6H4)-COOH, -(CH2)q(C6H4)-COOCH3 or -(CH2)q(CeH4)-COOCH2CH3, where q is an integer from one to 10. In certain cases, R1 is -CH2(C6H4)-COOH, -CH2(C6H4)-COOCH3 or -CH2(C6H4)-COOCH2CH3.

[00154] In certain cases, in formula II, R1 is an aryl group. In certain cases, R1 is a substituted aryl group. In certain cases, R1 is an aryl group substituted with a carboxyl group or a carboxyl ester. In certain cases, R1 is -(C6H4)-COOH, -(C6H4)-COOCH3, or -(CIH4)-COOCH2CH3.

[00155] In formula II, each R2 can be independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl, and aminoacyl. In certain cases, R2 is hydrogen or alkyl. In certain cases, R2 is hydrogen. In certain cases, R1 is alkyl. In certain cases, R2 is acyl. In certain cases, R2 is aminoacyl.

[00156] In formula II, each R3 can be independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl, and aminoacyl. In certain cases, R3 is hydrogen or alkyl. In certain cases, R3 is hydrogen. In certain cases, R3 is alkyl. In certain cases, R3 is acyl. In certain cases, R3 is aminoacyl.

[00157] In certain cases, R2 and R3 are hydrogen. In certain cases, R2 and R3 on the same carbon are both alkyl. In certain cases, R2 and R3 on the same carbon are methyl. In certain cases, R2 and R3 on the same carbon are ethyl.

[00158] In formula II, R2 and R3, together with the carbon to which they are attached, can form a cycloalkyl or substituted cycloalkyl group, or two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, can form a cycloalkyl or substituted cycloalkyl group. In certain cases, R2 and R3, together with the carbon to which they are attached, can form a cycloalkyl group. Thus, in certain cases, R2 and R3 on the same carbon form a spirocycle. In certain cases, R2 and R3, together with the carbon to which they are attached, can form a substituted cycloalkyl group. In certain cases, two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, can form a cycloalkyl group. In certain cases, two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, can form a substituted cycloalkyl group.

[00159] In certain cases, one of the R2 and R3 is an aminoacyl group.

[00160] In certain cases, one of the R2 and R3 groups is an aminoacyl group comprising phenylenediamine. In certain cases, one of the R2 and R3 groups is

[00161] wherein each R10 is independently selected from hydrogen, alkyl, substituted alkyl and acyl and R11 is alkyl or substituted alkyl. In certain cases, at least one of the R10 is acyl. In certain cases, at least one of the R10 is alkyl or substituted alkyl. In certain cases, at least one of the R10 is hydrogen. In certain cases, both R10 are hydrogen.

[00162] In certain cases, one of R2 and R3 is

[00163] where R10 is hydrogen, alkyl, substituted alkyl or acyl. In certain cases, R10 is acyl. In certain cases, R10 is alkyl or substituted alkyl. In certain cases, R10 is hydrogen.

[00164] In certain cases, R2 or R3 can modulate an intramolecular cyclization rate. R2 or R3 can accelerate an intramolecular cyclization rate, compared to the corresponding molecule in which R2 and R3 are both hydrogen. In certain cases, R2 or R3 comprise an electron-withdrawing group or an electron-donating group. In certain cases, R2 or R3 comprise an electron-withdrawing group. In certain cases, R2 or R3 comprise an electron-donating group.

[00165] Atoms and groups capable of functioning as electron-withdrawing substituents are well known in the field of organic chemistry. They include electronegative atoms and groups containing electronegative atoms. The function of these groups is to decrease the basicity or protonation state of a nucleophilic nitrogen in the beta position via inductive electron density withdrawal. These groups can also be positioned in other positions along the alkylene chain. Examples include halogen atoms (e.g., a fluorine atom), acyl groups (e.g., an alkanoyl group, an aroyl group, a carboxyl group, an alkoxycarbonyl group, an aryloxycarbonyl group, or an aminocarbonyl group (such as a carbamoyl, alkylaminocarbonyl, dialkylaminocarbonyl, or arylaminocarbonyl group)), an oxo (=O) substituent, a nitrile group, a nitro group, ether groups (e.g., an alkoxy group), and phenyl groups containing a substituent in the ortho position, in the para position, or both the ortho and para positions, wherein each substituent is independently selected from a halogen atom, a fluoroalkyl group (such as trifluoromethyl), a nitro group, a cyano group, and a carboxyl group. Each of the electron-withdrawing substituents may be selected independently of these.

[00166] In certain cases, -[C(R2)(R3)]n- is selected from -CH(CH2F)CH(CH2F)-; -CH(CHF2)CH(CHF2)-; -CH(CF3)CH(CF3)-; - CH2CH(CF3)-; -CH2CH(CHF2)-; -CH2CH(CH2F)-; -CH2CH(F)CH2-; - CH2C(F2)CH2-; -CH2CH(C(O)NR20R21)-; -CH2CH(C(O)OR22)-;-CH2CH(C(O)OH)-; -CH(CH2F)CH2CH(CH2F)-; -CH(CHF2)CH2CH(CHF2)- ; -CH(CF3)CH2CH(CF3)-; -CH2CH2CH(CF3)-; -CH2CH2CH(CHF2)-; -CH2CH2CH(CH2F)-; -CH2CH2CH(C(O)NR23R24)-; -CH2CH2CH(C(O)OR25)- , and -CH2CH2CH(C(O)OH)-, wherein R20, R21, R22 and R23, each independently, represent hydrogen or (1-6C)alkyl, and R24 and R25, each independently, represent (1-6C)alkyl.

[00167] In formula II, n represents an integer from 2 to 4. An example of a value for n is 2. An example of a value for n is 3. An example of a value for n is 4.

[00168] In formula II, in one embodiment, R4 represents -CH2CH2CH2NH(C=NH)NH2. In another embodiment, R4 represents -CH2CH2CH2CH2NH2.

[00169] An amino acid can be a naturally occurring amino acid. It will be appreciated that naturally occurring amino acids usually have the L configuration.

[00170] In formula II, with reference to R5, examples of particular values are:

[00171] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group;

[00172] for an amino acid: alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine or valine, and

[00173] for a dipeptide: a combination of any two amino acids selected independently from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine.

[00174] In formula II, examples of particular values for R5 are:

[00175] a hydrogen atom;

[00176] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group, and

[00177] for a residue of an amino acid, a dipeptide, or an N-acyl derivative of an amino acid or dipeptide: glycinyl or N-acetylglycinyl.

[00178] In formula II, in one embodiment, R5 represents N-acetyl, glycinyl or N-acetylglycinyl, such as N-acetyl.

[00179] In formula II, an example of the group represented by -C(O)-CH(R4)-NH(R5) is N-acetylarginyl or N-acetyllysinyl.

[00180] In formula II, in certain cases, R5 represents a substituted acyl group. In certain cases, R5 may be malonyl or succinyl.

[00181] In formula II, in certain cases, the group represented by -C(O)-CH(R4)-NH(R5) is N-malonylarginyl, N-malonyl-lysinyl, N-succinylarginyl and N-succinyl-lysinyl. Formula III

[00182] The embodiments provide a compound of general formula (III): X-C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5)(III)

[00183] or a pharmaceutically acceptable salt thereof, wherein:

[00184] X represents a residue of a phenolic opioid, in which the hydrogen atom of the phenolic hydroxyl group is replaced by a covalent bond, forming -C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5);

[00185] R1 represents a (1-4C)alkyl group;

[00186] R2 and R3, each independently, represent a hydrogen atom or a (1-4C)alkyl group;

[00187] n represents 2 or 3;

[00188] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[00189] R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or dipeptide.

[00190] In formula III, examples of values for the phenolic opioid as provided in X are oxymorphone, hydromorphone, and morphine.

[00191] In formula III, examples of values for R1 are methyl and ethyl groups.

[00192] In formula III, examples of values for each of R2 and R3 are hydrogen atoms. In formula III, an example of a value for n is 2.

[00193] In formula III, in one embodiment, R4 represents -CH2CH2CH2NH(C=NH)NH2. In another embodiment, R4 represents -CH2CH2CH2CH2NH2.

[00194] An amino acid can be a naturally occurring amino acid. It will be appreciated that naturally occurring amino acids usually have the L configuration.

[00195] In formula III, with reference to R5, examples of particular values are:

[00196] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group;

[00197] for an amino acid: alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine or valine, and

[00198] for a dipeptide: a combination of any two amino acids selected independently from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine.

[00199] In formula III, examples of particular values for R5 are:

[00200] a hydrogen atom;

[00201] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group, and

[00202] for a residue of an amino acid, a dipeptide or an N-acyl derivative of an amino acid or dipeptide: glycinyl or N-acetylglycinyl.

[00203] In formula III, in one embodiment, R5 represents N-acetyl, glycinyl or N-acetylglycinyl, such as N-acetyl.

[00204] In formula III, an example of the group represented by -C(O)-CH(R4)-NH(R5) is N-acetylarginyl or N-acetyllysinyl.

[00205] In formula III, in certain cases, R5 represents a substituted acyl group. In certain cases, R5 may be malonyl or succinyl.

[00206] In formula III, in certain cases, the group represented by -C(O)-CH(R4)-NH(R5) is N-malonylarginyl, N-malonyl-lysinyl, N-succinylarginyl and N-succinyl-lysinyl. Formula IV

[00207] The modalities provide a compound of general formula (IV):

[00208] or a pharmaceutically acceptable salt thereof, wherein:

[00209] Ra is hydrogen or hydroxyl;

[00210] Rb is oxo (=O) or hydroxyl;

[00211] the dashed line is a double link or a single link;

[00212] R1 represents a (1-4C)alkyl group;

[00213] R2 and R3, each independently, represent a hydrogen atom or a (1-4C)alkyl group;

[00214] n represents 2 or 3;

[00215] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[00216] R5 represents a hydrogen atom, an N-acyl group or a residue of an amino acid, a dipeptide, or an N-acyl derivative of an amino acid or dipeptide.

[00217] In formula IV, a certain example of Ra is hydrogen. In formula IV, a certain example of Ra is hydroxyl.

[00218] In formula IV, a certain example of Rb is oxo (=O). In formula IV, a certain example of Rb is hydroxyl.

[00219] In formula IV, a certain example of the dashed line is a double bond. In formula IV, a certain example of the dashed line is a single bond.

[00220] In formula IV, examples of values for R1 are methyl and ethyl groups.

[00221] In formula IV, examples of values for each of R2 and R3 are hydrogen atoms. In formula IV, an example of a value for n is 9 2. In formula IV, in one embodiment, R4 represents -CH2CH2CH2NH(C=NH)NH2. An amino acid can be a naturally occurring amino acid. It will be appreciated that naturally occurring amino acids usually have the L configuration.

[00222] In formula IV, with reference to R5, examples of particular values are:

[00223] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group;

[00224] for an amino acid: alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine or valine, and

[00225] for a dipeptide: a combination of any two amino acids selected independently from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine.

[00226] In formula IV, examples of particular values for R5 are:

[00227] a hydrogen atom;

[00228] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group, and

[00229] for a residue of an amino acid, a dipeptide, or an N-acyl derivative of an amino acid or dipeptide: glycinyl or N-acetylglycinyl.

[00230] In formula IV, in one embodiment, R5 represents N-acetyl, glycinyl or N-acetylglycinyl, such as N-acetyl.

[00231] In formula IV, an example of the group represented by -C(O)-CH(R4)-NH(R5) is N-acetylarginyl. Formula V

[00232] The modalities provide a compound of general formula (V):

[00233] or a pharmaceutically acceptable salt thereof, wherein:

[00234] Ra is hydrogen or hydroxyl;

[00235] Rb is oxo (=O) or hydroxyl;

[00236] the dashed line is a double link or a single link;

[00237] R1 is selected from alkyl, substituted alkyl, arylalkyl, substituted arylalkyl, aryl and substituted aryl;

[00238] each R2 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl and aminoacyl;

[00239] each R3 is independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl and aminoacyl;

[00240] or R2 and R3, together with the carbon to which they are attached, form a cycloalkyl or substituted cycloalkyl group, or two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, form a cycloalkyl or substituted cycloalkyl group;

[00241] n represents an integer from 2 to 4;

[00242] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[00243] R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or dipeptide.

[00244] In formula V, a certain example of Ra is hydrogen. In formula IV, a certain example of Ra is hydroxyl.

[00245] In formula V, a certain example of Rb is oxo (=O). In formula V, a certain example of Rb is hydroxyl.

[00246] In formula V, a certain example of the dashed line is a double bond. In formula V, a certain example of the dashed line is a single bond.

[00247] In formula V, R1 can be selected from alkyl, substituted alkyl, aryl alkyl, substituted aryl alkyl, aryl, and substituted aryl. In certain cases, R1 is (1-6C)alkyl. In other cases, R1 is (1-4C)alkyl. In other cases, R1 is methyl or ethyl. In other cases, R1 is methyl. In some cases, R1 is ethyl.

[00248] In certain cases, R1 is a substituted alkyl group. In certain cases, R1 is an alkyl group substituted with a carboxyl group or a carboxyl ester. In certain cases, R1 is -(CH2)5-COOH, -(CH2)5-COOCH3, or -(CH2)5-COOCH2CH3.

[00249] In certain cases, in formula V, R1 is an arylalkyl or substituted arylalkyl. In certain cases, in formula V, R1 is an arylalkyl. In certain cases, R1 is a substituted arylalkyl. In certain cases, R1 is an arylalkyl group substituted with a carboxyl or carboxyl ester. In certain cases, R1 is -(CH2)q(C1H4)-COOH, -(CH2)q(C6H4)-COOCH3 or -(CH2)q(C6H4)-COOCH2CH3, where q is an integer from one to 10. In certain cases, R1 is -CH2(C6H4)-COOH, -CH2(C6H4)-COOCH3 or -CH2(C6H4)-COOCH2CH3.

[00250] In certain cases, in formula V, R1 is an aryl group. In certain cases, R1 is a substituted aryl group. In certain cases, R1 is an aryl group substituted with a carboxyl group or a carboxyl ester. In certain cases, R1 is -(C6H4)-COOH, -(C6H4)-COOCH3, or -(C6H4)-COOCH2CH3.

[00251] In formula V, each R2 can be independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl, and aminoacyl. In certain cases, R2 is hydrogen or alkyl. In certain cases, R2 is hydrogen. In certain cases, R1 is alkyl. In certain cases, R2 is acyl. In certain cases, R2 is aminoacyl.

[00252] In formula V, each R3 can be independently selected from hydrogen, alkyl, substituted alkyl, aryl, substituted aryl, acyl, and aminoacyl. In certain cases, R3 is hydrogen or alkyl. In certain cases, R3 is hydrogen. In certain cases, R3 is alkyl. In certain cases, R3 is acyl. In certain cases, R3 is aminoacyl.

[00253] In certain cases, R2 and R3 are hydrogen. In certain cases, R2 and R3 on the same carbon are both alkyl. In certain cases, R2 and R3 on the same carbon are methyl. In certain cases, R2 and R3 on the same carbon are ethyl.

[00254] In formula V, R2 and R3, together with the carbon to which they are attached, can form a cycloalkyl or substituted cycloalkyl group, or two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, can form a cycloalkyl or substituted cycloalkyl group. In certain cases, R2 and R3, together with the carbon to which they are attached, can form a cycloalkyl group. Thus, in certain cases, R2 and R3 on the same carbon form a spirocycle. In certain cases, R2 and R3, together with the carbon to which they are attached, can form a substituted cycloalkyl group. In certain cases, two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, can form a cycloalkyl group. In certain cases, two R2 or R3 groups on adjacent carbon atoms, together with the carbon atoms to which they are attached, can form a substituted cycloalkyl group.

[00255] In certain cases, one of the R2 and R3 is an aminoacyl group.

[00256] In certain cases, one of the R2 and R3 groups is an aminoacyl group comprising phenylenediamine. In certain cases, one of the R2 and R3 groups is

[00257] wherein each R10 is independently selected from hydrogen, alkyl, substituted alkyl and acyl and R11 is alkyl or substituted alkyl. In certain cases, at least one of the R10 is acyl. In certain cases, at least one of the R10 is alkyl or substituted alkyl. In certain cases, at least one of the R10 is hydrogen. In certain cases, both R10 are hydrogen.

[00258] In certain cases, one of R2 and R3 is

[00259] where R10 is hydrogen, alkyl, substituted alkyl or acyl. In certain cases, R10 is acyl. In certain cases, R10 is alkyl or substituted alkyl. In certain cases, R10 is hydrogen.

[00260] In certain cases, R2 or R3 can modulate an intramolecular cyclization rate. R2 or R3 can accelerate an intramolecular cyclization rate, compared to the corresponding molecule in which R2 and R3 are both hydrogen. In certain cases, R2 or R3 comprise an electron-withdrawing group or an electron-donating group. In certain cases, R2 or R3 comprise an electron-withdrawing group. In certain cases, R2 or R3 comprise an electron-donating group.

[00261] Atoms and groups capable of functioning as electron-withdrawing substituents are well known in the field of organic chemistry. They include electronegative atoms and groups containing electronegative atoms. The function of these groups is to decrease the basicity or protonation state of a nucleophilic nitrogen in the beta position via inductive electron density withdrawal. These groups can also be positioned in other positions along the alkylene chain. Examples include halogen atoms (e.g., a fluorine atom), acyl groups (e.g., an alkanoyl group, an aroyl group, a carboxyl group, an alkoxycarbonyl group, an aryloxycarbonyl group, or an aminocarbonyl group (such as a carbamoyl, alkylaminocarbonyl, dialkylaminocarbonyl, or arylaminocarbonyl group)), an oxo (=0) substituent, a nitrile group, a nitro group, ether groups (e.g., an alkoxy group), and phenyl groups containing a substituent in the ortho position, in the para position, or both the ortho and para positions, wherein each substituent is independently selected from a halogen atom, a fluoroalkyl group (such as trifluoromethyl), a nitro group, a cyano group, and a carboxyl group. Each of the electron-withdrawing substituents may be selected independently of these.

[00262] In certain cases, -[C(R2)(R3)]n- is selected from -CH(CH2F)CH(CH2F)-; -CH(CHF2)CH(CHF2)-; -CH(CF3)CH(CF3)-; - CH2CH(CF3)-; -CH2CH(CHF2)-; -CH2CH(CH2F)-; -CH2CH(F)CH2-; - CH2C(F2)CH2-; -CH2CH(C(0)NR20R21)-; -CH2CH(C(0)0R22)-;-CH2CH(C(0)0H)-; -CH(CH2F)CH2CH(CH2F)-;-CH(CHF2)CH2CH(CHF2)-; -CH(CF3)CH2CH(CF3)-; -CH2CH2CH(CF3)-; -CH2CH2CH(CHF2)-; -CH2CH2CH(CH2F)-; -CH2CH2CH(C(0)NR23R24)-; -CH2CH2CH(C(0)0R25)-, and -CH2CH2CH(C(0)0H)-, wherein R20, R21, R22, and R23 each independently represent hydrogen or (1-6C)alkyl, and R24 and R25 each independently represent (1-6C)alkyl.

[00263] In formula V, n represents an integer from 2 to 4. An example of a value for n is 2. An example of a value for n is 3. An example of a value for n is 4. In formula V, in one embodiment, R4 represents -CH2CH2CH2NH(C=NH)NH2. In another embodiment, R4 represents -CH2CH2CH2CH2NH2. An amino acid can be a naturally occurring amino acid. It will be appreciated that naturally occurring amino acids usually have the L configuration.

[00264] In formula V, with reference to R5, examples of particular values are:

[00265] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group;

[00266] for an amino acid: alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine or valine, and

[00267] for a dipeptide: a combination of any two amino acids selected independently from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine.

[00268] In formula V, examples of particular values for R5 are:

[00269] a hydrogen atom;

[00270] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group, and

[00271] for a residue of an amino acid, a dipeptide, or an N-acyl derivative of an amino acid or dipeptide: glycinyl or N-acetylglycinyl.

[00272] In formula V, in one embodiment, R5 represents N-acetyl, glycinyl or N-acetylglycinyl, such as N-acetyl.

[00273] In formula V, an example of the group represented by -C(O)-CH(R4)-NH(R5) is N-acetylarginyl or N-acetyllysinyl.

[00274] In formula V, in certain cases, R5 represents a substituted acyl group. In certain cases, R5 may be malonyl or succinyl.

[00275] In formula V, in certain cases, the group represented by -C(O)-CH(R4)-NH(R5) is N-malonylarginyl, N-malonyl-lysinyl, N-succinylarginyl and N-succinyl-lysinyl. Formula VI

[00276] The modalities provide a compound of general formula (VI):

[00277] or a pharmaceutically acceptable salt thereof, wherein:

[00278] Ra is hydrogen or hydroxyl;

[00279] Rb is oxo (=O) or hydroxyl;

[00280] the dashed line is a double link or a single link;

[00281] R1 represents a (1-4C)alkyl group;

[00282] R2 and R3, each independently, represent a hydrogen atom or a (1-4C)alkyl group;

[00283] n represents 2 or 3;

[00284] R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, where the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and

[00285] R5 represents a hydrogen atom, an N-acyl group (including N-substituted acyl), an amino acid residue, a dipeptide, an N-acyl derivative (including N-substituted acyl derivative) of an amino acid or dipeptide.

[00286] In formula VI, a certain example of Ra is hydrogen. In formula VI, a certain example of Ra is hydroxyl.

[00287] In formula VI, a certain example of Rb is oxo (=O). In formula VI, a certain example of Rb is hydroxyl.

[00288] In formula VI, a certain example of the dashed line is a double bond. In formula VI, a certain example of the dashed line is a single bond.

[00289] In formula VI, examples of values for R1 are methyl and ethyl groups.

[00290] In formula VI, examples of values for each of R2 and R3 are hydrogen atoms. In formula VI, an example of a value for n is 2. In formula VI, in one embodiment, R4 represents -CH2CH2CH2NH(C=NH)NH2. In another embodiment, R4 represents -CH2CH2CH2CH2NH2. An amino acid can be a naturally occurring amino acid. It will be appreciated that naturally occurring amino acids usually have the L configuration.

[00291] In formula VI, with reference to R5, examples of particular values are:

[00292] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group;

[00293] for an amino acid: alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine or valine, and

[00294] for a dipeptide: a combination of any two amino acids selected independently from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine.

[00295] In formula VI, examples of particular values for R5 are:

[00296] a hydrogen atom;

[00297] for an N-acyl group: an N-(1-4C)alkanoyl group, such as acetyl, an N-aroyl group, such as N-benzoyl, or an N-piperonyl group, and

[00298] for a residue of an amino acid, a dipeptide or an N-acyl derivative of an amino acid or dipeptide: glycinyl or N-acetylglycinyl.

[00299] In formula VI, in one embodiment, R5 represents N-acetyl, glycinyl or N-acetylglycinyl, such as N-acetyl.

[00300] In formula VI, an example of the group represented by -C(O)-CH(R4)-NH(R5) is N-acetylarginyl or N-acetyllysinyl.

[00301] In formula VI, in certain cases, R5 represents a substituted acyl group. In certain cases, R5 may be malonyl or succinyl.

[00302] In formula VI, in certain cases, the group represented by -C(O)-CH(R4)-NH(R5) is N-malonylarginyl, N-malonyl-lysinyl, N-succinylarginyl and N-succinyl-lysinyl.

General Synthetic Procedures

[00303] Compounds of formula I are particular prodrugs described in WO 2007/140272, and the synthesis of compounds of formula I is described herein.

[00304] The schemes and synthesis procedure in WO 2007/140272 can also be used to synthesize compounds of formulas I-VI. The compounds described here can be obtained by the routes generally illustrated in Scheme 1.

[00305] The carrier moieties described herein can be prepared and linked to phenol-containing drugs by procedures known to those skilled in the art (see, for example, Green et al., "Protective Groups in Organic Chemistry," (Wiley, 2nd edition 1991); Harrison et al., "Compendium of Synthetic Organic Methods," Volumes 1-8 (John Wiley and Sons, 1971-1996); "Beilstein Handbook of Organic Chemistry," Beilstein Institute of Organic Chemistry, Frankfurt, Germany; Feiser et al., "Reagents for Organic Synthesis," Volumes 1-17, (Wiley Interscience); Trost et al., "Comprehensive Organic Synthesis," (Pergamon Press, 1991); "Theilheimer's Synthetic Methods of Organic Chemistry," Volumes 1-45, (Karger, 1991); March, "Advanced Organic Chemistry," (Wiley Interscience), 1991; Larock "Comprehensive Organic Transformations," (VCH Publishers, 1989); Paquette, "Encyclopedia of Reagents for Organic Synthesis," (John Wiley & Sons, 1995); Bodanzsky, "Principles of Peptide Synthesis," (Springer Verlag, 1984); Bodanzsky, "Practice of Peptide Synthesis," (Springer Verlag, 1984). Additionally, starting materials can be obtained from commercial sources or via well-established synthesis procedures, as described above.

[00306] Now with reference to Scheme 1 and formula I, above, wherein, for illustrative purposes, T is NH, Y is NR1, W is NH, p is one, R1, R4, and R5 are as previously defined, X is a phenolic opioid, P is a protecting group, and M is a leaving group, compound 1-1 can be acylated with an appropriate carboxylic acid or carboxylic acid equivalent to provide compound 1-2, which can then be deprotected to give compound 1-3. Subsequently, compound 1-3 reacts with an activated carbonic acid equivalent 1-4 to provide compound 1-5.

[00307] For compounds of formula ll-VI, -(C(R2)(Rs))n- corresponds to the portion -(CH2-CH2)- between Y and T. Thus, for the synthesis of compounds of formulas ll-VI, compound 1-1 will have the appropriate entities for -(C(R2)(R%))n- in order to result in the synthesis of compounds of formulas ll-Vl.

Trypsin inhibitors

[00308] As used herein, the term "trypsin inhibitor" refers to any agent capable of inhibiting the action of trypsin on a substrate. The ability of an agent to inhibit trypsin can be measured using well-known assays in the field. For example, in a typical assay, one unit corresponds to the amount of inhibitor that reduces trypsin activity by one unit of benzoyl-L-arginine ethyl ester (BAEE-U). One BAEE-U is the amount of enzyme that increases absorbance at 253 nm by 0.001 per minute at pH 7.6 and 25°C. See, for example, K. Ozawa, M. Laskowski, 1966, J. Biol. Chem. 241, 3955, and Y. Birk, 1976, Meth. Enzymol. 45, 700.

[00309] Many trypsin inhibitors are known in the field, both those specific to trypsin and those that inhibit trypsin and other proteases, such as chymotrypsin. Trypsin inhibitors can be derived from a variety of animal or vegetable sources: for example, soy, corn, lima beans and other beans, pumpkin, sunflower, bovine and other animal pancreas and lungs, chicken and turkey egg whites, soy-based infant formula, and mammalian blood. Trypsin inhibitors can also be of microbial origin: for example, antipain; see, for example, H. Umezawa, 1976, Meth. Enzymol. 45,678. A trypsin inhibitor may also be an arginine or lysine mimetic or another synthetic compound: for example, arylguanidine, benzamidine, 3,4-dichloroisocoumarin, diisopropyl fluorophosphate, gabexate mesylate, or phenylmethanesulfonyl fluoride. As used herein, an arginine or lysine mimetic is a compound that is capable of binding to the P1 trypsin pocket and/or interfering with the function of the trypsin active site.

[00310] In one embodiment, the trypsin inhibitor is derived from soy. Trypsin inhibitors derived from soy (Glycine max) are readily available and considered safe for human consumption. They include, but are not limited to, SBTI, which inhibits trypsin, and Bowman-Birk inhibitor, which inhibits both trypsin and chymotrypsin. These trypsin inhibitors are available, for example, from Sigma-Aldrich, St. Louis, MO, USA. It will be appreciated that the pharmaceutical composition according to the embodiments may additionally comprise one or more different trypsin inhibitors.

Small Molecule Trypsin Inhibitors

[00311] As stated above, a trypsin inhibitor may be an arginine or lysine mimetic or another synthetic compound. In certain embodiments, the trypsin inhibitor is an arginine mimetic or a lysine mimetic, wherein the arginine mimetic or lysine mimetic is a synthetic compound.

[00312] Certain trypsin inhibitors include compounds with the formula:

[00313] wherein: Q1 is selected from -O-Q4 or -Q4-COOE, wherein Q4 is C1-C4 alkyl; Q2 is N or CH, and Q3 is aryl or substituted aryl.

[00314] Certain trypsin inhibitors include compounds with the formula:

[00315] where: Q5 is -C(O)-COOH or -NH-Q6-Q7-SO2-C6H5, where Q6 is -(CH2)P-COOH; Q7 is -(CH2)r-C6H5, and

[00316] p is an integer from one to three, and

[00317] r is an integer from one to three.

[00318] Certain trypsin inhibitors include the following:

[00319] In certain embodiments the trypsin inhibitor is SBTI, BBSI, Compound 101, Compound 106, Compound 108, Compound 109 or Compound 110.

Pharmaceutical Compositions

[00320] As discussed above, the present description provides pharmaceutical compositions comprising a trypsin inhibitor and a phenolic opioid prodrug containing an unstable trypsin moiety which, when cleaved, facilitates the release of the phenolic opioid. Examples of compositions containing a phenolic opioid prodrug and a trypsin inhibitor are described below.

Combinations of Formulas I-VI and Trypsin inhibitor

[00321] The embodiments provide a pharmaceutical composition comprising a trypsin inhibitor and a compound of general formula (I), or a pharmaceutically acceptable salt thereof.

[00322] The embodiments provide a pharmaceutical composition comprising a trypsin inhibitor and a compound of general formulas (II)-(VI), or a pharmaceutically acceptable salt thereof. Certain embodiments provide a combination of a compound of Formula I and a trypsin inhibitor, in which the phenolic opioid of Formula I and the trypsin inhibitor are presented in the following table.

[00323] Certain embodiments provide a combination of a compound of formula I and a trypsin inhibitor, in which the phenolic opioid of formula I and the trypsin inhibitor are presented in the following table.

[00324] Certain embodiments provide a combination of a compound of formula I and a trypsin inhibitor, in which the phenolic opioid of formula I and the trypsin inhibitor are presented in the following table.

[00325] Certain embodiments provide a combination of a compound of formula I and a trypsin inhibitor, in which the phenolic opioid of formula I and the trypsin inhibitor are presented in the following table.

[00326] Certain embodiments provide a combination of a compound of formula II and a trypsin inhibitor, in which the phenolic opioid of formula II and the trypsin inhibitor are presented in the following table.

[00327] Certain embodiments provide a combination of a compound of formula II and a trypsin inhibitor, in which the phenolic opioid of formula II and the trypsin inhibitor are presented in the following table.

[00328] Certain embodiments provide a combination of a compound of formula II and a trypsin inhibitor, in which the phenolic opioid of formula II and the trypsin inhibitor are presented in the following table.

[00329] Certain embodiments provide a combination of a compound of formula II and a trypsin inhibitor, in which the phenolic opioid of formula II and the trypsin inhibitor are presented in the following table.

[00330] Certain embodiments provide a combination of a compound of formula III and a trypsin inhibitor, in which the phenolic opioid of formula III and the trypsin inhibitor are presented in the following table.

[00331] Certain embodiments provide a combination of a compound of formula III and a trypsin inhibitor, in which the phenolic opioid of formula III and the trypsin inhibitor are presented in the following table.

[00332] Certain embodiments provide a combination of a compound of formula III and a trypsin inhibitor, in which the phenolic opioid of formula III and the trypsin inhibitor are presented in the following table.

[00333] Certain embodiments provide a combination of a compound of formula III and a trypsin inhibitor, in which the phenolic opioid of formula III and the trypsin inhibitor are presented in the following table.

[00334] Certain embodiments provide a combination of Compound 1 and a trypsin inhibitor, in which the trypsin inhibitor is presented in the following table.

[00335] Certain embodiments provide a combination of Compound 2 and a trypsin inhibitor, in which the trypsin inhibitor is presented in the following table.

[00336] Certain embodiments provide a combination of Compound 3 and a trypsin inhibitor, in which the trypsin inhibitor is presented in the following table.

[00337] Certain embodiments provide a combination of Compound 4 and a trypsin inhibitor, in which the trypsin inhibitor is presented in the following table.

[00338] It should be appreciated that the invention also includes inhibitors of other enzymes involved in protein assimilation that can be used in combination with a prodrug having a formula of I-VI comprising an amino acid of alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine or valine.

[00339] The pharmaceutical composition according to the embodiments may also comprise a pharmaceutically acceptable carrier. The composition is conveniently formulated in a form suitable for oral administration (including buccal and sublingual), for example, in the form of a tablet, capsule, thin film, powder, suspension, solution, syrup, dispersion or emulsion. The composition may contain conventional components in pharmaceutical preparations, for example, one or more carriers, binders, lubricants, excipients (for example, to impart controlled-release characteristics), pH modifiers, sweeteners, bulking agents, coloring agents or other active agents.

[00340] The pharmaceutical composition according to the modalities is useful, for example, in the treatment of a patient who suffers from or is at risk of suffering from pain.

[00341] The patient can be a human or a non-human animal, for example, a companion animal such as a cat, dog or horse.

Management Methods

[00342] The amount of formula (l)-(VI) compound to be administered to a patient in order to be effective (i.e., to provide sufficient blood levels of phenolic opioid to be effective in the treatment or prophylaxis of pain) will depend on the bioavailability of the particular compound, the susceptibility of the particular compound to enzymatic activation in the intestines, the amount and potency of the trypsin inhibitor present in the composition, as well as other factors such as the species, age, weight, sex and condition of the patient, the method of administration and the assessment of the prescribing physician. In general, the dose may range from 0.01 to 20 milligrams per kilogram (mg/kg) of body weight. For example, a compound comprising a hydromorphone residue may be administered at a dose equivalent to the administration of free hydromorphone in the range of 0.02 to 0.5 mg/kg body weight, or 0.01 to 10 mg/kg body weight, or 0.01 to 2 mg/kg body weight. In one embodiment, the compound may be administered at a dose such that the blood level of phenolic opioid achieved is in the range of 0.5 ng/mL to 10 ng/mL.

[00343] The amount of a trypsin inhibitor to be administered to the patient in order to be effective (i.e., to attenuate the release of phenolic opioid when administration of a compound of formulas (I)-(VI) alone would lead to overexposure of the phenolic opioid) will depend on the effective dose of the particular compound and the potency of the particular inhibitor, as well as other factors, the species, age, weight, sex and condition of the patient, the mode of administration and the assessment of the prescribing physician. In general, the inhibitor dose may range from 0.05 to 50 mg per mg of compound of formulas (I)-(VI). In a certain embodiment, the inhibitor dose may range from 0.001 to 50 mg per mg of compound of formulas (I)-(VI).

Therapeutic Applications

[00344] In another aspect, the modalities provide a pharmaceutical composition as described above for use in the treatment of pain.

[00345] The present description provides the use of a phenolic opioid prodrug and a trypsin inhibitor in the treatment of pain.

[00346] The present description provides the use of a phenolic opioid prodrug and a trypsin inhibitor in the manufacture of a medicament for the treatment of pain.

[00347] In another aspect, the modalities provide a method of treating pain in a patient requiring treatment, which comprises administering an effective amount of a pharmaceutical composition as described above.

Counteracting abusive behaviors through trypsin-mediated release of phenolic opioids from prodrugs

[00348] The description provides a composition comprising a compound of formulas I-VI and a trypsin inhibitor that reduces the potential for drug dependence. A trypsin inhibitor can counteract a user's ability to apply trypsin to effect the release of a phenolic opioid from the phenolic opioid prodrug in vitro. For example, if a dependent individual attempts to incubate trypsin with a composition of the embodiments that includes a phenolic opioid prodrug and a trypsin inhibitor, the trypsin inhibitor can reduce the action of the added trypsin, thereby counteracting attempts to release the phenolic opioid for purposes of abuse.

Examples

[00349] The following examples are presented to provide those skilled in the art with a complete disclosure and description of how to manufacture and use the embodiments, and are not intended to limit the scope of what the inventors consider their invention, nor are they intended to represent that the experiments below are all or the only experiments performed. Efforts have been made to ensure accuracy with regard to the numbers used (e.g., quantities, temperature, etc.), but some experimental errors and deviations should be contemplated. Unless otherwise indicated, parts are parts by weight, molecular weight is weight-average molecular weight, temperature is in degrees Celsius, and pressure is equal to or close to atmospheric. Common abbreviations may be used.

Example 1: Oral administration of Compound 1 and the trypsin inhibitor SBTI to rats.

[00350] Hydromorphone 3-(N-Methyl-N-(2-N'-acetylarginylamino))ethylcarbamate (which may be produced as described in International Publication PCT No. WO 2007/140272, published on December 6, 2007, Example 3, hereinafter referred to as Compound 1) and SBTI (Glycine max (soybean) trypsin inhibitor (Catalog 93620, ~10,000 units per mg, Sigma-Aldrich)) were each dissolved in brine. Saline solutions of Compound 1 and SBTI were dosed as indicated in Table 1, via oral gavage, in jugular-cannulated male Sprague Dawley rats that had fasted for 1618 hours (hr) prior to oral dosing; 4 rats per group were dosed. When it was dosed SBTI was administered 5 minutes (min) before Compound 1. At the specified times, blood samples were collected, rapidly cooled in methanol, centrifuged at 14,000 rpm @ 4°C, and stored at -80°C until analysis by high-performance liquid chromatography/mass spectrometry (HPLC/MS). Table 1 shows the results for rats that received a constant amount of Compound 1 and varying amounts of SBTI. The results are reported as maximum blood concentration of hydromorphone (mean + standard deviation) for each group of 4 rats. Table 1. Maximum concentration (Cmax) of hydromorphone in rat blood

[00351] The lower limit of quantification was 1 nanogram per milliliter (ng/mL) for the first group and 5 ng/mL for the other groups.

[00352] The results in Table 1 indicate that SBTI attenuates the ability of Compound 1 to release hydromorphone in a dose-dependent manner, which can reach approximately 100% attenuation for higher concentrations of SBTI.

[00353] The data obtained from the rats represented in Table 1 are also provided in Figure 1, which compares mean blood concentrations (+ standard deviations) over time of hydromorphone after PO administration to rats of 20 mg/kg of Compound 1 (a) alone (solid line with closed black circles), (b) with 10 mg/kg of SBTI (dashed line with open white squares), (c) with 100 mg/kg of SBTI (dotted line with open white triangles), (d) with 500 mg/kg of SBTI (solid line with X symbols) or (e) with 1000 mg/kg of SBTI (solid line with closed black squares). The results in Figure 1 indicate that attenuation by means of SBTI of the ability of Compound 1 to release hydromorphone suppresses Cmax and delays Tmax of this hydromorphone release into the blood of rats that received Compound 1 and 10, 100, 500 or 1000 mg/kg of SBTI.

Example 2: Oral administration of Compound 1 and the trypsin inhibitor SBTI, in the presence of ovalbumin, to rats.

[00354] In an effort to understand the role of SBTI, ovalbumin was used as a non-trypsin inhibitor protein control. Chicken egg white albumin (ovalbumin) (Catalog No. A7641, Quality VII, lyophilized powder, Sigma-Aldrich) was dissolved in brine. Saline solutions of Compound 1 and SBTI (as described in Example 1) and ovalbumin were combined and dosed as indicated in Table 2, via oral gavage, in jugular vein cannulated male Sprague Dawley rats (4 per group) that had fasted for 16-18 hours before oral dosing. At specified times, blood samples were collected, plasma was centrifuged at 5400 rpm at 4°C for 5 min, and 100 microliters (μL) of plasma was transferred from each sample to a fresh tube containing 1 μL of formic acid. The tubes were vortexed for 5-10 seconds, immediately placed on dry ice, and then stored until analysis by HPLC/MS. Table 2 shows the results for rats that received Compound 1 with or without varying amounts of ovalbumin (OVA) and/or SBTI, as indicated. The results are reported as maximum plasma concentration of hydromorphone (mean + standard deviation) for each group of 4 rats. Table 2. Maximum concentration (Cmax) of hydromorphone in rat plasma

[00355] The lower limit of quantification was 12.5 picograms/mL (pg/mL) for the first group, 25 pg/mL for the last group, and 100 pg/mL for the other groups.

[00356] The results in Table 2 indicate that ovalbumin does not significantly affect the ability of Compound 1 to release hydromorphone or the ability of SBTI to attenuate this release. Data obtained from the rats represented in rows 1, 4, and 6 of Table 2 are also provided in Figure 2, which compares mean plasma concentrations (+ standard deviations) over time of hydromorphone after PO administration to rats of 20 mg/kg of Compound 1 (a) alone (solid line with circles), (b) with 500 mg/kg of OVA (dashed line with triangles), or (c) with 500 mg/kg of OVA and 500 mg/kg of SBTI (dotted line with squares). The results in Figure 2 indicate that the attenuation by SBTI of the ability of Compound 1 to release hydromorphone suppresses Cmax and delays Tmax of this hydromorphone in plasma, even in the presence of ovalbumin. Rats that received 20 mg/kg of Compound 1 with 500 mg/kg of OVA and 500 mg/kg of SBTI exhibited a plasma Tmax of 8.0 hours, whereas rats that received 20 mg/kg of Compound 1 alone exhibited a plasma Tmax of 2.3 hours. The results in Table 2 and Figure 2 also indicate that SBTI acts specifically by inhibiting trypsin, rather than in a non-specific manner.

Example 3: Oral administration of Compound 1 and BBSI inhibitor to rats.

[00357] Compound 1 and BBSI (Bowman-Birk trypsin-chymotrypsin inhibitor from Glycine max (soybean), Catalog No. T9777, Sigma-Aldrich) were each dissolved in brine.

[00358] The saline solutions of Compound 1 and BBSI were dosed as indicated in Table 3. The dosing, sampling and analysis procedures were as described in Example 1.

[00359] Table 3 shows the results for rats that received Compound 1 with or without BBSI. The results are reported as maximum blood concentration of hydromorphone (mean + standard deviation) for each group of 4 rats (n=4) as well as for 3 of the 4 rats that received Compound 1 and BBSI (n=3). Table 3. Maximum concentration (Cmax) of hydromorphone in rat blood

[00360] The lower limit of quantification was 1 ng/mL for both groups. The Cmax of the rat not included in the n=3 analysis was 43 ng/mL; the range for other rats was 6.8-17 ng/mL.

[00361] The results in Table 3 indicate that BBSI is able to attenuate the ability of Compound 1 to release hydromorphone.

[00362] The data obtained from the individual rats represented in Table 3, rows 1 and 3, are provided in Figure 3, which compares individual blood concentrations over time of hydromorphone after PO administration to rats of 20 mg/kg of Compound 1 (a) alone (solid lines) or (b) with 100 mg/kg of BBSI (dashed lines). The results in Figure 3 indicate that the attenuation by BBSI of the ability of Compound 1 to release hydromorphone suppresses Cmax and delays Tmax of this hydromorphone in blood, at least for 3 of the 4 rats that received Compound 1 and BBSI.

Example 4: Oral administration of Compound 2 and the trypsin inhibitor SBTI to rats.

[00363] Saline solutions of Compound 2 (which can be prepared as described in Example 11) and SBTI (which can be prepared as described in Example 1) were dosed as indicated in Table 4, via oral gavage, in jugular vein cannulated male Sprague Dawley rats (4 per group) that had fasted for 16-18 hours before oral dosing. When SBTI was dosed, it was administered 5 minutes before Compound 4. At specified times, blood samples were collected, processed, and analyzed as described in Example 2.

[00364] Table 4 and Figure 4 provide results for rats that received 20 mg/kg of Compound 2 with or without 500 mg/kg of SBTI, as indicated. The results in Table 4 are reported, for each group of 4 rats, as (a) maximum plasma concentration (Cmax) of hydromorphone (HM) (mean + standard deviation) and (b) time after administration of Compound 2, with or without SBTI, to reach maximum hydromorphone concentration (Tmax). Table 4. Cmax and Tmax of hydromorphone in rat plasma

[00365] The lower limit of quantification was 0.0125 ng/mL for both groups.

[00366] Figure 4 compares mean plasma concentrations (+ standard deviations) over the release time of hydromorphone after PO administration to rats of 20 mg/kg of Compound 2 alone (solid line) or with 500 mg/kg of SBTI (dashed line).

[00367] The results in Table 4 and Figure 4 indicate that SBTI attenuates the ability of Compound 2 to release hydromorphone, related to the suppression of Cmax and the delay of Tmax.

Example 5: Oral administration of Compound 3 and the trypsin inhibitor SBTI to rats.

[00368] Saline solutions of Compound 3 (which can be prepared as described in Example 12) and SBTI were dosed as indicated in Table 5. The dosing, sampling, and analysis procedures were as described in Example 4. Table 5 and Figure 5 provide results for rats that received 20 mg/kg of Compound 3 with or without 500 mg/kg of SBTI, as indicated. The results in Table 5 are reported as Cmax and Tmax of hydromorphone in plasma for each group of 4 rats. Table 5. Cmax and Tmax of hydromorphone in rat plasma

[00369] The lower limit of quantification was 0.100 ng/mL for both groups.

[00370] Figure 5 compares mean plasma concentrations (+ standard deviations) over the release time of hydromorphone after PO administration to rats of 20 mg/kg of Compound 3 alone (solid line) or with 500 mg/kg of SBTI (dashed line).

[00371] The results in Table 5 and Figure 5 indicate that SBTI attenuates the ability of Compound 3 to release hydromorphone, related to the suppression of Cmax and the delay of Tmax.

Example 6: Oral administration of Compound 4 and the trypsin inhibitor SBTI to rats.

[00372] Saline solutions of Compound 4 (which can be prepared as described in Example 13) and SBTI were dosed as indicated in Table 6. The dosing, sampling, and analysis procedures were as described in Example 4, except that Compound 4 without inhibitor was administered to 7 rats. Table 6 and Figure 6 provide results for rats that received 20 mg/kg of Compound 4 with or without 500 mg/kg of SBTI, as indicated. The results in Table 6 are reported as Cmax and Tmax of hydromorphone in plasma for each group of 4 rats. Table 6. Cmax and Tmax of HM in rat plasma

[00373] The lower limit of quantification was 0.500 ng/mL for both groups.

[00374] Figure 6 compares mean plasma concentrations (+ standard deviations) over the release time of hydromorphone after PO administration to rats of 20 mg/kg of Compound 4 alone (solid line) or with 500 mg/kg of SBTI (dashed line).

[00375] The results in Table 6 and Figure 6 indicate that SBTI attenuates the ability of Compound 4 to release hydromorphone, at least related to delaying Tmax.

Example 7: In vitro IC50 data

[00376] Several trypsin inhibitor candidates, namely Compounds 101-105, 107 and 108, have been produced as described in Examples 14-18, 19 and 20, respectively. Compound 106 (also called 4-aminobenzamidine), Compound 109 (also known as nafamostat mesylate) and Compound 110 (also known as pentamidine isethionate salt) are available from Sigma-Aldrich (St. Louis, MO).

[00377] The half-maximum inhibitory concentration (IC50 or IC50) values of each of Compounds 101-110, as well as SBTI and BBSI, were determined using a modified trypsin assay as described by Bergmeyer, HU et al., 1974, "Methods of Enzymatic Analysis" Volume 1, 2nd edition, 515-516, Bergmeyer, HU, editor, Academic Press, Inc. New York, NY.

[00378] Table 7 indicates the IC50 values for each of the designated trypsin inhibitors. Table 7 IC50 values of certain trypsin inhibitors

[00379] The results in Table 7 indicate that each of Compounds 101-110 exhibits trypsin inhibitory activity.

Example 8: Effect of trypsin inhibitors on trypsin-mediated, in vitro release of hydromorphone from Compound 4

[00380] Compound 4 (which can be produced as described in Example 13) was incubated with bovine pancreatic trypsin (Catalog No. T8003, Type I, 10000 BAEE units/mg protein, Sigma-Aldrich) in the absence or presence of one of the following trypsin inhibitors: SBTI, Compound 107, Compound 108, or Compound 109. When a trypsin inhibitor was part of the incubation mixture, Compound 4 was added 5 minutes after the other incubation components. Reactions were conducted at 37°C for 24 hours. Samples were collected at specified times, transferred to 0.5% formic acid in acetonitrile to stop trypsin activity, and stored at less than -70°C until analysis by LC-MS/MS. The final incubation mixtures consisted of the following components:

[00381] Figures 7A and 7B show the results of exposure to 0.51 mg/mL of Compound 4 for 22.8 ng/mL of trypsin in the absence of any trypsin inhibitor (diamond symbols) or in the presence of 10 mg/mL of SBTI (circle symbols), 1.67 mg/mL of Compound 107 (upward-pointing triangle symbols), 1.67 mg/mL of Compound 108 (square symbols), or 1.67 mg/mL of Compound 109 (downward-pointing triangle symbols). Specifically, Figure 7A illustrates the disappearance of Compound 4, and Figure 7B illustrates the appearance of hydromorphone, over time under these conditions.

[00382] The results in figures 7A and 7B indicate that a trypsin inhibitor of the modalities can counteract a user's ability to apply trypsin for the purpose of releasing hydromorphone from Compound 4.

Example 9: Oral administration of Compound 3 and the trypsin inhibitor Compound 101 to rats.

[00383] Saline solutions of Compound 3 (which can be prepared as described in Example 12) and Compound 101 (prepared as described in Example 14) were dosed as indicated in Table 8. The dosing, sampling, and analysis procedures were as described in Example 4, except that Compound 3 and Compound 101 were combined for dosing. Table 8 and Figure 8 provide results for rats that received 20 mg/kg of Compound 3 with or without 10 mg/kg of Compound 101, as indicated. The results in Table 8 are reported as Cmax and Tmax of hydromorphone in plasma for each group of 4 rats. Table 8. Cmax and Tmax of HM in rat plasma

[00384] The lower limit of quantification was 0.100 ng/mL for the first group and 0.500 ng/mL for the second group.

[00385] Figure 8 compares mean plasma concentrations (+ standard deviations) over the release time of hydromorphone after PO administration to rats of 20 mg/kg of Compound 3 alone (solid line) or with 10 mg/kg of Compound 101 (dashed line).

[00386] The results in Table 8 and Figure 8 indicate that Compound 101 attenuates the ability of Compound 3 to release hydromorphone, related to the suppression of Cmax and the delay of Tmax.

Example 10: Oral administration of Compound 4 and the trypsin inhibitor Compound 101 to rats.

[00387] Saline solutions of Compound 4 (which can be prepared as described in Example 13) and Compound 101 (prepared as described in Example 14) were dosed as indicated in Table 9. The dosing, sampling, and analysis procedures were as described in Example 4, except that Compound 4 and Compound 101 were combined for dosing, and Compound 4 without inhibitor was administered to 7 rats.

[00388] Table 9 and Figure 9 provide results for rats that received 20 mg/kg of Compound 4 with or without 10 mg/kg of Compound 101, as indicated. The results in Table 9 are reported as Cmax and Tmax of hydromorphone in plasma for each group of 4 rats. Table 9. Cmax and Tmax of HM in rat plasma

[00389] The lower limit of quantification was 0.500 ng/mL for both groups.

[00390] Figure 9 compares mean plasma concentrations (+ standard deviations) over the release time of hydromorphone after PO administration to rats of 20 mg/kg of Compound 4 alone (solid line) or with 10 mg/kg of Compound 101 (dashed line).

[00391] The results in Table 9 and Figure 9 indicate that Compound 101 attenuates the ability of Compound 4 to release hydromorphone, related to the suppression of Cmax and the delay of Tmax. Example 11 Synthesis of the hydromorphic ester of [2-((S)-2-amino-5-guanidino-pentanoylamino)-ethyl]-methyl-carbamic acid (Compound 2) Preparation 1 Synthesis of 2,2,2-trifluoro-N-(2-methylaminoethyl)-acetamide (A).

[00392] A solution of N-methylethylenediamine (27.0 g, 364.0 mmol) and ethyl trifluoroacetate (96.6 mL, 838.0 mmol) in a mixture of acetonitrile (350 mL) and water (7.8 mL, 436 mmol) was refluxed overnight with stirring. The solvents were then evaporated under vacuum. The residue was further evaporated with isopropanol (3 x 100 mL). The residue was dissolved in dichloromethane (500 mL) and left overnight at room temperature. The crystals formed were filtered, washed with dichloromethane and dried in vacuo, giving rise to compound A (96.8 g, 94%) in the form of a white solid powder.Preparation 2Synthesis of benzyl ester of {methyl-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-carbamic acid (B).

[00393] A solution of compound A (96.8 g, 340.7 mmol) and DIEA (59.3 mL, 340.7 mmol) in THF (350 mL) was cooled to ~5 °C, followed by the dropwise addition of a solution of N-(benzyloxycarbonyl)succinimide (84.0 g, 337.3 mmol) in THF (150 mL) over a period of 20 minutes. The temperature of the reaction mixture was increased to room temperature and stirring was continued for an additional 30 minutes, followed by evaporation of the solvents. The resulting residue was dissolved in EtOAc (600 mL). The EtOAc was extracted with 5% aqueous NaHCO3 (2 x 150 mL) and brine (150 mL). The organic layer was separated and evaporated, giving rise to compound B in the form of a yellowish oil (103.0 g, 340.7 mmols). LC-MS [M+H] 305.3 (C13H15F3N2O3 +H, calc: 305.3). Compound B was used without further purification. Preparation 3 Synthesis of benzyl ester of (2-aminoethyl)methylcarbamic acid (C).

[00394] A solution of compound B (103.0 g, 340.7 mmol) in MeOH (1200 mL) was added to a solution of LiOH (16.4 g, 681.4 mmol) in water (120 mL). The reaction mixture was stirred at room temperature for 3 hours. The solvents were evaporated to µ of the initial volume, followed by dilution with water (400 mL). The solution was extracted with EtOAc (2 x 300 mL). The organic layer was washed with brine (200 mL), dried over MgSO4, and evaporated in vacuo. The resulting residue was dissolved in ether (300 mL) and treated with 2 N HCl/ether (200 mL). The precipitate formed was filtered, washed with ether and dried in vacuo, giving rise to the hydrochloride salt of compound C (54.5 g, 261.2 mmols) in the form of a white solid. LC-MS [M+H] 209.5 (C11H16N2O2 +H, calc: 209.3). Preparation 4 Synthesis of tert-butyl ester of {(S)-4-({amino-[(E)-2,2!4!6,7- pentamethyl-2,3-dihydro-benzofuran-5-sulfonylimino]-methyl}-amino)-1-[2- (benzyloxycarbonyl-methyl-amino)-ethyl carbamoyl]-butyl}-carbamic acid (D).

[00395] A solution of Boc-Arg(Pbf)-OH (3.33 g, 6.32 mmol), HATU (2.88 g, 7.58 mmol) and DIEA (7.4 mL, 31.6 mmol) in DMF (40 mL) was kept at room temperature for 20 minutes, followed by the addition of compound C hydrochloride (1.45 g, 6.95 mmol). Stirring was continued for an additional 1 hour. The reaction mixture was diluted with EtOAc (500 mL) and extracted with water (3 x 75 mL) and brine (75 mL). The organic layer was dried over MgSO4 and then evaporated, yielding compound D (4.14 g, 5.77 mmol) as a yellowish amorphous solid. LC-MS [M+H] 717.6 (C35H52N6O8S +H, calc: 717.9).Preparation 5Synthesis of (2-methylamino-ethyl)-amide of (S)-2-amino-5-({amino-[(E)-2,2,4,6,7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonylimino]-methyl}-amino)-pentanoic acid (E).

[00396] Compound D (4.14 g, 5.77 mmol) and AcOH (330 μl, 5.77 mmol) were dissolved in methanol (40 mL), followed by the addition of a Pd/C suspension (5% by weight, 880 mg) in water (5 mL). The reaction mixture was hydrogenated (Parr apparatus, 517.11 kPa (75 psi)) at room temperature for 2.5 hours. The catalyst was filtered through a Celite pad in a sintered glass funnel and washed with methanol. The filtrate was evaporated in vacuo, yielding compound E (1.96 g, 3.2 mmol) as a yellowish amorphous solid. LC-MS [M+H] 483.2 (C22H38N6O4S +H, calc: 483.2).Preparation 6 Synthesis of tert-butyl ester of {(S)-4-({amino-[(E)-2,2,4,6,7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonUimino]-methyl}-amino)-1-[2-(hydromorphylcarbonyl-methyl-amino)-ethyl carbamoyl]-butyl}-carbamic acid (F).

[00397] A suspension of hydromorphone hydrochloride (332 mg, 1.03 mmol) and DIEA (179 μl, 1.03 mmol) in chloroform (4 mL) was sonicated in an ultrasonic bath at room temperature for 1 hour. This was followed by the addition of 4-nitrophenyl chloroformate (162 mg, 0.80 mmol). The reaction mixture was sonicated in an ultrasonic bath at room temperature for an additional 1 hour, followed by the addition of compound E solution (400 mg, 0.67 mmol) and 1-hydroxybenzotriazole (154 mg, 1.14 mmol) in DMF (4 mL). The reaction mixture was stirred overnight (~18 hours) at room temperature, followed by evaporation of the solvents in vacuo. The residue was dissolved in MeOH (5 mL) and precipitated with the addition of ether (500 mL). The precipitate formed was filtered and dried in vacuo, giving rise to compound F (520 mg, yield exceeded the quantitative) in the form of a nearly white solid. LC-MS [M+H] 894.6 (C45H63N7O10S +H, calc: 894.9). Synthesis of hydromorphic ester of [2-((S)-2-amino-5-guanidino-pentanoylamino)-ethyl]-methyl-carbamic acid (Compound 2).

[00398] Compound F (679 mg, 0.76 mmol) was dissolved in a 5% m-cresol/TFA mixture (10 mL). The reaction mixture was kept at room temperature for 1 hour, followed by dilution with ether (500 mL). The precipitate formed was filtered, washed with ether (100 mL) and dried in vacuo, yielding crude compound 2 (441 mg, yield exceeded the quantity) as a whitish solid. LC-MS [M+H] 542.4 (C27H39N7O5 +H, calc: 542).

[00399] Crude compound 2 was dissolved in water (10 mL) and subjected to purification by preparative reverse-phase HPLC. [Nanosyn-Pack Microsorb (100-10) C-18 column (50 x 300 mm); flow rate = 100 mL/minute; injection volume 10 mL; mobile phase A: 100% water, 0.1% TFA; mobile phase B: 100% acetonitrile, 0.1% TFA; isocratic elution to 0% B in 5 minutes, gradient elution to 6% B in 6 minutes, isocratic elution to 6% B in 23 minutes, gradient elution from 6% B to 55% B in 66 minutes; detection at 254 nm]. The portions containing the desired compound were combined and concentrated in vacuo. The residue was dissolved in i-PrOH (20 mL) and evaporated in vacuo (the procedure was repeated twice). The residue was dissolved in i-PrOH (2 mL) and treated with 2 N HCl/ether (100 mL, 200 mmol), yielding the hydrochloride salt of Compound 2 (80 mg, 17% yield, 98% purity) as a white solid. LC-MS [M+H] 542.0 (C27H39N7O5+H, calc: 542.9). Retention time*: 2.04 minutes.

[00400] * - [Chromolith SpeedRod RP-18e C18 column (4.6 x 50 mm); flow rate 1.5 mL/minute; mobile phase A: 0.1% TFA/water; mobile phase B 0.1% TFA/ACN; gradient elution from 5% B to 100% B in 9.6 minutes, detection 254 nm]Example 12Synthesis of (2-methylaminoethyl)amide hydromorphone ester of (S)-2-acetylamino-6-aminohexanoic acid (Compound 3) Preparation 7Synthesis of 9H-fluoren-9-ylmethyl ester of {(S)-1-[2-(benzyloxycarbonyl-methyl-amino)-ethylcarbamoyl]-5-tert-butoxycarbonylamino-pentylj-carbamic acid (G).

[00401] To a solution of Fmoc-Lys(Boc)-OH (2.0 g, 4.26 mmol) in DMF (50 mL) DIEA (2.38 mL, 13.65 mmol) was added and stirred for 15 minutes at room temperature. The reaction mixture was then cooled to ~5 °C, followed by the addition of HATU (1.95 g, 5.12 mmol), added in portions, and stirred for 30 minutes. CBZ-diamine (1.05 g, 4.26 mmol) was added to the reaction mixture and stirred at room temperature for 2 hours. The reaction mixture was diluted with EtOAc (250 mL), washed with water (250 mL) and brine (250 mL). The organic layer was separated and dried in Na2SO4, and removal of the solvent in vacuo gave rise to compound G (2.3 g, 82%). LC-MS [M+H] 659.6 (C37H46N4O7+H, calc: 659.7). Preparation 8 Synthesis of tert-butyl ester of {(S)-5-amino-5-[2-(benzyloxycarbonyl-methyl-amino)-ethylcarbamoyl]-pentyl}-carbamic acid (H).

[00402] To a solution of compound G (2.3 g, 3.49 mmol) in EtOAc (50 mL) was added piperidine (0.34 mL, 3.49 mmol). The reaction mixture was stirred for 18 hours at room temperature and then the solvents were removed in vacuo. The residue was dissolved in a minimal amount of EtOAc and then precipitated with Et2O. The precipitate was removed by filtration, washed with Et2O and dried, yielding compound H (1.4 g, 94%). LC-MS [M+H] 437.6 (C22H36N4O5+H, calc: 437.5).Preparation 9Synthesis of isopropyl ester of {(S)-5-acetylamino-5-[2-(benzyloxycarbonyl-methyl-amino)-ethyl carbamoyl]-pentyl}-carbamic acid (I).

[00403] To a solution of compound H (1.4 g, 3.21 mmol) in CHCl3 (10 mL) at room temperature, DIEA (2.6 mL, 15 mmol) was added followed by Ac2O (0.85 mL, 9.0 mmol). The reaction mixture was stirred at room temperature for 2 hours. The solvents were removed in vacuo and then the residue was dissolved in dichloromethane (100 mL). The organic layer was washed with 10% citric acid (75 mL), saturated NaHCO3 (75 mL) and brine (75 mL). The organic layer was separated and dried over Na2SO4 and the solvent was removed in vacuo, yielding compound I (1.45 g, 99%). LC-MS [M+H] 479.5 (C24H38N4O6+H, calc: 479.5).Preparation 10Synthesis of tert-butyl ester of [(S)-5-acetylamino-5-(2-methylamino-ethylcarbamoyl)-pentyl]-carbamic acid (,).

[00404] To a solution of compound I (1.4 g, 3.00 mmol) in MeOH (40 mL) was added 5% Pd/C (300 mg). This reaction mixture was hydrogenated at 70 psi for 2 hours. Then, the reaction mixture was filtered through a celite pad and the MeOH was removed in a rotary evaporator, yielding compound J (1.02 g, 98%). LC-MS [M+H] 344.9 (C16H32N4O4+H, calc: 345.4). Preparation 11 tert-butyl-hydromorphone diester of [(S)-5-acetylamino-5-(2-methylaminoethylcarbamoyl)-pentyl]-carbamic acid (L).

[00405] Hydromorphone HCl salt (1.24 g, 3.86 mmol) and DIEA (0.67 mL, 3.86 mmol) were suspended in CHCh (12 mL) and sonicated for 1 hour at room temperature. 4-Nitrophenyl chloroformate (600 mg, 2.97 mmol) was added to the reaction mixture and sonicated for 100 minutes. A solution of compound J (1.02 g, 2.97 mmol) and HOBt (0.52 g, 3.86 mmol) in DMF (12 mL) was added dropwise to the activated hydromorphone reaction mixture, and the system was stirred at room temperature overnight (~18 hours). The solvents were then removed in vacuo, and the residue was dissolved in a minimal amount of MeOH and precipitated with an excess of Et2O. The precipitate was removed by filtration, washed with Et2O and dried under vacuum, yielding compound L. LC-MS [M+H] 656.9 (C34H49N5O8+H, calc: 656.7). This crude product was purified by preparative reverse-phase HPLC. [Column: VARIAN, LOAD & LOCK, L&L 4002-2, Packing: Microsob 100-10 C18, Injection volume: ~15 mL, Injection flow rate: 20 mL/minute, 100% A (water/0.1% TFA), Flow rate: 100 mL/minute, Portion: 30 seconds (50 mL), Method: 0% B (MeCN/0.1% TFA)/2 minutes/75% B/96 minutes/100 mL/minute/254 nm]. The pure portions were combined, the solvents were removed in vacuum. The residue was dried via coevaporation with i-PrOH (4 x 100 mL), giving rise to compound L in the form of a yellow oil (0.90 g, 46%). Synthesis of hydromorphone ester of (2-methylaminoethyl)-amide of (S)-2-acetylamino-6-aminohexanoic acid (Compound 3).

[00406] Compound L (0.90 g, 1.37 mmol) was suspended in dioxane (~2 mL), sonicated, and treated with 4.0 N HCl/dioxane (~20 mL) at room temperature. A white precipitate formed immediately. The mixture was then diluted with Et2O (200 mL) and hexane (20 mL), and the precipitate was removed by filtration, washed with Et2O (100 mL) and hexane (100 mL), and dried under vacuum, yielding Compound 3 (0.67 g, 78% yield, 97.5% purity). LC-MS [M+H] 556.3 (C29H41N5O6+H, calc: 556.6). Example 13 Synthesis of hydromorphone ester of [2-((S)-2-acetylamino-5-guanidino-pentanoylamino)-ethyl]-ethyl-carbamic acid (Compound 4) Preparation 12 Synthesis of 2,2,2-trifluoro-N-(2-ethylaminoethyl)-acetamide (M).

[00407] A solution of N-ethylethylenediamine (10.0 g, 113.4 mmol) and ethyl trifluoroacetate (32.0 mL, 261 mmol) in a mixture of acetonitrile (110 mL) and water (2.5 mL, 139 mmol) was refluxed with stirring overnight (~18 hours). The solvents were evaporated in vacuo. The residue was evaporated again with i-PrOH (3 x 100 mL). The residue was dissolved in dichloromethane (500 mL) and left overnight at room temperature. The crystals formed were filtered, washed with dichloromethane (100 mL) and dried in vacuo, giving rise to compound M (24.6 g, 82.4 mmols) in the form of a white solid powder.Preparation 13Synthesis of benzyl ester of {ethyl-[2-(2,2,2-trifluoro-acetylamino)-ethyl]-carbamic acid (N).

[00408] A solution of compound M (24.6 g, 82.4 mmol) and DIEA (14.3 mL, 82.4 mmol) in THF (100 mL) was cooled to ~5 °C, followed by the dropwise addition of a solution of N-(benzyloxycarbonyl)succinimide (20.3 g, 81.6 mmol) in THF (75 mL) over 20 minutes. The temperature of the reaction mixture was increased to room temperature and stirring was continued for an additional 30 minutes. The solvents were evaporated and the residue was dissolved in EtOAc (500 mL). The organic layer was extracted with 5% aqueous NaHCO3 (2 x 100 mL) and brine (100 mL). The organic layer was evaporated, yielding compound N (24.9 g, 78.2 mmol) as a yellowish oil. LC-MS [M+H] 319.0 (C14H17F3N2O3 +H, calc: 319.2). Compound N was used without further purification.Preparation 14Synthesis of benzyl ester of (2-aminoethyl)ethylcarbamic acid (0).

[00409] A solution of compound N (24.9 g, 78.2 mmol) in MeOH (300 mL) was added to a solution of LiOH (3.8 g, 156 mmol) in water (30 mL). The reaction mixture was stirred at room temperature for 5 hours. Then, the solvents were evaporated to µ of the initial volume, followed by dilution with water (200 mL). The solution was extracted with EtOAc (200 mL x 2) and the organic layer was washed with brine (100 mL), dried over MgSO4 and evaporated in vacuo. The residue was dissolved in ether (200 mL) and treated with 2 N HCl/ether (200 mL). The precipitate formed was filtered, washed with ether and dried in vacuo, yielding the hydrochloride salt of Compound O (12.1 g, 46.7 mmol) as a white solid. LC-MS [M+H] 222.9 (C12H18N2O2 +H, calc: 223.2).Preparation 15Synthesis of benzyl ester of {2-[boc-Arg(Pbf)]-aminoethyl}-ethyl-carbamic acid (P).

[00410] A solution of Boc-Arg(Pbf)-OH (3.0 g, 5.69 mmol), compound O (1.62 g, 6.26 mmol), DIEA (3.17 mL, 18.21 mmol) and HATU (2.59 g, 6.83 mmol) in DMF (20 mL) was stirred at room temperature for 1 hour. The reaction mixture was diluted with EtOAc (300 mL) and extracted with water (3 x 75 mL) and brine (75 mL). The organic layer was dried over MgSO4, filtered and then evaporated, giving rise to compound P (5.97 g, yield exceeded the quantity) in the form of a yellowish oil. LC-MS [M+H] 731.5 (C36H54N6O8S +H, calc: 731.7). Compound P was used without further purification.Preparation 16Synthesis of benzyl ester of {2-[H-Arg(Pbf)]-aminoethyl}-ethyl-carbamic acid (Q).

[00411] Compound P (5.69 mmol) was dissolved in dioxane (20 mL) and treated with 4 N HCl/dioxane (100 mL, 70 mmol) at room temperature for 1 hour. The solvent was then removed in vacuo, followed by suspension in i-PrOH (50 mL) and finally the solvent was evaporated to remove residual solvents (the procedure was repeated twice). The crude reaction mixture was dried in vacuo, yielding compound Q (5.97, yield exceeded the quantity) as a yellowish solid. LC-MS [M+H] 631.5 (C31H46N6O6S +H, calc: 631.2). Compound Q was used without further purification.Preparation 17Synthesis of benzyl ester of {2-[Ac-Arg(Pbf)]-aminoethyl}-ethyl-carbamic acid (R).

[00412] A solution of compound Q (5.69 mmol), AC2O (649 μl, 6.83 mmol) and DIEA (2.97 mL, 17.07 mmol) in chloroform (20 mL) was stirred at room temperature for 1 hour. This was followed by the addition of 2M EtNH2/THF (1.71 mL, 3.41 mmol). The reaction mixture was stirred at room temperature for an additional 30 minutes, followed by dilution with EtOAc (300 mL). The organic layer was extracted with water (75 mL), 2% aqueous H2SO4 (75 mL), water (3 x 75 mL) and brine (75 mL). The organic layer was then dried over MgSO4 and evaporated, yielding compound R (3.99 g, yield exceeded quantity) as a yellowish solid. LC-MS [M+H] 673.6 (C33H48N6O7S +H, calc: 672.9). Compound R was used without further purification. Preparation 18 Synthesis of N-[Ac-Arg(Pbf)]-N'-ethyl-ethane-1,2-diamine (S).

[00413] Compound R (5.69 mmol) was dissolved in methanol (50 mL) followed by the addition of a Pd/C suspension (5% by weight, 1 g) in water (5 mL). The reaction mixture was hydrogenated (Parr apparatus, 551.58 kPa (80 psi)) at room temperature for 1 hour. After completion, the catalyst was filtered through a Celite pad into a sintered glass funnel and washed with methanol. The filtrate was evaporated in vacuo, yielding compound S (3.06 g, quantitative yield) as a colorless oil. LC-MS [M+H] 539.5 (C25H42N6O5S +H, calc: 539.9). Compound S was used without further purification. Synthesis of hydromorphone ester of [2-(2-acetylamino-5-guanidino-pentanoylamino)-ethyl]-ethyl-carbamic acid (Compound 4).

[00414] A suspension of hydromorphone hydrochloride (2.75 g, 8.54 mmol) and DIEA (1.49 mL, 8.54 mmol) in chloroform (8 mL) was sonicated in an ultrasonic bath at room temperature for 1 hour, followed by the addition of 4-nitrophenyl chloroformate (1.38 g, 6.83 mmol). The reaction mixture was sonicated in an ultrasonic bath at room temperature for an additional 1 hour, followed by the addition of compound S solution (3.06 g, 5.69 mmol) and 1-hydroxybenzotriazole (1.31 g, 9.67 mmol) in DMF (8 mL). The reaction mixture was stirred overnight (~18 hours) at room temperature, followed by solvent evaporation in vacuo. The crude reaction mixture was dissolved in MeOH (10 mL) and precipitated with ether (500 mL). The precipitate formed was filtered and dried in vacuo, yielding compound 4 protected with Pbf (6.96 g yield exceeded the quantitative) in the form of a whitish solid. LC-MS [M+H] 850.6 (C43H59N7O9S +H, requires 850.2).

[00415] Pbf-protected Compound 4 was dissolved in a 5% m-cresol/TFA mixture (100 mL). The reaction mixture was kept at room temperature for 1 hour, followed by dilution with ether (2 L). A precipitate formed, which was subsequently filtered through a sintered glass funnel, washed with ether (200 mL), and dried in vacuo, yielding crude compound 4 (5.2 g, 97%) as a whitish solid. Crude compound 4 (5.2 g, 5.54 mmol) was dissolved in water (50 mL) and purified by HPLC. [Nanosyn-Pack Microsorb (100-10) C-18 column (50 x 300 mm); flow rate = 100 mL/minute; injection volume 50 mL; mobile phase A: 100% water, 0.1% TFA; Mobile phase B: 100% acetonitrile, 0.1% TFA; isocratic elution at 0% B in 5 minutes, gradient elution to 6% B in 6 minutes, isocratic elution at 6% B in 13 minutes, gradient elution from 6% B to 55% B in 76 minutes; detection at 254 nm]. The portions containing the desired compound were combined and concentrated in vacuo. The residue was dissolved in i-PrOH (50 mL) and evaporated in vacuo (the procedure was repeated twice). The residue was dissolved in i-PrOH (50 mL) and treated with 2 N HCl/ether (200 mL, 400 mmol), yielding the hydrochloride salt of Compound 4 (1.26 g, 32% yield, 95.7% purity) as a white solid. LC-MS [M+H] 598.4 (C30H43N7O6+H, calc: 598.7). Retention time*: 2.53 minutes.

[00416] * - [Chromolith SpeedRod RP-18e C18 column (4.6 x 50 mm); flow rate 1.5 mL/minute; mobile phase A: 0.1% TFA/water; mobile phase B 0.1% TFA/ACN; gradient elution from 5% B to 100% B in 9.6 minutes, detection 254 nm].Example 14Synthesis of (S)-ethyl 4-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperazino-1-carboxylate (Compound 101) Preparation 19Synthesis of tert-butyl ester of 4-[(S)-5-({amino-[(E)-2,2!4!6,7- pentamethyl-2,3-dihydro-benzofuran-5-sulfonylimino]-methyl}-amino)-2-(9H-fluoren-9-ylmethoxy-carbonylamino)-pentanoyl]-piperazino-1-carboxylic acid (T).

[00417] To a solution of Fmoc-Arg(Pbf)-OH 1 (25.0 g, 38.5 mmol) in DMF (200 mL) at room temperature, Dl-EA (13.41 mL, 77.1 mmol) was added. After stirring at room temperature for 10 minutes, the reaction mixture was cooled to ~5 °C. HATU (16.11 g, 42.4 mmol) was added to the reaction mixture in portions, the system was stirred for 20 minutes, and a solution of tert-butyl-1-piperazine carboxylate (7.18 g, 38.5 mmol) in DMF (50 mL) was added dropwise. The reaction mixture was stirred at ~5 °C for 5 minutes. The reaction mixture was then allowed to warm to room temperature and stirred for 2 hours. The solvent was removed under vacuum and the residue was dissolved in EtOAc (500 mL) and washed with water (2 x 750 mL), 1% H2SO4 (300 mL) and brine (750 mL). The organic layer was separated and dried over Na2SO4 and the solvent was removed under vacuum to a total volume of 100 mL. Compound T was collected for the next step as a solution in EtOAc (100 mL). LC-MS [M+H] 817.5 (C43H56N6O8S+H, calc: 817.4).Preparation 20Synthesis of tert-butyl ester of 4-[(S)-2-amino-5-({amino-[(E)- 2# 2# 4# 6#7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonylimino]-methyl}-amino)-pentanoyl]-piperazino-1-carboxylic acid (U).

[00418] To a solution of compound T (46.2 mmol) in EtOAc (175 mL) at room temperature, piperidine (4.57 mL, 46.2 mmol) was added, and the reaction mixture was stirred for 18 hours at room temperature. Then, the solvent was removed in vacuo, the resulting residue was dissolved in a minimal amount of EtOAc (~50 mL), and hexane (~1 L) was added. The crude precipitate was removed by filtration and recrystallized again with EtOAc (~30 mL) and hexane (~750 mL). The precipitate was removed by filtration, washed with hexane, and dried in vacuo, yielding compound U (28.0 g, 46.2 mmol). LC-MS [M+H] 595.4 (C28H46N6O6S+H, calc: 595.3).Preparation 21Synthesis of tert-butyl ester of 4-[(S)-5-({amino-[(E)-2,2!4!6,7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonylimino]-methyl}-amino)-2-(naphthalene-2-sulfonyl-amino)-pentanoyl]-piperazino-1-carboxylic acid (V).

[00419] To a solution of compound U (28.0 g, 46.2 mmol) in THF (250 mL) was added 1 N aqueous NaOH (171 mL). The reaction mixture was cooled to ~5 °C, and a solution of 2-naphthalenesulfonyl chloride (26.19 g, 115.6 mmol) in THF (125 mL) was added dropwise. The reaction mixture was stirred at ~5 °C for 10 minutes, with continued stirring at room temperature for 2 hours. The reaction mixture was diluted with EtOAc (1 L) and washed with 1 N aqueous NaOH (1 L), water (1 L), and brine (1 L). The organic layer was separated and dried over Na2SO4, and solvent removal was performed in vacuo, yielding compound V (36.6 g, 46.2 mmol). LC-MS [M+H] 785.5 (C38H52N6O8S2+H, calc: 785.9).Preparation 22Synthesis of 1-amino-1-[(S)-4-(naphthalene-2-sulfonylamino)-5-oxo-5-piperazin-1-yl-pentylamino]-methyl-(E)-ylideneamide of 2,2,4,6,7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonic acid (4).

[00420] To a solution of compound V (36.6 g, 46.2 mmol) in dioxane (60 mL) was added dropwise 4 M HCl in dioxane (58 mL). The reaction mixture was stirred at room temperature for 1.5 hours. Et2O (600 mL) was added to the reaction mixture, the precipitated product was removed by filtration, washed with Et2O and finally dried under vacuum, yielding compound W (34.5 g, 46.2 mmol). LC-MS [M+H] 685.4 (C33H44N6O6S2+H, calc: 685.9). Compound W was used without further purification.Preparation 23Ethyl ester synthesis of 4-[(S)-5-({amino-[(E)-2,2!4!6,7-pentamethyl-2#3-dihydro-benzofuran-5-sulfonyl-imino]-methyl}-amino)-2-(naphthalene-2-sulfonylamino)-pentanoyl]-piperazino-1-carboxylic acid (X).

[00421] To a solution of compound W (8.0 g, 11.1 mmol) in CHCl3 (50 mL) DIEA (4.1 mL, 23.3 mmol) was added at room temperature and the system was stirred for 15 minutes. The mixture was cooled to ~5 °C, and ethyl chloroformate (1.06 mL, 11.1 mmol) was added dropwise. After stirring at room temperature overnight (~18 hours), the solvent was removed under vacuum. The residue was dissolved in MeOH (25 mL) and Et2O (500 mL) was added. The crude precipitated product was removed by filtration, washed with Et2O, and dried under vacuum, yielding compound X (8.5 g, 11.1 mmol). LC-MS [M+H] 757.6 (C36H48N6O8S2+H, calc: 757.9). Compound X was used without further purification. Synthesis of (S)-ethyl 4-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperazino-1-carboxylate (Compound 101)

[00422] A 5% m-cresol/TFA solution (50 mL) was added to compound X (8.5 g, 11.1 mmol) at room temperature. After stirring for 1 hour, the reaction mixture was precipitated with Et2O (500 mL). The precipitate was filtered and washed with Et2O and dried under vacuum, yielding the crude product. The crude product was purified by preparative reversed-phase HPLC. [Column: VARIAN, LOAD & LOCK, L&L 4002-2, Packing: Microsob 100-10 C18, Injection, Volume: 15 mL x 2, Injection flow rate: 20 mL/minute, 100% A, (water/0.1% TFA), Flow rate: 100 mL/minute, Fraction: 30 seconds (50 mL), Method: 0% B (MeCN/0.1% TFA)-60% B/60 minutes/100 mL/minute/254 nm]. Solvents were removed from the pure fractions in vacuo. Traces of water were removed by coevaporation with 2 x i-PrOH (50 mL). The residue was dissolved in a minimal amount of i-PrOH and the product was precipitated with 2 M HCl in Et2O. The product was removed by filtration, washed with Et2O and dried under vacuum, yielding Compound 101 in the form of the salt HCl 7 (3.78 g, 63% yield, 99.4% purity). LC-MS [M+H] 505.4 (C38H52N6O8S2+H, calc: 505.6). Example 15 Synthesis of (S)-ethyl 4-(5-guanidino-2-(2,4,6-triisopropylphenylsulfonamido)pentanoyl)piperazino-1-carboxylate (Compound 102) Preparation 24Ethyl ester synthesis of 4-[(S)-5-({amino-[(E)-2,2!4!6,7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonyl-imino]-methyl}-amino)-2-tert-butoxycarbonylamino-pentanoyl]-piperazino-1-carboxylic acid (I).

[00423] To a solution of Boc-Arg(Pbf)-OH (13.3 g, 25.3 mmol) in DMF (10 mL) DIEA (22.0 mL, 126.5 mmol) was added at room temperature and the system was stirred for 15 minutes. The reaction mixture was then cooled to ~5 °C, HATU (11.5 g, 30.3 mmol) was added in portions and the system was stirred for 30 minutes, followed by the dropwise addition of ethyl-1-piperazine carboxylate (4.0 g, 25.3 mmol) in DMF (30 mL). After 40 minutes, the reaction mixture was diluted with EtOAc (400 mL) and poured into H2O (1 L). It was extracted with EtOAc (2 x 400 mL) and washed with H2O (800 mL), 2% H2SO4 (500 mL), H2O (2 x 800 mL) and brine (800 mL). The organic layer was separated and dried over MgSO4, and the solvent was removed in vacuo. The resulting oily residue was dried in vacuo, yielding compound Y (16.4 g, 24.5 mmol) as a foamy solid. LC-MS [M+H] 667.2 (C31H50N6O8S+H, calc: 667.8). Compound Y was used without further purification.Preparation 25Ethyl ester synthesis of 4-[(S)-2-amino-5-({amino-[(E)- 2# 2# 4# 6#7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonilimino]-methyl}-amino)-pentanoyl]-piperazino-1-carboxylic acid (Z).

[00424] A solution of compound Y (20.2 g, 30.2 mmol) in dichloromethane (90 mL) was treated with 4.0 N HCl in 1,4-dioxane (90 mL, 363.3 mmol) and stirred at room temperature for 2 hours. Then, most of the dichloromethane was removed in vacuo and Et2O (~1 L) was added. The resulting precipitate was removed by filtration, washed with Et2O and dried in vacuo, yielding compound Z (17.8 g, 30.2 mmol). LC-MS [M+H] 567.8 (C26H42N6O6S+H, calc: 567.8).Preparation 26Ethyl ester synthesis of 4-[(S)-5-({amino-[(E)-2#2#4#6#7-pentamethyl-2#3-dihydro-benzofuran-5-sulfonyl-imino]-methyl}-amino)-2-(2#4#6-tri-isopropyl-benzenesulfonylamino)-pentanoyl]-piperazino-1-carboxylic acid (AA).

[00425] A solution of compound Z (1.0 g, 1.8 mmol) in THF (7 mL) was added to 3.1 N aqueous NaOH (4.0 mL) and the system was stirred for 5 minutes. The reaction mixture was cooled to ~5 °C, then a solution of tripsyl chloride (2.2 g, 7.3 mmol) in THF (5 mL) was added dropwise and the system was stirred at room temperature overnight (~18 hours). The reaction mixture was diluted with H2O (130 mL), acidified with 2% H2SO4 (15 mL) and extracted with EtOAc (3 x 80 mL). The organic layer was combined and washed with H2O (2 x 400 mL), saturated NaHCO3 (100 mL), H2O (200 mL) and brine (200 mL). The organic layer was separated and dried over MgSO4, and the solvent was removed in vacuo, yielding (2.9 g) of the crude product. This was purified by fast normal-phase chromatography (5-10% MeOH/DCM), yielding compound AA (0.52 g, 1.0 mmol). LC-MS [M+H] 833.8 (C41H64N6O8S2+H, calc: 834.1). Synthesis of (S)-ethyl 4-(5-guanidino-2-(2,4,6-triisopropylphenylsulfonamido)pentanoyl)piperazino-1-carboxylate (Compound 102)

[00426] A 5% m-cresol/TFA solution (40 mL) was added to compound AA (3.73 g, 3.32 mmol) at room temperature. After stirring for 45 minutes, the solvents were removed under vacuum. The residue was dissolved in dichloromethane (100 mL) and washed with H2O (3 x 200 mL) and brine (200 mL). The organic layer was separated and dried over MgSO4, and then the solvent was removed under vacuum. The residue was dissolved in dichloromethane (~5 mL), then hexane (~250 mL) was added, and a precipitate formed. This was washed with hexane and dried under vacuum, yielding the crude product (1.95 g). The crude product was purified by reversed-phase HPLC [Column: VARIAN, LOAD & LOCK, L&L 4002-2, Packing: Microsob 100-10 C18, Injection Volume: ~ 15 mL, Injection Flow Rate: 20 mL/minute, 100% A, (water/ 0.1% TFA), Flow Rate: 100 mL/minute, Fraction: 30 seconds (50 mL), Method: 25% B (MeCN / 0.1% TFA)/ 70% B/ 98 minutes/ 100 mL/minute/ 254 nm]. Solvents were removed from the pure fractions in vacuo. Traces of water were removed by co-evaporation with 2 x i-PrOH (50 mL). The residue was dissolved in a minimal amount of i-PrOH and the product was precipitated with 2 M HCl in Et2O. The product was removed by filtration, washed with Et2O and dried under vacuum, yielding the product in the form of the HCl salt of Compound 102 (0.72 g, 35% yield, 99.8% purity). LC-MS [M+H] 581.6 (C28H48N6O5S+H, calc: 581.7). Example 16 Synthesis of HCl salt of (S)-ethyl 1-(5-/uanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperidino-4-carboxylate (Compound 103) Preparation 27 Synthesis of ethyl ester of 1-[boc-Arg(Pbf)]-piperidine-4-carboxylic acid (BB)

[00427] To a solution of Boc-Arg(Pbf)-OH (3.4 g, 6.36 mmol) and HATU (2.9 g, 7.63 mmol) in DMF (15 mL) DIEA (7.4 mL, 42.4 mmol) was added and the reaction mixture was stirred for 10 minutes at room temperature. A solution of ethyl isonipecotate (1.0 g, 6.36 mmol) in DMF (6 mL) was added to the reaction mixture dropwise. The reaction mixture was stirred at room temperature for 1 hour, then diluted with ethyl acetate (150 mL) and poured into water (500 mL). The product was extracted with ethyl acetate (2 x 100 mL). The organic layer was washed with 0.1 N aqueous HCl (200 mL), 2% aqueous sodium bicarbonate (200 mL), water (200 mL), and brine (200 mL). The organic layer was dried over sodium sulfate and filtered, then evaporated in vacuo. The resulting oily product was dried in vacuo overnight, yielding compound BB (3.7 g, 5.57 mmol) as a viscous solid. LC-MS [M+H] 666.5 (C32H51N5O8 S+H, calc: 666.7). Compound BB was used without further purification. Preparation 28 Synthesis of HCl salt of ethyl ester of 1-[Arg(Pbf)]-piperidine-4-carboxylic acid (CC)

[00428] To a solution of compound BB (4.7 g, 7.07 mmol) in dichloromethane (25 mL) was added 4 N HCl in dioxane (25.0 mL, 84.84 mmol) and the reaction mixture was stirred at room temperature for 2 hours. The reaction mixture was concentrated in vacuo to ~20 mL of solvent, and then diluted with diethyl ether (250 mL) to produce a fine white precipitate. The reaction mixture was stirred for 1 hour, the solid was washed with ether (50 mL) and dried under high vacuum overnight, yielding compound CC (4.3 g, 7.07 mmol) in the form of a fine powder. LC-MS [M+H] 566.5 (C27H43N5O6 S+H, calc: 566.7). Preparation 29 Synthesis of ethyl ester of 1-[5(S)-(N'-Pbf-guanidino)-2-(naphthaIeno-2-suIfoniIamino)-pentanoiI]-piperidino-4-carboxylic acid (DD)

[00429] To a solution of CC compound (1.1 g, 1.6 mmol) and NaOH (260 mg, 5.9 mmol) in a mixture of THF (5 mL) and water (3 mL), a solution of 2-naphthalosulfonyl chloride (0.91 g, 2.5 mmol) in THF (10 mL) was added dropwise with stirring at ~5 °C. The reaction mixture was stirred at room temperature for 1 hour and then diluted with water (5 mL). 1 N aqueous HCl (5 mL) was added to obtain a pH ~3. An additional amount of water was added (20 mL) and the product was extracted with ethyl acetate (3 x 50 mL). The organic layer was removed and then washed with 2% aqueous sodium bicarbonate (50 mL), water (50 mL) and brine (50 mL). The extract was dried over anhydrous sodium sulfate, filtered, and evaporated under vacuum. The oily product formed was dried under vacuum overnight, yielding compound DD (1.3 g, 1.6 mmol) as an oily foamy solid. LC-MS [M+H] 756.5 (C37H49N5O8S2+H, calc: 756.7). Synthesis of HCl salt of (S)-ethyl 1-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperidino-4-carboxylate (Compound 103)

[00430] DD compound (1.3 g, 1.6 mmol) was added to a flask and then treated with 5% m-cresol/TFA (10 mL). The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was then concentrated in vacuo to a volume of 5 mL. Diethyl ether (200 mL) was then added to the residue, forming a fine white precipitate. The precipitate was removed by filtration and washed with ether (2 x 25 mL). The resulting solid was dried in vacuo overnight, yielding a crude material that was purified by preparative reversed-phase HPLC. [Nanosyn-Pack Microsorb (100-10) C-18 column (50 x 300 mm); flow rate = 100 mL/minute; injection volume 12 mL (DMSO-water, 1:1, v/v); Mobile phase A: 100% water, 0.1% TFA; mobile phase B: 100% ACN, 0.1% TFA; gradient elution from 25% B to 55% B in 90 minutes, detection at 254 nm. The fractions containing the desired compound were combined and concentrated in vacuo. The residue was dissolved in i-PrOH (50 mL) and evaporated under vacuum (repeated twice). Then, the residue was dissolved in i-PrOH (5 mL) and treated with 2 N HCl/ether (100 mL, 200 mmol), resulting in a white precipitate. It was dried in vacuo overnight, yielding Compound 103 (306 mg, 31% yield, 95.7% purity) as a white solid. LC-MS [M+H] 504.5 (C24H33N5O5S+H, calc: 504.6). Example 17 Synthesis of HCl salt of (S)-ethyl 1-(5-guanidino-2-(2,4,6-triisopropylphenylsulfonamido)pentanoyl)piperidino-4-carboxylate (Compound 104) Preparation 30 Synthesis of ethyl ester of 1-[5(S)-(N'-Pbf-guanidino)-2-(2,4,6-tri-isopropyl-benzenesulfonylamino)-pentanoyl]-piperidine-4-carboxylic acid (EE)

[00431] To a solution of CC compound (1.0 g, 1.6 mmol) and NaOH (420.0 mg, 10.4 mmol) in a mixture of THF (5 mL) and water (4 mL), a solution of 2,4,6-triisopropylbenzenesulfonyl chloride (2.4 g, 8.0 mmol) was added dropwise with stirring and kept at ~5 °C. The reaction mixture was then stirred at room temperature for 1 hour, the progress of the reaction was monitored, then it was diluted with water (20 mL) and acidified with 1 N aqueous HCl (5 mL) to pH ~3. An additional amount of water was added (30 mL) and the product was extracted with ethyl acetate (3 x 50 mL). The organic layer was washed with 2% aqueous sodium bicarbonate (50 mL), water (50 mL) and brine (50 mL). The organic layer was dried over anhydrous sodium sulfate, filtered, and evaporated under vacuum. The oily residue formed was dried under vacuum overnight, yielding compound EE (1.0 g, 1.2 mmol) as an oily material. LC-MS [M+H] 832.8 (C42H65N5O8S2+H, calc: 832.7). Synthesis of HCl salt of 1-(5-guanidino-2-(2,4,6-triisopropylphenylsulfonamido)pentanoyl)piperidino-4-ethyl carboxylate (Compound 104)

[00432] Compound EE (2.3 g, 2.8 mmol) was added to a flask and then treated with 5% m-cresol/TFA (16 mL). The reaction mixture was stirred at room temperature for 1 hour. The reaction mixture was then concentrated in vacuo to a volume of 5 mL. Hexane (200 mL) was added to the residue and removed by decantation, resulting in an oily precipitate. The product was purified by preparative reversed-phase HPLC. [Nanosyn-Pack Microsorb (100-10) C-18 column (50 x 300 mm); flow rate = 100 mL/minute; injection volume 15 mL (DMSO-water, 1:1, v/v); mobile phase A: 100% water, 0.1% TFA; mobile phase B: 100% ACN, 0.1% TFA; [Gradient elution from 35% B to 70% B in 90 minutes, detection at 254 nm]. The fractions containing the desired compound were combined and concentrated in vacuo. The residue was dissolved in i-PrOH (100 mL) and evaporated in vacuo (repeated twice). The residue was dissolved in i-PrOH (5 mL) and treated with 2 N HCl/ether (100 mL, 200 mmol), resulting in an oily residue. It was dried in vacuo overnight, yielding Compound 104 (1.08 g, 62.8%) as a viscous solid. LC-MS [M+H] 580.6 (C29H49N5O5S+H, calc: 580.8). Example 18 Synthesis of (S)-6-(4-(5-guanidino-2-(naphthalene-2-sulfonami-do)pentanoyl)piperazin-1-yl)-6-oxo-hexanoic acid (Compound 105) Preparation 31 Synthesis of methyl ester of 6-{4-[(S)-5-({amino-[(E)-2!2,4!6,7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonylimino]-methyl}-amino)-2-(naphthalene-2-sulfonylamino)-pentanoyl]-piperazin-1-yl}-6-oxo-hexanoic acid (FF)

[00433] To a solution of compound W (1.5 g, 2.08 mmol) in CHCl3 (50 mL) was added DIEA (1.21 mL, 4.16 mmol), followed by adipoyl chloride (0.83 mL, 6.93 mmol) dropwise. The reaction mixture was stirred at room temperature overnight (~18 hours). The solvents were removed in vacuo and the residue was dried under vacuum, yielding compound FF (2.1 g, quantity exceeds quantitative). LC-MS [M+H] 827.5 (C40H54N6O9S2+H, calc: 827.3). Compound FF was used without further purification. Preparation 32 Synthesis of 6-{4-[(S)-5-({amino-[(E)-2,2!4!6!7-pentamethyl-2,3-dihydro-benzofuran-5-sulfonylimino]-methyl}-amino)-2-(naphthalene-2-sulfonylamino)-pentanoyl]-piperazin-1-yl}-6-oxohexanoic acid (GG)

[00434] To a solution of compound FF (2.1 g, 2.08 mmol) in THF (5 mL), H2O (5 mL) was added 2 M aqueous LiOH (6 mL). The reaction mixture was stirred at room temperature for 2 hours. The solvents were removed in vacuo, then the residue was dissolved in water (50 mL), acidified with saturated aqueous NaHSO4 (~100 mL) and extracted with EtOAc (2 x 100 mL). The organic layer was dried over Na2SO4 and solvent removal yielded compound GG (1.72 g, 2.08 mmol). LC-MS [M+H] 813.5 (C39H52N6O9S2+H, calc: 813.3). Compound GG was used without further purification. Synthesis of (S)-6-(4-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperazin-1-yl)-6-oxohexanoic acid (Compound 105)

[00435] A 5% m-cresol/TFA solution (25 mL) was added to compound GG (1.72 g, 2.08 mmol) at room temperature. After stirring for 30 minutes, the reaction mixture was precipitated with the addition of Et20 (~200 mL). The precipitate was filtered, washed with Et20, and dried under vacuum, yielding the crude product. The crude product was purified by preparative reverse-phase HPLC [Column: VARIAN, LOAD & LOCK, L&L 4002-2, Packing: Microsob 100-10 C18, Injection Volume: ~ 25 mL, Injection Flow Rate: 20 mL/minute, 95% A, (water/ 0.1% TFA), Flow Rate: 100 mL/minute, Fraction: 30 seconds (50 mL), Method: 5% B (MeCN / 0.1% TFA)/ 5 minutes/ 25% B/ 20 minutes/ 25% B/ 15 minutes/ 50% B/ 25 minutes/ 100 mL/minute/ 254 nm]. Solvents were removed from the pure fractions in vacuo. Traces of water were removed by coevaporation with i-PrOH (25 mL) (repeated twice). The residue was dissolved in a minimal amount of i-PrOH, then 2 M HCl in Et2O (~50 mL) was added and the system was diluted with Et2O (~250 mL). The precipitate formed was removed by filtration, washed with Et2O and dried under vacuum, yielding the product in the form of the HCl salt of Compound 105 (0.74 g, 59% yield, 98.9% purity). LC-MS [M+H] 561.4 (C26H36N6O6S+H, calc: 561.2). Example 19 Synthesis of 3-(4-carbamimidoylphenyl)-2-oxopropanoic acid (Compound 107)

[00436] Compound 107, that is, 3-(4-carbamimidoylphenyl)-2-oxopropanoic acid, can be produced using methods known to those skilled in the art, such as that described by Richter P et al., Pharmazie, 1977, 32, 216-220, and references thereto. The purity of Compound 107 used in Example 7 was estimated at 76%, an estimate due to the low UV absorbance of this compound via HPLC. Mass spectrometry data: LC-MS [M+H] 207.0 (C10H10N2O3+H, calc: 207.1). Example 20 Synthesis of (S)-5-(4-carbamimidoylbenzylamino)-5-oxo-4-((R)-4-phenyl-2-(phenylmethylsulfonamido)butanamido)pentanoic acid (Compound 108) Preparation 33Synthesis of benzyl ester of (S)-4-tert-butoxycarbonylamino-4-(4-cyano-benzylcarbamoyl)-butyric acid (HH).

[00437] A solution of Boc-Glu(OBzl)-OH (7.08 g, 21.0 mmol), BOP (9.72 g, 22.0 mmol) and DIEA (12.18 mL, 70.0 mmol) in DMF (50 mL) was kept at room temperature for 20 minutes, followed by the addition of 4-(aminomethyl)benzonitrile hydrochloride (3.38 g, 20.0 mmol). The reaction mixture was stirred at room temperature for an additional 1 hour and diluted with EtOAc (500 mL). The resulting solution was extracted with water (100 mL), 5% aqueous NaHCO3 (100 mL) and water (2 x 100 mL). The organic layer was dried in MgSO4, evaporated, and dried in vacuo, giving rise to compound HH (9.65 g, yield exceeded the quantity) in the form of a yellowish oil. LC-MS [M+H] 452.0 (C25H29N3O5 +H, calc: 452.4). Compound HH was used without further purification. Preparation 34 Synthesis of benzyl ester of (S)-4-tert-butoxy-carbonylamino-4-[4-(N-hydroxycarbamimidoyl)-benzylcarbamoyl]-butyric (II) acid.

[00438] A solution of compound HH (9.65 g, 20.0 mmol), hydroxylamine hydrochloride (2.10 g, 30.0 mmol) and DIEA (5.22 mL, 30.0 mmol) in ethanol (absolute, 150 mL) was refluxed for 6 hours. The reaction mixture was allowed to cool to room temperature and stirred for a further 16 hours, then the solvents were evaporated in vacuo. The resulting residue was dried in vacuo, yielding compound II (14.8 g, yield exceeded the quantitative) in the form of a yellowish oil. LC-MS [M+H] 485.5 (C25H32N4O6 +H, calc: 485.8). Compound II was used without further purification.Preparation 35Synthesis of benzyl ester of (S)-4-tert-butoxy-carbonylamino-4-[4-(N-acetyl-hydroxycarbamimidoyl)-benzylcarbamoyl]-butyric acid (JJ).

[00439] A solution of compound II (14.8 g, 20.0 mmol) and acetic anhydride (5.7 mL, 60.0 mmol) in acetic acid (100 mL) was stirred at room temperature for 45 minutes and then the solvent was evaporated in vacuo. The resulting residue was dissolved in EtOAc (300 mL) and extracted with water (2 x 75 mL) and brine (75 mL). The organic layer was then dried over MgSO4, evaporated and dried in vacuo, yielding compound JJ (9.58 g, 18.2 mmol) as a yellowish solid. LC-MS [M+H] 527.6 (C27H34N4O7 +H, calc: 527.9). Compound JJ was used without further purification. Preparation 36 Benzyl ester synthesis of (S)-4-[4-(N-acetylhydroxycarbamimidoyl)-benzyl carbamoyl]-butyric acid (KK).

[00440] Compound JJ (9.58 g, 18.2 mmol) was dissolved in 1,4-dioxane (50 mL) and treated with 4 N HCl/dioxane (50 mL, 200 mmol) at room temperature for 1 hour. The solvent was then evaporated in vacuo. The resulting residue was ground with ether (200 mL). The precipitate obtained was filtered, washed with ether (100 mL) and hexane (50 mL), and dried in vacuo, yielding compound KK (9.64 g, yield exceeded the quantity) as a whitish solid. LC-MS [M+H] 426.9 (C22H26N4O5 +H, calc: 427.3). Compound KK was used without further purification.Preparation 37Synthesis of (R)-4-phenyl-2-phenylmethanesulfonylaminobutyric acid (LL).

[00441] A solution of D-homophenylalanine (10.0 g, 55.9 mmol) and NaOH (3.35 g, 83.8 mmol) in a mixture of 1,4-dioxane (80 mL) and water (50 mL) was cooled to ~5 °C, followed by alternating addition of α-toluenesulfonyl chloride (16.0 g, 83.8 mmol; 5 portions per 3.2 g) and 1.12 M NaOH (50 mL, 55.9 mmol; 5 portions per 10 mL) maintaining a pH > 10. The reaction mixture was acidified with 2% aqueous H2SO4 to pH = ~2. The resulting solution was extracted with EtOAc (2 x 200 mL). The organic layer was washed with water (3 x 75 mL) and dried over MgSO4, and then the solvent was evaporated in vacuo. The resulting residue was dried in vacuo, yielding compound LL (12.6 g, 37.5 mmol) as a white solid. LC-MS [M+H] 334.2 (C17H19NO4S+H, calc: 333.4). Compound LL was used without further purification. Preparation 38 Synthesis of benzyl ester of (S)-4-[4-(N-acetylhydroxycarbamimidoyl)-benzylcarbamoyl]-4-((R)-4-phenyl-2-phenylmethanesulfonylaminobutyrylamino)-butyric acid (MM).

[00442] A solution of compound LL (5.9 g, 17.8 mmol), compound KK dihydrochloride (18.0 mmol), BOP (8.65 g, 19.6 mmol) and DIEA (10.96 mL, 19.6 mmol) in DMF (250 mL) was stirred at room temperature for 2 hours. The reaction mixture was diluted with EtOAc (750 mL) and extracted with water (200 mL). The precipitate formed was filtered, washed with EtOAc (200 mL) and water (200 mL) and dried at room temperature overnight (~18 hours), yielding compound MM (8.2 g, 11.0 mmol) as a whitish solid. LC-MS [M+H] 743.6 (C39H43N5O8S +H, calc: 743.9). Compound MM was used without further purification. Synthesis of (S)-5-(4-carbamimidoylbenzylamino)-5-oxo-4-((R)-4-phenyl-2-(phenylmethylsulfonamido)butanamido)pentanoic acid (Compound 108)

[00443] Compound MM (8.0 g, 10.77 mmol) was dissolved in acetic acid (700 mL), followed by the addition of Pd/C (5% by weight, 3.0 g) as a suspension in water (50 mL). The reaction mixture was hydrogenated (Parr apparatus, 34.47 kPa (5 psi)) at room temperature for 3 hours. The catalyst was filtered through a Celite pad onto a sintered glass filter and washed with methanol. The filtrate was evaporated in vacuo, yielding compound 108 as a colorless oil. LC-MS [M+H] 594.2 (C30H35N5O6S +H, calc: 594). The oil obtained was dissolved in water (150 mL) and purified by HPLC. [Nanosyn-Pack YMC-ODS-A (100-10) C-18 column (75 x 300 mm); flow rate = 250 mL/minute; injection volume 150 mL; mobile phase A: 100% water, 0.1% TFA; mobile phase B: 100% acetonitrile, 0.1% TFA; isocratic elution at 10% B in 4 minutes, gradient elution to 24% B in 18 minutes, isocratic elution at 24% B in 20 minutes, gradient elution from 24% B to 58% B in 68 minutes; detection at 254 nm]. The fractions containing the desired compound were combined and concentrated in vacuo. The residue was dissolved in i-PrOH (75 mL) and evaporated in vacuo (the procedure was repeated twice), yielding Compound 108 (4.5 g, 70% yield, 98.0% purity) as a white solid. LC-MS [M+H] 594.2 (C30H35N5O6S +H, calc: 594). Retention time*: 3.55 minutes.

[00444] * - [Chromolith SpeedRod RP-18e C18 column (4.6 x 50 mm); flow rate 1.5 mL/minute; mobile phase A: 0.1% TFA/water; mobile phase B: 0.1% TFA/acetonitrile; gradient elution from 5% B to 100% B in 9.6 minutes, detection at 254 nm].

[00445] Notwithstanding the fact that the present invention has been described with reference to specific embodiments thereof, it should be understood by those skilled in the art that various alterations may be made and equivalents may be substituted without departing from the true spirit and scope of the invention. Additionally, many modifications may be made to adapt a particular situation, material, material composition, process, step or process steps to the objective, spirit and scope of the present invention. It is intended that all such modifications fall within the scope of the appended claims.

Claims (12)

1. Pharmaceutical composition, characterized in that it comprises a trypsin inhibitor and a compound of general formula (II) which has a general formula (V): or a pharmaceutically acceptable salt thereof, wherein: Ra is hydrogen or hydroxyl; Rb is oxo (=O) or hydroxyl; the dashed line is a double bond or single bond; R1 represents a (C1-4)alkyl group; each R2 represents a hydrogen or a (C1-4)alkyl group; each R3 represents a hydrogen or a (C1-4)alkyl group; n represents an integer from 2 to 4; R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and R5 represents a hydrogen atom, an N-acyl group, an N-substituted acyl group, an amino acid residue, a dipeptide, an N-acyl derivative of an amino acid or dipeptide, or an N-substituted acyl derivative of an amino acid or dipeptide, wherein: the N-acyl group is selected from acetyl, benzoyl, malonyl, piperonyl, succinyl; the amino acid is selected from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine; and the dipeptide is a combination of two amino acids selected from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine, wherein the trypsin inhibitor is: a) a compound with the formula: where: Q1 is selected from -O-Q4 or -Q4-COOH, in which Q4 is C1C4 alkyl; Q2 is N or CH; and Q3 is aryl or substituted aryl; b) a compound of formula: where: Q5 is -C(O)-COOH or -NH-Q6-Q7-SO2-C6H5, in which Q6 is -(CH2)p-COOH; Q7 is -(CH2)r-C6H5; ep is an integer from one to three, er is an integer from one to three; or c) a compound selected from the group consisting of: 4-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperazino-1-carboxylate of (S)-ethyl, 4-(5-guanidino-2-(2,4,6-triisopropylphenylsulfonamido)pentanoyl)piperazino-1-carboxylate of (S)-ethyl, 1-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperidino-4-carboxylate of (S)-ethyl, 1-(5-guanidino-2-(2,4,6-triisopropylphenylsulfonamido)pentanoyl)piperidino-4-carboxylate of (S)-ethyl, acid (S)-6-(4-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperazin-1-yl)-6-oxohexanoic acid,3-(4-carbamimidoylphenyl)-2-oxopropanoic acid, 6-carbamimidoylnaphthalen-2-yl (S)-5-(4-carbamimidoylbenzyl-amino)-5-oxo-4-((R)-4-phenyl-2-(phenylmethylsulfonamido)butanamido)pentanoic acid,4-(diaminomethylene-amino)benzoate; e4,4'-(pentane-1,5-diylbis(oxy))dibenzimidamide. 2. Pharmaceutical composition, characterized in that it comprises a trypsin inhibitor and a compound of general formula (III): XC(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5)(III) or a pharmaceutically acceptable salt thereof, wherein: X represents a residue of a phenolic opioid, in which the hydrogen atom of the phenolic hydroxyl group is covalently substituted for -C(O)-NR1-(C(R2)(R3))n-NH-C(O)-CH(R4)-NH(R5); R1 represents a (1-4C)alkyl group; R2 and R3 each independently represent a hydrogen atom or a (1-4C)alkyl group; n represents 2 or 3; R4 represents -CH2CH2CH2NH(C=NH)NH2 or -CH2CH2CH2CH2NH2, the configuration of the carbon atom to which R4 is attached corresponds to that of an L-amino acid, and R5 represents a hydrogen atom, an N-acyl group, an N-substituted acyl group, an amino acid residue, a dipeptide, an N-acyl derivative of an amino acid or dipeptide, or an N-substituted acyl derivative of an amino acid or dipeptide, wherein: the N-acyl group is selected from acetyl, benzoyl, malonyl, piperonyl, succinyl; the amino acid is selected from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine and valine; The dipeptide is a combination of two amino acids selected from alanine, arginine, asparagine, aspartic acid, cysteine, glutamic acid, glutamine, glycine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, proline, serine, threonine, tryptophan, tyrosine, and valine, wherein the trypsin inhibitor is: a) a compound with the formula: where: Q1 is selected from -O-Q4 or -Q4-COOH, in which Q4 is C1C4 alkyl; Q2 is N or CH; and Q3 is aryl or substituted aryl; b) a compound of formula: where: Q5 is -C(O)-COOH or -NH-Q6-Q7-SO2-C6H5, in which Q6 is -(CH2)p-COOH; Q7 is -(CH2)r-C6H5; ep is an integer from one to three, er is an integer from one to three; or c) a compound selected from the group consisting of: 4-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperazino-1-carboxylate of (S)-ethyl, 4-(5-guanidino-2-(2,4,6-triisopropylphenylsulfonamido)pentanoyl)piperazino-1-carboxylate of (S)-ethyl, 1-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperidino-4-carboxylate of (S)-ethyl, 1-(5-guanidino-2-(2,4,6-triisopropylphenylsulfonamido)pentanoyl)piperidino-4-carboxylate of (S)-ethyl, acid (S)-6-(4-(5-guanidino-2-(naphthalene-2-sulfonamido)pentanoyl)piperazin-1-yl)-6-oxohexanoic acid,3-(4-carbamimidoylphenyl)-2-oxopropanoic acid, 6-carbamimidoylnaphthalen-2-yl (S)-5-(4-carbamimidoylbenzyl-amino)-5-oxo-4-((R)-4-phenyl-2-(phenylmethylsulfonamido)butanamido)pentanoic acid,4-(diaminomethylene-amino)benzoate; e4,4'-(pentane-1,5-diylbis(oxy))dibenzimidamide. 3. Pharmaceutical composition according to claim 1 or 2, characterized in that n is 2. 4. Pharmaceutical composition, according to claim 1 or 2, characterized in that R1 is methyl or ethyl. 5. Pharmaceutical composition, according to claim 1 or 2, characterized in that R5 is acetyl, benzoyl, malonyl, piperonyl, succinyl, N-acetylarginine, or N-acetyl-lysine. 6. Pharmaceutical composition, according to claim 1 or 2, characterized in that R2 and R3 are hydrogens. 7. Pharmaceutical composition, according to claim 1 or 2, characterized in that R2 and R3, which are on the same carbon, are alkyl groups. 8. Pharmaceutical composition, according to claim 1 or 2, characterized in that the compound is the hydromorphone of 3-(N-methyl-N-(2-N'-acetylarginylamine)) ethylcarbamate, or a pharmaceutically acceptable salt thereof. 9. Pharmaceutical composition, according to any one of claims 1 to 8, characterized in that the trypsin inhibitor is 4-(diaminomethylene-amino)benzoate of 6-carbamimidoylnaphthalen-2-yl 10. Pharmaceutical composition, according to any one of claims 1 to 9, characterized in that it is for the treatment of the human or animal body. 11. Pharmaceutical composition, according to any one of claims 1 to 9, characterized in that it is for use in the treatment or prevention of pain. 12. Use of a pharmaceutical composition, as defined in any one of claims 1 to 9, characterized in that it is for the preparation of a medicament for the treatment or prevention of pain.