Compounds and methods for amine oxidase imaging - Patents.com

Abstract

The present invention relates to functionalizable, enzyme-activatable fluorescent probes and methods for monitoring the activity of amine oxidases, which catalyze the oxidative deamination of, for example, the ε-amine of lysine to an aldehyde, which can form a covalent bond with an adjacent side chain, for example, in the context of collagen crosslinking. Amine oxidase activity may be correlated with collagen-related diseases such as pulmonary and hepatic fibrosis, cardiomyopathy, and tumor metastasis.

Description

[Technical field]

[0001] The present invention relates to functionalizable, enzyme-activatable fluorescent probes and methods for monitoring the activity of amine oxidases, which can, for example, catalyze the oxidative deamination of the ε-amine of lysine to an aldehyde, which can then form a covalent bond with an adjacent side chain in the context of collagen crosslinks. Amine oxidase activity may be correlated with collagen-related diseases, including pulmonary and hepatic fibrosis, cardiomyopathy, and tumor metastasis. [Background technology]

[0002] Amine oxidases are enzymes that catalyze the oxidative cleavage of alkylamines to aldehydes and ammonia, e.g., the oxidative deamination of the ε-amine of lysine to an aldehyde. There are two subfamilies of amine oxidases that are characterized by the cofactors they contain. Representative amine oxidases that use copper as a cofactor are lysyl oxidase (LOX), primary amine oxidases (e.g., AOC2, AOC3), and diamine oxidase (AOC1). Representative monoamine oxidases that use flavin as a cofactor are monoamine oxidases A and B (MAOA and MAOB).

[0003] Collagen is an essential protein that provides mechanical strength to skin and other tissues. Collagen cross-linking generates many of the macroscopic structural properties of collagen, such as tensile strength and resistance to proteolysis. This cross-linking occurs post-translationally by amine oxidases, such as LOX, which form covalent bonds between adjacent amino acid chains of collagen. Cross-linking can occur as a normal mechanism, for example, during tissue maturation, growth, and development, and in wound healing. However, increased amine oxidase activity, such as LOX activity, is correlated with many diseases and disorders, such as fibrosis, cardiomyopathy, and cancer.

[0004] Existing probes for amine oxidases, e.g., LOX, do not allow the synthetic functionalization required to control the localization, spatial accumulation, and site of action of the probe, e.g., in cells or tissues and in diagnostic contexts. Moreover, existing fluorescent probes for other enzymes both have low quantum yields.

[0005] Aslam et al. (Chemical Science 2015, 6, 4946-4953) disclose a fluorescent probe for visualizing LOX activity. However, this probe shows insufficient increase in fluorescence upon unmasking and cannot be functionalized for diagnostic purposes. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] U.S. Patent Application Serial No. 09 / 902,822 [Patent Document 2] PCT / US01 / 21860 [Patent Document 3] U.S. Provisional Patent Application No. 60 / 313,527 [Patent Document 4] U.S. Provisional Patent Application No. 60 / 339,249 [Non-patent literature]

[0007] [Non-Patent Document 1] Aslam et al., (Chemical Science 2015, 6, 4946-4953) [Non-Patent Document 2] AH Palamakumbura, PC Trackman, Analytical Biochemistry 2002, 300, pp. 245-251 [Non-Patent Document 3] H. C. Ansel and N. G. Popovish, Pharmaceutical Dosage Forms and Drug Delivery Systems, 5th Edition, Lea and Febiger (1990)

Non-Patent Document 4

Non-Patent Document 5

Non-Patent Document 6

Non-Patent Document 7

Non-Patent Document 8

Non-Patent Document 9

Non-Patent Document 10

Non-Patent Document 11

Summary of the Invention

Problems to be Solved by the Invention

[0008] The technical problem underlying the present invention is the provision of new compounds and methods for detecting amine oxidase activity. [Means for solving the problem]

[0009] In a first aspect, the present invention provides a compound of formula I

[0010] [ka]

[0011] (In the formula, a and b are independently an integer from 0 to 10, optionally 1 or 2; A is,

[0012] [ka]

[0013] is a structure selected from the group consisting of: R 3 and R 4 teeth, --NHR 9' , -OR 9' , S.R. 9' ,

[0014] [ka]

[0015] , if a is 0 to 3, optional -SR 9' ; - Optionally, when a and / or b are 0, linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl, (C 2~10 ) alkenyl and (C 2~10 ) alkynyl, (C 3~10 ) carbocycles, optionally substituted or unsubstituted carbocycles selected from the group consisting of aromatic (C6) carbocycles; - substituted or unsubstituted triphenylphosphines connected via phosphorus; - N-maleimidyl; and - halogen, optionally F, Cl, Br, and I, optionally R 3 or R 4 is a proteinogenic amino acid, a non-proteinogenic amino acid, or a peptide, R 3 or R 4 the other is halogen, optionally F, Cl, Br or I independently selected from the group consisting of: L is absent or a linker, optionally - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally propyl, butyl and pentyl, (C 2~10 ) alkenyl and (C 2~10 ) alkynyl, - containing one or more amide functions in the alkyl chain (C 1~10 ) alkyl, optionally

[0016] [ka]

[0017] where c and d are independently selected from 1, 2, 3, 4, and 5; and - Linear or branched, substituted or unsubstituted (C 1~20 ) alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; (C 2~10 ) alkenyl ether, (C 2~10 ) alkynyl ethers and (C 4~10 ) carbocyclic ethers, is a linker selected from the group consisting of: X, Y, and Z are independently selected from the group consisting of O, N, and S, optionally Z is selected from the group consisting of O and N, optionally Z is O; R 9' teeth, - Hydrogen; - linear or branched, substituted or unsubstituted, optionally sulfonated, (C 1~10 ) alkyl, optionally methyl, ethyl and propyl, (C 2~10 ) alkenyl and (C 2~10 ) alkynyl, - Linear or branched, substituted or unsubstituted (C 1~20 ) alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; (C 2~10 ) alkenyl ether, (C 2~10 ) alkynyl ethers and (C 4~10 ) carbocyclic ethers; - (C 3~10 ) a substituted or unsubstituted carbocycle selected from the group consisting of a carbocycle, optionally aromatic (C6) carbocycle, optionally substituted with a substituent selected from the group consisting of Cl, F, Br, substituted or unsubstituted methyl, optionally -(CF3), ethyl, propyl and cyclopropyl; - substituted or unsubstituted triphenylphosphines connected via phosphorus; - Substituted or unsubstituted (C 3~6 ) Heterocycles and (C7-C 10 ) Carbocyclic or bicyclic rings each having 1 to 3 heteroatoms independently selected from N, O, and S (C 10 ) heterobicycles, optionally substituted or unsubstituted (C7)heterobicycles having two heteroatoms selected from N and S; - proteinogenic amino acids, non-proteinogenic amino acids, optionally aminooxy or hydrazide derivatives of proteinogenic or non-proteinogenic amino acids, optionally lysine, proline, glycine, 4-hydroxyproline, 4-aminoproline, and 4-aminooxyproline; - a peptide optionally comprising 1-2000, optionally 1-100, optionally 1-25 amino acids, optionally comprising 1-10, optionally 7 [proline]-[4-hydroxyproline]-[glycine] units; - collagen peptides, fibronectin peptides, fibrillin peptides, elastin peptides and RGD (arginylglycylaspartic acid) peptides; and - antibodies, optionally anti-collagen, anti-elastin, anti-fibronectin and anti-fibrillin antibodies, selected from the group consisting of; R 9 is R 9' is selected from the residues defined for - further selected from the group consisting of azido, N-maleimidyl, -NH2, -OH and -SH2; R 9'' teeth, - halogens, optionally F, Cl, Br, and I, substituted or unsubstituted triphenylphosphines connected via phosphorus, azide, -SR 10 ,

[0018] [ka]

[0019] selected from the group consisting of; R 10 teeth, - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl, (C 2~10 ) alkenyl and (C 2~10 ) alkynyl; and - Linear or branched, substituted or unsubstituted (C 1~20 ) alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; (C 2~10 ) alkenyl ether, (C 2~10 ) alkynyl ethers and (C 4~10 ) carbocyclic ethers, selected from the group consisting of; R 5 teeth, - hydrogen and -OH; and halogens, optionally F, Cl, Br and I; selected from the group consisting of; R 6 and R 8 is independently selected from the group consisting of hydrogen, -OH, halogen, optionally F, Cl, Br, and I; and / or R 7 teeth, - Hydrogen; - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl, (C 2~10 ) or (C 3~10 ) alkenyl and (C 2~10 ) or (C 3~10 ) alkynyl; - Linear or branched, substituted or unsubstituted (C 1~20 ) alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; (C 2~10 ) alkenyl ether, (C 2~10 ) alkynyl ethers and (C 4~10 ) carbocyclic ethers; - -N2 which forms an azide with the nitrogen atom of A; and - Boc, Fmoc, Cbz, Ac, TFA, phthalimide, Bn, Tr, benzylidene, and Ts (each independently selected from the group consisting of and salts and solvates thereof, optionally physiologically acceptable salts and solvates.

[0020] Without wishing to be bound by theory, the cleavable masking group AZ of the compounds of the invention masks the fluorescence of the fluorescent probe until cleavage by an amine oxidase (see scheme below for a representative example, LOX = lysyl oxidase). Furthermore, upon cleavage, a shift in the emission wavelength of the compound allows one to distinguish between the intact probe and the probe deaminated by the amine oxidase.

[0021] [ka]

[0022] For example, the enzyme-cleavable masking group AZ of the compounds described herein drastically reduces the quantum yield, e.g., by more than 10-fold, while the feasibility of functionalization provides a platform for introducing fluorescent probes into collagen model peptides, e.g., for in vivo imaging of collagen-related processes (e.g., wound healing, see Example 16 and Figures 31 and 40-44). Furthermore, the compounds can also be attached to specific targeting moieties, such as, for example, antibodies, that direct the compound to a desired location.

[0023] The compounds described herein are characterized by a significant shift in excitation wavelength. For example, upon masking, the wavelength shifts from the maximum wavelength to shorter wavelengths, and thus the fluorescent properties of the compounds are effectively masked, for example, under the usual excitation wavelengths (e.g., 400-405 nm) used in fluorescence microscopy or other detection means. Upon unmasking, for example, by an amine oxidase enzyme, the excitation wavelength essentially shifts to the excitation wavelength of the fluorophore core of the compound, thereby allowing the determination of enzyme activity and localization (see, for example, Figures 1 and 2). Compared to state-of-the-art probes (e.g., the oLOX probes of Aslam et al., Chemical Science 2015 6, 4946-4953), the compounds of the present invention unexpectedly show a significantly higher (e.g., 4-fold higher) increase in fluorescence upon unmasking by an amine oxidase without showing any background fluorescence before unmasking (see, for example, Figure 13).

[0024] Furthermore, functionalization of the compounds can be easily performed due to the presence of functional groups, e.g., esters, that can react with any molecule that exhibits nucleophilicity, e.g., alcohols or amines. Alternatively, the nucleophilic residues of the compounds described herein can be reacted with electrophilic groups of targets, e.g., amino acid sequences or proteins. The compounds according to the invention specifically detect the amine oxidase activity of different amine oxidases in vitro (see, e.g., Figures 11, 12, 13 and 34), ex vivo (see, e.g., Figures 3-5, 8-10, 13 and 35-39), and in vivo (see, e.g., Figures 31 and 40-44).

[0025] The compounds of the invention may contain residues, e.g., functional groups, that are protected by protecting groups used, e.g., in organic or peptide chemistry. The compounds described herein can be installed into peptides, e.g., synthetic peptides, by applying common peptide synthesis strategies, such as, but not limited to, Fmoc / t-Bu peptide synthesis protocols, which are compatible with all stages of peptide synthesis.

[0026] In one embodiment, the compound of the invention is A,

[0027] [ka]

[0028] The compound is a compound having a structure selected from the group consisting of:

[0029] In one embodiment, the compound of the invention is R 3 but,

[0030] [ka]

[0031] It is a compound wherein

[0032] In one embodiment, the compound of the invention is R 3 but,

[0033] [ka] It is a compound wherein

[0034] In one embodiment, the compound of the invention is a and b are independently an integer from 0 to 3, optionally 1 or 2; A is,

[0035] [ka]

[0036] is a structure selected from the group consisting of: R 3 and R 4 teeth, --NHR 9' , -OR 9' , S.R. 9' ,

[0037] [ka]

[0038] , if a is 0 to 3, optional -SR 9' ; - Optionally, when a and / or b are 0, linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl; - triphenylphosphine connected through phosphorus; - N-maleimidyl; and - halogen, optionally F, Cl, Br, and I, optionally R 3 or R 4 is a proteinogenic amino acid, a non-proteinogenic amino acid, or a peptide, R 3 or R 4the other is halogen, optionally F, Cl, Br or I independently selected from the group consisting of: L is absent or a linker, optionally - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally propyl, butyl and pentyl;

[0039] [ka]

[0040] where c and d are independently selected from 1, 2, 3, 4, and 5; and - Linear or branched, substituted or unsubstituted (C 1~20 ) an alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; is a linker selected from the group consisting of: X, Y, and Z are independently selected from the group consisting of O, N, and S, optionally Z is selected from the group consisting of O and N, optionally Z is O; R 9' teeth, - Hydrogen; - linear or branched, substituted or unsubstituted, optionally sulfonated, (C 1~10 ) alkyl, optionally methyl, ethyl and propyl, (C 2~10 ) alkenyl and (C 2~10 ) alkynyl, - Linear or branched, substituted or unsubstituted (C 1~20 ) an alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; - triphenylphosphine connected through phosphorus; - proteinogenic amino acids, non-proteinogenic amino acids, optionally aminooxy or hydrazide derivatives of proteinogenic or non-proteinogenic amino acids, optionally lysine, proline, glycine, 4-hydroxyproline, 4-aminoproline, and 4-aminooxyproline; - a peptide optionally comprising 1-2000, optionally 1-100, optionally 1-25 amino acids, optionally comprising 1-10, optionally 7 [proline]-[4-hydroxyproline]-[glycine] units; - collagen peptides, fibronectin peptides, fibrillin peptides, elastin peptides and RGD (arginylglycylaspartic acid) peptides; and - antibodies, optionally anti-collagen, anti-elastin, anti-fibronectin and anti-fibrillin antibodies, selected from the group consisting of; R 9 is R 9' is selected from the residues defined for - further selected from the group consisting of azido, N-maleimidyl, -NH2, -OH and -SH2; R 9'' teeth, -Azide, -SR 10 ,

[0041] [ka]

[0042] selected from the group consisting of; R 10 teeth, - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl; and - Linear or branched, substituted or unsubstituted (C 1~20 ) an alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; selected from the group consisting of; R 5teeth, - hydrogen and -OH; and halogens, optionally F, Cl, Br and I; selected from the group consisting of; R 6 and R 8 are independently selected from the group consisting of hydrogen, halogen, optionally F, Cl, Br, and I; and / or R 7 teeth, - Hydrogen; - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl; - Linear or branched, substituted or unsubstituted (C 1~20 ) an alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; - -N2 which forms an azide with the nitrogen atom of A; and - Boc, Fmoc, Cbz, Ac, TFA, phthalimide, Bn, Tr, benzylidene, and Ts are compounds each independently selected from the group consisting of:

[0043] In a further embodiment, the compound of the invention is a is an integer from 0 to 3, optionally 1 or 2; b is 0; A is,

[0044] [ka]

[0045] is a structure selected from the group consisting of: R 3 teeth,

[0046] [ka]

[0047] and; R 4 is hydrogen, straight or branched chain, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl; L is absent or a linker, optionally - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally propyl, butyl and pentyl; -

[0048] [ka]

[0049] where c and d are independently selected from 1, 2, 3, 4, and 5; and - Linear or branched, substituted or unsubstituted (C 1~10 ) an alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; is a linker selected from the group consisting of: X is selected from the group consisting of O, N, and S; Z is O; R 9 teeth, - Hydrogen, N-maleimidyl; - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl, (C 2~10 ) alkenyl and (C 2~10 ) alkynyl; - Linear or branched, substituted or unsubstituted (C 1~20 ) an alkyl ether, optionally a polyethylene glycol (PEG) chain, optionally a PEG chain having 1 to 10 ethylene oxide moieties; - triphenylphosphine connected through phosphorus; - proteinogenic amino acids, non-proteinogenic amino acids, optionally lysine, proline, glycine, 4-hydroxyproline, 4-aminoproline, and 4-aminooxyproline; - a peptide optionally comprising 1-2000, optionally 1-100, optionally 1-25 amino acids, optionally comprising 1-10, optionally 7 [proline]-[4-hydroxyproline]-[glycine] units; - Collagen peptides, fibronectin peptides, fibrillin peptides, elastin peptides and RGD (arginylglycylaspartic acid) peptides; - antibodies, optionally anti-collagen, anti-elastin, anti-fibronectin and anti-fibrillin antibodies; and - azide, -NH2, -OH and -SH2, selected from the group consisting of; R 9'' teeth, -Azide, -SR 10 ,

[0050] [ka]

[0051] selected from the group consisting of; R 10 is linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl, (C 2~10 ) alkenyl and (C 2~10 ) alkynyl; R 5 teeth, - hydrogen and -OH; and halogens, optionally F, Cl, Br and I; selected from the group consisting of; R 6 and R 8 are independently selected from the group consisting of hydrogen, halogen, optionally F, Cl, Br, and I; and / or R 7 teeth, - Hydrogen; - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl; - -N2 which forms an azide with the nitrogen atom of A; and - Boc, Fmoc, Cbz, are compounds independently selected from the group consisting of:

[0052] In another embodiment, the compound of the invention is a compound according to Formula II.

[0053] [ka]

[0054] (In the formula, a is 1 or 2; b is 0; A is,

[0055] [ka]

[0056] is a structure selected from the group consisting of: R 3 teeth,

[0057] [ka]

[0058] and; R 4 is hydrogen, straight or branched chain, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl; L does not exist,

[0059] [ka]

[0060] (wherein c and d are independently selected from 1, 2, 3, 4, and 5); X is selected from the group consisting of O and N; Z is O; R 9 teeth, - Hydrogen, N-maleimidyl; - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl; - triphenylphosphine connected through phosphorus; - proteinogenic amino acids, non-proteinogenic amino acids, optionally lysine, proline, glycine, 4-hydroxyproline, 4-aminoproline, and 4-aminooxyproline; - collagen peptides, fibronectin peptides, fibrillin peptides, elastin peptides and RGD (arginylglycylaspartic acid) peptides; and - azide, -NH2, -OH and -SH2, selected from the group consisting of; R 9'' teeth, - Azide,

[0061] [ka]

[0062] selected from the group consisting of; and / or R 7 teeth, - hydrogen; and - Linear or branched, substituted or unsubstituted (C 1~10 ) alkyl, optionally methyl, ethyl and propyl; (each independently selected from the group consisting of

[0063] Where the compounds described herein exist in various tautomeric forms, the recited compounds are not limited to any one particular tautomer, but rather are intended to encompass all tautomers.

[0064] The recited compounds are further intended to include compounds in which one or more atoms have been replaced with an isotope, i.e., atoms having the same atomic number but different mass numbers. As general examples, isotopes of hydrogen include tritium and deuterium, isotopes of carbon include 11 C. 13 C, and 14 These include, but are not limited to, C.

[0065] As used herein, a "substituent" or "residue" or "R" refers to a molecular moiety that is covalently bonded to an atom in a molecule of interest. For example, a "substituent", "R" or "residue" can be a moiety such as a halogen, an alkyl group, a haloalkyl group, or any other substituent described herein, that is covalently bonded to an atom, optionally a carbon or nitrogen atom, that forms part of the molecule of interest.

[0066] The term "substituted" as used herein means that any one or more hydrogens on the specified atom or group are replaced with an atom different from hydrogen, optionally with a halogen, optionally with a fluorine, with a sulfonyl group, with a tertiary or quaternary amine, or with one selected from the indicated substituents, provided that the normal valence of the specified atom is not exceeded and the replacement results in a stable compound, i.e., a compound that can be isolated and characterized using conventional means. Optionally, as used herein, "substitution" means that any one or more hydrogens on the specified atom or group are replaced with a group that increases the solubility of the compounds described herein, for example, in water, aqueous buffers, or physiological fluids, such as serum, saliva, or blood. For example, the substitution can be in the form of oxygen bonded to any chemical atom other than carbon, for example, a hydroxyl group, or in the form of an oxygen anion. When the substituent is oxo, i.e., =O, two hydrogens on the atom are replaced. An oxo group that is a substituent of an aromatic carbon atom results in the conversion of -CH- to -C(=O)-, resulting in loss of aromaticity. For example, a pyridyl group substituted with oxo is a pyridone.

[0067] For all aspects and embodiments of the present invention, 3 , R 4 and / or R 9 can be selected, for example, from maleimides, resulting in the following exemplary structures:

[0068] [ka]

[0069] where all residues R, Z, A and integers a, b and c are as defined herein; R 3 The residues shown for 4 is selected.

[0070] The linker L as used herein in all aspects and embodiments can be, for example, a substituted or unsubstituted (C 1~10 ) alkyl, i.e., at least two carbon atoms are linked to each other via an amide bond, such as:

[0071] [ka]

[0072] (wherein e is selected from 1, 2, or 3, and c and d are independently selected from 1, 2, 3, 4, and 5). The linker L can be attached to the structure of formula I or II either at the amine side of the amide or at the carbonyl side of the amide, optionally at the amine side, and can be attached to the structure of formula I or II either at the carbonyl side of the amine or at the amine side of the amide, optionally at the carbonyl side, by R 9 can be combined into, for example,

[0073] [ka]

[0074] where X is as defined herein or optionally N, or

[0075] [ka]

[0076] It is.

[0077] R 3 , R 4 , R 9 , R 9' and R 9'' The triphenylphosphine moiety is attached to the phosphorus atom as follows:

[0078] [ka]

[0079] and, for example, a halogen anion, optionally Cl - , Br - , I - or trifluoroacetate. Triphenylphosphine may also be substituted or unsubstituted, i.e., the phenyl ring of the triphenylphosphine moiety may be substituted or unsubstituted, and may optionally be substituted with a halogen. Exemplary structures of triphenylphosphine-containing compounds according to the present invention are shown below:

[0080] [ka]

[0081] where X is as defined herein or optionally N, or for example

[0082] [ka]

[0083] It is.

[0084] The term "proteinogenic amino acid" as used herein refers to any amino acid that is biosynthetically incorporated into a protein during translation, optionally lysine, proline, and glycine. The term "non-proteinogenic amino acid" as used herein refers to any amino acid that is not naturally encoded in the genetic code of any organism, e.g., D-amino acids, non-alpha amino acids, amino acids lacking a hydrogen on the alpha carbon. Optionally, for example, for R9' in all aspects of the invention, proline, lysine, glycine or hydroxy, amino, aminooxy or hydrazide derivatives of proteinogenic or non-proteinogenic amino acids are included for use in the present invention. Optionally, the derivative is hydroxyproline, 4-hydroxyproline, cis-4-hydroxyproline, trans-4-hydroxyproline, 3-hydroxyproline, cis-3-hydroxyproline, trans-4-hydroxyproline, aminoproline, 4-aminoproline, cis-4-aminoproline, trans-4-aminoproline, 3-aminoproline, cis-3-aminoproline, trans-4-aminoproline, aminoxyproline, 4-aminooxyproline, cis-4-aminooxyproline, trans-4-aminooxyproline, 3-aminooxyproline, cis-3-aminooxyproline, trans-4-aminooxyproline, cis- or trans-4-hydrazinecarbonylproline derivatives, optionally

[0085] [ka]

[0086] and a cis- or trans-4-hydrazine carboxylate proline derivative, optionally

[0087] [ka]

[0088] , may be selected from the group consisting of:

[0089] Optionally, for example for R9', in all aspects of the invention, one or more proline-hydroxyproline-glycine or 4-proline-hydroxyproline-glycine units are preferred to mimic the structure of collagen.

[0090] The term "heteroatom" as used herein should be understood to mean atoms other than carbon and hydrogen, such as, for example, O, N, S and P.

[0091] In the context of the present invention, it is understood that antecedent terms such as "straight or branched chain", "substituted or unsubstituted" and the like indicate that each of the following terms is to be construed as modified by said preceding term. For example, the scope of the term "straight or branched chain, substituted or unsubstituted alkyl, alkenyl, alkynyl, carbocycle" includes straight or branched chain, substituted or unsubstituted alkyl; straight or branched chain, substituted or unsubstituted alkenyl; straight or branched chain, substituted or unsubstituted alkynyl; straight or branched chain, substituted or unsubstituted alkylidene; and straight or branched chain, substituted or unsubstituted carbocycle. For example, "(C 2~10 The term "alkenyl, alkynyl, or alkylidene" refers to a group of compounds containing from 2 to 10 carbons and an alkenyl, alkynyl, or alkylidene functional group.

[0092] The term "alkyl" refers to a saturated straight or branched chain hydrocarbon group containing the indicated number of carbon items, e.g., "(C 1~10 )Alkyl" means a hydrocarbon residue containing 1 to 10 carbon atoms, for example, methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, n-hexyl, 2,2-dimethylbutyl.

[0093] The term "alkenyl" refers to an at least partially unsaturated, substituted or unsubstituted, straight or branched chain hydrocarbon group containing the indicated number of carbon atoms, e.g., "(C2~10 By "alkenyl" is meant a hydrocarbon residue containing 2 to 10 carbon atoms, such as an ethenyl (vinyl), propenyl (allyl), isopropenyl, butenyl, isoprenyl or hex-2-enyl group, or a hydrocarbon group containing a methylene chain interrupted by one double bond, such as found in monounsaturated fatty acids, or a hydrocarbon group containing a methylene-interrupted polyene, such as found in polyunsaturated fatty acids, containing two or more of the following structural units -[CH=CH-CH2]-. An alkyl group may have one or more, such as 1, 2, 3, 4, 5 or 6 double bonds.

[0094] The term "alkynyl" refers to an at least partially unsaturated, substituted or unsubstituted straight or branched chain hydrocarbon group containing the indicated number of carbon atoms, e.g., "(C 2~10 "Alkynyl" means a hydrocarbon residue containing 2 to 10 carbon atoms, for example, an ethynyl, propynyl, butynyl, acetylenyl, or propargyl group. Optionally, the alkynyl group has one or two (for example, one) triple bonds.

[0095] The terms "alkyl", "alkenyl", and "alkynyl" also refer to groups in which one or more hydrogen atoms have been replaced, optionally by a halogen atom, such as F, Cl, or Br, such as 2,2,2-trichloroethyl, tribromoethyl, or trifluoromethyl groups.

[0096] The term "carbocycle" refers to a ring having the indicated number of carbon elements, e.g., "(C 3-10 ) carbocycle" or substituted or unsubstituted aliphatic hydrocarbon rings containing 3 to 10, optionally 3 to 6, carbon atoms, optionally 5 or 6 carbon atoms. These carbocycles can be either aromatic or non-aromatic systems. Non-aromatic ring systems can be mono- or polyunsaturated.

[0097] The term "carbobicycle" refers to a carbocycle as defined above containing multiple rings, optionally two rings. Exemplary carbocycles and carbobicycles include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclopentenyl, cyclohexyl, cyclohexenyl, cycloheptanyl, cycloheptenyl, phenyl, indanyl, indenyl, benzocyclobutanyl, dihydronaphthyl, tetrahydronaphthyl, naphthyl, decahydronaphthyl, benzocycloheptanyl, benzocycloheptenyl, spiro[4,5]decanyl, norbornyl, decalinyl, bicyclo[4.3.0]nonyl, tetralin, or cyclopentylcyclohexyl. Carbocyclic and / or carbobicyclic residues can be attached to the rest of the complete molecule by any atom of the ring that results in a stable structure.

[0098] The term "carbocycle" should also be understood to include "cycloalkyl," which optionally means an aliphatic hydrocarbon-containing ring having from 3 to 12 carbon atoms. These non-aromatic ring systems can be mono- or polyunsaturated, i.e., the term encompasses cycloalkenyl and cycloalkynyl.

[0099] The term "heterocycle" refers to a stable substituted or unsubstituted, aromatic or non-aromatic, optionally 3-10 membered, optionally 3-6 membered, optionally 5 or 6 membered, monocyclic, heteroatom-containing ring. Each heterocycle consists of carbon atoms and optionally one or more, optionally 1-4, optionally 1-3 heteroatoms selected from nitrogen, oxygen, and sulfur. A heterocycle may include the number of carbon atoms in addition to the non-carbon atoms, as indicated: "(C 3~6 ) Heterocycle" means having 3 to 6 carbon atoms in addition to the given number of heteroatoms.

[0100] The term "heterobicycle" refers to a heterocycle as defined above containing more than one ring, optionally two rings.

[0101] The heterocyclic and / or heterobicyclic residues can be attached to the remainder of the intact molecule by any atom of the ring that results in a stable structure. Exemplary heterobicycles and heterobicycles include pyrrolidinyl, pyrrolinyl, morpholinyl, thiomorpholinyl, thiomorpholinyl sulfoxide, thiomorpholinyl sulfone, dioxalanyl, piperidinyl, piperazinyl, tetrahydrofuran, 1-oxo-λ4-thiomorpholinyl, 13-oxa-11-aza-tricyclo[7.3.1.0-2,7]-tridecy-2,4,6-triene, tetrahydropyranyl, 2-oxo-2H-pyranyl, tetrahydrofuran, 1,3-dioxolanone, 1,3-dioxanone, 1,4-dioxanyl, 8-oxa-3 -aza-bicyclo[3.2.1]-octanyl, 2-oxa-5-aza-bicyclo[2.2.1]heptanyl, 2-thia-5-aza-bicyclo[2.2.1]heptanyl, piperidinonyl, tetrahydro-pyrimidonyl, pentamethylene sulfide, pentamethylene sulfoxide, pentamethylene sulfone, tetramethylene sulfide, tetramethylene sulfoxide and tetramethylene sulfone, indazolyl, benzimidazolyl, benzodioxolyl, imidazolyl, 1,3-benzodioxolyl and pyrazolyl.

[0102] The term "alkyl / alkenyl / alkynyl ether" refers to a saturated or unsaturated, straight or branched chain hydrocarbon group containing the indicated number of carbon atoms. For example, "(C 1~20) alkyl ether" refers to a hydrocarbon residue containing 1 to 20 carbon atoms and any suitable number of oxygen atoms resulting in an ether structure. An alkyl / alkenyl / alkynyl ether group as used herein should be understood to mean any linear or branched, substituted or unsubstituted alkyl / alkenyl / alkynyl chain containing an oxygen atom as an ether motif, i.e., either an oxygen atom bonded by two carbons. An exemplary alkyl ether is a polyethylene glycol (PEG) chain. The term polyethylene glycol as used herein refers to a chain of substituted or unsubstituted ethylene oxide monomers. For example, a PEG chain may contain 1 to 10 ethylene oxide monomers. The ether residue can be attached to the formula provided in the present invention through either the carbon atom or the oxygen atom of the ether residue.

[0103] As used herein, "substituent" or "residue" or "R" is optionally R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 9' , R 9'' , and / or R 10 can be attached to the formula provided herein directly or, optionally, through a linker, which may also be in the form of PEG.

[0104] When multiple residues R are attached to a given atom of the formulas described herein, the other residues R may not be present if the attachment of one R results in the complete valence of the atom. For example, R 7 can be -N2. This allows R 7 In this case, an azide is formed at the nitrogen to which one R is attached, as shown below. 7 Only R is bonded to the nitrogen. 7 does not exist.

[0105] [ka]

[0106] As used herein, the terms "nitrogen" or "N" and "sulfur" or "S" include any oxidized form of nitrogen and sulfur, and the quaternized form of any basic nitrogen, so long as the resulting compound is chemically stable. For example, -SC 1~6 For alkyl groups, -S(O)-C 1~6 Alkyl and -S(O)2-C 1~6 It should be understood to include alkyl.

[0107] As used herein, phrases defining the limits of a length range, e.g., "from 1 to 5" or "(C 1~5 ) means any integer from 1 to 5, e.g., 1, 2, 3, 4, and 5. In other words, any range defined by two integers explicitly recited is meant to include and disclose any integers defining said limits, as well as any integers subsumed within said range.

[0108] The scope of the present invention includes analogues of the above and claimed compounds characterized by the replacement of one or more carbon-bonded hydrogens, optionally one or more aromatic carbon-bonded hydrogens, with a halogen atom such as F, Cl or Br, optionally F. For example, one or more carbon-bonded hydrogens can be replaced by fluorine for reasons of metabolic stability and / or pharmacokinetic and physicochemical properties.

[0109] In another aspect, the invention relates to a composition, optionally a pharmaceutical composition, comprising as an active substance a compound for use as described herein or a pharma- ceutically acceptable derivative thereof, optionally in combination with an excipient and / or carrier.

[0110] The present invention includes pharmaceutically acceptable salts or solvates of the compounds of formula (I) and (II) of the present invention. "Pharmaceutically acceptable salts or solvates" refers to any pharmaceutically acceptable salt, solvate or ester, or any other compound, or a pharmacologically active metabolite or pharmacologically active residue thereof, that can provide (directly or indirectly) a compound of the present invention upon administration to a patient. A pharmacologically active metabolite should be understood to mean any compound of the present invention that can be metabolized enzymatically or chemically. This includes, for example, hydroxylated or oxidized derivative compounds of the present invention.

[0111] Pharmaceutically acceptable salts include those derived from pharma- ceutically acceptable inorganic and organic acids and bases. Examples of suitable acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfuric acid, tartaric acid, acetic acid, citric acid, methanesulfonic acid, formic acid, benzoic acid, malonic acid, naphthalene-2-sulfuric acid, and benzenesulfonic acid. Other acids, such as oxalic acid, are not themselves pharma- ceutically acceptable, but may be used in the preparation of salts useful as intermediates in obtaining compounds and their pharma- ceutically acceptable acid addition salts. Salts derived from appropriate bases include alkali metal salts (e.g., sodium salts), alkaline earth metal salts (e.g., magnesium salts), ammonium salts, and N-(C1-C4 alkyl)4 salts. + Salt is one example.

[0112] The terms "compound (according to the invention)" and "probe" are used interchangeably herein.

[0113] The terms "polypeptide" and "peptide" are used interchangeably and, as used herein, are meant to include peptides, polypeptides, oligopeptides, and proteins comprising two or more amino acids covalently linked via peptide bonds. The term does not refer to a specific length of the product. Polypeptides include post-translational modifications of the polypeptide, such as glycosylation, acetylation, phosphorylation, disulfide bridges, truncations, etc. The terms peptide and polypeptide also encompass polypeptide analogs, (poly)peptides containing unnatural amino acids, peptidomimetics, β-amino acids, etc.

[0114] The terms "peptide" and "polypeptide" as used herein also encompass isolated and purified glycosylated or non-glycosylated polypeptides.

[0115] R of all aspects and embodiments 9 Exemplary peptides include, for example, [ChaR]3 (see Example 18), -[ProHypGly]3-AopProGly-[ProHypGly]3, -[ProHypGly]7 or -[ProProGly]7 (see Examples 11-13), with or without a linker as defined herein.

[0116] In another aspect, the present invention relates to a compound as described herein, or a pharma- ceutically acceptable salt thereof, for use in the diagnosis of an amine oxidase-related disorder and / or a collagen- or elastin-related disorder.

[0117] The term "amine oxidase" as used herein is meant for all embodiments to describe any enzyme, i.e. a polypeptide having the catalytic activity of at least catalyzing the oxidative cleavage of alkylamines to aldehydes and ammonia, and optionally the oxidative deamination of the ε-amine of lysine to an aldehyde. Tests and assays for determining whether a given enzyme is an amine oxidase as defined herein are well known in the art (see, for example, AH Palamakumbura, PC Trackman, Analytical Biochemistry 2002, 300, 245-251). Alternatively or additionally, the test of Example 2 below can be used to determine whether a given enzyme is an amine oxidase as defined herein. Optionally, a change in fluorescence emission of at least 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75% or 80% in maximum emission of the fluorescent cleavage probe is indicative of amine oxidase activity as used herein. Optionally, with reference to the figures and examples (and corresponding conditions) provided herein, a change in fluorescence emission of at least 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000, 6500, 7000, 7500, 8000, 8500, 9000, 9500, 10000, 10500, 11000, 12000, 13000, 14000, 15000, 16000, 17000, 18000, 19000, or 20000 RFU in maximum emission of the fluorescent cleavage probe is indicative of amine oxidase activity as used herein. Optionally, any enzyme that cleaves 95, 90, 85, 80, 75, 70, 65, 60, 55, 50, 45, 40, 35, 30, 25, or 20% of a provided alkylamine to aldehyde and ammonia under known conditions or under conditions described in Example 2 is an amine oxidase as used herein.

[0118] Optionally, the amine oxidase described herein is optionally an animal or human amine oxidase, optionally a mammalian amine oxidase, optionally a human amine oxidase.

[0119] The term "catalyst" as used herein means that the amine oxidases described herein increase the rate of a reaction towards a desired product, i.e., towards the oxidative cleavage of an alkylamine to an aldehyde and ammonia, and optionally towards the deamination of the ε-amine of lysine to an aldehyde, to a greater extent compared to the rate of the reaction in the absence of the amine oxidase.

[0120] An "amine oxidase-related disorder" as used herein is any disorder that is directly or indirectly caused or exacerbated by amine oxidase. The disorder may be caused or exacerbated, for example, by defective expression of amine oxidase, underexpression of amine oxidase, overexpression of amine oxidase, degradation or accumulation of amine oxidase, increased or decreased amine oxidase activity, or other effects on amine oxidase that lead to unhealthy, e.g., non-standard or pathological, amine oxidase function. The amine oxidase-related disorder may be a disorder that is caused by, leads to, or is associated with a collagen or elastin-related disorder.

[0121] The term "collagen or elastin-related disorder" as used herein in all aspects is a disorder caused by or leading to defects in collagen or elastin structure, e.g., the extracellular matrix. Optionally, the collagen or elastin-related disorder described herein is based on or reflected by defects in collagen or elastin cross-linking, e.g., due to increased or decreased amine oxidase activity.

[0122] In another aspect, the present invention relates to the use of the compounds described herein in the in vitro or ex vivo diagnosis of amine oxidase related disorders and / or collagen or elastin related disorders.

[0123] The diagnosis of amine oxidase, collagen or elastin related diseases is based on the detection of the catalytic activity of amine oxidase by the compounds described herein.The fluorescent signal of the compounds can be used to detect, localize and / or quantify amine oxidase activity.Based on the detection, localization or quantification of amine oxidase activity, amine oxidase, collagen or elastin related diseases can be diagnosed or detected, and the progression of such diseases can be monitored.

[0124] The diagnosis of any aspect described herein or the diagnostic uses and methods described herein can be carried out in vitro, in vivo or ex vivo. In vivo applications include, for example, a mouthwash containing a compound according to the invention, where the mouthwash is suitable for diagnosing amine oxidase activity in the oral cavity of a subject by shining light on the oral cavity of the subject and evaluating the fluorescence emitted by the compound. Another diagnostic method may include using a compound according to the invention to treat tissue locally or to contact a tissue removal site of a patient, for example during surgery, to determine whether tissue with elevated amine oxidase activity is still present in the patient. In general, fluorescence can be detected by any method known in the art, including fluorescence microscopy or the naked eye, for example, to qualitatively determine the presence or absence of elevated amine oxidase activity.

[0125] Optionally, the compounds or pharmaceutical compositions described herein are for diagnostic use in animals or humans, optionally in mammals, optionally in humans.

[0126] The term "patient sample" as used in this context is meant to include any tissue, eg, biopsy, bodily fluids, eg, blood, serum, brain or cerebrospinal fluid.

[0127] For use in diagnosis, the compounds described herein can be administered in any conventional manner in any conventional dosage form.Routes of administration include, but are not limited to, oral administration, such as mouthwash, topical administration, such as liquid, cream, gel, spray or dispersion, dermal administration, inhalation administration, intravenous, intramuscular and subcutaneous injection.Exemplary modes of administration are topical, intravenous, intradermal or subcutaneous.

[0128] The compounds can be administered alone or in certain embodiments in combination with adjuvants, including other active ingredients, that enhance the stability of the compounds, facilitate administration of pharmaceutical compositions containing them, delay or enhance release of the compounds, provide increased dissolution or dispersion, increase activity, etc.

[0129] The compounds described herein can be physically combined with conventional therapeutic agents, diagnostic agents, or other adjuvants into a single pharmaceutical composition. In this regard, reference can be made to Cappola et al.: U.S. Patent Application No. 09 / 902,822, PCT / US01 / 21860, and U.S. Provisional Patent Application No. 60 / 313,527, each of which is incorporated herein by reference in its entirety. Advantageously, the compounds can then be administered together in a single dosage form. In some embodiments, pharmaceutical compositions containing such combinations of compounds contain at least about 5%, and optionally at least about 20% (w / w) of the compounds of the present invention. The optimal percentage (w / w) of the compounds of the present invention may vary and is within the skill of the art. Alternatively, the compounds can be administered separately (sequentially or in parallel). Separate administration allows for greater flexibility in dosing schedules.

[0130] As noted above, dosage forms of the compounds described herein in all aspects include pharma- ceutically acceptable carriers and adjuvants known to those skilled in the art. Methods for preparing such dosage forms are known (see, for example, HC Ansel and NG Popovish, Pharmaceutical Dosage Forms and Drug Delivery Systems, 5th ed., Lea and Febiger (1990)). Dosage levels and requirements are well recognized in the art and may be selected by the skilled artisan from available methods and techniques suitable for a particular patient. In some embodiments, dosage levels range from 1-500 mg / dose for a 70 kg patient. Although once a day may be sufficient, up to 5 times a day may be administered. For oral administration, up to 2500 mg / day may be required. For topical administration, for example, 1 cm 2 A dose of 10 to 10,000 μM per dose can be used. In this regard, reference can also be made to US Provisional Patent Application No. 60 / 339,249. As will be appreciated by those skilled in the art, lower or higher doses may be required depending on certain factors. For example, the specific dose and diagnostic regimen will vary depending on factors such as the patient's general health profile, the severity and course or nature of the patient's disorder, and the judgment of the treating physician.

[0131] In one embodiment, the compounds for use or uses described herein are (i) a lysyl oxidase (LOX), optionally having at least 80%, optionally at least 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 1-3, optionally over the entire sequence; (ii) lysyl oxidase homolog 1, 2, 3, or 4 (LOXL1, 2, 3, or 4); (iii) a primary amine oxidase, optionally AOC2 or AOC3; (iv) a diamine oxidase, optionally AOC1; (v) A compound for use or use, which is a disease associated with an amine oxidase selected from the group consisting of a monoamine oxidase, optionally MAOA or MAOB, optionally having at least 80%, optionally at least 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 4 to 7, optionally over the entire sequence.

[0132] The percentage identity of the amino acid sequences described herein can be determined with the help of known methods. In general, special computer programs are used that use algorithms adapted to the specific needs of this task. An exemplary method for determining amino acid identity begins with the generation of the maximum degree of identity among the sequences to be compared. Exemplary computer programs for determining identity between two amino acid sequences include, but are not limited to, TBLASTN, BLASTP, BLASTX, TBLASTX (Altschul et al., J. Mol. Biol., 215, 403-410, 1990), or ClustalW (Larkin MA et al., Bioinformatics, 23, 2947-2948, 2007). BLAST programs are available from the National Center for Biotechnology Information (NCBI) and other sources (BLAST handbook, Altschul et al., NCB NLM NIH Bethesda, MD 20894). The ClustalW program is available from http: / / www.clustal.org.

[0133] The amine oxidases described herein that can be monitored by the compounds of the invention encompass any functional derivative or functional fragment thereof. The term "functional derivative" of an enzyme or peptide as described in the context of the present invention is meant to include any enzyme or peptide or fragment thereof that has been chemically, post-translationally or genetically modified in its amino acid sequence, e.g. by addition, substitution and / or deletion of amino acid residues, and / or chemically modified in at least one of its atoms and / or functional chemical groups, e.g. by glycosylation, addition, deletion, rearrangement, oxidation, reduction, disulfide bridges, etc., so long as the derivative still has at least some amine oxidase activity, to a measurable extent, e.g. at least about 1-10%, optionally at least about 20-50%, of the amine oxidase activity of an unmodified peptide, e.g. an amine oxidase, LOX, LOXL, primary amine oxidase, diamine oxidase, monoamine oxidase, or an amine oxidase comprising any one of SEQ ID NOs: 1-7.

[0134] In this context, a "functional fragment" as used herein is one which forms part of a peptide or derivative referred to in the context of the present invention and which still has at least some amine oxidase activity, measurably, e.g. at least about 1-10%, optionally at least about 20-50% of the amine oxidase activity of the unmodified peptide. For example, the lysyl oxidase according to SEQ ID NO: 1 is cleaved intracellularly into a signal peptide (amino acids 1-21), a propeptide (amino acids 22-168), a long form of lysyl oxidase (amino acids 169-417; SEQ ID NO: 2) and a short form of lysyl oxidase (amino acids 219-417; SEQ ID NO: 3). Both the long form and the short form are functional fragments of the lysyl oxidase according to SEQ ID NO: 1.

[0135] Inappropriate expression of amine oxidases has been observed in many human diseases, many of which are associated with fibrotic reactions, especially cancer (Barker et al., 2012, Nature Reviews Cancer, 12, 540-552).

[0136] In one embodiment, the compound for use or use as described herein is one in which the disease is selected from the group consisting of fibrosis, optionally pulmonary and hepatic fibrosis, cardiomyopathy, Occupational Horn Syndrome (OHS), Menkes syndrome, myocardial ischemia, atherosclerosis, scleroderma, keloid disorders, liver cirrhosis, Alzheimer's and non-Alzheimer's dementia, Wilson's disease, primary biliary cirrhosis, chronic venous insufficiency, pseudoexfoliation syndrome, glaucoma, pelvic organ prolapse, endometriosis, intracranial aneurysms, heart failure, and cancer, including tumor metastasis.

[0137] In one embodiment, the compounds for use or uses described herein are those wherein the cancer is selected from the group consisting of colorectal cancer, bladder cancer, pancreatic cancer, breast cancer, head and neck squamous cell carcinoma (HNSCC), laryngeal cancer, lung cancer, gastric cancer, prostate cancer, esophageal squamous cell carcinoma, endometrial cancer, testicular seminoma cancer, hepatocellular carcinoma, renal clear cell carcinoma, and basal and squamous cell carcinoma.

[0138] In another aspect, the present invention relates to the use of a compound as described herein for detecting an amine oxidase activity, optionally for detecting the activity of an amine oxidase selected from the group consisting of: (i) a lysyl oxidase (LOX), optionally having at least 80%, optionally at least 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 1-3, optionally over the entire sequence; (ii) lysyl oxidase homolog 1, 2, 3, or 4 (LOXL1, 2, 3, or 4); (iii) a primary amine oxidase, optionally AOC2 or AOC3; (iv) a diamine oxidase, optionally AOC1; (v) a monoamine oxidase, optionally MAOA or MAOB, optionally having at least 80%, optionally at least 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 4-7, optionally over the entire sequence.

[0139] In another aspect, the present invention provides a method for producing a pharmaceutical composition comprising the steps of: (a) providing a compound described herein; (b) providing and contacting a cell, tissue or body fluid with the compound of step (a), optionally in vivo, in vitro or ex vivo, under conditions permitting amine oxidase activity; (c) measuring the fluorescence in the cells, tissues, or body fluids, optionally by fluorescence microscopy; and (d) determining the activity and / or location of the amine oxidase based on the fluorescence measured in step (c), optionally in vivo, in vitro or ex vivo.

[0140] In another aspect, the present invention relates to a method for the diagnosis of an amine oxidase related disorder and / or a collagen or elastin related disorder in a patient, optionally a mammal, or in a patient sample in need thereof, comprising the steps of: (a) providing a compound described herein; (b) contacting a patient or a patient sample in need thereof with an effective amount of the compound of step (a), wherein the effective amount is effective to detect amine oxidase activity; (c) measuring the fluorescence of the patient or a patient sample; and (d) determining the activity and / or location of the amine oxidase based on the fluorescence measured in step (c).

[0141] In one embodiment the method described herein is a method wherein the amine oxidase is selected from the group consisting of: (i) a lysyl oxidase (LOX), optionally having at least 80%, optionally at least 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 1-3, optionally over the entire sequence; (ii) lysyl oxidase homolog 1, 2, 3, or 4 (LOXL1, 2, 3, or 4); (iii) a primary amine oxidase, optionally AOC2 or AOC3; (iv) a diamine oxidase, optionally AOC1; (v) a monoamine oxidase, optionally MAOA or MAOB, optionally having at least 80%, optionally at least 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 4-7, optionally over the entire sequence.

[0142] In one embodiment, the method described herein is a method, wherein the disease is selected from the group consisting of fibrosis, optionally pulmonary and hepatic fibrosis, cardiomyopathy, Occupital Horn Syndrome (OHS), Menkes syndrome, myocardial ischemia, atherosclerosis, scleroderma, keloid disorders, liver cirrhosis, Alzheimer's and non-Alzheimer's dementia, Wilson's disease, primary biliary cirrhosis, chronic venous insufficiency, pseudoexfoliation syndrome, glaucoma, pelvic organ prolapse, endometriosis, intracranial aneurysm, heart failure, and cancer, including tumor metastasis, optionally selected from the group consisting of colorectal cancer, bladder cancer, pancreatic cancer, breast cancer, head and neck squamous cell carcinoma (HNSCC), laryngeal cancer, lung cancer, gastric cancer, prostate cancer, esophageal squamous cell carcinoma, endometrial cancer, testicular seminoma cancer, hepatocellular carcinoma, renal clear cell carcinoma, and basal and squamous cell carcinoma.

[0143] In another aspect, the invention relates to the use of the compounds described herein in the methods described herein.

[0144] The following figures and examples are used to illustrate the present invention and are not intended to limit the scope of the invention as set forth in the appended claims. As used herein, the term "Pacific Blue" or "PB" corresponds to 3-acetoxy-6,8-difluoro-7-hydroxycoumarin. [Brief description of the drawings]

[0145] [Figure 1] 1 shows absorbance scans (excitation) of compound 2 and 3-acetoxy-6,8-difluoro-7-hydroxycoumarin (Pacific Blue). [Diagram 2] Fluorescence emission scans of compound 2 and 3-acetoxy-6,8-difluoro-7-hydroxycoumarin (Pacific Blue) are shown. Samples are excited at their respective absorbance maxima (313 nm for compound 2, 360 nm for Pacific Blue) and emission is monitored from 333 nm or 380 nm to 600 nm. [Diagram 3] Figure 1 shows the fluorescence signal obtained from Compound 2 treated with mouse tissue homogenates of isolated lung, skin, and liver samples. The fluorescence signal generated by 100 μM Compound 2 during 18 h incubation at 37° C. with tissue homogenates is shown as relative fluorescence units (RFU) excited at 405 nm and measured at 460 nm (Ex / Em, respectively). Background controls of buffer, 100 μM untreated Compound 2, and untreated homogenate samples are measured simultaneously with treated samples (100 μM Compound 2). The most significant increase in fluorescence signal was obtained with skin tissue type (p<0.0001, one-way ANOVA). Error bars represent SD, n≧3. [Figure 4](a) Fluorescence signal obtained from masked fluorescent probe cleaved by amine oxidase according to Example 2. (b) Fluorescence signal obtained from compound 2 treated with tissue homogenate of isolated mouse skin. The signal generated by 100 μM compound 2 during 1 h incubation with tissue homogenate at 37° C. is shown as relative fluorescence units (RFU) when excited at 405 nm and measured at 460 nm (Ex / Em, respectively). Background controls of buffer, 100 μM untreated compound 2, and untreated homogenate samples are measured simultaneously with all samples treated with 100 μM compound 2. Error bars represent SD, n≧3. (c) HPLC-MS trace showing enzymatic conversion of compound 1 to PB with acrolein elimination and loss of propylamine (C3H7N, [M]=57) after 18 h incubation using UV detection at 254 nm. Reverse phase HPLC, gradient over 10 min from 10% to 90% CH3CN in H2O containing 1% CH3CN and 0.1% TFA, (1) tR = 4.9 min and [M+H]+ = 328.1; (PB) tR = 5.4 min and [M+H]+ = 271.1 and [2M+Na]+ = 563.0. [Diagram 5] Figure 1 shows the fluorescence signal obtained from Compound 2 treated with mouse isolated skin tissue homogenates with and without heat denaturation (90°C for 1.5 hours). The fluorescence signal generated by 100 μM Compound 2 during incubation with treated tissue homogenates for 1 hour at 37°C is shown as relative fluorescence units (RFU) excited at 405 nm and measured at 460 nm (Ex / Em, respectively). Buffer, 100 μM untreated Compound 2, and background controls of untreated samples are measured simultaneously with all treated samples. Incubation with heat denatured homogenates resulted in a significant decrease in the fluorescence signal of Compound 2 (p<0.0001, one-way ANOVA). Error bars represent SD, n>3. [Figure 6]Fluorescence signal obtained from Compound 2 treated with tissue homogenates of isolated mouse skin with and without the addition of BAPN, a lysyl oxidase inhibitor. Skin homogenates were pretreated with 100 μM BAPN and incubated at 37° C. for 2 h before Compound 2 was added. Fluorescence signal generated by 100 μM Compound 2 during 1 h incubation at 37° C. with treated tissue homogenates is shown as relative fluorescence units (RFU) excited at 405 nm and measured at 460 nm (Ex / Em, respectively). All samples and controls are measured simultaneously with all treated samples (100 μM Compound 2). Incubation with BAPN-inhibited homogenates resulted in a significant decrease in the fluorescence signal of Compound 2 (p<0.0001, one-way ANOVA). Error bars represent SD, n≧3. [Figure 7] Fluorescence emission scans of the experiment from Figure 6 obtained from compound 2 alone and from compound 2 treated with tissue homogenates of isolated mouse skin with and without the addition of BAPN, a lysyl oxidase inhibitor. Skin homogenates were pretreated with 100 μM BAPN and incubated at 37 °C for 2 h before compound 2 was added. The fluorescent signal generated by 100 μM compound 2 during 1 h incubation at 37 °C with treated tissue homogenates is shown as (Ex / Em, respectively) relative fluorescence units (RFU) when excited at 360 nm and measured at 400-600 nm. All samples and controls were measured simultaneously with all treated samples. Traces represent the average of duplicates (n=3). [Figure 8] Fluorescence signals obtained in tissue homogenates of isolated mouse skin from specimens representing two different age groups are shown. Skin homogenates from 6-day and 9-week-old mice were treated with 10 μM compound 2 and incubated at 37° C. for 18 h before measurement. Fluorescence signals generated are shown as relative fluorescence units (RFU) when excited at 405 nm and measured at 460 nm (Ex / Em, respectively). All samples and controls were measured in parallel assays as treated samples (10 μM probe). Error bars represent SD, n≧3. [Figure 9] Fluorescence signals obtained in tissue homogenates of wounded or non-wounded isolated mouse skin tissues are shown. Wounds were made 5 days prior to sample collection and activity analysis. Skin homogenates were treated with 10 μM compound 2 and incubated at 37° C. for 18 h before measurement. Fluorescence signals generated are shown as relative fluorescence units (RFU) when excited at 405 nm and measured at 460 nm (Ex / Em, respectively). All samples and controls are measured simultaneously with treated samples (10 μM compound 2). Error bars represent SD, n≧3. [Figure 10] (a) Fluorescence signals obtained with tissue homogenates of isolated skin from male and female mice. Skin homogenates were treated with 10 μM of compound 2 and incubated at 37° C. for 18 hours before measurement. The fluorescence signals generated are shown as relative fluorescence units (RFU) excited at 405 nm and measured at 460 nm (Ex / Em, respectively). All samples and controls are measured in parallel assays with treated samples (10 μM of compound 2). Error bars represent SD, n≧3. (b) Fluorescence signals obtained from compound 5 treated with homogenates of isolated mouse skin at a total protein concentration of 1 mg / mL. The signals generated by 20 μM of compound 5 during 1 hour incubation at 37° C. with tissue homogenates are shown as relative fluorescence units (RFU) excited at 405 nm and measured at 460 nm (Ex / Em, respectively). Background controls of untreated homogenate and 20 μM untreated Compound 5 are measured simultaneously with all treated samples. Error bars represent SD, n≧3. [Figure 11]Figure 1 shows the fluorescence signal obtained from compound 2 treated with recombinant human lysyl oxidase-like 2 (LOX-L2). Fluorescence signals were measured after 24 h incubation at 37°C and normalized to the background signal of empty wells. Values ​​are shown as relative fluorescence units (RFU) excited at 360 nm and measured at 460 nm (Ex / Em, respectively). All samples, including buffer, 100 μM untreated compound 2, and LOX-L2, were treated and measured simultaneously with the treated samples (100 μM compound 2). Error bars represent SD, n > 3. [Figure 12] Figure 1 shows the fluorescence signal obtained from compound 2 treated with recombinant human monoamine oxidases A (MAO-A) and B (MAO-B). The fluorescence signal generated during 2 h incubation at 37°C is shown as relative fluorescence units (RFU) excited at 405 nm and measured at 460 nm (Ex / Em, respectively). Buffer, 100 μM untreated compound 2, and background controls of MAO-A and MAO-B are measured simultaneously with treated samples (100 μM compound 2). Incubation with MAO-B resulted in a significant increase in the fluorescence signal of compound 2 compared to the control after 2 h incubation (p<0.0001, one-way ANOVA). Error bars represent SD, n>3. [Figure 13](a) Fluorescence signal obtained from compound 2 treated with tissue homogenates of isolated mouse skin compared to 3',6'-bis(3-aminopropoxy)-3H-spiro[2-benzofuran-1,9'-xanthen]-3-one dihydrochloride (oLOX probe). Fluorescence signal generated by 10 μM compound 2 and 10 μM oLOX after 24 h incubation with tissue homogenates at 37° C. is shown as relative fluorescence units (RFU). Samples were excited and measured at the maximum absorbance and emission of each system (oLOX Ex. 450 nm / Em. 540 nm, and compound 2 Ex. 360 nm / Em. 460 nm). All controls are measured simultaneously with the respective treated samples with otherwise identical instrument settings. When incubated with homogenates, a significant increase in the fluorescent signal was obtained with probe compound 2 (****, p<0.0001, one-way ANOVA), but not with oLOX (ns, p=0.2696, one-way ANOVA). Error bars represent SD, n≧3. (b) 1H NMR spectrum (400 MHz) of 3-carboxymethyl-6,8-dichloro-7-hydroxycoumarin in CD3CN. [Figure 14] (a) 1H NMR spectrum (400 MHz) of 3-methylacetate-6,8-difluoro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin in CD3CN. (b) 1H NMR spectrum (400 MHz) of 3-methylacetate-6,8-dichloro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin in CD3CN. [Figure 15] 13C NMR spectrum (101 MHz) of 3-methylacetate-6,8-difluoro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin in CD3CN. [Figure 16] Figure 2 shows the F NMR HF decoupled spectrum (376 MHz) of 3-methylacetate-6,8-difluoro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin in CDCN. [Figure 17](a) 1H NMR spectrum (400 MHz) of compound 1 in DMSO-d6. (b) 1H NMR spectrum (400 MHz) of MRA_3102 in CDCl3. [Figure 18] 13C NMR spectrum (101 MHz) of compound 1 in DMSO-d6. [Figure 19] 19F NMR FH decoupled spectrum (376 MHz) of compound 1 in DMSO-d6. [Figure 20] 19F NMR FH coupling spectrum (376 MHz) of compound 1 in DMSO-d6. [Figure 21] (a) 1H NMR spectrum (400 MHz) of compound 2 in DMSO-d6. (b) 1H NMR spectrum (400 MHz) of 3-methylacetate-6,8-dichloro-7-(4-ammoniumpropoxy)-coumarine chloride in DO. [Figure 22] 13C NMR spectrum (101 MHz) of compound 2 in d6-DMF. [Diagram 23] 19F NMR FH decoupled spectrum (376 MHz) of compound 2 in CDCl3. [Figure 24] 19F NMR FH coupling spectrum (376 MHz) of compound 2 in CDCl3. [Diagram 25] 1H NMR spectrum (400 MHz) of MRA_3068 in CDCl3 is shown. [Figure 26] 13C NMR spectrum (101 MHz) of MRA_3068 in CDCl3. [Figure 27] Figure 2 shows the 19F NMR (FH decoupling) spectrum (376 MHz) of MRA_3068 in CDCl3. [Figure 28] Figure 2 shows the 19F NMR spectrum (376 MHz) of MRA_3068 in CDCl3. [Figure 29] The CD spectra of compounds 3, 4, and 5 are shown. [Diagram 30]The thermal denaturation curves of compounds 3, 4, and 5 are shown. [Diagram 31] (a)-(c) show the analysis of skin tissue after in vivo administration of (a) compound 3, (b) compound 4 and (c) compound 5 in mice. In these images of wound cross-sections, fluorescence can be clearly visualized at the positions where the peptide-bound probe was unmasked by in vivo reaction with amine oxidase (blue) at and around the injection site. In Fig. 31(a), the top image shows the context of the three images below. Fig. 31(a) (middle) demonstrates that the compound localizes the probe to collagen and elastin at the site of LOX-mediated cross-linking during new tissue growth and maturation in wounds in vivo, demonstrating that the compound according to the invention can be used in vivo for targeting and analysis of collagen cross-linking in the extracellular matrix, such as those occurring during fibrosis. [Diagram 32] 1H NMR spectrum (400 MHz) of MRA_3100 in CDCl3 is shown. [Diagram 33] 1H NMR spectrum (600 MHz) of MRA_3103 in CD3CN. [Diagram 34] (a) Fluorescence signals of MAO probes are shown, showing the efficiency of fluorescence quenching for probe 1 and control 2 vs. Pacific Blue. Fluorescence signals are shown as relative fluorescence units (RFU) when excited at 405 nm and measured at 460 nm (Ex / Em, respectively). Error bars represent SD, n=3. (b) Absorbance scans from 300 to 600 nm of MAO probes are shown, showing the efficiency of quenching as the absorbance maximum shifts from 360 nm to approximately 315 nm for probe 1 and control 2 vs. Pacific Blue, as well as the decrease in absorbance. Transmittance is shown as absorbance units (AU). [Diagram 35]Confocal microscopy images of cells after treatment with MAO probe are shown. Top: MCF-7 cells were treated with MRA_3100 (10 mM) for 3 h and then stained with MitoTracker Red. (A) Blue channel (405 nm) shows probe activation and fluorescence, (B) overlay of blue and red channels (405 nm and 561 nm) shows colocalization of probe and dye in mitochondria. Bottom: SY5Y cells were treated with MRA_3100 (10 mM) for 3 h and then stained with MitoTracker Red. (C) Blue channel (405 nm) shows probe activation and fluorescence, (D) overlay of blue and red channels (405 nm and 561 nm) shows colocalization of probe and dye in mitochondria. [Diagram 36] Confocal microscopy images of MCF-7 cells after treatment with a peptide MAO probe (10 mM) for 1 hour and staining with MitoTracker Red are shown: (A) the blue channel (405 nm) shows the activation and fluorescence of the probe, (B) the red channel (561 nm) shows the location of mitochondria within the cells, (C) an overlay of the blue and red channels (405 nm and 561 nm) shows the colocalization of the probe and dye in mitochondria, and (D) a magnified view of the inset of (C). [Figure 37] Confocal microscopy images of MCF-7 cells after treatment with PB peptide (10 mM) for 1 hour and staining with MitoTracker Red are shown: (A) blue channel (405 nm) shows the fluorescence of the probe, (B) red channel (561 nm) shows the location of mitochondria within the cells, (C) overlay of blue and red channels (405 nm and 561 nm) shows the lack of colocalization between the probe and the red dye located in mitochondria, and (D) magnification of the inset of (C). [Figure 38]Confocal microscopy images of MCF-7 cells after treatment with a peptide MAO probe (10 mM) for 1 hour and staining with MitoTracker Red are shown: (A) the blue channel (405 nm) shows no fluorescence from the probe as it cannot be activated by the MAO enzyme, (B) the red channel (561 nm) shows the location of mitochondria within the cells, and (C) an overlay of the blue and red channels (405 nm and 561 nm) shows only the red dye located in mitochondria. [Figure 39] Confocal microscopy images of MCF-7 cells after 1 hour pretreatment with the MAO-B inhibitor (R)-N,α-dimethyl-N-2-propynylphenethylamine (selegiline) and then 1 hour treatment with a peptide MAO probe (10 mM), followed by staining with MitoTracker Red: (A) the blue channel (405 nm) shows little fluorescence from the probe, (B) the red channel (561 nm) shows the location of mitochondria within the cells, and (C) an overlay of the blue and red channels (405 nm and 561 nm) shows poor colocalization between the probe and the red dye located in mitochondria. [Diagram 40] A collage of 10x images ((a)) displaying an entire section of an SSC13 ear tumor after (b) staining with compound 3 (blue), (c) immunostaining for collagen I (green), and (d) nuclear staining with propidium iodide (red). Single channel images show a black and white representation of the fluorescence. The blue fluorescence of the probe is seen at the periphery of the tumor but is strongest near the attachment side to the cartilage (arrows). [Diagram 41] Merged 20x images ((a) and (b)) and single channel representations ((c)-(e)) of SSC13 ear tumors after staining are shown. Nuclear staining with propidium iodide ((c), red), immunostaining for collagen I ((d), green), and blue fluorescence from the probe ((e)) seen at the periphery of the tumor is strongest near the attachment side to the cartilage but does not appear in the dermis opposite the cartilage. Fluorescence of the cartilage (cart) and hair follicles (hf) can also be observed. Scale bar = 100 μm. [Diagram 42]A collage of 10x images ((a)) displaying an entire section of an SSC13 ear tumor after staining with compound 6 ((b) blue), immunostaining for collagen I ((c) green), and nuclear staining with propidium iodide ((d) red). Single channel images show a black and white representation of the fluorescence. Fluorescence of the probe in the blue channel (top right) can be seen at the periphery of the tumor (upper arrow), but is also clearly visible in the "healthy" dermis (lower arrow) opposite the cartilage. [Diagram 43] Merged 20x images ((a) and (b)) and single channel images ((c)-(e)) of an SSC13 ear tumor after staining with compound 6 are shown. Nuclear staining with propidium iodide ((c), red), immunostaining for collagen I ((d), green), and blue fluorescence from compound 6 ((e)) are seen in the fibrous tissue near the attachment side to the cartilage, but are also abundant in the outer dermis of the tumor opposite the cartilage. Fluorescence of cartilage (cart) and hair follicles (hf) can also be observed. 20x scale bar = 100 μm. [Diagram 44] (a)-(i) show 10x images of healthy skin after staining with compounds 3 ((a)-(c)), 6 ((d)-(f)), 4 ((g)), and 5 ((h)), as well as PBS ((i)). Nuclear staining with propidium iodide (red), immunostaining for collagen I ((b) and (e), green), or collagen III ((c) and (f), green), and blue fluorescence from compound 6 can be seen throughout the dermis. Autofluorescence of hair follicles (hf) can also be observed. 10x scale bar = 100 μm. EXAMPLES

[0146] Example 1 General Experimental Mode Materials and reagents were of the highest grade commercially available and were used without further purification. They were purchased from Acros Organics (Switzerland), Sigma Aldrich (Switzerland), Fischer (Switzerland), Bachem (Switzerland), Chem-Impex (USA), and TCI (Germany). Water used for peptide preparation and purification was nanopure with a resistivity of 18.2 MΩ*cm, prepared with a Sartorius Arium611VF (Switzerland) water purification system or redistilled, and water was purchased from AppliChem Panreac (Switzerland). For small molecule synthesis, reactions were monitored by thin layer chromatography using Merck Millipore (Switzerland) silica gel 60 F254 glass backing plates. Visualization of compounds was achieved by UV-Vis or staining with KMnO4. Flash chromatography and plug filtration were performed using silica gel (60 Å pore size, 230-400 mesh particle size, Sigma Aldrich, Switzerland). Solvents for extraction and chromatography were of technical quality and were distilled before use. 1 H and 13C NMR spectra were recorded on a Bruker DRX 400, a Bruker AV III 400 (400 MHz / 100 MHz), or a Bruker AV III 600 (600 MHz / 150 MHz). All spectra were recorded at 25 °C unless otherwise stated. Chemical shifts (δ) are reported in parts per million (ppm) relative to tetramethylsilane (TMS) or residual solvent signals. Signals were assigned COSY, HSQC, HMBC, and NOESY spectra. Solid-phase peptide synthesis (SPPS) was performed on Rink amide ChemMatrix resin from Biotage (Sweden) and automated peptide synthesis was performed on a Syro I peptide synthesizer (MultiSynTech GmbH, Witten Germany). High resolution mass spectrometry (HRMS) was performed by the Molecular and Biomolecular Analysis (MoBiAs) service of D-CHAB, ETH Zurich using a Bruker Daltons maXis with an ESI (electrospray ionization) source and a Q-TOF ion analyzer or a Bruker Daltonics SOLARIX with a MALDI (matrix-assisted laser desorption / ionization) / ESI source and a Q-TOF ion analyzer. α-Cyano-4-hydroxycinnamic acid (CHCA) was used as the MALDI-MS matrix. Analytical reversed-phase high performance liquid chromatography (RP HPLC) was performed on a Dionex UHPLC, Ultimate 3000 (Thermo Fisher Scientific, Waltham / USA). Preparative RP HPLC purification was performed on a Dionex UHPLC, Ultimate 3000 (Thermo Fisher Scientific, Waltham / USA). Circular dichroism (CD) spectra were recorded on a Chirascan plus spectrometer (Applied Photophysics Ltd, Leatherhead / UK) equipped with a Nitropack nitrogen generator (Parker Balston, Haverhill / USA) and a temperature controller TC 125 (Quantum Northwest).Solutions were measured in quartz cuvettes (Hellma 110-QS) with a path length of 1.0 mm. Peptides were dried by lyophilization in a Christ Alpha 2-4 LD plus (Kuhnner AG, Birsfelden / CH) freeze dryer. Absorbance emission spectroscopy, UV-visible spectra were acquired on a Cary-500 scanning spectrophotometer. Spectra were measured in quartz cuvettes (ThorLabs, CV10Q3500, Newton, NJ USA). Samples were exposed to approximately 2 mW cm. -2 The fluorescence was measured with a LED transilluminator (Roithner Lasertechnik, LED405-06V, Vienna, Austria) emitting at the wavelengths as stated, with an incident intensity of 100 nm and measured with a power meter (ThorLabs, PM100D, Newton, NJ USA) equipped with a Si photodiode detector (ThorLabs, S120VC, Newton, NJ USA). Fluorescence spectroscopy was measured with a Fluorolog 3 fluorometer (Horiba Jobin-Yvon, Germany) equipped with a cuvette sample changer (ThorLabs, CV10Q3500F-E, Newton, NJ USA) for quartz cuvettes. To avoid photoactivation, all measurements were performed at 25 °C in 50 mM PBS pH 7.4 buffer under red light for ambient illumination. Quantum yields were determined in PBS and correlated, where necessary, with the known reference of Pacific Blue (Φ F = 0.884 measurements vs. Φ F =0.89). Ex vivo fluorescence measurements were performed at room temperature using a Tecan Spark 10M multimode microplate reader (Tecan, Mannedorf, Switzerland) with excitation / emission filters (e.g., 405 / 460 nm, respectively) unless otherwise stated, and values ​​are expressed in relative fluorescence units (RFU) for each experiment. All experiments were performed in triplicate. Graphical presentation and statistical analysis were performed using Graphpad Prism7 software (GraphPad Software, Inc., San Diego, USA.) and statistical significance was determined by ordinary one-way ANOVA with multiple comparisons at p<0.05.

[0147] Example 2 General and exemplary experimental protocols for determining amine oxidase activity Fluorescence experiments to determine the amine oxidase activity of a given enzyme are performed using a fluorescence plate reader and standard 96-well black polystyrene plates with clear flat bottoms. 150 μL of the following solutions are added to each well of the plate, for example in duplicate at least three times: buffer (e.g., 25 mM at pH 7.5), a solution of an amine oxidase-reactive probe, e.g., Compound 2 (e.g., 100 μM), in buffer, a buffered solution of a sample containing an amine oxidase at the desired concentration (e.g., 40 μg / mL), and a buffered solution of a sample containing an amine oxidase plus a probe, e.g., Compound 2 (e.g., 100 μM). Before measurements are taken, the plate is incubated in the dark at 37° C. for between 15 minutes and 24 hours. Fluorescence is measured by exciting the unmasked probe at maximum absorbance (e.g., 360 nm) or 405 nm and measuring the fluorescence at the maximum emission of the probe, e.g., 460 nm. Enzyme activity is observed as fluorescence measured relative to an untreated control, e.g., expressed in relative fluorescence units (RFU). The activation of the probe, and thus the amine oxidase activity, can be displayed graphically (see FIG. 4a). A change in fluorescence, optionally a statistically relevant change, compared to an untreated control indicates amine oxidase activity.

[0148] Example 3 Synthesis of 3-carboxymethyl-6,8-difluoro-7-hydroxycoumarin ("Pacific Blue" ester)

[0149] [ka]

[0150] Pacific blue (ester) was prepared by Peckmann condensation of 2,4-difluororesorcinol and dimethylacetylsuccinate as previously reported in Chang, D.; Kim, KT; Lindberg, E.; Winssinger, N., Bioconj. Chem. 2018, 29, 158-163. 1 H NMR (400 MHz, DMF-d7) δ 7.61 (dd, J = 11.8, 2.2 Hz, 1H), 3.79 (s, 2H), 3.71 (s, 3H), 2.48 (s, 3H).

[0151] Example 3a Synthesis of 3-carboxymethyl-6,8-dichloro-7-hydroxycoumarin

[0152] [ka]

[0153] 2,4-Dichlororesorcinol (1.68 g, 9.4 mmol) was dissolved in dimethylacetylsuccinate (1.77 g, 9.5 mmol) by trituration. Concentrated H2SO4 (1.5 mL) was added and the solution was stirred at room temperature for 24 h. The resulting viscous liquid was homogenized by the addition of methanol (1-2 mL) and then poured onto crushed ice (100 mL). The precipitate was collected by filtration and recrystallized from methanol / water to give 3-carboxymethyl-6,8-dichloro-7-hydroxycoumarin as a purple crystalline solid. 1 H NMR (400 MHz, CD3CN) δ 7.71 (s, 1H), 3.66 (s, 5H), 2.35 (s, 3H). 13 C NMR (101 MHz, CD3CN) δ 171.54, 161.19, 152.23, 149.77, 149.10, 125.08, 118.93, 115.46, 109.76, 52.75, 33.51, 15.88. HRMS (ESI): m / z C 13 H 10Calculated value for Cl2NaO5: 338.9797 [M+Na] + ; Measured Value: 338.9795.

[0154] Example 4 3-Methylacetate-6,8-difluoro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin

[0155] [ka]

[0156] Pacific blue (5.4 mmol, 1.56 g) and 3-(tert-butoxycarbonylamino)-propyl bromide (11 mmol, 2.6 g) were dissolved in 18 mL of anhydrous DMF. CsCO3 (8.1 mmol, 2.64 g) was added and the resulting solution was heated to 60 °C for 2 h. The solution was cooled to room temperature and the reaction was quenched with 200 mL of aqueous ammonium chloride, extracted with EtOAc (3 x 50 mL), dried over Na2SO4, and concentrated under reduced pressure. The crude mixture was purified by chromatography on silica gel (1-5% gradient of MeOH in dichloromethane) and concentrated under reduced pressure to yield 3-methylacetate-6,8-difluoro-7-(4-((tert-butoxycarbonyl))amino)propoxy)-coumarin ("Boc-PB-LOX-OMe") as a clear oil (2.3 g, 96%). 1 H NMR (400 MHz, CD3CN) δ 7.27 (dd, J = 12.0, 2.2 Hz, 1H), 5.38 (s, 1H), 4.25 (t, J = 6.2 Hz, 2H), 3.60 (s, 5H), 3.16 (q, J = 6.6 Hz, 2H), 2.26 (s, 3H), 1.84 (p, J = 6.4 Hz, 2H), 1.32 (s, 9H). 13C NMR (101 MHz, CD3CN) δ 171.31, 160.66, 156.92, 152.40 (dd, J = 243.1, 4.6 Hz), 149.54 (t, J = 2.8 Hz), 143.70 (dd, J = 249.3, 6.3 Hz), 139.43 (dd, J = 10.3, 2.5 Hz), 138.78 (dd, J = 16.0, 10.9 Hz), 120.63 (d, J = 0.8 Hz), 116.21 (dd, J = 9.2, 1.3 Hz), 107.68 (dd, J = 22.9, 3.6 Hz), 79.12, 73.76, 52.76, 37.83, 33.62, 31.21, 28.60, 15.96. 19 F NMR, FH decoupling (377 MHz, CD3CN) δ -133.72 (d, J = 5.4 Hz), -150.18 (d, J = 5.4 Hz). 19 F NMR (377 MHz, CD3CN) δ -133.72 (dd, J = 11.9, 5.4 Hz), -150.18 (d, J = 5.4 Hz). HRMS (ESI): m / z C 21 H 25 Calculated value for F2NNaO7: 464.1491 [M+Na] + ; Measured value: 464.1494.

[0157] Example 4a Synthesis of 3-methylacetate-6,8-dichloro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin

[0158] [ka]

[0159] 3-Carboxymethyl-6,8-dichloro-7-hydroxycoumarin (0.71 mmol, 227 mg) and 3-(tert-butoxycarbonylamino)-propyl bromide (1.45 mmol, 347 mg) were dissolved in 2.4 mL of anhydrous DMF. CsCO3 (1.07 mmol, 349 mg) was added and the resulting solution was stirred and heated to 60 °C for 2 h. The solution was cooled to room temperature and the reaction was quenched with 100 mL of aqueous ammonium chloride solution, extracted with EtOAc (3 x 50 mL), dried over Na2SO4, and concentrated under reduced pressure. The crude mixture was purified by chromatography on silica gel (1-5% gradient of MeOH in dichloromethane) and concentrated under reduced pressure to yield 3-methylacetate-6,8-dichloro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin as a white solid (175 mg, 52%). 1 H NMR (400 MHz, CD3CN) δ 7.70 (s, 1H), 5.44 (s, 1H), 4.09 (t, J = 6.2 Hz, 2H), 3.66 (s, 2H), 3.66 (s, 3H), 3.28 (q, J = 6.6 Hz, 2H), 2.34 (s, 3H), 2.01 - 1.94 (m, 2H), 1.40 (s, 9H). 13 C NMR (101 MHz, CD3CN) δ 171.27, 160.85, 156.93, 154.33, 149.34, 148.90, 125.42, 125.26, 120.79, 118.99, 117.72, 73.13, 52.78, 38.20, 33.62, 33.50, 31.19, 28.63, 15.99. HRMS (ESI): m / z C 21 H 25 Calculated value for Cl2NNaO7: 496.0900 [M+Na] + ; Measured value: 496.0901.

[0160] Example 4b Synthesis of 3-methylacetate-6,8-dichloro-7-(4-ammoniumpropoxy)-coumarine chloride

[0161] [ka]

[0162] 3-Methylacetate-6,8-dichloro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin (0.042 mmol, 20 mg) was dissolved in 250 μL of 4N HCl in dioxane at room temperature and stirred for 1 h. The solvent was then removed from the resulting slurry by evaporation with a gentle stream of compressed air and the white solid was dried under high vacuum overnight to yield the HCl salt 3-methylacetate-6,8-dichloro-7-(4-ammoniumpropoxy)-coumarin chloride (17 mg, quantitative) as a white solid. 1 H NMR (400 MHz, heavy water) δ 7.77 (s, 1H), 4.26 (t, J = 5.7 Hz, 2H), 3.81 (s, 2H), 3.77 (s, 3H), 3.39 (t, J = 7.5 Hz, 2H), 2.39 (s, 3H), 2.26 (dt, J = 12.7, 6.1 Hz, 2H). 13 C NMR (126 MHz, D2O) δ 173.01, 162.07, 152.47, 151.06, 146.91, 124.69, 124.52, 118.74, 117.94, 116.36, 71.80, 52.88, 37.41, 32.71, 27.32, 15.10.

[0163] Example 5 3-Carboxy-6,8-difluoro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin (Compound 1)

[0164] [ka]

[0165] Boc-PB-LOX-OMe (5.2 mmol, 2.3 g) was dissolved in 8 mL of 1:1 THF:MeOH, NaOH (15.6 mmol, 624 mg) was added as an aqueous solution in 1 mL of HO, and the resulting solution was heated to 50° C. for 5 h. Upon completion as observed by thin layer chromatography (Rf=0.1 for 5% MeOH in DCM), the solution was cooled to room temperature and the reaction was carefully acidified to pH 2 with 1 M HCl, immediately extracted with EtOAc (3×50 mL), dried over NaSO, and concentrated under reduced pressure to give compound 1 (2.2 g, 99%) as a white solid. 1 H NMR (400 MHz, DMSO-d6) δ 12.53 (s, 1H), 7.66 (d, J = 12.5 Hz, 1H), 6.89 (t, J = 5.7 Hz, 1H), 4.29 (t, J = 6.3 Hz, 2H), 3.63 (s, 2H), 3.11 (q, J = 6.5 Hz, 2H), 2.38 (s, 3H), 1.84 (p, J = 6.6 Hz, 2H), 1.37 (s, 9H). 13 C NMR (101 MHz, DMSO) δ 171.06, 159.32, 155.61, 152.13, 152.08, 149.71, 149.67, 148.34, 143.36, 143.29, 140.88, 140.82, 137.90, 137.88, 137.80, 137.78, 137.32, 137.21, 137.16, 137.05, 119.92, 115.16, 115.07, 107.40, 107.18, 77.53, 72.55, 36.41, 32.94, 29.97, 28.21, 15.39. 19 F NMR, FH decoupling (376 MHz, DMSO-d6) δ -132.45 (d, J = 5.0 Hz), -149.31 (d, J = 5.1 Hz). 19 F NMR (376 MHz, DMSO-d6) δ -132.45 (dd, J = 11.9, 4.9 Hz), -149.31 (d, J = 4.7 Hz). HRMS (ESI): m / z C 20 H23 Calculated value for F2NNaO7: 450.1335 [M+Na] + ; Measured value: 450.1340.

[0166] Example 5a Synthesis of MRA_3102

[0167] [ka]

[0168] MRA_3102.2 (0.68 mmol, 300 mg) was dissolved in 3.4 mL of THF and N-hydroxysuccinimide (0.68 mmol, 78 mg) was added. The resulting solution was cooled in an ice batch and DCC was added (0.68 mmol, 140.2 mg). The reaction mixture was stirred on ice for 30 min and then warmed to room temperature overnight. The DCC was filtered and the solution was concentrated by rotary evaporation. The crude residue was resuspended in EtOAc, filtered and concentrated again by rotary evaporation, and this process was repeated twice until no precipitate was observed in the solution. The crude NHS ester was then concentrated under reduced pressure to yield MRA_3102 (353 mg, 99%) as a white solid. 1 H NMR (400 MHz, CDCl3) δ 7.17 (dd, J = 11.2, 2.2 Hz, 1H), 4.80 (s, 1H), 4.35 (t, J = 6.0 Hz, 2H), 4.08 (s, 2H), 3.36 (q, J = 6.2 Hz, 2H), 2.83 (s, 4H), 2.41 (s, 3H), 2.00 (p, J = 6.4 Hz, 2H), 1.44 (s, 9H). 13C NMR (101 MHz, CDCl3) δ 168.81, 165.57, 159.57, 156.18, 151.73 (dd, J = 246.0, 4.2 Hz), 149.64 (t, J = 2.7 Hz), 143.15 (dd, J = 253.2, 5.7 Hz), 138.97 (dd, J = 10.3, 2.6 Hz), 138.70 (dd, J = 15.6, 10.7 Hz), 119.85 (d, J = 13.4 Hz), 117.63, 114.98 (d, J = 8.8 Hz), 106.25 (dd, J = 22.4, 3.7 Hz), 77.36, 73.09, 37.66, 30.41, 30.08, 28.52, 25.70, 16.06. 19 F NMR, FH decoupling (376 MHz, CDCl3) δ -131.48 (d, J = 5.8 Hz), -146.94 (d, J = 5.8 Hz). 19 F NMR (376 MHz, CDCl3) δ -131.48 (dd, J = 10.4, 5.5 Hz), -146.94 (d, J = 4.8 Hz).

[0169] Example 6 3-Carboxy-6,8-difluoro-7-(4-((tert-butoxycarbonyl)amino)propoxy)-coumarin (compound 2)

[0170] [ka]

[0171] Compound 1 (0.047 mmol, 20 mg) was dissolved in 250 μL of 4N HCl in dioxane at room temperature and stirred for 1 h. The solvent was then removed from the resulting slurry by evaporation with a gentle stream of compressed air, and the white solid was dried under high vacuum overnight to yield the HCl salt of compound 2 (17 mg, quantitative) as a white solid. 1H NMR (400 MHz, DMSO-d6) δ 8.13 (s, 3H), 7.67 (dd, J = 12.1, 2.1 Hz, 1H), 4.37 (t, J = 6.1 Hz, 2H), 3.62 (s, 2H), 2.97 (t, J = 7.5 Hz, 2H), 2.36 (s, 3H), 2.05 (p, J = 6.4 Hz, 2H). 13 C NMR (101 MHz, DMSO-d6) δ 171.14, 159.38, 150.92 (dd, J = 242.9, 4.2 Hz), 148.43 (t, J = 2.5 Hz), 142.14 (dd, J = 249.1, 6.2 Hz), 137.87 (dd, J = 10.1, 2.3 Hz), 136.95 (dd, J = 16.3, 11.0 Hz), 120.14, 115.41 (d, J = 9.3 Hz), 107.44 (dd, J = 22.5, 3.4 Hz), 71.92, 66.43, 35.84, 33.06, 27.65, 15.51. 19 F NMR, FH decoupling (376 MHz, DMSO-d6) δ -132.51 (d, J = 4.9 Hz), -149.30 (d, J = 4.9 Hz). 19 F NMR (376 MHz, DMSO-d6) δ -132.51 (dd, J = 12.0, 4.9 Hz), -149.30 (d, J = 5.1 Hz). HRMS (ESI): m / z C 15 H 16 Calculated value for F2NO5: 328.0991 [M+H] + ; Measured value: 328.0989.

[0172] Example 7 3-Methylacetate-6,8-difluoro-7-(4-((tert-butoxycarbonyl)amino)butoxy)-coumarin (MRA_3068)

[0173] [ka]

[0174] 3-Carboxymethyl-6,8-difluoro-7-hydroxycoumarin (Pacific Blue) (1 mmol, 284 mg) and 3-(tert-butoxycarbonylamino)-butyl bromide (2 mmol, 504 mg) were dissolved in 2 mL of anhydrous DMF. CsCO3 (1.5 mmol, 489 mg) was added and the resulting solution was heated to 60 °C for 2 h. The solution was cooled to room temperature and the reaction was quenched with 100 mL of aqueous ammonium chloride, extracted with EtOAc (3 × 50 mL), dried over Na2SO4, and concentrated under reduced pressure. The crude mixture was purified by chromatography on silica gel (1-5% gradient of MeOH in dichloromethane) and concentrated under reduced pressure to yield MRA_3068 (341 mg, 75%) as a white solid. 1 H NMR (400 MHz, CDCl3) δ 7.13 (dd, J = 11.4, 2.3 Hz, 1H), 4.60 (s, 1H), 4.29 (tt, J = 6.2, 1.0 Hz, 2H), 3.73 (s, 2H), 3.71 (s, 3H), 3.19 (s, 2H), 2.34 (s, 3H), 1.86 - 1.78 (m, 2H), 1.73 - 1.65 (m, 2H), 1.43 (s, 9H). 13 C NMR (101 MHz, CDCl3) δ 170.35, 160.05, 156.13, 151.71 (dd, J = 245.5, 4.5 Hz), 148.03 (t, J = 2.7 Hz), 143.14 (dd, J = 253.0, 6.0 Hz), 138.76 (dd, J = 10.5, 2.5 Hz), 138.33 (dd, J = 15.7, 10.7 Hz), 119.76, 115.20 (dd, J = 8.8, 1.0 Hz), 106.03 (dd, J = 22.4, 3.7 Hz), 74.73 (t, J = 3.5 Hz), 52.54, 33.01, 28.53, 27.34, 26.54, 15.81. 19F NMR, FH decoupling (376 MHz, CDCl3) δ -131.81 (d, J = 5.6 Hz), -147.25 (d, J = 5.8 Hz). 19 F NMR (376 MHz, CDCl3) δ -131.81 (dd, J = 11.4, 5.7 Hz), -147.25 (dd, J = 5.7, 2.0 Hz). HRMS (ESI): m / z C 22 H 31 Calculated value for F2N2O7: 473.2094 [M+NH4] + ; Measured value: 473.2087.

[0175] Example 8 3-Carboxy-6,8-difluoro-7-(4-((tert-butoxycarbonyl)amino)butoxy)-coumarin (MRA_3069)

[0176] [ka]

[0177] MRA_3068 (0.5 mmol, 241 mg) was dissolved in 1.3 mL of 1:1 THF:MeOH, NaOH (1.6 mmol, 64 mg) was added as an aqueous solution in 0.6 mL of HO, and the resulting solution was heated to 50° C. for 5 h. Upon consumption of starting material as observed by thin layer chromatography (Rf=0.1 for 5% MeOH in DCM), the solution was cooled to room temperature and the reaction was carefully acidified to pH 2 with 1 M HCl, immediately extracted with EtOAc (3×50 mL), dried over NaSO and concentrated under reduced pressure to yield MRA_3069 (231 mg, 97%) as a clear oil. 11H NMR (400 MHz, methanol-d4) δ 7.40 (dd, J = 11.8, 2.2 Hz, 1H), 4.30 (t, J = 6.2 Hz, 2H), 3.70 (s, 2H), 3.11 (t, J = 6.8 Hz, 2H), 2.38 (s, 3H), 1.85 - 1.76 (m, 3H), 1.71 - 1.63 (m, 3H), 1.42 (s, 9H). 13 13C NMR (101 MHz, methanol-d4) δ 173.51, 161.70, 158.51, 153.00 (dd, J = 244.3, 4.4 Hz), 150.12 (t, J = 2.7 Hz), 144.08 (dd, J = 250.5, 6.2 Hz), 139.63 (dd, J = 10.3, 2.5 Hz), 139.28 (dd, J = 16.0, 10.9 Hz), 121.06, 116.66 (d, J = 9.1 Hz), 107.66 (dd, J = 22.9, 3.6 Hz), 79.85, 75.86 (t, J = 3.4 Hz), 40.86, 33.62, 28.77, 28.34, 27.25, 15.68. 19 19F NMR, F-H decoupling (376 MHz, methanol-d4) δ -133.41 (d, J = 5.5 Hz), -150.56 (d, J = 5.5 Hz). 19 19F NMR (377 MHz, methanol-d4) δ -133.41 (dd, J = 11.8, 5.4 Hz), -150.55 (d, J = 5.4 Hz). HRMS (ESI): m / z C 21 H 23 Calculated for C - H

[0178] (Example 9) Ex vivo experiment - General aspects

[0179] Skin tissues used for enzymatic analysis of LOX activity were harvested (1 cm x 1 cm) and placed in 2 ml of ice-cold PBS. Skin tissues were homogenized using an Ultra-TURRAX homogenizer (KA®-Werke GmbH & CO. KG, Staufen, Germany) and samples were spun at 2000 x g to pellet tissue debris. The supernatant was further purified using a 0.2 μm filter. Tissue homogenates were then assessed for total protein recovery in solution using nanospec absorbance measurements (Pierce™, Thermo Fisher Scientific Inc, Waltham, USA) standardized to 1 mg / mL BSA. Homogenates were diluted to appropriate concentrations (e.g., 1 mg / ml total protein) in 25 mM PIPES, 0.5% Triton X-100 (Sigma Aldrich, St Louis, MA, USA) and pre-incubated with the probes (10 μM or 100 μM) for the indicated time periods at 37° C. in the dark before fluorescence measurements. The resulting fluorescence was then measured using a Tecan Spark 10M multimode microplate reader (Tecan, Mannedorf, Switzerland) at room temperature with experimentally defined excitation / emission filters (e.g., 405 / 460 nm, respectively). Activity was observed as measured fluorescence and expressed in relative fluorescence units (RFU). Graphical presentation and statistical analysis were performed using Graphpad Prism7 software (GraphPad Software, Inc., San Diego, USA.) and statistical significance was determined as p<0.05.

[0180] Example 10 General protocol for synthesizing collagen model peptides 3, 4, and 5 Protocol A - General procedure for swelling: Prior to automated peptide synthesis, the resin was swelled in CH2Cl2 for 15 min with shaking. The resin was then drained, washed with DMF (3x6 mL) and drained again. Protocol B - General protocol for automated peptide synthesis: For automated peptide synthesis, a SyroI peptide synthesizer (Biotage, Sweden) was used. Couplings were carried out using either the appropriate Fmoc-protected amino acid or the trimer Fmoc-Pro-Hyp-Gly-OH. After swelling of the resin with DMF in the synthesizer, i-Pr2NEt (9 eq., as a 3M solution in NMP (N-methyl-2-pyrrolidone)), HATU (3 eq., 0.5M in DMF) and Fmoc-amino acid / Fmoc-tripeptide (3 eq., 0.5M in DMF) were added to the resin. The mixture was reacted at 1 min intervals, stirred for 5 min, left to stand for 30 min (2x), and then washed with DMF (5x). Fmoc-deprotection was performed by adding a solution of 40% (v / v) piperidine in DMF and reacting for 1 min. This step was repeated 4 times. The resin was then washed with DMF (5x). Tripeptide coupling and Fmoc-deprotection were repeated until the desired peptide was obtained. No acylation (capping) was performed in the automated synthesis of CMP. Protocol C- For N-terminal functionalization of the resin with compound 1: Functionalization was performed manually at room temperature on solid support-bound peptides. Compound 1 (2.0 equiv), HATU (1.9 equiv) and i-Pr2NEt (4 equiv) were dissolved in DMF (1-2 mL). After 5 min preactivation, the coupling mixture was added to the resin and stirred for 1-2 h. The resin was washed with CHCl (3×), DMF (3×), CHCl (3×), and petroleum ether (2×). The reaction was monitored by a qualitative color test of the beads or by LC-MS after test cleavage (see protocol E). Protocol D- Cleavage from the resin: The resin was shaken in a mixture of TFA / (i-Pr)Si-H / H0 (92.5:2.5:2.5) for 1 h and washed with pure TFA (2×). The peptide in solution was collected by filtration in a conical flask. Upon addition of ice-cold Et0, the peptide was obtained as a white precipitate.The solid was isolated by centrifugation followed by decantation. The solid was suspended in Et2O, sonicated, centrifuged again and the supernatant was decanted. The remaining white solid was dissolved in water / CH3CN, frozen and lyophilized to give a white foam. Protocol E- Purification and analysis by RP HPLC: CH3CN (A) and H2O containing 1% CH3CN and 0.1% TFA (B) were used as eluents. A flow rate of 6 mL / min was used for semi-preparative HPLC, a flow rate of 1 mL / min for analytical HPLC and a flow rate of 0.5 mL / min for LC-MS. After purification by semi-preparative HPLC, all collected fractions were analyzed by analytical HPLC or LC-MS and only pure fractions were combined. The amine containing CMP was desalted on a VariPure cartridge and then lyophilized. Preparative column: Reprosil Gold 120 C18, 150 x 10 mm. Analytical column: Phenomenex, Jupiter 5 μm, 300 Å, 250×4.6 mm. LC-MS: Reprosil Gold C18 5 μm, 125×3 mm. Protocol F- Gel Permeation Chromatography: Nanopure H2O was used as eluent. A flow rate of 0.1 mL / min was used at room temperature.

[0181] Example 11 Synthesis of 1-[ProHypGly]3-AopProGly-[ProHypGly]3-NH2 (Compound 3)

[0182] [ka]

[0183] Compound 3 was synthesized on Rink amide ChemMatrix resin (approximately 0.5 mmol / g). The resin was swollen according to protocol A. Fmoc-γ-aminooxyproline (Aop)-OH, Fmoc-ProHypGly-OH, Fmoc-Pro-OH, Fmoc-Gly-OH, and Fmoc-Ahx-OH were coupled according to protocol B, and compound 1 was coupled to the resin using manual protocol C. Compound 3 was cleaved from the solid support according to protocol D and purified according to protocol E using a gradient of 92% B to 72% B over 20 min, tR=14.0 min. After desalting and lyophilization, compound 3 was obtained as a white foam and stored at −20° C. in the dark. Analytical reversed-phase HPLC: 91% to 60% B over 20 min, tR=8.99 min; purity as determined by analytical HPLC using UV detection at 214 nm: >97%. HRMS(MALDI):[C 105 H 148 F2N 25 O 33 ] + m / z calculated for: 2325.0634; Found: 2325.0668 [M+H] + .

[0184] Example 12 Synthesis of 1-Ahx-[ProHypGly]7-NH2 (Compound 4)

[0185] [ka]

[0186] Compound 4 was synthesized on Rink amide ChemMatrix resin (approximately 0.5 mmol / g). The resin was swollen according to protocol A. Fmoc-ProHypGly-OH and Fmoc-Ahx-OH were coupled to the resin according to protocol B, and compound 1 was coupled to the resin using protocol C. Compound 4 was cleaved from the solid support according to protocol D and eluted with a gradient of 92% B to 60% B over 20 min, t R= 15.1 min. After desalting and lyophilization, compound 4 was obtained as a white foam and stored in the dark at -20 °C. Analytical reversed-phase HPLC: 91% to 45% B over 20 min, t R = 7.47 min; Purity as determined by analytical HPLC with UV detection at 214 nm: >97%. HRMS (MALDI): [C 105 H 147 F2N 24 O 33 ] + m / z calculated for: 2310.0525; Found: 2310.0537 [M+H] + .

[0187] Example 13a Synthesis of 1-Ahx-[ProProGly]7-NH2 (compound 5)

[0188] [ka]

[0189] Compound 5 was synthesized on Rink amide ChemMatrix resin (approximately 0.5 mmol / g). The resin was swollen according to protocol A. Fmoc-ProProGly-OH and Fmoc-Ahx-OH were coupled to the resin according to protocol B, and compound 1 was coupled to the resin using protocol C. Compound 5 was cleaved from the solid support according to protocol D and eluted with a gradient of 92% B to 62% B over 20 min, t R = 14.8 min. After desalting and lyophilization, compound 5 was obtained as a white foam and stored in the dark at -20 °C. Analytical reverse phase HPLC: 91% B to 60% B over 20 min, t R = 13.7 min; Purity as determined by analytical HPLC with UV detection at 214 nm: >99%. HRMS (MALDI): [C 105 H 147 F2N 24 O 26 ] + m / z for: 2198.0881; Measured value: 2198.0909 [M+H]+ .

[0190] Example 13b Synthesis of 1-Ahx-[ProProGly]7-NH2 (Compound 6)

[0191] [ka]

[0192] Compound 6 was synthesized on Rink amide ChemMatrix resin (approximately 0.5 mmol / g). The resin was swollen according to protocol A. Fmoc-(4S)aminooxyproline (Boc)-OH and Fmoc-Ahx-OH were coupled according to protocol B, and compound 1 was coupled to the resin using the manual protocol C. The peptide was cleaved from the solid support according to protocol D and purified according to protocol E with a gradient of 98% B to 50% B over 20 min, t R = 10.1 min. After desalting and lyophilization, compound 6 was obtained as a white foam and stored in the dark at -20 °C. Analytical reversed-phase HPLC: 98% B to 40% B over 20 min, t R = 8.1 min; Purity as determined by analytical HPLC with UV detection at 214 nm: >99%.

[0193] Example 14 CD spectra of compounds 3, 4, and 5 CD spectra (see FIG. 29) of 0.2 mM solutions of compounds 3, 4 and 5 in PBS buffer (pH=7.4) at 7° C. were recorded. The solutions were allowed to equilibrate for >24 hours at 5° C. before measurement. Spectra were recorded from 190 nm to 260 nm.

[0194] Example 15 T m Basic steps for determining value Thermal denaturation experiments were carried out using 0.2 mM solutions of compounds 3, 4, and 5 equilibrated in 50 mM PBS buffer (pH=7.4) at 5° C. for at least 24 hours. Samples were heated at a heating rate of 1° C. / 100 sec while monitoring the molar ellipticity at 224 nm. Experiments were repeated at least three times for each sample. The data obtained from the thermal denaturation experiments were fitted to an all-or-nothing transition in which three single strands combine into three helices as previously reported by J. Engel, HT Chen, DJ Prokop, H. Klump, Biopolymers 1997, 16, 601-622 and S. Frank, RA Kammerer, D. Mechling, T. Schulthess, R. Landwehr, J. Bann, Y. Guo, A. Lustig, HP Bachinger, J. Engel, J. Mol. Biol. 2001, 308, 1081-1089. The fitting was performed using Micromath Scientist 3.0 with initial values ​​of H=-500000 and Tm=40. The model used is shown below: / / 2-state model; IndVars:TEMP; DepVars:F, CD, K; Params:H, DEU, REFU, DEN, REFN, Tm; R=8.31; K=EXP(H / (R*(TEMP+273.15))*((TEMP+273.15) / (Tm+273.15)-1)-ln(0.75*0.0002^2)); P=1 / (3*K*(0.0002^2)); U=(-P / 2+(P^2 / 4+P^3 / 27)^(1 / 2))^(1 / 3); V=-(P / 2+(P^2 / 4+P^3 / 27)^(1 / 2)) ^(1 / 3); F= U+V+1; CDU=REFU+DEU*(TEMP+273.15); CDN=REFN+DEN*(TEMP+273.15); CD=F*(CDN-CDU)+CDU. The thermal denaturation curves of compounds 3, 4, and 5 are shown in FIG.

[0195] Example 16 General procedure for in vivo assay Animals: Mice were kept and fed according to Swiss guidelines and all animal experiments were approved by the local veterinary authorities (Kantonales Veterinaramt Zurich). All mice used in these experiments are on a C57BL / 6 background. Dermal administration of compounds 3, 4, and 5: 100 μM peptide solution was allowed to come to room temperature for 30 min. Eight-week-old male or female mice were anesthetized by intraperitoneal injection of ketamine / xylazine (100 mg ketamine / 5–10 mg xylazine per kg body weight), shaved, and 50 μL was injected intradermally into the dorsal skin. Mice were kept for a further 5 days and the skin was harvested for histological analysis.

[0196] This example demonstrates the successful design and synthesis of representative activity-based fluorescent compounds according to the present invention that allow real-time quantification of lysyl oxidase activity in fibrotic conditions. The activation of the compounds by the enzyme was confirmed in an ex vivo tissue homogenate model (see above) and demonstrated in vivo via conjugation to peptides that specifically and precisely target collagen and elastin undergoing real-time cross-linking and remodeling in the extracellular matrix. Furthermore, the compounds according to the present invention plus conjugated peptides show utility for the analysis of enzyme-mediated tissue remodeling in relevant models of fibrosis.

[0197] (Example 17) Synthesis of (5-((3-((Boc)amino)propyl)amino)-5-oxopentyl)triphenyl-phosphonium bromide

[0198] [ka]

[0199] (4-Carboxybutyl)triphenyl-phosphonium bromide (2.26 mmol, 1 g) and 1,1'-carbonyldiimidazole (2.26 mmol, 366 mg) were dissolved in anhydrous DMF (15 mL) and stirred at room temperature for 30 min. N-Boc-1,3-propanediamine (2.26 mmol, 393 mg) was added to the suspension and stirred overnight. The crude mixture was concentrated by rotary evaporation and purified by silica gel chromatography eluting with a gradient of 2-10% methanol in dichloromethane to give (5-((3-((Boc)amino)propyl)amino)-5-oxopentyl)triphenyl-phosphonium bromide (860 mg, 73%) as a white foam. 1 H NMR (400 MHz, CDCl3) δ 8.47 (t, J = 6.0 Hz, 1H), 7.85 - 7.65 (m, 15H), 5.54 (t, J = 5.7 Hz, 1H), 3.75 - 3.58 (m, 2H), 3.20 (q, J = 6.2 Hz, 2H), 3.00 (q, J = 6.2 Hz, 2H), 2.59 (t, J = 6.8 Hz, 2H), 1.93 (q, J = 7.0, 6.6 Hz, 2H), 1.68 - 1.55 (m, 4H), 1.39 (s, 9H). 13 C NMR (101 MHz, CDCl3) δ 173.30, 156.29, 135.23 (d, J = 3.1 Hz), 133.77 (d, J = 10.0 Hz), 130.60 (d, J = 12.6 Hz), 118.12 (d, J = 86.1 Hz), 78.62, 50.70, 36.53 (d, J = 138.0 Hz), 34.17, 29.49, 28.59, 26.21 (d, J = 17.0 Hz), 22.45 (d, J = 50.7 Hz), 21.09 (d, J = 4.2 Hz). 31 P NMR (162 MHz, CDCl3) δ 24.67. HRMS (ESI): m / z C 31 H 40 Calculated value of N2O3P: 519.2771 [M] + ; Measured value: 519.2764.

[0200] Example 17a Synthesis of (5-((3-ammoniopropyl)amino)-5-oxopentyl)triphenyl-phosphonium chloride

[0201] [ka]

[0202] (4-Carboxybutyl)triphenyl-phosphonium bromide (2.26 mmol, 1 g) and 1,1'-carbonyldiimidazole (2.26 mmol, 366 mg) were dissolved in anhydrous DMF (15 mL) and stirred at room temperature for 30 min. N-Boc-1,3-propanediamine (2.26 mmol, 393 mg) was added to the suspension and stirred overnight. The crude mixture was concentrated by rotary evaporation and purified by silica gel chromatography eluting with a gradient of 2-10% methanol in dichloromethane to give (5-((3-ammoniopropyl)amino)-5-oxopentyl)triphenyl-phosphonium chloride (860 mg, 73%) as a white solid. 1 H NMR (400 MHz, MeOD) δ 7.92 - 7.72 (m, 15H), 3.50 - 3.41 (m, 2H), 3.22 (t, J = 6.7 Hz, 2H), 2.91 (t, J = 7.4 Hz, 2H), 2.29 (t, J = 7.4 Hz, 2H), 1.89 - 1.77 (m, 4H), 1.76 - 1.64 (m, 2H). 13 C NMR (101 MHz, MeOD) δ 175.94, 136.31 (d, J = 3.1 Hz), 134.85 (d, J = 10.0 Hz), 131.55 (d, J = 12.6 Hz), 119.86 (d, J = 86.5 Hz), 38.27, 36.92, 35.74, 28.67, 27.63 (d, J = 17.4 Hz), 23.12 (d, J = 4.1 Hz), 22.55 (d, J = 51.6 Hz). 31P NMR (162 MHz, MeOD) δ 23.74. HRMS (ESI): m / z C 26 H 32 Calculated N2OP: 419.2247 [M] + ; Measured value: 419.2241.

[0203] Example 17b Synthesis of MRA_3103

[0204] [ka]

[0205] (5-((3-Ammoniopropyl)amino)-5-oxopentyl)triphenylphosphonium chloride (0.21 mmol, 89 mg) was dissolved in anhydrous DMF (1.4 mL) and cooled to 0 °C with stirring. N,N-Diisopropylethylamine (0.424 mmol, 74 μL) was added followed by MRA_3102 (0.19 mmol, 100 mg) and the solution was allowed to warm to room temperature and stirred overnight. The crude mixture was concentrated by rotary evaporation and purified by RP-HPLC with a gradient of 90-10% acetonitrile in water with 0.1% TFA. Fractions containing the product as determined by HPLC-MS were combined, flash frozen, and lyophilized to give MRA_3103 as a white solid (14 mg, 9%). 1H NMR (600 MHz, CD3CN) δ 7.88 - 7.82 (m, 3H), 7.72 - 7.65 (m, 12H), 7.37 (dd, J = 12.0, 2.2 Hz, 1H), 6.94 (t, J = 5.9 Hz, 1H), 6.77 (t, J = 5.6 Hz, 1H), 5.46 (s, 1H), 4.30 (t, J = 6.2 Hz, 2H), 3.52 (s, 2H), 3.24 - 3.16 (m, 4H), 3.06 (dq, J = 12.9, 6.3 Hz, 4H), 2.35 (s, 3H), 2.13 (t, J = 7.1 Hz, 2H), 1.89 (p, J = 6.5 Hz, 2H), 1.72 (p, J = 7.2 Hz, 2H), 1.60 (dq, J = 15.8, 7.7 Hz, 2H), 1.49 (p, J = 6.6 Hz, 2H), 1.38 (s, 9H). 19 F NMR (F-Hデカップリング) (376 MHz, CD3CN) δ -133.94 (d, J = 5.2 Hz), -150.46 (d, J = 6.0 Hz). 19 F NMR (376 MHz, CD3CN) δ -133.94 (dd, J = 12.7, 5.8 Hz), -150.46 (d, J = 6.3 Hz). 31 P NMR (162 MHz, CD3CN) δ 23.50. 13C NMR (151 MHz, CD3CN) δ 173.51, 170.28, 161.20, 157.08, 152.47 (dd, J = 242.9, 4.5 Hz), 149.91, 143.78 (dd, J = 248.9, 6.2 Hz), 139.49 (dd, J = 10.1, 1.9 Hz), 138.66 (dd, J = 16.1, 11.0 Hz), 136.11 (d, J = 3.1 Hz), 134.61 (d, J = 10.0 Hz), 131.26 (d, J = 12.6 Hz), 121.60, 119.56, 118.99, 116.69 (d, J = 9.2 Hz), 107.71 (dd, J = 22.7, 3.5 Hz), 79.27, 73.85, 37.85, 37.21, 36.93, 35.68, 35.49, 31.20, 30.07, 28.60, 27.05 (d, J = 17.3 Hz), 22.44 (d, J = 4.1 Hz), 22.40 (d, J = 51.8 Hz), 16.10. HRMS (ESI): m / z C 46 H 53 Calculated value of F2N3O7P: 828.3584 [M] + ; Measured value: 828.3582.

[0206] (Example 18) A general protocol for the synthesis of mitochondrial targeting peptides Protocol A - Basic swelling procedure: Prior to automated peptide synthesis, the resin was swelled in CH2Cl2 with shaking for 15 min, then drained, washed with DMF (3x6 mL), and drained again.

[0207] Protocol B - General protocol for automated peptide synthesis: For automated peptide synthesis a SyroI peptide synthesizer (Biotage, Sweden) was used. Couplings were carried out using either the appropriate Fmoc-protected amino acid or the trimer Fmoc-Pro-Hyp-Gly-OH. After swelling of the resin with DMF in the synthesizer, i-Pr2NEt (9 eq., as a 3 M solution in NMP (N-methyl-2-pyrrolidone)), HATU (3 eq., 0.5 M in DMF) and Fmoc-amino acid / Fmoc-tripeptide (3 eq., 0.5 M in DMF) were added to the resin. The mixture was reacted for 1 min intervals, stirred for 5 min, left to stand for 30 min (2x) and then washed with DMF (5x). Fmoc-deprotection was carried out by adding a solution of 40% (v / v) piperidine in DMF and reacting for 1 min. This step was repeated 4 times. The resin was then washed with DMF (5x). The tripeptide coupling and Fmoc deprotection were repeated until the desired peptide was obtained. No acylation (capping) was performed in the automated synthesis of CMP.

[0208] Protocol C - N-Terminal Functionalization of Resin with Compound 1 Functionalization was performed manually on solid support-bound peptides at room temperature. Compound 1 (2.0 equiv.), HATU (1.9 equiv.) and i-Pr2NEt (4 equiv.) were dissolved in DMF (1-2 mL). After 5 min preactivation, the coupling mixture was added to the resin and stirred for 1-2 h. The resin was washed with CHCl2 (3×), DMF (3×), CHCl2 (3×), and petroleum ether (2×). The reaction was monitored by a qualitative color test of the beads or by LC-MS after test cleavage (see protocol E).

[0209] Protocol D - Cleavage from the resin The resin was shaken in a mixture of TFA / (i-Pr2)3Si-H / H2O (92.5:2.5:2.5) for 1 h and washed with pure TFA (2x). The peptide in solution was collected by filtration into a conical flask. Ice-cold Et2O was added, giving the peptide as a white precipitate. The solid was isolated by centrifugation followed by decanting. The solid was suspended in Et2O, sonicated, centrifuged again, and the supernatant was decanted. The remaining white solid was dissolved in water / CH3CN, frozen, and lyophilized to give a white foam.

[0210] Protocol E - Purification and analysis by RP HPLC For semi-preparative HPLC, HO (A) and CHCN (B) containing 1% CHCN and 0.1% TFA were used as eluents. Flow rates of 6 mL / min were used for semi-preparative HPLC, 1 mL / min for analytical HPLC, and 0.5 mL / min for LC-MS. After purification by semi-preparative HPLC, all collected fractions were analyzed by analytical HPLC or LC-MS, and only pure fractions were combined. For analytical HPLC, CHCN (A) and HO (B) containing 1% CHCN and 0.1% TFA were used as eluents. The amine containing CMP was desalted on a VariPure cartridge and then lyophilized. Preparative column: Reprosil Gold 120 C18, 150 x 10 mm. Analytical column: Phenomenex, Jupiter 5 μm, 300 Å, 250 x 4.6 mm. LC-MS: Reprosil Gold C18, 125 x 3 mm.

[0211] Example 18a Synthesis of 1-Ahx-[Char]3-NH2

[0212] [ka]

[0213] Peptides were synthesized on Rink amide resin (approximately 0.5 mmol / g). The resin was swollen according to protocol A. Fmoc-D-Arginine(Boc)-OH, Fmoc-Cha-OH, and Fmoc-Ahx-OH were coupled to the resin according to protocol B, and compound 1 was coupled to the resin using the manual protocol C. The peptide was cleaved from the solid support according to protocol D and eluted with a gradient of 15% B to 40% B over 20 min, t R = 11.4 min. After desalting and lyophilization, the peptide was obtained as a white foam and stored in the dark at -20°C. Analytical reversed-phase HPLC: 90% to 10% B over 20 min, t R = 8.3 min; Purity as determined by analytical HPLC with UV detection at 214 nm: >99%. HRMS (MALDI): [C 66 H 110 F2N 18 O 11 ] 2+ m / z calculated for: 684.4279; Found: 684.4269 [M+H] 2+ .

[0214] Example 18b Synthesis of MRA_3069-Ahx-(ChaR)3-NH2

[0215] [ka]

[0216] Peptides were synthesized on Rink amide resin (approximately 0.5 mmol / g). The resin was swollen according to protocol A. Fmoc-D-Arginine(Boc)-OH, Fmoc-Cha-OH, and Fmoc-Ahx-OH were coupled to the resin according to protocol B, and MRA_3069 was coupled to the resin using the manual protocol C. The peptide was cleaved from the solid support according to protocol D and eluted with a gradient of 15% to 48% B over 20 min, t R= 15.0 min. After desalting and lyophilization, the peptide was obtained as a white foam and stored in the dark at -20°C. Analytical reversed-phase HPLC: 10% to 90% B over 20 min, t R = 8.7 min; Purity as determined by analytical HPLC with UV detection at 214 nm: >95%. HRMS (MALDI): [C 66 H 110 F2N 18 O 11 ] 2+ m / z calculated for: 691.4358; Found: 691.4343 [M+H] 2+ .

[0217] (Example 18c) Synthesis of PB-Ahx-(ChaR)3-NH2

[0218] [ka]

[0219] Peptides were synthesized on Rink amide resin (approximately 0.5 mmol / g). The resin was swollen according to protocol A. Fmoc-D-Arginine(Boc)-OH, Fmoc-Cha-OH, and Fmoc-Ahx-OH were coupled to the resin according to protocol B. Pacific Blue (PB) NHS was coupled to the resin directly using manual protocol C but without the addition of HATU. Peptides were cleaved from the solid support according to protocol D and eluted with a gradient of 15% B to 60% B over 20 min, t R = 13.9 min. After desalting and lyophilization, the peptide was obtained as a white foam and stored in the dark at -20°C. Analytical reversed-phase HPLC: 10% to 90% B over 20 min, t R = 9.56 min; Purity as determined by analytical HPLC with UV detection at 214 nm: >99%. HRMS (MALDI): [C 63 H 103 F2N 17 O 11 ] 2+m / z calculated for: 655.899; Found: 655.8979 [M+H] 2+ .

[0220] (Example 19) A cellular assay to determine monamine oxidase activity from MAO A / MAO B Cell culture: MCF-7 and SY5Y cells were obtained from the Health Protection Agency (www.HPA.org.uk) or the American Type Culture Collection. Cells were grown in a humidified 5% CO2 atmosphere at 37°C using Kaighn's modification of Ham's F12 (F-12KTM) supplemented with L-glutamine (4 mM), penicillin (100 U / mL penicillin), streptomycin (100 μg / mL), and 10% fetal serum (FCS) superior (standardized). Culture media DMEM high glucose, F-12KTM, L-glutamine (200 mM), penicillin (10.000 U / mL), streptomycin (10 mg / mL), and trypan blue solution were purchased from Sigma, Invitrogen, ATCC, or BioConcept. Trypsin-EDTA (0.05% / 0.02%) in Ca2+- and Mg2+-deficient phosphate buffered saline (PBS) (1:250) was purchased from Amimed. PBS (pH 7.4) was purchased from Invitrogen. FCS Superior was purchased from Oxoid AG and Biochrom AG. Cell culture flasks and serological pipettes were purchased from BD Biosciences and Sarstedt. Ethylenediaminetetraacetic acid (EDTA) was purchased from Sigma-Aldrich. Hoechst 33342 was purchased from Invitrogen. MitoTracker Red mitochondrial staining dye and live cell imaging solution were purchased from ThermoFisher.

[0221] Confocal microscopy: Fluorescent images of cells were collected using a Nikon Eclipse T1 microscope equipped with a Yokogawa spinning disk confocal scanner unit CSU-W1-T2, two sCMOS cameras (Orca Flash 4.0 V2), and a LUDL BioPrecision2 stage with piezo focus. Blue channel emission was filtered with a 450 / 50 bandpass filter, green channel emission was filtered with a 525 / 50 bandpass filter, and far-red channel emission was filtered with a 700 / 75 bandpass filter. Fluorescent images were acquired using a 100x1.49 magnification CFI Apo TIRF oil immersion objective. The microscope was operated using VisiVIEW (Metamorph).

[0222] Prior to measurements, cells were seeded at a density of 10000 cells / well in ibidi 8-well plates and grown for 1 day in DMEM (10% FCS) at 37°C. Afterwards, cells were washed with PBS and untreated DMEM was added. Compounds were added to cells as stock solutions in PBS to reach a final concentration of 10 mM. Cells were incubated for 2 h at 37°C, washed with PBS, and incubated with MitoTracker Red for 5 min at 37°C based on the manufacturer's recommended procedure. Cells were washed with PBS and live cell imaging solution (200 μL) was added. Live cells were then immediately examined under a confocal microscope (Vistron Spinning Disk). A 405 nm laser line was used for detection of coumarin activation and a 561 nm laser line for MitoTracker.

[0223] (Example 20) Biochemical and histological analysis of skin treated with compounds 3-6 Skin tissues used for histology were harvested and immediately embedded and frozen in Tissue Freezing Medium® (Leica Biosystems, Wetzlar, Germany). Tissue sections (7 μM) were fixed for 10 min at −20° C. using ice-cold acetone and stained with propidium iodide to visualize nuclei. Images were taken using a 20× objective. Immunofluorescence analysis of collagens used antibodies against collagen I (Southern Biotech, Birmingham, AL, USA; 1310-01) or collagen III (Abcam, Cambridge, UK, ab7778). Fixed tissue sections were incubated with 1% bovine serum albumin (BSA) in PBS for 30 min, followed by incubation with collagen I (1:400 dilution in 1% BSA) or collagen III (1:800 dilution in 1% BSA) antibodies for 60 min at room temperature. After washing (3 x 5 min PBS), secondary antibodies (anti-rabbit AF488 (711-547-003) or anti-goat AF488 (705-545-147), Jackson ImmunoResearch) were incubated on the tissues for 30 min at room temperature (both diluted 1:400 in 1% BSA) and nuclei were visualized using propidium iodide. All images were taken using a 20x objective.

[0224] Example 21 Biochemical and histological analysis of tissue sections treated with compounds 3-6 Skin tissues used for histology were harvested and immediately embedded and frozen in Tissue Freezing Medium® (Leica Biosystems, Wetzlar, Germany). Tissue sections (7 uM) were processed by pipetting 50 μL of each compound (compounds 3, 4, 5, or 6) at 100 μM in PBS to completely cover the surface of the section, incubating at 37° C. for 4 hours in the dark, and washing twice with PBS. Sections were fixed with ice-cold acetone at −20° C. for 10 minutes and stained with propidium iodide to visualize nuclei. Images were taken using a 20x objective.

[0225] For collagen immunofluorescence analysis, antibodies against collagen I (Southern Biotech, 1310-01) or collagen III (Abcam, ab7778) were used. Fixed tissue sections were incubated with collagen I or collagen III antibodies for 60 min at room temperature. After washing, secondary antibodies (anti-rabbit AF488 (711-547-003) or anti-goat AF488 (705-545-147), Jackson ImmunoResearch) were incubated on the tissue for 30 min at room temperature and nuclei were visualized using propidium iodide. All images were taken using a 20x objective.

Claims

1. Formula I 【Chemistry 1】 and salts and solvates thereof, and physiologically acceptable salts and solvates thereof, a is an integer from 0 to 3; b is 0; A is, 【Chemistry 2】 is a structure selected from the group consisting of: R3 is 【Chemistry 3】 and; R 4 is straight or branched chain, substituted or unsubstituted (C 1-10 ) alkyl, methyl, ethyl, and propyl; L is absent or - straight or branched chain, substituted or unsubstituted (C 1-10 ) alkyl, propyl, butyl and pentyl; - 【Chemistry 4】 (wherein c and d are independently selected from 1, 2, 3, 4, and 5); - linear or branched, substituted or unsubstituted (C 1-10 ) alkyl ether, PEG chains having 1 to 10 ethylene oxide moieties; is a linker selected from the group consisting of: X is selected from the group consisting of O, N, and S; Z is O; R9 is - Hydrogen, N-maleimidyl; - straight or branched chain, substituted or unsubstituted (C 1-10 ) alkyl, methyl, ethyl, propyl, (C 2-10 ) alkenyl and (C 2-10 ) alkynyl; - linear or branched, substituted or unsubstituted (C 1-20 ) alkyl ether, PEG chains having 1 to 10 ethylene oxide moieties; - triphenylphosphine connected through phosphorus; - proteinogenic and non-proteinogenic amino acids, lysine, proline, glycine, 4-hydroxyproline, 4-aminoproline, and 4-aminooxyproline; - peptides, peptides containing 1-2000, 1-100 or 1-25 amino acids, peptides containing 1-10 [proline]-[4-hydroxyproline]-[glycine] units; - Collagen peptides, fibronectin peptides, fibrillin peptides, elastin peptides and RGD (arginylglycylaspartic acid) peptides; - antibodies, anti-collagen antibodies, anti-elastin antibodies, anti-fibronectin antibodies and anti-fibrillin antibodies; and - azide, -NH2, -OH and -SH2; selected from the group consisting of; R 9 '' is -Azide, -SR 10 , 【Chemistry 5】 selected from the group consisting of; R 10 is selected from the group consisting of straight or branched chain, substituted or unsubstituted (C 1-10 ) alkyl, methyl, ethyl, propyl, (C 2-10 ) alkenyl, and (C 2-10 ) alkynyl; R5 is - hydrogen and -OH; and - the halogens F, Cl, Br and I; selected from the group consisting of; R 6 and R 8 are independently selected from the group consisting of hydrogen, halogen, F, Cl, Br, and I; and R7 is - Hydrogen; - straight or branched chain, substituted or unsubstituted (C 1-10 ) alkyl, methyl, ethyl and propyl; and - Boc, Fmoc and Cbz, or both R 7 together with the N to which they are attached form an azide; The compound, as well as its salts and solvates, and its physiologically acceptable salts and solvates.

2. The compound according to claim 1, which is a compound according to formula II 【Chemistry 6】 (In the formula, a is 1 or 2; b is 0; A is, 【Chemistry 7】 is a structure selected from the group consisting of: R 3 teeth, 【Chemistry 8】 and; R 4 is linear or branched, substituted or unsubstituted (C 1～10 ) alkyl, methyl, ethyl and propyl; L does not exist or 【Chemistry 9】 (wherein c and d are independently selected from 1, 2, 3, 4, and 5); X is selected from the group consisting of O and N; Z is O; R 9 teeth, - Hydrogen, N-maleimidyl; - Linear or branched, substituted or unsubstituted (C 1～10 ) alkyl, methyl, ethyl and propyl; - triphenylphosphine connected through phosphorus; - proteinogenic and non-proteinogenic amino acids, lysine, proline, glycine, 4-hydroxyproline, 4-aminoproline, and 4-aminooxyproline; - collagen peptides, fibronectin peptides, fibrillin peptides, elastin peptides and RGD (arginylglycylaspartic acid) peptides; and -Azide, -NH 2 , -OH and -SH 2 , selected from the group consisting of; R 9'' teeth, - Azide, 【Chemistry 10】 is selected from the group consisting of: R 7 teeth, - hydrogen; and - Linear or branched, substituted or unsubstituted (C 1～10 ) alkyl, methyl, ethyl and propyl; (each independently selected from the group consisting of:

3. 3. A pharmaceutical composition for use in a method for the diagnosis of an amine oxidase-related disorder and / or a collagen- or elastin-related disorder, comprising a compound according to claim 1 or 2 or a pharma- ceutically acceptable salt thereof.

4. The method comprises the steps of: (a) providing a compound according to claim 1 or 2; (b) contacting a patient, mammal, or patient sample in need thereof with an effective amount of the compound of step (a), wherein the effective amount is an amount effective to detect amine oxidase activity; (c) measuring the fluorescence of the patient or a patient sample; and (d) determining the activity and / or location of the amine oxidase based on the fluorescence measured in step (c), 4. The pharmaceutical composition according to claim 3.

5. 3. Use of a compound according to claim 1 or 2 in the manufacture of a medicament for use in the in vitro or ex vivo diagnosis of an amine oxidase related disorder and / or a collagen or elastin related disorder.

6. 3. Use of a compound according to claim 1 or 2 in the in vitro or ex vivo detection of amine oxidase activity.

7. The amine oxidase-associated disease is (i) lysyl oxidase (LOX), or a lysyl oxidase having at least 80%, 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 1-3; (ii) lysyl oxidase homolog 1, 2, 3, or 4 (LOXL1, 2, 3, or 4); (iii) a primary amine oxidase, AOC2, or AOC3; (iv) diamine oxidase, or AOC1, and (v) a monoamine oxidase having at least 80%, 85%, 90% or 95% sequence identity to a monoamine oxidase, MAOA, MAOB, or at least one of SEQ ID NOs: 4-7; The pharmaceutical composition according to claim 3 or 4, wherein the disease is an amine oxidase-associated disease selected from the group consisting of:

8. The amine oxidase-associated disease is (i) lysyl oxidase (LOX), or a lysyl oxidase having at least 80%, 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 1-3; (ii) lysyl oxidase homolog 1, 2, 3, or 4 (LOXL1, 2, 3, or 4); (iii) a primary amine oxidase, AOC2, or AOC3; (iv) diamine oxidase, or AOC1, and (v) a monoamine oxidase having at least 80%, 85%, 90% or 95% sequence identity to a monoamine oxidase, MAOA, MAOB, or at least one of SEQ ID NOs: 4-7; 6. The use according to claim 5, wherein the disease is an amine oxidase-associated disease selected from the group consisting of:

9. 8. The pharmaceutical composition of claim 3, 4 or 7, wherein the disease is selected from the group consisting of fibrosis, pulmonary fibrosis, hepatic fibrosis, cardiomyopathy, Occupative Horn's Syndrome (OHS), Menkes syndrome, myocardial ischemia, atherosclerosis, scleroderma, keloid disorders, liver cirrhosis, Alzheimer's and non-Alzheimer's dementia, Wilson's disease, primary biliary cirrhosis, chronic venous insufficiency, pseudoexfoliation syndrome, glaucoma, pelvic organ prolapse, endometriosis, intracranial aneurysm, heart failure, and cancer including tumor metastasis.

10. 9. The use according to claim 5 or 8, wherein the disease is selected from the group consisting of fibrosis, pulmonary fibrosis, hepatic fibrosis, cardiomyopathy, Occupatory Horn's Syndrome (OHS), Menkes syndrome, myocardial ischemia, atherosclerosis, scleroderma, keloid disorders, liver cirrhosis, Alzheimer's and non-Alzheimer's dementia, Wilson's disease, primary biliary cirrhosis, chronic venous insufficiency, pseudoexfoliation syndrome, glaucoma, pelvic organ prolapse, endometriosis, intracranial aneurysm, heart failure, and cancer, including tumor metastasis.

11. 10. The pharmaceutical composition of claim 9, wherein the cancer is selected from the group consisting of colorectal cancer, bladder cancer, pancreatic cancer, breast cancer, head and neck squamous cell carcinoma (HNSCC), laryngeal cancer, lung cancer, gastric cancer, prostate cancer, esophageal squamous cell carcinoma, endometrial cancer, testicular seminoma cancer, hepatocellular carcinoma, renal clear cell carcinoma, and basal and squamous cell carcinoma.

12. 11. The use of claim 10, wherein the cancer is selected from the group consisting of colorectal cancer, bladder cancer, pancreatic cancer, breast cancer, head and neck squamous cell carcinoma (HNSCC), laryngeal cancer, lung cancer, gastric cancer, prostate cancer, esophageal squamous cell carcinoma, endometrial cancer, testicular seminoma cancer, hepatocellular carcinoma, renal clear cell carcinoma, and basal and squamous cell carcinoma.

13. The following steps: (a) providing a compound according to claim 1 or 2; (b) providing and contacting a cell, tissue or body fluid with a compound of step (a), in vitro or ex vivo, under conditions that permit amine oxidase activity; (c) measuring the fluorescence in the cells, tissues, or body fluids; and (d) determining the activity and / or location of the amine oxidase based on the fluorescence measured in step (c).

14. A composition for use in an in vivo method for the detection of amine oxidase activity, comprising a compound according to claim 1 or 2, The method comprises the steps of: (a) providing a compound according to claim 1 or 2; (b) providing and contacting in vivo a cell, tissue or body fluid with the compound of step (a) under conditions that permit amine oxidase activity; (c) measuring the fluorescence in the cells, tissues, or body fluids; and (d) determining the activity and / or location of the amine oxidase based on the fluorescence measured in step (c).

15. The amine oxidase is (i) lysyl oxidase (LOX), or a lysyl oxidase having at least 80%, 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 1-3; (ii) lysyl oxidase homolog 1, 2, 3, or 4 (LOXL1, 2, 3, or 4); (iii) a primary amine oxidase, AOC2, or AOC3; (iv) diamine oxidase, or AOC1, and (v) a monoamine oxidase having at least 80%, 85%, 90% or 95% sequence identity to a monoamine oxidase, MAOA, MAOB, or at least one of SEQ ID NOs: 4-7; 14. The method of claim 13, selected from the group consisting of:

16. The amine oxidase is (i) lysyl oxidase (LOX), or a lysyl oxidase having at least 80%, 85%, 90% or 95% sequence identity to at least one of SEQ ID NOs: 1-3; (ii) lysyl oxidase homolog 1, 2, 3, or 4 (LOXL1, 2, 3, or 4); (iii) a primary amine oxidase, AOC2, or AOC3; (iv) diamine oxidase, or AOC1, and (v) a monoamine oxidase having at least 80%, 85%, 90% or 95% sequence identity to a monoamine oxidase, MAOA, MAOB, or at least one of SEQ ID NOs: 4-7; 15. The composition of claim 14, selected from the group consisting of:

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