COMPOUND - DIAGNOSTIC MARKER OF KIDNEY CANCER, METHOD OF DETECTING ENZYME ACTIVITY, METHOD OF DIAGNOSIS OF KIDNEY CANCER, KIT CONTAINING THE COMPOUND, USE VARIANTS OF THE COMPOUND AND METHOD OF TREATMENT OF KIDNEY CANCER

Claims

1. Compound of formula 1 X1 1 -Leu 2 -Pro 3 -Gly 4 -Thr 5 -X2 6 (formula 1), where X1 contains or consists of a molecule C1, and X2 contains or consists of a molecule C2, wherein the said pair of molecules C1 and C2 represents a pair of a fluorescence donor and a fluorescence acceptor, and wherein the said compound undergoes enzymatic cleavage into fragments X1-Leu-Pro-Gly-Thr-OH (fragment 1) and X2 (fragment 2) with the generation of a measurable optical signal upon spatial separation of the molecules C1 and C2.

2. The compound according to claim 1, characterized in that said compound undergoes hydrolytic, preferably proteolytic cleavage.

3. The compound according to claim 1 or 2, characterized in that in said compound the pair of molecules C1 and C2 is selected from the group consisting of: 2-aminobenzoic acid (ABZ) / 5-amino-2-nitrobenzoic acid (ANB), (ABZ) / pNA, ABZ / ANB-NH2, ABZ / DNP, ABZ / EDDNP, EDANS / DABCYL, TAM / DANSYL, ABZ / Tyr(3-NO2), preferably said pair of C1 and C2 is ABZ / pNA or ABZ / ANB-NH2.

4. A compound according to any one of claims 1-3, characterized in that said compound is a compound of formula 2 ABZ-Leu-Pro-Gly-Thr-ANB-NH2(formula 2) or a compound of formula 3 ABZ-Leu-Pro-Gly-Thr-pNA (formula 3).

5. The compound according to claim 4, characterized in that said compound undergoes hydrolytic cleavage to form the following fragment 1: ABZ-Leu-Pro-Gly-Thr-OH and fragment 2: ANB-NH2.

6. A method for in vitro detection of enzymatic activity present in a biological fluid of a subject, in particular originating from kidney cancer cells, comprising: a) bringing a sample of the said biological fluid into contact with a compound of formula 1 X1 1 -Leu 2 -Pro 3 -Gly 4 -Thr 5 -X2 6 (formula 1), where X1 contains or consists of a molecule C1, and X2 contains or consists of a molecule C2, wherein the said pair of molecules C1 and C2 represents a pair of a fluorescence donor and a fluorescence acceptor, and wherein said compound undergoes enzymatic cleavage into fragments X1-Leu-Pro-Gly-Thr-OH (fragment 1) and X2 (fragment 2), and b) detection of a measurable optical signal generated by spatial separation of C1 and C2 molecules.

7. The in vitro method according to claim 6, characterized in that said enzymatic activity is hydrolytic activity, preferably proteolytic activity.

8. The in vitro method according to claim 6 or 7, characterized in that the compound used as the said compound is a compound of formula 2 ABZ-Leu-Pro-Gly-Thr-ANB-NH2(formula 2) or a compound of formula 3 ABZ-Leu-Pro-Gly-Thr-pNA (formula 3).

9. The method according to any one of paragraphs 6-8, characterized in that urine, preferably human urine, is used as the said biological fluid.

10. A method for in vitro diagnostics of kidney cancer, wherein the presence or absence of kidney cancer in a subject is detected by measuring an enzymatic activity specific to kidney cancer in a sample of biological fluid from the subject being examined, and wherein the absence of said enzymatic activity indicates the absence of kidney cancer, and the presence of said enzymatic activity indicates the presence of kidney cancer, and wherein the measurement of said enzymatic activity is performed using a compound of formula 1 X1 1 -Leu 2 -Pro 3 -Gly 4 -Thr 5 -X2 6 (formula 1), where X1 contains or consists of a molecule of C1, and X2 contains or consists of a molecule of C2, wherein the said pair of molecules C1 and C2 represents a pair of a fluorescence donor and a fluorescence acceptor, and in this case the said compound undergoes enzymatic cleavage into fragments X1-Leu-Pro-Gly-Thr-OH (fragment 1) and X2 (fragment 2) with the generation of a measurable optical signal upon spatial separation of the molecules C1 and C2.

11. The method according to claim 10, characterized in that said detection of enzymatic activity is carried out by the method according to any of claims 6-9.

12. The method according to any one of claims 10, 11, characterized in that said sample of biological fluid is incubated with said compound in a measurement buffer with a neutral or alkaline pH, preferably physiological, in a range of sample to measurement buffer ratio from 1:2 to 1:10, preferably 1:

5.

13. The method according to any one of paragraphs 10-12, characterized in that the said compound is used in a concentration of 0.1-10 mg / ml, in particular 0.25-7.5 mg / ml.

14. The method according to paragraphs 10-13, characterized in that the specified compound is a compound of formula 2 ABZ-Leu-Pro-Gly-Thr-ANB-NH2(formula 2) or a compound of formula 3 ABZ-Leu-Pro-Gly-Thr-pNA (formula 3).

15. The method according to any one of paragraphs 10-14, characterized in that the said sample is a urine sample, preferably human urine.

16. The method according to any one of claims 10-15, characterized in that measuring said enzymatic activity comprises measuring the absorption intensity in the range of 300-500 nm, preferably 380-430 nm, in particular 405 nm, for 40-60 minutes in the temperature range of 25-40°C, preferably 36-38°C.

17. A kit comprising a compound according to any one of claims 1-5 and a measurement buffer.

18. The kit according to claim 17, characterized in that said compound is a compound of formula 2 ABZ-Leu-Pro-Gly-Thr-ANB-NH2(formula 2) or a compound of formula 3 ABZ-Leu-Pro-Gly-Thr-pNA (formula 3).

19. Use of a compound according to any one of claims 1 to 5 for detecting enzymatic activity specific for kidney cancer.

20. A compound according to any one of claims 1-5 for use in diagnosing kidney cancer.

21. A compound for use according to claim 20, wherein said diagnosing kidney cancer comprises detecting primary kidney cancer, detecting minimal residual disease after surgical resection of kidney cancer, and / or detecting recurrence of kidney cancer.

22. A compound for use according to any one of claims 20, 21, wherein said compound is a compound of formula 2 ABZ-Leu-Pro-Gly-Thr-ANB-NH2(formula 2) or a compound of formula 3 ABZ-Leu-Pro-Gly-Thr-pNA (formula 3).

23. A compound according to any one of claims 1 to 5 for use as a diagnostic marker for detecting kidney cancer.

24. A compound for use according to claim 23, wherein said compound is a compound of formula 2 ABZ-Leu-Pro-Gly-Thr-ANB-NH2(formula 2) or a compound of formula 3 ABZ-Leu-Pro-Gly-Thr-pNA (formula 3).

25. A compound according to any one of claims 1 to 5 for use in a method for treating kidney cancer, wherein a) detecting the presence of an enzymatic activity specific for kidney cancer by the method of any one of paragraphs 6-9 in a sample of a biological fluid of the subject being examined, and (b) the said subject is treated for kidney cancer if the said sample is found to contain the said enzymatic activity.

26. A compound for use according to claim 25, wherein after completion of treatment according to point b), said enzymatic activity specific to kidney cancer is monitored at predetermined time intervals.

27. A compound for use according to claim 25 or 26, characterized in that said sample is a urine sample, preferably human urine.

28. A compound for use according to any one of paragraphs 25-27, wherein said compound is a compound of formula 2 ABZ-Leu-Pro-Gly-Thr-ANB-NH2(formula 2) or a compound of formula 3 ABZ-Leu-Pro-Gly-Thr-pNA (formula 3).