3,6,9, 15-tetraaza-bicyclo [9.3.1]pentadeca-1(14), 11(15), 12-triene based compounds and their application as ligands of essential metal ion based MRI and 52MN based PET contrast agents

Abstract

A compound of the following general formula (I), and isomers, physiologically acceptable salts, and complexes thereof, for use as a contrast agent in diagnostic imaging: ##STR00001##

Claims

1. A compound of general formula (I) ##STR00042## wherein R.sub.a is a Q group selected from: ##STR00043## where R.sub.2 is a H atom, carboxyl or alkyl-carbonyl; ##STR00044## R.sub.b is H-atom or alkyl group; X is H-atom; and Y means NH, >NC(O)-Q group where Q is defined as above, or >NCH2-Z group where Z selected from: ##STR00045## where z=0-18, ##STR00046## where z=0-18 and R.sub.2 is H atom, alkyl, or (CH.sub.2).sub.nCOOH group, where n=1-10, ##STR00047## where R.sub.2b is H atom, CH.sub.3, OCH.sub.3, CF.sub.3, COOH, COON(CO).sub.2(CH.sub.2).sub.2, NO.sub.2, NH.sub.2, or NCS group, ##STR00048## where z=0-18 and R.sub.2c is H atom, NO.sub.2, NH.sub.2, or NCS group, ##STR00049## where z=0-18, and x=1-5, ##STR00050## where z=1-5, and x=1-5, R.sub.2d is H atom, CH.sub.3, OCH.sub.3, CF.sub.3, NO.sub.2, NH.sub.2, NCS COOH, or COON(CO).sub.2(CH.sub.2).sub.2 group.

2. The compound according to claim 1, wherein the compound is 3,9-bis[2-oxo-2-(piperidine-1-yl)ethyl]-3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene.

3. A complex comprising the compound of claim 1 and a metal selected from Mn(II), Fe(II), Fe(III), Co(II), or Ni(II).

4. A complex comprising the compound of claim 2 and a metal selected from Mn(II), Fe(II), Fe(III), Co(II), or Ni(II).

5. A method of imaging, comprising: applying a compound according to claim 1 as a contrast agent in diagnostic imaging, and performing diagnostic imaging.

6. A method of imaging, comprising: applying a compound according to claim 2 as a contrast agent in diagnostic imaging, and performing diagnostic imaging.

7. A contrast agent kit comprising: a compound according to claim 1.

8. A contrast agent kit comprising: a compound according to claim 2.

9. A method of imaging, comprising: applying a complex according to claim 3 as a contrast agent in diagnostic imaging, and performing diagnostic imaging.

10. A contrast agent kit comprising: a complex according to claim 3.

11. A compound of general formula (I) ##STR00051## wherein R.sub.a is OH group; R.sub.b is H-atom or alkyl group; X is CH.sub.3, COOH, OH, OCH.sub.3, alkoxy-, NO.sub.2, NH.sub.2, NCS, NHS-activated ester, alkyl, or aryl, wherein alkyl and aryl may be optionally substituted with hydroxyl, hydroxyalkyl, nitro, amino, or isothiocyanate group; and Y means NH, >NC(O)-Q group, or >NCH.sub.2Z group, where Q group is selected from: ##STR00052## where R.sub.2 is a H atom, carboxyl or alkyl-carbonyl; ##STR00053## where Z group is selected from: ##STR00054## where z=0-18, ##STR00055## where z=0-18 and R.sub.2 is H atom, alkyl, or (CH.sub.2).sub.nCOOH where n=1-10, ##STR00056## where R.sub.2b is H atom, CH.sub.3, OCH.sub.3, CF.sub.3, COOH, COON(CO).sub.2(CH.sub.2).sub.2, NO.sub.2, NH.sub.2, or NCS group, ##STR00057## where z=0-18 and R.sub.2c is H atom, NO.sub.2, NH.sub.2, or NCS group, ##STR00058## where z=0-18, and x=1-5, ##STR00059## where z=1-5, and x=1-5, R.sub.2d is H atom, CH.sub.3, OCH.sub.3, CF.sub.3, NO.sub.2, NH.sub.2, NCSCOOH, or COON(CO).sub.2(CH.sub.2).sub.2 group.

12. A complex comprising the compound of claim 11 and a metal selected from Mn(II), Fe(II), Fe(III), Co(II), or Ni(II).

13. A method of imaging, comprising: applying a compound according to claim 11 as a contrast agent in diagnostic imaging, and performing diagnostic imaging.

14. A contrast agent kit comprising: a compound according to claim 11.

15. A method of imaging, comprising: applying a complex according to claim 12 as a contrast agent in diagnostic imaging, and performing diagnostic imaging.

16. A contrast agent kit comprising: a complex according to claim 12.

Description

EXAMPLE 1

Synthesis of tPC2AM.SUP.Pyp

(1) a.) 2-bromo-1-(piperidine-1-yl)ethanone: Bromoacetyl bromide (3.56 g, 17.6 mmol, 1.55 ml, 1.5 equivalent), dry CH.sub.2Cl.sub.2 (50 ml) and K.sub.3PO.sub.4 (6.41 g, 30.2 mmol, 2.5 equ.) was mixed in a flask of 250 ml under N.sub.2 atmosphere. Piperidine (1.00 g, 11.7 mmol, 1.0 equivalent) was dissolved in dry CH.sub.2Cl.sub.2 (20 ml) and was added dropwise to dichloromethane solution of bromoacetyl bromide at 0 C. in 30 minutes, then the reaction mixture was stirred for additional 12 hours at room temperature under N.sub.2 atmosphere before the aqueous HCl solution (0.5 M, 30 ml) was added to the reaction mixture. After the addition of HCl solution, the reaction mixture was stirred for additional 5 minutes, then the two phases were separated using a separatory funnel. The aqueous phase was washed with CH.sub.2Cl.sub.2 (115 ml) and then the unified organic phases were washed with KHCO.sub.3 solution (230 ml, 10 m/m %) and saturated NaCl solution (130 ml). The organic phase was dried with MgSO.sub.4, then dichloromethane was evaporated at reduced pressure, and the crude product was stored at 20 C. until further use. Yield: 1.73 g (70%).

(2) ##STR00015##

(3) .sup.1H NMR [360 MHz, CDCl.sub.3] 1.59 (2H, m, (CH.sub.2) ring), 1.67 (4H, m, (CH.sub.2) ring), 3.45 (2H, t, (CH.sub.2) ring), 3.59 (2H, t, (CH.sub.2) ring), 3.87 (2H, s, (CH.sub.2)),

(4) .sup.13C NMR [100 MHz, CDCl.sub.3] 25.4 2pcs CH.sub.2 ring; 26.0 CH.sub.2 ring; 27.2 CH.sub.2Br; 44.2 2pcs CH.sub.2 ring; 169.5 C(O);

(5) b.) tPC2AM.sup.Pyp synthesis: The 2-bromo-1-(piperidin-1-yl)ethanone obtained as described above (0.22 g, 1.06 mmol, 2.2 equivalent) was dissolved in dry acetonitrile and added dropwise to the acetonitrile solution (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene (0.10 g, 0.48 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.44 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and acetonitrile was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.14 g (64%).

(6) ##STR00016##

(7) NMR [360 MHz, D.sub.2O] 1.40-1.70 (12H, m, (6pcs CH.sub.2)), 3.30 (8H, m, (4pcs CH.sub.2)), 3.40-3.55 (8H, m, (4pcs CH.sub.2)), 4.32 (4H, s, (2pcs CH.sub.2)), 4.62 (4H, s, (2pcs CH.sub.2)), 7.63 (2H, d, (CH) aromatic), 8.16 (1H, t, (CH) aromatic);

(8) .sup.13C NMR [100 MHz, D.sub.2O] 23.5 2pcs CH.sub.2; 25.2 2pcs CH.sub.2; 25.6 2pcs CH.sub.2; 43.7 2pcs CH.sub.2; 44.8 2pcs CH.sub.2; 45.9 2pcs CH.sub.2; 53.0 2pcs CH.sub.2; 57.5 2pcs CH.sub.2; 58.5 2pcs CH.sub.2; 123.6 2pcs CH aromatic; 144.3 CH aromatic; 153.0 2pc C aromatic; 167.7 2pcs C(O); MS (ESI) m/z 457.520(M+H).sup.+100%; 479.50(M+Na).sup.+8%;

(9) IR: 1636 cM.sup.1 (>CO); 2162, 2010 (Aromatic >CC)

EXAMPLE 2

Synthesis of tPC2AM.SUP.Pro

(10) a.) Tert-butyl 1-(2-bromoacetyl)pyrrolidine-2-carboxylate: Bromoacetyl bromide (1.44 g, 7.2 mmol, 0.63 ml, 1.5 equivalent), dry CH.sub.2Cl.sub.2 (30 ml) and K.sub.3PO.sub.4 (2.55 g, 12.0 mmol, 2.5 equ.) was mixed in a flask of 250 ml and stirred under N.sub.2 atmosphere. D-proline tert-butyl ester hydrochloride (1.00 g, 4.8 mmol, 1.0 equivalent) was dissolved in dry CH.sub.2Cl.sub.2 (20 ml) and was added dropwise to dichloromethane solution of bromoacetyl bromide at 0 C. in 30 minutes, then the reaction mixture was stirred for additional 12 hours at room temperature under N.sub.2 atmosphere before the aqueous HCl solution (0.5 M, 20 ml) was added to the reaction mixture. After the addition of HCl solution, the reaction mixture was stirred for additional 5 minutes, then the two phases were separated using a separatory funnel. The aqueous phase was washed with CH.sub.2Cl.sub.2 (110 ml) and then the unified organic phases were washed with KHCO.sub.3 solution (220 ml, 10 m/m %) and saturated NaCl solution (120 ml). The organic phase was dried with MgSO.sub.4, then dichloromethane was evaporated at reduced pressure, and the crude product was stored at 20 C. until further use. Yield: 1.05 g (75%).

(11) ##STR00017##

(12) .sup.1H NMR [360 MHz, CDCl.sub.3] 1.60 (9H, s, (CH.sub.3)), 2.15 (2H, m, (CH.sub.2) ring), 2.43 (2H, m, (CH.sub.2)ring), 3.72 (2H, m, (CH.sub.2) ring), 4.00 (2H, s, (CH.sub.2)), 4.55 (1H, m, (CH) ring);

(13) .sup.13C NMR [100 MHz, CDCl.sub.3] 25.0 CH.sub.2 ring; 27.0 CH.sub.2Br; 28.0 (3C CH.sub.3); 29.2 CH.sub.2 ring; 47.5 CH.sub.2 ring; 60.2 CH ring; 81.8 CH t-butyl; 165.2 C(O); 170.9 C(O);

(14) b.) tPC2AM.sup.Pro synthesis: The tert-butyl 1-(2-bromoacetyl)pyrrolidine-2-carboxylate obtained as described above (0.32 g, 1.08 mmol, 2.2 equivalent) was dissolved in dry acetonitrile (20 ml) and added dropwise to the acetonitrile solution of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11 (15),12-triene (0.10 g, 0.49 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.47 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and acetonitrile was evaporated at reduced pressure. The obtained yellowish oil was dissolved in CH.sub.2Cl.sub.2 (10 ml), then trifluoroacetic acid is added to it (0.23 ml, 6 equivalent) and the reaction mixture was refluxed for 24 hours. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.13 g (52%).

(15) ##STR00018##

(16) .sup.1H NMR [360 MHz, D.sub.2O] 2.00 (6H, m, (CH.sub.2)), 2.28 (2H, m, (CH.sub.2)), 3.16 (2H, m, (CH.sub.2)), 3.52 (10H, m, (CH.sub.2)), 4.45 (8H, m, (CH.sub.2)), 4.53 (2H, m, (CH), 7.37 (1H, d, (CH) aromatic), 7.44 (1H, d, (CH) aromatic) 7.92 (1H, t, (CH) aromatic);

(17) .sup.13C NMR [100 MHz, D.sub.2O] 24.3 2 pcs CH.sub.2; 28.9 2 pcs CH.sub.2; 46.5 2pcs CH.sub.2; 51.4 2pcs CH.sub.2; 56.1 2pcs CH.sub.2, 59.6 2pcs CH.sub.2; 60.2 2pcs CH; 121.8 1pcs CH aromatic; 122.7 1pc CH aromatic; 140.0 1 pc CH aromatic; 149.2 1pc C aromatic; 149.7 1 pc C aromatic; 163.8 2pcs C(O); 171.5 2pcs C (COOH);

(18) MS (ESI) m/z 517.67(M+H).sup.+100%; 539.67(M+Na).sup.+35%;

(19) IR: 1721, 1652 cm.sup. (>CO); 2158, 2025 (Aromatic >CC)

EXAMPLE 3

Synthesis of tPC2AM.SUP.Morf

(20) The commercially available 4-(bromoacetyl)morpholine (0.22 g, 1.06 mmol, 2.2 equivalent) was dissolved in dry acetonitrile, then added dropwise to the acetonitrile suspension (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene (0.10 g, 0.48 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.44 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and the solvent was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.11 g (50%).

(21) ##STR00019##

(22) .sup.1H NMR [360 MHz, D.sub.2O] 2.40-2.66 (8H, m, (4pcs CH.sub.2)), 3.32 (4H, s, (2pcs CH.sub.2)), 3.46-3.71 (8H, m, (4pcs CH.sub.2)), 4.10 (4H, s, (2pcs CH.sub.2)), 7.61 (2H, d, (CH) aromatic), 8.12 (1H, t, (CH) aromatic);

(23) .sup.13C NMR [100 MHz, D.sub.2O] 44.5 2pcs CH.sub.2; 48.8 4pcs CH.sub.2; 56.6 2pcs CH.sub.2; 59.8 2pcs CH.sub.2; 69.2 2pcs CH.sub.2; 118.3 2pcs C aromatic; 133.2 1pcs C aromatic; 159.0 2pcs C aromatic; 168.0 2pcs C(O)

EXAMPLE 4

Synthesis of tPC2AM.SUP.PipAc

(24) The commercially available 1-acetyl-4-(bromoacetyl)morpholine piperazine (0.26 g, 1.04 mmol, 2.15 equivalent) was dissolved in dry acetonitrile (5.00 cm.sup.3), then added dropwise to the acetonitrile suspension (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene (0.10 g, 0.48 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.44 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and the filtrate was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.12 g (46%).

(25) ##STR00020##

(26) .sup.1H NMR [360 MHz, D.sub.2O] 2.2 (6H, (2pcs CH.sub.3) 2.43-2.69 (8H, m, (4pcs CH.sub.2)), 3.30 (4H, s, (2pcs CH.sub.2)), 3.56-3.66 (8H, m, (4pcs CH.sub.2)), 4.11 (4H, s, (2pcs CH.sub.2)), 7.41 (2H, d, (CH) aromatic), 7.91 (1H, t, (CH) aromatic);

(27) .sup.13C NMR [100 MHz, D.sub.2O] 23.3 2pcs CH.sub.3; 43.3 2pcs CH.sub.2; 45.1 8pcs CH.sub.2; 59.1 2pcs CH.sub.2; 59.6 2pcs CH.sub.2; 69.2 2pcs CH.sub.2; 119.1 2pcs C aromatic; 132.3 1pc C aromatic; 161.2 2pcs C aromatic; 168.0 2pcs C(O); 172.3 2pcs C(O)

EXAMPLE 5

Synthesis of tPC2AM.SUP.Sar

(28) a.) N-(bromoacetyl)sarcosine tert-butyl ester: Bromoacetyl bromide (3.56 g, 17.6 mmol, 1.55 ml, 1.5 equivalent), dry CH.sub.2Cl.sub.2 (50 ml) and K.sub.3PO.sub.4 (6.41 g, 30.2 mmol, 2.5 equ.) was mixed in a flask of 250 ml under N.sub.2 atmosphere. Sarcosine tert-butyl ester (1.7 g, 11.7 mmol, 1.0 equivalent) was dissolved in dry CH.sub.2Cl.sub.2 (20 ml) and was added dropwise to dichloromethane solution of bromoacetyl bromide at 0 C. in 30 minutes, then the reaction mixture was stirred for additional 12 hours at room temperature under N.sub.2 atmosphere before the aqueous HCl solution (0.5 M, 30 ml) was added to the reaction mixture. After the addition of HCl solution, the reaction mixture was stirred for additional 5 minutes, then the two phases were separated using a separatory funnel. The aqueous phase was washed with CH.sub.2Cl.sub.2 (115 ml) and then the unified organic phases were washed with KHCO.sub.3 solution (230 ml, 10 m/m %) and saturated NaCl solution (130 ml). The organic phase was dried with MgSO.sup.4, then dichloromethane was evaporated at reduced pressure, and the crude product was stored at 20 C. until further use. Yield: 2.01 g (65%).

(29) ##STR00021##

(30) .sup.1H NMR [360 MHz, CDCl.sub.3] 1.6 (9H, s, CH.sub.3) 2.8 (3H, s, CH.sub.3), 4.01 (2H, s, CH.sub.2), 4.4 (2H, s, CH.sub.2) b). Synthesis of tPC2AM.sup.Sar: The N-(bromoacetyl)sarcosine tert-butyl ester obtained as described above (0.29 g, 1.08 mmol, 2.2 equivalent) was dissolved in dry acetonitrile (20 ml) and added dropwise to the acetonitrile solution of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11 (15),12-triene (0.10 g, 0.49 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.47 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and acetonitrile was evaporated at reduced pressure. The obtained yellowish oil was dissolved in CH.sub.2Cl.sub.2 (10 ml), then trifluoroacetic acid is added to it (0.25 ml, 6 equivalent) and the reaction mixture was refluxed for 24 hours. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.14 g (60%). 1H NMR [360 MHz, D2O] 7.91 (1H, t, aromatic), 7.35 (2H, d, aromatic) 4.97 (4H, s, CH.sub.2) 4.12 (4H, s, CH.sub.2) 3.66 (6H, s, CH.sub.3) 3.21 (4H, s, CH.sub.2) 2.67 (4H, m, CH.sub.2) 2.55 (4H, m, CH.sub.2).

(31) ##STR00022##

EXAMPLE 6

Synthesis of tPC2AM.SUP.PypCOOH

(32) a.) N-(bromoacetyl)piperidine-4-carboxylic acid tert-butyl ester: Bromoacetyl bromide (3.56 g, 17.6 mmol, 1.55 ml, 1.5 equivalent), dry CH.sub.2Cl.sub.2 (50 ml) and K.sub.3PO.sub.4 (6.41 g, 30.2 mmol, 2.5 equivalent) was mixed in a flask of 250 ml under N.sub.2 atmosphere. Piperidine tert-butyl ester (2.2 g, 11.7 mmol, 1.0 equivalent) was dissolved in dry CH.sub.2Cl.sub.2 (20 ml) and was added dropwise to dichloromethane solution of bromoacetyl bromide at 0 C. in 30 minutes, then the reaction mixture was stirred for additional 12 hours at room temperature under N.sub.2 atmosphere before the aqueous HCl solution (0.5 M, 30 ml) was added to the reaction mixture. After the addition of HCl solution, the reaction mixture was stirred for additional 5 minutes, then the two phases were separated using a separatory funnel. The aqueous phase was washed with CH.sub.2Cl.sub.2 (115 ml) and then the unified organic phases were washed with KHCO.sub.3 solution (230 ml, 10 m/m %) and saturated NaCl solution (130 ml). The organic phase was dried with MgSO.sub.4, then dichloromethane was evaporated at reduced pressure, and the crude product was stored at 20 C. until further use. Yield: 2.1 g (59%).

(33) ##STR00023##

(34) .sup.1H NMR [360 MHz, CDCl.sub.3] 1.50 (9H, s, CH.sub.3) 2.50 (1H, s, CH), 4.01 (2H, s, (CH.sub.2), 3.5-1.6 (811, m, CH.sub.2), 4.31 (2H, s, CH.sub.2)

(35) b). preparation of tPC2AMPypCOOH: The N-(bromoacetyl)piperidine tert-butyl ester obtained as described above (0.33 g, 1.08 mmol, 2.2 equivalent) was dissolved in dry acetonitrile (20 ml) and added dropwise to the acetonitrile solution of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene (0.10 g, 0.49 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.47 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and acetonitrile was evaporated at reduced pressure. The obtained yellowish oil was dissolved in CH.sub.2Cl.sub.2 (10 ml), then trifluoroacetic acid is added to it (0.25 ml, 6 equivalent) and the reaction mixture was refluxed for 24 hours. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.19 g (70%). .sup.1H NMR [360 MHz, D.sub.2O] 7.91 (1H, t, aromatic), 7.33 (2H, d, aromatic) 4.11 (4H, s, CH.sub.2) 3.75-3.22 (8H, m, CH.sub.2) 3.29 (4H, s, CH.sub.2) 2.67 (4H, m, CH.sub.2) 2.44 (4H, m,) 2.44 (2H, m, CH) 1.96-1.44 (8H, m, CH.sub.2)

(36) ##STR00024##

EXAMPLE 7

Synthesis of tPC2AM.SUP.PypCOONHS

(37) c). tPC2AMi.sup.PypCOONHS preparation: The tPC2AM.sup.PypCOOH(0.20 g, 0.37 mmol, 1.0 equivalent) obtained as described above was dissolved in dry DMF, then DCC (0.15 g, 0.74 mmol, 2 equ.) was added at room temperature and the reaction mixture was stirred at room temperature for 2 hours. Then NHS (N-Hydroxysuccinimide) (0.085 g, 0.74 mmol, 2 equ.) was added and the reaction mixture was stirred for additional 20 hours. When the reaction time was elapsed, the precipitate was filtered, and the filtrate was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.17 g (63%). .sup.1H NMR [360 MHz, D.sub.2O] 7.92 (1H, t, aromatic), 7.31 (2H, d, aromatic) 4.04 (4H, s, CH.sub.2) 3.59-3.10 (8H, m, CH.sub.2) 3.35 (4H, s, CH.sub.2) 2.71 (4H, m, CH.sub.2) 2.60 (8H, s, CH.sub.2) 2.37 (4H, m, CH.sub.2) 2.34 (2H, m, CH) 1.90-1.59 (8H, m, CH.sub.2).

(38) ##STR00025##

EXAMPLE 8

Synthesis of tPC2AM.SUP.PypCH2OH

(39) a.) N-bromoacetyl-4-hydroxymethyl piperidine: Bromoacetyl bromide (3.56 g, 17.6 mmol, 1.55 ml, 1.5 equivalent), dry CH.sub.2Cl.sub.2 (50 ml) and K.sub.3PO.sub.4 (6.41 g, 30.2 mmol, 2.5 equ.) was mixed in a flask of 250 ml under N.sub.2 atmosphere. 4-hydroxymethyl piperidine (1.34 g, 11.7 mmol, 1.0 equivalent) was dissolved in dry CH.sub.2Cl.sub.2 (20 ml) and was added dropwise to dichloromethane solution of bromoacetyl bromide at 0 C. in 30 minutes, then the reaction mixture was stirred for additional 12 hours at room temperature under N.sub.2 atmosphere before the aqueous HCl solution (0.5 M, 30 ml) was added to the reaction mixture. After the addition of HCl solution, the reaction mixture was stirred for additional 5 minutes, then the two phases were separated using a separatory funnel. The aqueous phase was washed with CH.sub.2Cl.sub.2 (115 ml) and then the unified organic phases were washed with KHCO.sub.3 solution (230 ml, 10 m/m %) and saturated NaCl solution (130 ml). The organic phase was dried with MgSO.sub.4, then dichloromethane was evaporated at reduced pressure, and the crude product was stored at 20 C. until further use. Yield: 1.85 g (67%).

(40) ##STR00026##

(41) .sup.1H NMR [360 MHz, CDCl.sub.3] 1.61 (1H, m, CH) 1.71-1.30 (8H, m, CH.sub.2), 3.52 (2H, m, (CH.sub.2), 4.21 (2H, s, CH.sub.2)

(42) b.) Synthesis of tPC2AM.sup.PypCH2OH: The N-bromoacetyl-4-hydromethyl piperidine obtained as described above (0.29 g, 1.23 mmol, 2.5 equivalent) was dissolved in dry acetonitrile and added dropwise to the acetonitrile solution (30 ml) of (0.10 g, 0.49 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.47 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and acetonitrile was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.17 g (69%).

(43) ##STR00027##

(44) .sup.1H NMR [360 MHz, D.sub.2O] 8.05 (1H, t, aromatic), 7.32 (2H, d, aromatic) 4.10 (4H, s, CH.sub.2) 3.62 (4H, d, CH.sub.2) 3.55 (4H, m, CH.sub.2) 3.49-3.21 (8H, m, CH.sub.2) 3.38 (4H, s, CH.sub.2) 2.61 (4H, t, CH.sub.2), 1.63-1.33 (8H, m, CH.sub.2) 1.61 (2H, m, CH)

EXAMPLE 9

Manufacture of tPC2AM.SUP.PypBn

(45) a.) N-bromoacetyl-4-benzylpiperidine: Bromoacetyl bromide (3.56 g, 17.6 mmol, 1.55 ml, 1.5 equivalent), dry CH.sub.2Cl.sub.2 (50 ml) and K.sub.3PO.sub.4 (6.41 g, 30.2 mmol, 2.5 equ.) was mixed in a flask of 250 ml under N.sub.2 atmosphere. 4-benzylpiperidine (2.05 g, 11.7 mmol, 1.0 equivalent) was dissolved in dry CH.sub.2Cl.sub.2 (20 ml) and was added dropwise to dichloromethane solution of bromoacetyl bromide at 0 C. in 30 minutes, then the reaction mixture was stirred for additional 12 hours at room temperature under N.sub.2 atmosphere before the aqueous HCl solution (0.5 M, 30 ml) was added to the reaction mixture. After the addition of HCl solution, the reaction mixture was stirred for additional 5 minutes, then the two phases were separated using a separatory funnel. The aqueous phase was washed with CH.sub.2Cl.sub.2 (115 ml) and then the unified organic phases were washed with KHCO.sub.3 solution (230 ml, 10 m/m %) and saturated NaCl solution (130 ml). The organic phase was dried with MgSO.sub.4, then dichloromethane was evaporated at reduced pressure, and the crude product was stored at 20 C. until further use. Yield: 1.85 g (67%).

(46) ##STR00028##

(47) .sup.1H NMR [360 MHz, CDCl.sub.3] 1.91 (1H, m, CH) 1.69-1.24 (8H, m, CH.sub.2) 2.66 (2H, m, CH.sub.2) 4.23 (2H, s, (CH.sub.2), 7.05-7.33 (5H, m, aromatic)

(48) b.) Synthesis of tPC2AM.sup.PypBn: The N-bromoacetyl-4-benzylpiperidine obtained as described above (0.32 g, 1.08 mmol, 2.2 equivalent) was dissolved in dry acetonitrile and added dropwise to the acetonitrile solution (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1 (14),11 (15),12-triene (0.10 g, 0.49 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.47 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and acetonitrile was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.21 g (66%).

(49) .sup.1H NMR [360 MHz, D.sub.2O] 7.92 (1H, t, aromatic), 7.40 (4H, m, aromatic) 7.30-7.25 (6H, m, aromatic) 7.31 (2H, d, aromatic) 3.95 (4H, s, CH.sub.2) 3.45-3.22 (8H, m, CH.sub.2) 3.29 (4H, s, CH.sub.2) 2.67 (4H, m, CH.sub.2) 2.59 (4H, d, CH.sub.2) 2.52 (4H, m, CH.sub.2) 2.1 (2H, m, CH) 1.69-1.23 (8H, m, CH.sub.2)

(50) ##STR00029##

EXAMPLE 10

Synthesis of tPC2AM.SUP.PypBnNO2

(51) a.) N-bromoacetyl-4-(4-nitrobenzyl)piperidine: Bromoacetyl bromide (3.56 g, 17.6 mmol, 1.55 ml, 1.5 equivalent), dry CH.sub.2Cl.sub.2 (50 ml) and K.sub.3PO.sub.4 (6.41 g, 30.2 mmol, 2.5 equ.) was mixed in a flask of 250 ml under N.sub.2 atmosphere. 4-(4-nitrobenzyl)piperidine (2.57 g, 11.7 mmol, 1.0 equivalent) was dissolved in dry CH.sub.2Cl.sub.2 (20 ml) and was added dropwise to dichloromethane solution of bromoacetyl bromide at 0 C. in 30 minutes, then the reaction mixture was stirred for additional 12 hours at room temperature under N.sub.2 atmosphere before the aqueous HCl solution (0.5 M, 30 ml) was added to the reaction mixture. After the addition of HCl solution, the reaction mixture was stirred for additional 5 minutes, then the two phases were separated using a separatory funnel. The aqueous phase was washed with CH.sub.2Cl.sub.2 (115 ml) and then the unified organic phases were washed with KHCO.sub.3 solution (230 ml, 10 m/m %) and saturated NaCl solution (130 ml). The organic phase was dried with MgSO.sub.4, then dichloromethane was evaporated at reduced pressure, and the crude product was stored at 20 C. until further use. Yield: 2.43 g (61%).

(52) ##STR00030##

(53) .sup.1H NMR [360 MHz, CDCl.sub.3] 1.90 (1H, m, CH) 1.62-1.33 (8H, m, CH.sub.2) 2.56 (2H, m, CH.sub.2) 4.21 (2H, s, (CH.sub.2), 7.40-8.3 (4H, m, aromatic)

(54) b.) Synthesis of tPC2AM.sup.PypBnNO2: The N-bromoacetyl-4-(4-nitrobenzyl)piperidine obtained as described above (0.37 g, 1.08 mmol, 2.2 equivalent) was dissolved in dry acetonitrile and added dropwise to the acetonitrile solution (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene (0.10 g, 0.49 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.2 g, 1.47 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and acetonitrile was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.25 g (69%).

(55) .sup.1H NMR [360 MHz, D.sub.2O] 8.21 (4H, m, aromatic) 7.93 (1H, t, aromatic), 7.75 (4H, m, aromatic) 7.31 (2H, d, aromatic) 4.11 (4H, s, CH.sub.2) 3.50-3.20 (8H, m, CH.sub.2) 3.41 (4H, s, CH.sub.2) 2.71 (4H, m, CH.sub.2) 2.66 (4H, d, CH.sub.2) 2.59 (4H, m, CH.sub.2) 2.12 (2H, m, CH) 1.70-1.35 (8H, m, CH.sub.2)

(56) ##STR00031##
c). Synthesis of tPC2AM.sup.PypBnNH2: The above obtained tPC2AM.sup.PypBnNO2 (0.4 g, 0.55 mmol) was dissolved in dry methanol, 0.04 g Pd-carbon catalyst was added, and then the mixture was reduced under 1 bar hydrogen pressure at room temperature for 2 hours. Then filter the catalyst was removed by filtration, the filtrate was evaporated at reduced pressure. Yield: 0.33 g (89%).

(57) .sup.1H NMR [360 MHz, D.sub.2O] 7.92 (1H, t, aromatic), 7.30 (2H, d, aromatic) 7.12 (4H, m, aromatic) 6.44 (4H, m, aromatic) 4.11 (4H, s, CH.sub.2) 3.48-3.21 (8H, m, CH.sub.2) 3.31 (4H, s, CH.sub.2) 2.70 (4H, m, CH.sub.2) 2.68 (4H, d, CH.sub.2) 2.56 (4H, m, CH.sub.2) 1.92 (2H, m, CH) 1.66-1.21 (8H, m, CH.sub.2)

(58) ##STR00032##
d). Synthesis of tPC2AM.sup.PypBnNH2(0.20 g, 0.30 mmol, 1.0 equivalent) obtained as described above was dissolved in chloroform (50 ml) and cooled to 0 C., then solution of K.sub.2CO.sub.3 prepared with 30 ml water was added (0.09 g, 0.60 mmol, 2 equivalent) and thiophosgene (0.07 g, 0.60 mmol, equivalent) solution prepared with 30 ml chloroform was also added. Then the mixture was allowed to warm up to room temperature for 5 hours. The organic phase was separated, washed with water (110 ml), dried on MgSO.sub.4, and then the chloroform was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.13 g (56%). .sup.1H NMR [360 MHz, D.sub.2O] 7.91 (1H, t, aromatic), 7.28 (2H, d, aromatic) 7.40-7.18 (8H, m, aromatic) 3.95 (4H, s, CH.sub.2) 3.39-3.29 (8H, m, piperidine CH.sub.2) 3.33 (4H, s, CH.sub.2) 2.65 (4H, m, CH.sub.2) 2.54 (4H, d, CH.sub.2) 2.51 (4H, m, CH.sub.2) 1.98 (2H, m, CH) 1.65-1.33 (811, m, CH.sub.2)

(59) ##STR00033##

EXAMPLE 11

Synthesis of tPC2AMP.SUP.Pyp.EA

(60) The tert-butyl N-(2-bromoethyl)carbamate (0.09 g, 0.40 mmol, 1.3 equivalent) was dissolved in dry acetonitrile, and then added dropwise at room temperature to the acetonitrile solution (30 ml) of the tPC2AMP.sup.Pyp ligand manufactured as per Example 1 (0.14 g, 0.31 mmol, 1 equivalent) (prepared on the basis of Organic Preparations and Procedures International, 2009, 41(4), 301-307 publication by Li, Hongbo; Hao, Meng-an; Wang, Liping; Liang, Wu; Chen, Kai From) and K.sub.2CO.sub.3 (0.09 g, 0.62 mmol, 2 equivalent) within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and the filtrate was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. The oily material obtained after purification (10 ml) was dissolved in CH2Cl2, then trifluoroacetic acid was added to it (0.07 ml, 3 equivalent) and the reaction mixture was refluxed for 24 hours. Then the solvent was evaporated at reduced pressure, once 6 M (10 ml), and twice water (10-10 ml) was evaporated from the solid to remove trifluoroacetic acid excess. Yield: 0.09 g (58%).

(61) ##STR00034##

(62) .sup.1H NMR [360 MHz, D.sub.2O] 1.4-1.7 (12H, m, (6pcs CH.sub.2)), 2.68 (4H, d, 2pcs CH.sub.2), 3.32 (8H, m, (4pcs CH.sub.2)), 3.4-3.6 (8H, m, (4pcs CH.sub.2)), 4.32 (4H, s, (2pcs CH.sub.2)), 4.62 (4H, s, (2pcs CH.sub.2)), 7.63 (2H, d, (CH) aromatic), 8.16 (1H, t, (CH) aromatic);

(63) .sup.13C NMR [100 MHz, D.sub.2O] 23.2 2pcs CH.sub.2; 25.1 2pcs CH.sub.2; 25.4 2pcs CH.sub.2; 40.2 1pc CH.sub.2; 43.5 2pcs CH.sub.2; 44.3 2pcs CH.sub.2; 45.7 2pcs CH.sub.2; 53.0 2pcs CH.sub.2; 55.1 1pc CH.sub.2; 57.5 2pcs CH.sub.2; 58.3 2pcs CH.sub.2; 123.1 2pcs CH aromatic; 143.2 CH aromatic; 153.1 2pcs C aromatic; 169.6 2pcs C(O);

EXAMPLE 12

Synthesis of tPC2AMP.SUP.Pyp.DPA

(64) The 2-(chloromethyl)pyridine (0.088 g, 0.69 mmol, 2.3 equivalent) was converted into the commercially available 2-(chloromethyl)pyridine hydrochloride free base form was dissolved in dry acetonitrile and was added dropwise to the acetonitrile solution (30 ml) of the tPC2AMPPypEA ligand (0.15 g, 0.30 mmol, 1 equivalent) and N,N-diisopropylethylamine (0.12 g, 0.90 mmol, 3 equivalent) at room temperature within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours and was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.12 g (60%).

(65) ##STR00035##

(66) .sup.1H NMR [360 MHz, D.sub.2O] 1.4-1.7 (12H, m, (6pcs CH.sub.2)), 2.51 (4H, d, 2pcs CH.sub.2) 3.32 (8H, m, (4pcs CH.sub.2)), 3.4-3.6 (8H, m, (4pcs CH.sub.2)), 4.11 (4H, s, (2pcs CH.sub.2)), 4.32 (4H, s, (2pcs CH.sub.2)), 4.62 (4H, s, (2pcs CH.sub.2)), 7.26 (4H, t, (CH) aromatic), 7.64 (2H, d, (CH) aromatic), 7.72 (2H, t, (CH) aromatic), 8.16 (1H, t, (CH) aromatic), 8.29 (2H, d, (CH) aromatic),

(67) .sup.13C NMR [100 MHz, D.sub.2O] 24.8 2pcs CH.sub.2; 25.3 2pcs CH.sub.2; 25.8 2pcs CH.sub.2; 41.0 1pc CH.sub.2; 44.4 2pcs CH.sub.2; 44.9 2pcs CH.sub.2; 45.9 2pcs CH.sub.2; 53.3 2pcs CH.sub.2; 56.3 1pc CH.sub.2; 58.3 2pcs CH.sub.2; 58.9 2pcs CH.sub.2; 119.1 2pcs CH aromatic; 123.1 2pcs CH aromatic; 126.0 2pcs CH aromatic; 141.1 2pcs CH aromatic; 143.2 CH aromatic; 149.1 2pcs CH aromatic; 153.1 2pcs C aromatic; 158.1 2pcs CH aromatic; 169.6 2pcs C(O);

EXAMPLE 13

Synthesis of tPC2AMP.SUP.Pyp.TPA

(68) The 1-(6-(chloromethyl)pyridine-2-yl)-N,N-bis(pyridine-2-ylmethyl)methaneamine (0.15 g, 0.43 mmol, 1.3 equivalent, manufactured on the basis of Dalton Trans., 2006, 3108-3113 publication by Simon J. A. Pope, Rebecca H. Laye) was dissolved in dry acetonitrile, and then added dropwise at room temperature to the acetonitrile solution (30 ml) of the tPC2AMP tPC2AM.sup.Pyp ligand (0.15 g, 0.30 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.12 g, 0.90 mmol, 3 equivalent) within 30 minutes. Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and the reaction mixture was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Yield: 0.15 g (65%).

(69) ##STR00036##

(70) .sup.1H NMR [360 MHz, D.sub.2O] 1.4-1.7 (12H, m, (6pcs CH.sub.2)), 2.68 (4H, d, 2pcs CH.sub.2) 3.32 (8H, m, (4pcs CH.sub.2)), 3.4-3.6 (8H, m, (4pcs CH.sub.2)), 4.14 (8H, s, 4pcs CH.sub.2) 4.62 (4H, s, (2pcs CH.sub.2)), 7.63 (2H, d, (CH) aromatic), 7.71 (2H, d, (CH) aromatic), 7.88 (4H, m, (CH) aromatic), 8.16 (1H, t, (CH) aromatic), 8.32 (3H, t, (CH) aromatic);

(71) .sup.13C NMR [100 MHz, D.sub.2O] 23.2 2pcs CH.sub.2; 25.1 2pcs CH.sub.2; 25.4 2pcs CH.sub.2; 40.2 1pc CH.sub.2; 43.5 2pcs CH.sub.2; 44.3 2pcs CH.sub.2; 53.0 2pcs CH.sub.2; 55.1 1pc CH.sub.2; 57.5 2pcs CH.sub.2; 58.3 2pcs CH.sub.2; 62.1 3pcs CH.sub.2; 63.9 1pc CH.sub.2; 123.1 2pcs CH aromatic; 122.7 4pcs CH aromatic; 124.9 2pcs CH aromatic; 143.2 2pcs CH aromatic; 146.4 2pcs CH aromatic; 149.0 2pcs CH aromatic; 153.1 2pcs C aromatic; 154.1 2pcs C aromatic; 158.4 2pcs C aromatic; 173.3 2pcs C(O);

EXAMPLE 14

Synthesis of pOH-tPC2A

(72) The commercially available tert-butyl bromoacetate (0.19 g, 0.99 mmol, 0.145 ml, 2.2 equivalent) was dissolved in dry acetonitrile, then added dropwise to the acetonitrile solution (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene-13-ol (0.10 g, 0.45 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.19 g, 1.35 mmol, 3 equivalent) at room temperature within 30 minutes (the macrocycle was manufactured based on the K. M. Lincoln, M. E. Offutt, T, D. Hayden, R. E. Saunders, K. N. Green, Inorg, Chem., 2014, 53(3), 1406-1416, and K. M. Lincoln, P. Gonzalz, T. E: Richardson, D. A. Julovich, R. Saunders, J. W. Simpkins, K. N. Green, Chem. Commun., 2013, 49(26), 2712-2714 publications). Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and acetonitrile was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. The obtained yellowish oil was dissolved in 10 ml CH.sub.2Cl.sub.2, then trifluoroacetic acid was added to it (0.21 ml, 6 equivalent) and the reaction mixture was refluxed for 24 hours. Then the solvent was evaporated at reduced pressure, once 6 M (10 ml), and twice water (10-10 ml) was evaporated from the solid to remove trifluoroacetic acid excess. Yield: 0.11 g (72%).

(73) ##STR00037##

(74) .sup.1H NMR [360 MHz, D.sub.2O] 3.23 (2H, m, (CH.sub.2)), 3.3-3.5 (8H, m, (4pcs CH.sub.2)), 3.43 (4H, s, (2pcs CH.sub.2), 3.81 (2H, m, (CH.sub.2)), 6.40 (2H, s, (CH) aromatic);

(75) .sup.13C NMR [100 MHz, D.sub.2O] 46.2 2pcs CH.sub.2; 57.5 2pcs CH.sub.2; 60.0 2pcs CH.sub.2; 64.3 2pcs CH.sub.2; 113.3 2pcs CH aromatic; 153.0 C(OH) aromatic; 158.2 2pcs C aromatic; 161.6 2pcs C(O);

EXAMPLE 15

Synthesis of pOH-tPC2AM.SUP.Pyp

(76) The 2-bromo-1-(piperidine-1-yl)ethanone manufactured as per Example 1 (0.21 g, 0.99 mmol, 2.2 equivalent) was dissolved in dry acetonitrile, then added dropwise to the acetonitrile solution (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene-13-ol (0.10 g, 0.45 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.19 g, 1.35 mmol, 3 equivalent) at room temperature within 30 minutes (the macrocycle was manufactured based on the K. M. Lincoln, M. E. Offutt, T. D. Hayden, R. E. Saunders, K. N. Green, Inorg, Chem., 2014, 53(3), 1406-1416, and K. M. Lincoln, P. Gonzalz, T. E: Richardson, D. A. Julovich, R. Saunders, J. W. Simpkins, K. N. Green, Chem. Commun., 2013, 49(26), 2712-2714 publications). Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and the mother liquor was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACM acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. Then the solvent was evaporated at reduced pressure, once 6 M (10 ml) hydrochloric acid, and twice water (10-10 ml) was evaporated from the solid to remove trifluoroacetic acid excess. Yield: 0.11 g (57%).

(77) ##STR00038##

(78) .sup.1H NMR [360 MHz, D.sub.2O] 1.52 (12H, m, (6pcs CH.sub.2)), 2.84 (4H, m, (2pcs CH.sub.2)), 3.23 (2H, m, (CH.sub.2)), 3.3-3.6 (8H, m, (4pcs CH.sub.2)), 3.87 (2H, m, (CH.sub.2)), 4.84 (4H, s, (2pcs CH.sub.2), 6.38 (2H, s, (CH) aromatic);

(79) .sup.13C NMR [100 MHz, D.sub.2O] 24.3 2pcs CH.sub.2; 24.7 2pcs CH.sub.2; 25.8 2pcs CH.sub.2; 43.9 4pcs CH.sub.2; 45.1 2pcs CH.sub.2; 46.4 2pcs CH.sub.2; 58.0 2pcs CH.sub.2; 60.3 2pcs CH.sub.2; 114.0 2pcs CH aromatic; 157.0 C(OH) aromatic; 159.4 2pcs C aromatic; 159.0 2pcs C(O);

EXAMPLE 16

Synthesis of tPC2AM.SUP.OMe

(80) The commercially available tert-butyl bromoacetate (0.27 g, 1.40 mmol, 0.20 ml, 2.2 equivalent) was dissolved in dry acetonitrile, then added dropwise to the acetonitrile solution (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene-13-methoxy (0.15 g, 0.63 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.26 g, 1.89 mmol, 3 equivalent) at room temperature within 30 minutes (the macrocycle was manufactured based on the K. M. Lincoln, M. E. Offutt, T. D. Hayden, R. E. Saunders, K. N. Green, Inorg, Chem., 2014, 53(3), 1406-1416, and K. M. Lincoln, P. Gonzalz, T. E: Richardson, D. A. Julovich, R. Saunders, J. W. Simpkins, K. N. Green, Chem. Commun., 2013, 49(26), 2712-2714 publications). Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and the filtrate was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. The obtained yellowish oil was dissolved in CH.sub.2Cl.sub.2 (10 ml), then trifluoroacetic acid is added to it (0.24 ml, 6 equivalent) and the reaction mixture was refluxed for 24 hours. Then the solvent was evaporated at reduced pressure, once 6 M (10 ml), and twice water (10-10 ml) was evaporated from the solid to remove trifluoroacetic acid excess. Yield: 0.13 g (59%).

(81) ##STR00039##

(82) .sup.1H NMR [360 MHz, D.sub.2O] 3.21 (2H, m, (CH.sub.2)), 3.24-3.47 (8H, m, (4pcs CH.sub.2)), 3.41 (4H, s, (2pcs CH.sub.2), 3.78 (2H, m, (CH.sub.2)), 3.81 (3H, s, CH.sub.3), 6.42 (2H, s, (CH) aromatic);

(83) .sup.13C NMR [100 MHz, D.sub.2O] 47.1 2pcs CH.sub.2; 56.2 1pc CH.sub.3 57.8 2 pcs CH.sub.2; 61.1 2pcs CH.sub.2; 63.9 2pcs CH.sub.2; 112.1 2pcs CH aromatic; 157.9 2pcs C aromatic; 161.1 C(OCH.sub.3) aromatic; 169.7 2pcs C(O);

EXAMPLE 17

Synthesis of tPC2AMP.SUP.Pyp.OMe

(84) The 2-bromo-1-(pyperidine-1-yl)ethanone manufactured as per Example 1 (0.29 g, 1.40 mmol, 2.2 equivalent) was dissolved in dry acetonitrile, then added dropwise to the acetonitrile solution (30 ml) of 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(14),11(15),12-triene-13-methoxy (0.15 g, 0.63 mmol, 1 equivalent) and K.sub.2CO.sub.3 (0.26 g, 1.89 mmol, 3 equivalent) at room temperature within 30 minutes (the macrocycle was manufactured based on the K. M. Lincoln, M. E. Offutt, T. D. Hayden, R. E. Saunders, K. N. Green, Inorg, Chem., 2014, 53(3), 1406-1416, and K. M. Lincoln, P. Gonzalez, T. E: Richardson, D. A. Julovich, R. Saunders, J. W. Simpkins, K. N. Green, Chem. Commun., 2013, 49(26), 2712-2714 publications). Then the reaction mixture was refluxed in N.sub.2 atmosphere for 24 hours. After 24 hours, K.sub.2CO.sub.3 was filtered from the hot solution, and the filtrate was evaporated at reduced pressure. The obtained crude product was purified with HPLC (Luna 10u-Prep C18(2) 100A (25021.20 mm; 10 m) column), ACN:H.sub.2O/TFA-t was applied as eluent [ACN: acetonitrile; TFA: trifluoroacetic acid]. TFA was contained only in water in 0.005 M concentration. The fraction containing the product was evaporated at reduced pressure, once 6 M hydrochloric acid (10 ml), and twice water (10-10 ml) was evaporated from the solid to remove trifluoroacetic acid excess. Yield: 0.14 g (46%).

(85) ##STR00040##

(86) .sup.1H NMR [360 MHz, D.sub.2O] 1.52 (12H, m, (6pcs CH.sub.2)), 2.84 (4H, m, (2pcs CH.sub.2)), 3.23 (2H, m, (CH.sub.2)), 3.3-3.6 (8H, m, (4pcs CH.sub.2)), 3.87 (2H, m, (CH.sub.2)), 4.84 (4H, s, (2pcs CH.sub.2), 6.38 (2H, s, (CH) aromatic);

(87) .sup.13C NMR [100 MHz, D.sub.2O] 24.5 2pcs CH.sub.2; 24.8 2pcs CH.sub.2; 25.9 2pcs CH.sub.2; 44.2 4pcs CH.sub.2; 45.5 2pcs CH.sub.2; 46.0 2pcs CH.sub.2; 57.2 1pc CH.sub.3, 57.6 2pcs CH.sub.2; 60.1 2pcs CH.sub.2; 112.8 2pcs CH aromatic; 155.5 C(OCH.sub.3) aromatic; 161.4 2pcs C aromatic; 170.1 2pcs C(O);

EXAMPLE 18

Efficacy Data

(88) During the physico-chemical tests of tPC2AM.sup.Pyp and tPC2AM.sup.Pro compounds synthesized according to Example 1 and 2, their protonation constant, as well as equilibrium behaviour and kinetic inertism of their Mn(II) complexes was studied in detail, and the characteristic relaxivity values of the complexes were determined in the presence and absence of HSA (Human Serum Albumin), at 25 and 37 C. and physiological pH. All studies were performed in the presence of 0.15 M NaCl, the same concentration as that of the electrolyte under physiological conditions.

(89) The results of equilibrium study are summarized in Table 1, in addition to the protonation constants, total basicity of ligands and stability constants of their Mn(II) complexes, the pMn value calculated for complexes are also represented in the table.

(90) TABLE-US-00001 TABLE 1 Protonation constants and total basicity of the studied ligands, stability constants of their Mn(II) complexes and calculated pMn values (25 C., 0.15 M NaCl). logK.sub.1 logK.sub.2 logK.sub.3 logK.sub.4 logK.sub.i.sup.H logK.sub.MnL pMn tPC2AM.sup.Pyp 11.31(1) 3.94(3) 15.26 13.09(2) 7.09 tPC2AM.sup.Pro 9.80(2) 7.96(4) 3.65(5) 2.26(4) 23.67 13.77(3) 7.85 pMn values were calculated by using the equilibrium constants at pH = 7.4 and cMn = cL = 10.sup.5 M.
Based on the pMn values represented in Table 1 (calculated using the equilibrium constants at pH=7.4 and cMn=cL=10.sup.5 M), it can be concluded that the studied Mn(II) complexes are formed in 100% at physiological pH, which is an essential aspect of the practical use.

(91) An important parameter of using Mn(II) containing contrast agents in vivo is the low kinetic reactivity of the complex. The kinetic reactivity is generally tested with metal ion exchange reactions, where the replacing metal ion is Zn(II) or Cu(II) in most of the cases. The application of Cu(II) is advantageous for more reasons, in one hand the complexes with ligands are of great thermodynamic stability, so relatively small excess of Cu(II) ion leads to complete replacement, on the other hand molar absorbance values of Cu(II) complexes both in UV and visible range are sufficiently high to enable spectrophotometric method for examinations even at low concentrations. Moreover, the endogenic characteristic of the Cu(II) ion provides additional information on in vivo processes. Replacement reactions were executed with at least 10-fold excess Cu(II) ion concentration to ensure pseudo-primary conditions.

(92) Dissociation reactions of Mn(II) complexes may take place in several pathways as represented below.

(93) ##STR00041##

(94) The k.sub.0, k.sub.H, k.sub.H.sup.H, k.sub.Cu and k.sub.Cu.sup.H, rate constants indicate the spontaneous, proton associated, metal assisted and proton-metal assisted (when the replacing metal ion attacks the protonated complex) reaction pathways of the complex. The K.sub.MnHL, K.sub.MnH.sub.2.sub.L and K.sub.MnLCu are stability constants of the protonated and binuclear intermediate complexes.

(95) In case of metal complexes formed with macrocyclic ligands the above detailed mechanism involves only proton associated dissociation pathways (in some instances spontaneous dissociation may have some role), since the formation of binuclear complexes are inhibited (denticity of rigid ligands does not exceed the coordination number of the metal ion, Mn.sup.2+). Due to this reason, replacement reactions were executed in 2.0-5.0 pH range with only 10-fold Cu(II) replacement metal ion excess.

(96) In general the k.sub.obs pseudo-primary rate constants obtained in each reaction are given with the following equation, where the stability constants of each reaction pathway and that of the forming intermediate are also considered:

(97) $\begin{array}{cc}{k}_{\mathrm{obs}}=\frac{k_0+k_1\left[H^{+}\right]+{k_2\left[H^{+}\right]}^2+k_3\left[{\mathrm{Cu}}^{2+}\right]+k_4\left[{\mathrm{Cu}}^{2+}\right]\left[H^{+}\right]}{1+K_{\mathrm{MnHL}}\left\{H^{+}\right]+{K_{{\mathrm{MnH}}_{2}L}\left[H^{+}\right]}^2+K_{\mathrm{MnLCu}}\left[{\mathrm{Cu}}^{2+}\right]},& \left(1\right)\end{array}$
whereas K.sub.MnHL=[Mn(HL)]/[Mn(L)][H.sup.+], K.sub.MnH.sub.2.sub.L=[Mn(H.sub.2L)]/[Mn(HL)][H.sup.+], K.sub.MnLCu=[Mn(L)M]/[Mn(L)][M], k.sub.1=k.sub.H.Math.K.sub.MnHL, k.sub.2=k.sub.H.sup.H.Math.K.sub.MnHL.Math.K.sub.MnH.sub.2.sub.L, k.sub.3=k.sub.Cu.Math.K.sub.MnLCu k.sub.4=k.sub.Cu.sup.H.Math.K.sub.MnHL

(98) Results of the kinetic study showed that in the dissociation of [Mn(tPC2AM.sup.Pyp)].sup.2+ complexes, the proton associated dissociation, characterized with k.sub.1 plays an important role. Using these rate constants the half-life (t.sub.1/2) of [Mn(tPC2AM.sup.Pyp)].sup.2+ and [Mn(tPC2AM.sup.Pro)].sup.2+ complex dissociation may be calculated at physiologic pH, being 352 and 215 hours.

(99) In order to estimate the quantity of complex decomposing in the body, it is useful to handle elimination and complex dissociation as parallel, primary reaction characterized by the (2) equation set for Gd.sup.3+ complexes [F. K. Klmn and G. Tircs, Inorg. Chem., 2012, 51, 10065]:

(100) $\begin{array}{cc}{\left[\mathrm{GdL}\right]}_d={\frac{k_d}{k_d+k_{\mathrm{ex}}}\left[\mathrm{GdL}\right]}_0\left(1-e^{-\left(k_d+k_{\mathrm{ex}}\right)t}\right)& \left(2\right)\end{array}$
The equation indicates that the extent of the dissociation of the complex depends on the ratio of rate constants. For the (renal) elimination of contrast agent 1.6 hour half life can be given in general, characterized by a k.sub.ex=0.433 11.sup.1 rate constant. Using the k.sub.d values of Mn(II) complexes and the k.sub.ex values characteristic for elimination, one can calculate the percentile ratio of injected complex dissociated in vivo until complete elimination (12-24 hours). Calculation verified, that less than 0.8% of the complexes would dissociate before the elimination of the complex. Considering the essential character of Mn(II) ion and its complexes the given amount cannot cause significant burden for MRI tested patients. Considering these new results, during the in vivo dissociation (37 C.) of [Gd(DTPA)].sup.2 complex (Magnevist) applied in practice, 2.2% Gd(III) ion releases being 4.4-fold of the value calculated on the basis of experiments at 25 C. (0.5%). [Sarka L. et al, Chem. Eur. J., 2000, 6, 719]. Using this finding, the quantity of in vivo releasing Mn(II) for the presented Mn(II) complexes is approximately 3%. This value is better than some of those applied for Gd containing contrast agents in practice [Baranyai Z et al, Chem. Eur. J., 2015, 21, 4789]

(101) In addition to appropriately low kinetic reactivity, complexes shall also have suitable relaxivity for the purpose of practical use (relaxivity (mM.sup.1 s.sup.1): relaxation rate increase of 1 mM solution of the paramagnetic substance compared to the measured value under diamagnetic conditions [Tth ., et. al., The Chemistry of Contrast Agents in Medical Magnetic Resonance Imaging, Chichester: John Wiley & Sons, 2001.]). Higher complex relaxivity results in higher contrast increasing effect, meaning that the same image quality is obtained by introducing less amount of complex with higher relaxivity. The relaxivity value of both complexes were determined at pH=7.4 and 25 and 37 C. in the presence and absence of HSA (Human Serum Albumin, c=0.7 mM) to better stimulate conditions of the in vivo application. Relaxivity values of the complexes are presented in Table 2. Comparing the data in Table 2 with the relaxivity values of DOTAREM ([Gd(DOTA)].sup. complex, r.sub.1=3.83 mM.sup.1 s.sup.1) and MAGNEVIST ([Gd(DTPA)].sup.2 complex, r.sub.1=4.02 mM.sup.1 s.sup.1) [Powell, D. H., Ni Dhubhghaill, O. M., Pubanz, D. et al. (1996) J. Am. Chem. Soc., 118, 9333-9346] applied in practice under the same conditions, the Mn(II) complexes presented herein obviously have higher relaxivities and thus higher contrast increasing effect.

(102) TABLE-US-00002 TABLE 2 Relaxivity values of the tested complexes (pH = 7.4) Complex T ( C.) r.sub.1 (mM.sup.1s.sup.1) r.sub.1 (mM.sup.1s.sup.1) HSA [Mn(tPC2AM.sup.Pyp)].sup.2+ 25 4.90 18.34 37 3.96 14.07 [Mn(tPC2AM.sup.Pro)] 25 4.43 5.53 37 3.44 4.36