KR19980701397A - Novel Peptides, Their Production and Use - Google Patents

Abstract

The compounds represented by the following formula (1) and their preparation are described.

Formula 1

In the formula,

R is H or OH,

X is Ala, Arg, Gly or Val,

Y is Pro or His,

Z is H, F, Cl, Br or I,

Provided that when X is Gly, Y is Pro, and R is OH, then Z is not H. This compound is suitable for controlling disease.

Description

Novel peptides, their preparation and uses

The present invention relates to novel peptides, their preparation and their use for disease control.

Heart rhythm denial is known to represent one of the most common causes of death in western industrialized countries. In addition, arrhythmias are particularly important with regard to coronary heart disease, ischemia and old age. In addition, the mechanisms causing arrhythmia vary widely, and in some cases are still unknown. For example, it is certain that surviving Purkinge fibers in the infarct region can maintain arrhythmias as external pacemakers. Likewise, the outflow of potassium in the structure of cardiac ischemia may lead to depolarization of the fiber, which may not be partially excited, resulting in unidirectional blockage of stimulus induction. In addition to many other mechanisms, there is a local difference (dispersion) in the duration of action potentials in the infarct structure, which can then trigger a reintroduction circuit. Then, there is a possibility that ventricular vibration or fibrillation occurs from this circulation induction. Another mechanism cell separation that can lead to this dispersion of action potential duration is because it can no longer compensate for the potential difference between cells (Circ. Res. 65 (1989) 1426). This separation may, on the one hand, increase with age, for example due to connective tissue infiltration (Circ. Res. 62 (1988) 811), or on the other hand, for example, an increase in pCO ₂ and pH and ATP content. Closure of intercellular connections due to the drop can occur in infarct structures (Am. J. Physiol. 248 , (1985) H753-H764, Circ. Res 45, (1979) 324).

Antiarrhythmic agents used to date have been suitable ion channel blockers to block transmembrane ion channels (sodium channels, calcium channels and / or potassium channels) and to treat acute existing cardiac rhythm negatives.

However, problems arise when trying to use these substances for the prevention of arrhythmias. These ion channel blockers, at least when administered prophylactically, present a high total venous risk (Drugs 29 , (1985) Suppl. 4, 33-34, New England J. Med. 324 , (1991) 781). This contradicts the arrhythmia precisely caused by the administration of classic antiarrhythmic agents. This is why these materials are only suitable under a number of limitations as a preventive measure.

Therefore, research is now being conducted on substances that can be administered for prophylaxis, have a novel principle of action, and no longer exhibit the above allergic effect. One novel principle is the improvement of cell connectivity.

Clinical and experimental results using conventional antiarrhythmics have demonstrated that prevention of arrhythmia is rarely possible due to apparent allergic vein side effects, which could also be detected in vitro (New England J. Med. 324 . (1991) 781), Circulation 87, (1993) 617).

An antiarrhythmic peptide AAP10 has been proposed (Naunyn Schmiedeberg's Arch. Pharmacol. 350 (1994) 174), and has been proposed as a novel principle to improve cell connectivity, reduce local differences in duration of action potential and stabilize the epicardial induction pattern. . This material shows virtually no risk of total vein in in vitro testing on isolated rabbit hearts and is very effective against ischemia associated with ischemia. The main effect of this material is to reduce the dispersion of dislocation durations.

The present invention relates to compounds of formula (I) and the use of these peptides for disease control.

In the formula,

R is H or OH,

X is Ala, Arg, Gly or Val,

Y is Pro or His,

Z is H, F, Cl, Br or I,

Provided that when X is Gly, Y is Pro, and R is OH, then Z is not H. Hyp in the formula (1) means 4-hydroxyproline.

In formula (1), Z is preferably a glycine residue, Y is preferably a proline residue and Z is in particular at the position 2 and preferably at position 3 as a halogen atom, preferably iodine.

This compound can be prepared by conventional peptide chemistry methods. Particularly suitable methods for preparation are described using the Fmoc method. Atherton R.C. Sheppard (1989; Solid phase peptide synthesis, IRL-Press, Oxford)] is a solid phase synthesis of an insoluble resin phase by a modified Merrifield process.

In this case, amino acid activation can be generated by the formation of anhydrides, 1-hydroxybenzotriazole esters or pentafluorophenyl esters.

The particular effect of the novel peptides is to eliminate local differences in the duration of action potentials and irritation-induced irregularities, all of which occur, for example, within the structure of myocardial infarction or with increasing age.

The present invention enables the prophylactic treatment of ischemia-related and age-related cardiac rhythms. Moreover, this material exhibited a negligible total venous risk in vitro, as opposed to conventional antiarrhythmics. Compared with known materials, this novel peptide can exhibit better efficacy and higher minimum effects.

1. Preparation of Fluorenylmethoxycarbonyl-iodotricin

Tyrosine and phthalic anhydride were reacted together in glacial acetic acid for 20 hours corresponding to the Gabriel synthesis method, and the reaction product was reacted with iodine and silver acetate (II) to obtain N-phthaloyl-L-moniodotryptocin. The protecting group was then removed with phenylhydrazine. The reaction product was reacted with N-fluoroenylmethoxycarbonyloxysuccinimide in the presence of Na ₂ CO ₃ , water and acetone. After acidification with HCl the required Fmoc-iodotyrosine was obtained.

2. Synthesis of Peptide (1) H ₂ N-Gly-Ala-Gly-Hyp-Pro-3-iodotryptocinamide

The synthesis protocol for the Fmoc method published by Atherton and Shepard was used. 272.7 mg of the link resin with a load of 0.55 mmol / g was swollen with dimethylformamide and then the protecting group was removed with 20% piperidine in dimethylformamide (DMF). After washing with DMF, dicyclohexylcarbodiimide (DCC) in DMF was added to 0.9 mmol of Fmoc-iodotyrosine. After washing with DMF and methanol, the protecting group was removed with 20% piperidine in DMF and after further washing, the amino acids were then linked. For this purpose, Fmoc-proline-OH was added to O- (1H-benzotriazol-1-yl) -N, N, N ', N'-tetramethyluronium tetrafluoroborate (TBTU), 1-hydric in DMF. The peptide was reacted with oxybenzotriazole (HOBT) and diisopropylethylamine (DIPEA). After washing, the protecting group was once again removed with piperidine and DMF, and after further washing, the reaction product was reacted with Fmoc-hydroxyproline-OH as described above. Fmoc-glycine-OH, Fmoc-alanine-OH and Fmoc-glycine-OH were then linked in this manner after washing and removal. The protecting group was then removed with 20% piperidine in DMF and after washing, dried at 0.1 mbar for 6 hours. Finally, the resin was separated with 5% aqueous trifluoroacetic acid solution for 2 hours and the product was washed, distilled in a rotary distillation, dissolved in glacial acetic acid and precipitated with diethyl ether. The precipitate was filtered off by suction and purified by conventional methods by semipreparative HPLC. 7.6 mg of new peptide was obtained (molecular weight: 701.6).

3. The following peptides were obtained in a similar manner to Examples 1 and 2.

(2) H ₂ N-Gly-Ala-Gly-Hyp-Pro-3-fluorotyrosineamide

(3) H ₂ N-Gly-Ala-Gly-Hyp-Pro-3-chlorotyrosineamide

(4) H ₂ N-Gly-Ala-Gly-Hyp-Pro-3-bromotyrosineamide

(5) H ₂ N-Arg-Ala-Gly-Hyp-Pro-tyrosineamide

(6) H ₂ N-Val-Ala-Gly-Hyp-Pro-tyrosineamide

(7) H ₂ N-Ala-Ala-Gly-Hyp-Pro-tyrosineamide

(8) H ₂ N-Gly-Ala-Gly-Hyp-His-tyrosineamide

(9) H ₂ N-Gly-Ala-Gly-Hyp-Pro-phenylalanineamide.

Usage:

Increasing the concentration (10 ^-10 , 10 ^-9 , 10 ^-8 , 10 ^-7 mol /) in the isolated rabbit heart sprayed with Tyrode solution by the Langendorf method under normal pressure (70 cm H ₂ O) The fusion between the tubes was performed with the peptide (1) of l), and at the same time the outer core potential mapping was performed. References [J. Pharmacol. Methods 22 , (1989), 197, Circulation 87 , (1933) 617).

In this study, unipolar electrocardiograms were simultaneously recorded at 256 sites on the epicardial surface of the heart to allow measurement of local epicardial action potential duration at a total of 256 sites. The distribution of the action potential duration near the mean and the change by this substance in comparison with AAP10 were investigated using these data. Narrow increases in the curves were observed for both materials, ie more values approached the mean as the concentrations of both AAP10 and novel peptides increased. Thus, under control conditions, 50% of all values are in the range of ± 5 ms around the mean, up to 74% (10 ⁻⁸ mol / l) for AAP10, but 90% (for new peptides) 10 ^-7 mol / l). This means that the novel peptides significantly reduce the dispersion of epicardial action potential duration, and this reduction is more pronounced than that achieved by AAP10.

Concentration-dependent effect on dispersion of epicardial potential duration by antiarrhythmic peptide AAP10 and novel peptides Log concentration AAP10 New Peptides (1) contrast 7.8 ± 0.9 6.0 ± 1.0 -10 6.5 ± 0.4 4.5 ± 0.9 -9 6.2 ± 0.4 5.0 ± 1.1 -8 5.2 ± 0.4 4.6 ± 0.9 -7 6.1 ± 0.1 3.8 ± 0.6

In particular, investigating the number of values for other epicardial potential durations (ARI) from less than ± 5 ms from the mean will clarify the good effect of the novel peptide. The ratio of ARI values within ± 5 ms intervals near the mean is 54% for new peptides and 71% at 10 ⁻¹ 0 mol / l, 73% at 10 ⁻⁹ mol / l and 10 ⁻⁸ mol / l under control conditions 75% and 90% at 10 ⁻⁷ mol / l. In the case of AAP10, values above 70% were not reached and concentrations did not exceed 74% until the concentration exceeded 10 ⁻⁸ mol / l.

The novel materials of the present invention are effective even at low concentrations and have a higher maximum achieved effect than AAP10. 90% for new material versus 74% for AAP10 (value%: number of epicardial action potentials representing duration in the range of ± 5 ms near the mean, corresponding to a decrease in dispersion). This means that the new peptides are not only more potent but also more effective than AAP10 in terms of maximal effect, thus making progress compared to AAP10.

Total Vein Risk

As with AAP10, the novel materials of the present invention exhibit extremely low total vein risk compared to conventional chemotherapy (Table 2).

Changes in total venous risk (similarity in the field of vector) in normal treatment concentrations (free plasma concentrations) of AAP10, new peptides, Lidocaine and Flecainide (details in the parameters) (detailed description of parameters and classic antiarrhythmic agents) For results, see Cirulation 87 (1993) 617). Log concentration AAP10 AAP13TT Lidocaine Flecaineid contrast 29 ± 3 29 ± 6 23 ± 2 23 ± 2 -10 24 ± 3 23 ± 4 -9 23 ± 2 18 ± 5 -8 22 ± 3 17 ± 5 -7 21 ± 2 17 ± 6 26 ± 2 -6.3 19 ± 3 -5.698 26 ± 2 -5.824 10 ± 2 -5.3 23 ± 1 -5 17 ± 3

If the field of vector shows less similarity, the material of the present invention shows a greater total venous effect. All concentrations correspond to normal untreated plasma concentrations. Among conventional Class I antiarrhythmic agents, lidocaine is known to have a relatively low risk of venous vein, whereas flecaid is believed to have a very high risk of venous vein.

Claims (3)

A compound of formula 1 Formula 1 In the formula, R is H or OH, X is Ala, Arg, Gly or Val, Y is Pro or His, Z is H, F, Cl, Br or I, Provided that when X is Gly, Y is Pro, and R is OH, then Z is not H. H ₂ N-Gly-Ala-Gly-Hyp-Pro-3-iodotyrosineamide. The compound of formula 1 according to claim 1 used for disease control.