Ganirelix Acetate

Originator

Antagon,Organon

Definition

ChEBI: Ganirelix is a polypeptide.

Manufacturing Process

The abbreviations for common aminoacids are those recommended by IUPACIUB Comission on Biochemical Nomenclature. Other abbreviations useful in describing the replacements of aminoacids in the natural LH-RH peptide are following:
Nal(2) - 3-(2-naphthyl)alanyl; p-Cl-Phe - 3-(p-chlorophenyl)alanyl; Pal(3) - 3- (3-pyridyl)alanyl; ; hArg(Et)2 - NG,NG - bis(ethtyl)homoarginyl; Boc - tbutyloxycarbonyl.
Ganirelix (N-Ac-Nal(2)-D-pCl-Phe-D-Pal(3)-Ser-Tyr-D-hArg(Et)2-Leu-hArg(Et)2- Pro-Ala-NH2) was prepared using the following side chain protection protocol: salt protection for L- and D-hArg(Et)2 (as the chloride) and t-butyl protection for serine.
Amino acids were added to the Nα-Boc-D-Ala-O-Resin (1.0 mmol of resin was replaced in the reaction vessel of 5.0 L Vega 296 automated solid phase peptide synthesizer; in the following sequence:
Acetic anhydride
An acetylation (capping of the resin) was done after Ala, Pro and Leu with N,N'-diisopropyl carbodiimide - 1-hydroxybenztriazole (HBt). Excess HBt (2 equiv.) was used for the coupling of the basic amino acids, hArg(Et)2 and Pal(3).
The following protocols were used to remove the Nα-protecting group following each addition.
Program A: The resin was first washed with CH2Cl2 1 times/1 min, TFA-CH2Cl2 (40/60) 1 times/1 min, TFA-CH2Cl2 (40/60) 1 times/30 min, CH2Cl2 2 5 times/1 min, Et3N-CH2Cl2 (5/95) 3 times/1 min, CH2Cl2 4 times/1 min.
Program B: The resin was first washed with CH2Cl2 1 times/1 min, 4-4.5 N HCl in CH2Cl2/i-PrOH (1/1) 1 times/1 min, 4-4.5 N HCl in CH2Cl2/i-PrOH (1/1) 1 times/30 min, CH2Cl2 3 times/1 min, DMF 1 times/1 min, Et3N-CH2Cl2 (5/95) 3 times/1 min, DMF 1 times/1 min, CH2Cl2 4 times/1 min.
After each deprotecting and washing step, following protocol A or B, the next amino acid in sequence was added and the resin washed with CH2Cl2 3 times/1 min, MeOH 4 times/1 min, DMF 2 times/1 min and CH2Cl2 4 times/1 min.
Program A was used for the removal of the protecting groups on Ala, Pro, LhArg(Et)2, Leu and D-Nal(2).
Program B was used for the removal of the protecting groups on D-hArg(Et)2, Tyr, Ser, D-Pal(3) and p-Cl-Phe.
The crude peptide was first dissolved in 2 M acetic acid and converted to its acetate salt by passage through a column of AG3-X4A resin (Bio-Rad). The acetate was subjected to chromatography on a silica gel column (CH2Cl2/iPrOH/MeOH/H2O/HOAc solvent); the acetate fractions dissolved in H2O and loaded onto a reversed-phase column (Vydec C-18, 15-20 μ), and purified using acetonitrile/TEAP (pH 3). Fractions of the desired purity were combined and diluted with water and reloaded on a reversed-phase HPLC column, then washed with 1% acetic acid in water. The peptide was stripped with a mixture of MeOH/CH3CN/HOAc/H2O (44/50/1/5). The residue was dissolved in acetic acid and precipitated over ether, filtered, washed with ether and dried under vacuum. Amino acid analyses were performed on a Beckman 119CL amino acid analyzer. Samples for amino acid analyses were hydrolyzed with 6 N HCl at 110°C for 20 hrs. Analytical HPLC was performed on a Spectra Physics 8800 chromatograph. Synthesis of ganirelix was confirmed by the presence of a main peak at rt 18 min; no other peak over 1% was noted at rt 16 min.

Therapeutic Function

LHRH antagonist

Clinical Use

Ganirelix acetate is an analog of GnRH with substitutions at residues 1, 2, 3, 6, 8, and 10. It is not a superagonist but, rather, is a synthetic decapeptide with high antagonist activity and the first GnRH antagonist to be marketed. It is approved for the suppression of LH surges in women who are undergoing ovarian hyperstimulation fertility treatment; LH surges normally promote ovulation. The goal of this drug is to significantly reduce the number of medication days necessary to suppress the LH surge, thereby maintaining eggs in the ovaries. In vitro fertilization (IVF) treatment cycles were historically initiated by the administration of leuprolide acetate to suppress the premature release of LH. This inhibits ovulation so that the eggs remain available for retrieval by a fertility specialist. For this purpose, leuprolide acetate usually is injected for as many as 26 days. Clinical studies have shown that ganirelix acetate can shut down the LH surge in only 5 days of treatment, that the suppression of LH is more pronounced than that of FSH, and that the shorter treatment time minimizes unpleasant side effects, such as hot flashes and headaches.

Description

Ganirelix acetate is a synthetic decapeptide with high antagonistic activity against naturally occurring gonadotropin-releasing hormone (GnRH). Ganirelix acetate is derived from native GnRH with substitutions of amino acids at positions 1, 2, 3, 6, 8, and 10 to form the following molecular formula of the peptide: N-acetyl-3-(2-napthyl)-D-alanyl-4-chloro-D-phenylalanyl-3-(3-pyridyl)-D-alanyl-L-seryl-L-tyrosyl-N 9 ,N 10 -diethyl- D-homoarginyl-L-leucyl-N 9 ,N 10 -diethyl-L-homoarginyl-L-prolyl-D-alanylamide acetate. The molecular weight for Ganirelix acetate is 1570.4 Dalton as an anhydrous free base.