Nafarelin Acetate

Description

Nafarelin acetate is a synthetic decapeptide agonist analog of the natural gonadotropin-releasing hormone, differing in the amino acid at position 6 where D-naphthylalanine has been substituted for glycine. This substitution gives nafarelin greater potency and a substantially longer half-life than the endogenous hormone. Nafarelin acetate administered as a nasal spray, suppresses estrogen levels on repeated dosing and is indicated for the management of endometriosis. It also has orphan drug status for use in precocious puberty.

Originator

Syntex (USA)

Uses

Nafarelin Acetate is a gonadotropin-releasing hormone agonist. Treatment with Nafarelin improves sleep disturbances and anxiety-depression. Nafarelin is also used in the treatment of estrogen-dependent gynecologic disorders such as endometriosis and uterine leiomyomas.

Uses

LHRH agonist.

Manufacturing Process

In the reaction vessel of a Beckman 990 Peptide Synthesizer was placed 0.8 g (0.8 mmol) of benzhydrylamino-polystyrene-divinylbenzene resin (Lab Systems, Inc.) as described by Rivaille, supra. Amino acids were added sequentially to this resin by means of the usual methods of Boc-strategy of peptide synthesis on above copolymer.
The resin was coupled sequentially with a 2.5 molar excess of each protected amino acid and DCC. Thus, the resin was treated during successive coupling cycles with 0.433 g Boc-Gly-OH, 0.432 g Boc-Pro-OH, 0.857 g Boc-Arg(Tosyl)- OH, 0.462 g Boc-Leu-OH, 0,504 g Boc-3-(2-naphthyl)-D-alanine and 0.272 g 1-hydroxybenzotriazole, 0.724 g N-Boc, O-2-bromobenzoyloxycarbonyl-Ltyrosine, 0.59 g Boc-Ser(Benzyl)-OH, 0.608 g Boc-Trp-OH, 0.654 g BocHis(Tosyl)-OH and 0.524 g pyroglutamic acid. A coupling cycle for one amino acid and completeness of the reaction is checked by the ninhydrin method of E. Kaiser, et al., Anal. Biochem., 34, 595 (1970).
The resin was removed from the reaction vessel, washed with CH2Cl2, and dried in vacuo to yield 2.0 g of protected polypeptide resin.
The polypeptide product was simultaneously removed from the resin and completely deprotected by treatment with anhydrous liquid HF. A mixture of 2.0 g of protected polypeptide resin and 2 mL of anisole (scavenger) in a KelF reaction vessel was treated with 20 mL of redistilled (from CoF3) anhydrous liquid HF at 0°C for 30 minutes. The HF was evaporated under vacuum and the residue of (pyro)-Glu-His-Trp-Ser-Tyr-3-(2-naphthyl)-D-alanyl-Leu-ArgPro-Gly-NH2,as its HF salt, was washed with ether. The residue was then extracted with glacial acetic acid. The acetic acid extract was lyophilized to yield 0.8 g of crude material. The crude polypeptide was loaded on a 4x40 cm. Amberlite XAD-4 column (polystyrene-4% divinylbenzene copolymer) and eluted with a concave gradient from water (0.5 L) to ethanol (1 L). The tubes containing fractions from effluent volume 690 mL to 1,470 mL were pooled and stripped to dryness to yield 490 mg of partially purified polypeptide.
A 150 mg sample of the partially purified product was subjected to partition chromatography on a 3 times 50 cm. column of Sephadex G-25 using the solvent system 1-butanol/toluene/acetic acid/water containing 1.5% pyridine in the ratios 10:15:12:18. The pure fractions were pooled on the basis of thin layer chromatography (silica gel; BuOH/H2O/HOAc/EtOAc; 1:1:1:1) and HPLC (5 micron, reverse phase, octadecylsilyl packing; 40% 0.03 M NH4OAc/60% acetonitrile). The desired product came off the column in fractions from effluent volume 1,000 mL to 1,400 mL (Rf 0.1). The pure fractions were pooled, stripped to dryness, taken up in H2O, and lyophilized to yield 57 mg of pure pyro-glutamyl-histidyl-tryptophylseryl-tyrosyl-3-(2-naphthyl)-D-alanylleucyl-arginylprolyl-glycinamide, as its acetic acid addition salt, [α]D25-27.4° (c 0.9, HOAc), m.p. 185°-193°C (dec.).

brand name

Synarel (Searle).

Therapeutic Function

Gonadotropic

Clinical Use

Nafarelin acetate, contains D Nal(2)6 [Nal = 3-(2-naphthyl)-Ala] in place of Gly6, but the C-terminus, Gly10-NH2, is identical with natural GnRH. Nafarelin acetate is available as a 0.2% nasal spray for the relief of endometriosis. Estrogen, of course, is needed for the growth of endometrial tissue; thus, decreased estrogen leads to shrinkage of errant endometrial tissue. The observed side effects of nafarelin acetate are related to falling estrogen levels and include decreased libido, amenorrhea, hot flashes, and vaginal dryness. When used consistently, nafarelin acetate will inhibit ovulation and stop menstruation.
Nafarelin acetate also is used in children, male and female, for the treatment of central precocious puberty. By suppressing the release of LH, the estradiol or testosterone levels fall to prepubertal levels; early secondary sexual development is arrested, linear growth and skeletal maturation are slowed, and in girls, menstruation stops.