Gastrin releasing peptide antagonists with improved potency and stability

Abstract

Gastrin releasing peptide (GRP) is a 27 amino acid peptide hormone which is homologous to the amphibian peptide bombesin. Two series of novel GRP antagonists were developed by C-terminal modification of N-acetyl-GRP-20-27 amide. Peptide derivatives within each series resist enzymatic degradation in serum and exhibit strong affinity for the GRP receptor. The first series of compounds replaces the Leu26-Met27 region of GRP with an alkyl ether N-acetyl-GRP-20-25-NH-[(S)-1-ethoxy-4-methyl-2-pentane], specifically blocked radiolabeled GRP binding with an IC50 of 6 nM. In the second series of antagonists the oxygen of the ether moiety is replaced with a methylene group, resulting in GRP antagonists which are equipotent to native GRP in receptor binding assays (IC50 = 2 nM) and are also resistant to proteolytic degradation in vitro. All of the C-terminally modified peptides tested blocked GRP-stimulated mitogenesis in Swiss 3T3 mouse fibroblasts. Representative compounds also blocked GRP-induced elevation of [Ca2+]i in human SCLC cells, and inhibited GRP-independent release of gastrin in vivo.

Development of potent gastrin-releasing peptide antagonists having a D-Pro-psi(CH2NH)-Phe-NH2 C terminus.

“…Cells were maintained by serial passage at 37°C in a humidified atmosphere containing 5% C02/95% air in Dulbecco's modified Eagle's medium/Ham's nutrient mixture F-12 (DMEM/F-12; Mediatech, Herndon, VA) supplemented with 10% (vol/vol) fetal bovine serum (lot 882050; Biologos, Naperville, IL) on plastic ware. Our passages [7][8][9][10][11][12][13][14][15][16][17][18][19][20] were employed in these studies. Cells were judged free of mycoplasma by coculture on Vero cells followed by Hoechst stain and examination by fluorescence microscopy.…”

“…Bombesin is a 14-residue amphibian peptide that reacts approximately equivalently to GRP in studies ofmammalian responses. The C-terminal Met is naturally amidated and is required for full biologic activity, although it does not appear to be required for binding (12)(13)(14)(15)(16)(17)(18). Coy et al (14,15) replaced the terminal peptide bond with a reduced form, T(CH2NH), yielding a reasonably potent antagonist, [13T14,CH2NH]bombesin, and have further explored the use of the T bond to make more potent antagonists (11).…”

“…Coy et al (14,15) replaced the terminal peptide bond with a reduced form, T(CH2NH), yielding a reasonably potent antagonist, [13T14,CH2NH]bombesin, and have further explored the use of the T bond to make more potent antagonists (11). Other antagonists have been made by replacing the terminal Met-NH2 with either an alkyl ester (16) or an alkyl amide (17,18). Specific members of these three classes of antagonists have potencies 21 nM, which is the potency of the natural hormone.…”

International Union of Pharmacology. LXVIII. Mammalian Bombesin Receptors: Nomenclature, Distribution, Pharmacology, Signaling, and Functions in Normal and Disease States

“…A final group of potent antagonists in this class were synthesized by J. Martínez's group, with the most potent being JMV641 and JMV594 (Azay et al, 1996;Lamharzi et al, 1998). JMV641 [H-D-Phe,Gln,Trp,Ala, Val,Gly,His-NH-*CH[CH 2 -CH(CH 3 ) 2 ]-**CHOH-(CH 2 ) 3 -CH 3 [where * is (S) and ** is 92% of (S isomer)], contains a pseudopeptide bond that mimics the transition state analog (K i murine BB 2 0.85 nM) (Azay et al, 1996) and has a Ͼ3000-fold selectivity for the BB 2 over the BB 1 receptor (Tokita et al, 2001b (where statine ϭ 4-amino-3-hydroxy-6-methylhepatanonoic acid) also has a high affinity for the murine BB 2 receptor (K i 0.60 nM) (Azay et al, 1998;Llinares et al, 1999) and has Ͼ5000-fold selectivity for the BB 2 over the BB 1 receptor (Tokita et al, 2001b) ( Table 2 Jensen et al, 1993;de Castiglione and Gozzini, 1996), but vary widely in chemical groups attached, including desMet amides (Heimbrook et al, 1989;Wang et al, 1990a,b), alkylamides (Camble et al, 1989;Heimbrook et al, 1989;Wang et al, 1990a,b), esters (Heimbrook et al, 1989;Wang et al, 1990b;Coy et al, 1992b), hydrazides (Wang et al, 1990b), and with other COOH-terminal groups attached (Heimbrook et al, 1989(Heimbrook et al, , 1991 (Fig. 1; Table 2).…”

Theranostic Perspectives in Prostate Cancer with the Gastrin-Releasing Peptide Receptor Antagonist NeoBOMB1: Preclinical and First Clinical Results

“…In the quest of radioligands with higher stability, our attention was attracted by the improved stability, and potency, reported for the well-characterized GRPR antagonist Ac-His-Trp-Ala-Val-GlyHis-NH-CH[CH 2 -CH(CH 3 ) 2 ] 2 (26). By replacing the C-terminal -His 12 -Leu 13 NHEt of SB3 by -His 12 -NH-CH[CH 2 -CH(CH 3 ) 2 ] 2 , we produced NeoBOMB1, which we labeled with 68 Ga (for PET), 111 In (for SPECT), and 177 Lu (for radionuclide therapy).…”