The neuropeptide y y(1) receptor: a diagnostic marker? Expression in mcf-7 breast cancer cells is down-regulated by antiestrogens in vitro and in xenografts.

The neuropeptide Y (NPY) Y(1) receptor (Y(1)R) has been suggested as a tumor marker for in vivo imaging and as a therapeutic target. In view of the assumed link between estrogen receptor (ER) and Y(1)R in mammary carcinoma and with respect to the development of new diagnostic tools, we investigated the Y(1)R protein expression in human MCF-7 cell variants differing in ER content and sensitivity against antiestrogens. ER and Y(1)R expression were quantified by radioligand binding using [(3)H]-17ß-estradiol and the Y(1)R selective antagonist [(3)H]-UR-MK114, respectively. The latter was used for cellular binding studies and for autoradiography of MCF-7 xenografts. The fluorescent ligands Cy5-pNPY (universal Y(1)R, Y(2)R and Y(5)R agonist) and UR-MK22 (selective Y(1)R antagonist), as well as the selective antagonists BIBP3226 (Y(1)R), BIIE0246 (Y(2)R) and CGP71683 (Y(5)R) were used to identify the NPY receptor subtype(s) by confocal microscopy. Y(1)R functionality was determined by mobilization of intracellular Ca(2+). Sensitivity of MCF-7 cells against antiestrogen 4-hydroxytamoxifen correlated directly with the ER content. The exclusive expression of Y(1)Rs was confirmed by confocal microscopy. The Y(1)R protein was up-regulated (100%) by 17ß-estradiol (EC(50) 20 pM) and the predominant role of ERα was demonstrated by using the ERα-selective agonist "propylpyrazole triol". 17ß-Estradiol-induced over-expression of functional Y(1)R protein was reverted by the antiestrogen fulvestrant (IC(50) 5 nM) in vitro. Furthermore, tamoxifen treatment of nude mice resulted in an almost total loss of Y(1)Rs in MCF-7 xenografts. In conclusion, the value of the Y(1)R as a target for therapy and imaging in breast cancer patients may be compromised due to Y(1)R down-regulation induced by hormonal (antiestrogen) treatment.

The bivalent ligand approach leads to highly potent and selective acylguanidine-type histamine H2 receptor agonists.

Bivalent histamine H(2) receptor (H(2)R) agonists were synthesized by connecting pharmacophoric 3-(2-amino-4-methylthiazol-5-yl)-, 3-(2-aminothiazol-5-yl)-, 3-(imidazol-4-yl)-, or 3-(1,2,4-triazol-5-yl)propylguanidine moieties by N(G)-acylation with alkanedioic acids of various chain lengths. The compounds were investigated for H(2)R agonism in GTPase and [(35)S]GTPγS binding assays at guinea pig (gp) and human (h) H(2)R-Gsα(S) fusion proteins including various H(2)R mutants, at the isolated gp right atrium, and in GTPase assays for activity on recombinant H(1), H(3), and H(4) receptors. The bivalent ligands are H(2)R partial or full agonists, up to 2 orders of magnitude more potent than monovalent acylguanidines and, with octanedioyl or decanedioyl spacers, up to 4000 times more potent than histamine at the gpH(2)R. In contrast to their imidazole analogues, the aminothiazoles are highly selective for H(2)R vs other HR subtypes. Compounds with (theoretically) sufficient spacer length (20 CH(2) groups) to simultaneously occupy two orthosteric binding sites in H(2)R dimers are nearly inactive, whereas the highest potency resides in compounds with considerably shorter spacers. Thus, there is no evidence for interaction with H(2)R dimers. The high agonistic potency may result from interaction with an accessory binding site at the same receptor protomer.