Bifunctional [2',6' -dimethyl -L -tyrosine1]endomorphin-2 analogues substituted at position 3 with alkylated phenylalanine derivatives yield potent mixed μ -agonist/δ -antagonist and dual μ-agonist/δ-agonist opioid ligands

Endomorphin-2 (H-Tyr-Pro-Phe-Phe-NH2) and [Dmt1]EM-2 (Dmt = 2‘,6‘-dimethyl-l-tyrosine) analogues, containing alkylated Phe3 derivatives, 2‘-monomethyl (2, 2‘), 3‘,5‘- and 2‘,6‘-dimethyl (3, 3‘, and 4‘, respectively), 2‘,4‘,6‘-trimethyl (6, 6‘), 2‘-ethyl-6‘-methyl (7, 7‘), and 2‘-isopropyl-6‘-methyl (8, 8‘) groups or Dmt (5, 5‘), had the following characteristics:  (i) [Xaa3]EM-2 analogues exhibited improved μ- and δ-opioid receptor affinities. The latter, however, were inconsequential (Kiδ = 491−3451 nM). (ii) [Dmt1,Xaa3]EM-2 analogues enhanced μ- and δ-opioid receptor affinities (Kiμ = 0.069−0.32 nM; Kiδ = 1.83−99.8 nM) without κ-opioid receptor interaction. (iii) There were elevated μ-bioactivity (IC50 = 0.12−14.4 nM) and abolished δ-agonism (IC50 > 10 μM in 2‘, 3‘, 4‘, 5‘, 6‘), although 4‘ and 6‘ demonstrated a potent mixed μ-agonism/δ-antagonism (for 4‘, IC50μ = 0.12 and pA2 = 8.15; for 6‘, IC50μ = 0.21 nM and pA2 = 9.05) and 7‘ was a dual μ-agonist/δ-agonist (IC50μ = 0.17 nM; IC50δ = 0.51 nM).