Synthesis of novel basic head‐to‐side‐chain cyclic dynorphin A analogs: Strategies and side reactions

Abstract

Novel N-terminus-to-side-chain cyclic analogs of the opioid peptide dynorphin (Dyn) A-(1-11)NH(2) were prepared that retain the basicity of the N-terminal amine and restrict the backbone conformation around the important Tyr(1) residue. Cyclic peptides were synthesized in which the N-terminal amine and the N(epsilon)-amine of a Lys at position 3 or 5 were attached to the alpha-carbon and carbonyl of an acetyl group, respectively. Several synthetic strategies were explored with detailed analysis of the side reactions in order to obtain the desired cyclic peptides. One of the side reactions observed involved premature loss of the N-terminal 9-fluorenylmethoxycarbonyl (Fmoc) group during the neutralization step following deprotection of the Mtt (4-methyltrityl) protecting group from the side chain of Lys. The successful strategy involved the synthesis of the linear peptide up through Gly(2) and functionalization through the N(epsilon)-amine of Lys. A linear N-terminal alkylated analog was prepared by alkylation of the peptide on the resin with an equimolar amount of bromoacetamide, followed by treatment of the peptide with Fmoc-OSu prior to cleavage from the resin to facilitate separation by reversed phase high performance liquid chromatography of unreacted peptide from the desired alkylated product. The novel N-terminal cyclic Dyn A analogs and the linear analog were evaluated for their opioid receptor affinities. These peptides exhibited large losses in affinity for opioid receptors; the low affinity of the linear N-terminal alkylated peptide suggested that the alpha-acetamide group on the N-terminal amine resulted in unfavorable interactions with opioid receptors.

Amino Acid-Protecting Groups

“…The N α -Fmoc protecting group can be prematurely removed by a primary amine of sufficient basicity, such as the ε-amino group of Lys and to a lesser extent the δ-amino of Orn and the γ-amino of Dab, present in the peptide. , This side reaction is not promoted by either the β-amino side chain of the Dap residue or the α-amino group. These results are consistent with the p K a values of these amino functions in the model compounds shown in Table .…”

Cyclic Endomorphin Analogs in Targeting Opioid Receptors to Achieve Pain Relief

“…Over the years a great number of cyclic analogs of DOR-selective enkephalins and deltorphins [44,[50][51][52][53] and KOR-selective dynorphins [54,55] have been synthesized through a variety of strategies, including several types of sulfur-containing bridges, carbonyl linkages, guanidine bridges [see 56 for review].…”

Cyclic Opioid Peptides

“…L/DTrp 3 and Pro 3 substitution in the cyclic variant decreased potency by up to 390-fold arbitrated by the adenylyl cyclase assay. In order to maintain basicity at the N-terminus and constrain Tyr 1 in DYN A, an acetyl linker between the N-terminus and the side-chain of Lys was implemented in Dyn A (1–11)-NH 2 [78]. Considering that positions 3 and 5 of Dyn A have been shown to not be critical for opioid activity, cyclizations spanned from the head to the 3 rd or 5 th position which result in 15- and 21-membered ring, respectively.…”