Irina Zeltser

Backbone Cyclic Peptide Antagonists, Derived from the Insect Pheromone Biosynthesis Activating Neuropeptide, Inhibit Sex Pheromone Biosynthesis in Moths

We describe an application of the backbone cyclization and cycloscan concept for the design and synthesis of pheromone biosynthesis activating neuropeptide (PBAN) antagonists capable of inhibiting sex pheromone biosynthesis in Heliothis peltigera female moths. Two backbone cyclic (BBC) sub-libraries were designed and synthesized. The structure of the first sub-library ([Arg27]PBAN27–33NH2, termed the Ser sub-library) was based on the active C-terminal hexapeptide sequence (Tyr-Phe-Ser-Pro-Arg-Leu-NH2) of PBAN1–33NH2, which was found to comprise its active core. The second sub-library ([Arg27,d-Phe30]PBAN27–33NH2, termed the d-Phe sub-library) was based on the sequence of the lead antagonist Arg-Tyr-Phe-(d)Phe-Pro-Arg-Leu-NH2. In both sub-libraries the Pro residue was replaced by anN α(ω-amino-alkyl)Gly building unit having various lengths of the alkyl chain. All the cyclic peptides in each sub-library had the same primary sequence and the same location of the ring. The members of each library differed from each other by the bridge size and bridge chemistry. Screening of the two libraries for pheromonotropic antagonists resulted in the disclosure of four compounds that fully inhibited sex pheromone biosynthesis at 1 nmol and were devoid of agonistic activity. All antagonistic peptides originated from the d-Phe sub-library. Substitution of thed-Phe30 amino acid with a Ser resulted in a loss of antagonistic activity. Agonistic activities were exhibited by peptides from both sub-libraries.

Discovery of a linear lead antagonist to the insect pheromone biosynthesis activating neuropeptide (PBAN).

We report the discovery of a linear lead antagonist for the insect pheromone biosynthesis activating neuropeptide (PBAN) which inhibits sex pheromone biosynthesis in the female moth Heliothis peltigera. Two approaches have been used in attempting to convert PBAN agonists into antagonists. The first involved omission of the C-terminal amide and reduction of the sequence from the N-terminus in a linear library based on PBAN 1-33NH(2.) The second involved replacement of L amino-acids by the D hydrophobic amino acid D-Phe in a linear library based on PBAN28-33NH(2.) Screening of the two libraries for pheromonotropic antagonists resulted in the disclosure of one compound out of the D-Phe library (Arg-Tyr-Phe-D-Phe-Pro-Arg-Leu-NH(2)) which inhibited sex pheromone production by 79 and 64% at 100 pmol in two moth colonies and exhibited low agonistic activity. Omission of the C-terminal amide in PBAN 1-33NH(2) and its shorter analogs did not lead to the discovery of an antagonistic compound.

Advances in the application of neuropeptides in insect control

Abstract The development of a new approach for the generation of a novel type of putative insect control agents based on backbone cyclic peptidomimetic antagonists of insect-neuropeptides is reported. The approach, termed the backbone cyclic neuropetide based on autogonist (BBC-NBA) was applied to the insect pyrokinin/pheromone biosynthesis activating neuropeptide (PBAN) family as a model, and led to the discovery of a potent linear lead antagonist and several highly potent, metabolically stable BBC peptidomimetic antagonists, devoid of agonistic activity, which inhibited in vivo PBAN-mediated activities in moths.

Inhibition of PK/PBAN-mediated functions in insects: Discovery of selective and non-selective inhibitors☆

The antagonistic properties of a few linear and backbone cyclic (BBC) conformationally constraint peptide libraries and their analogs, were tested for the ability to inhibit pyrokinin/pheromone biosynthesis activating neuropeptide (PK/PBAN) mediated functions: sex pheromone biosynthesis in Heliothis peltigera female moths, cuticular melanization in Spodoptera littoralis larvae, pupariation in the fleshfly Neobellieria bullata and hindgut contraction in Leucophaea maderae, elicited by exogenously injected PBAN, pheromonotropin (PT), leucopyrokinin (LPK), myotropin (MT) or by the endogenous peptides. The data revealed differential inhibitory patterns within the same assay with different elicitors (in both the pheromonotropic and melanotropic assays) and among the different functions and disclosed selective antagonists, hinting at the possibility that the receptors that mediate those functions may differ from one another structurally.

Backbone cyclic pheromone biosynthesis activating neuropeptide (PBAN) antagonists: inhibition of melanization in the moth Spodoptera littoralis (Insecta, Lepidoptera).

Antagonistic and agonistic activities of backbone cyclic (BBC) pheromone biosynthesis activating neuropeptide (PBAN) analogues were evaluated in an attempt to identify potent melanotropic antagonists, to gain an insight into their structure-activity relationship (SAR), and to discover molecules with selective and non-selective melanotropic and pheromonotropic properties. Eight potent melanotropic BBC antagonists and seven agonists were disclosed. SAR studies revealed that the structural requirements of the melanotropic and pheromonotropic agonists and antagonists are different. The cyclic structure of the BBC peptides was unimportant for antagonistic activity, and linearization retained their melanotropic and pheromonotropic antagonistic properties. Comparison of the antagonistic activities of the BBC and precyclic peptides with respect to both functions revealed eight selective antagonists (six that were selective melanotropic antagonists and two selective pheromonotropic antagonists) and four non-selective (melanotropic and pheromonotropic) antagonists. The selective melanotropic antagonists exhibited both, pure or mixed agonistic/antagonistic activities. The selective pheromonotropic compounds were pure antagonists. All non-selective compounds were pure antagonists. Comparison of the agonistic activities of the BBC peptides with respect to both functions revealed six selective melanotropic agonists and one non-selective agonistic compound. All compounds (whether selective or non-selective) exhibited pure agonistic activity. Discovery of the selective compounds hints at the possibility that the receptors that mediate the respective activities may have different properties.

Pyrokinin/PBAN radio-receptor assay: development and application for the characterization of a putative receptor from the pheromone gland of Heliothis peltigera<

A radio-receptor assay (RRA) for the insect pyrokinin/PBAN family has been developed. The development involved examination of the ligand (3H-tyrosyl-PBAN28-33NH2)-receptor interaction under various incubation conditions and variations on sex pheromone gland membrane preparation. Application of the RRA for a partial characterization of the putative pyrokinin/PBAN receptor in the pheromone gland of H. peltigera revealed age-dependence of its expression. Pharmacological characterization revealed a high correlation between the binding-affinity to the receptor of various PBAN-derived peptides and their in vivo pheromonotropic bioactivity, and shed light on the interaction of backbone cyclic and linear ([Arg27,D-Phe30]PBAN28-33NH2) PBAN antagonists with the receptor.

Rationally designed neuropeptide antagonists: A novel approach for generation of environmentally friendly insecticides

We report our approach for the generation of a novel type of putative insecticides based on backbone cyclic peptidomimetic antagonists of insect neuropeptides using pheromone biosynthesis activating neuropeptide (PBAN) as a model. This approach, called the backbone cyclic neuropeptide based antagonist (BBC-NBA), includes the following steps: (i) elucidation of the active sequence of the chosen insect neuropeptide; (ii) disclosure of a lead antagonist based on the sequence found in step (i); (iii) design and synthesis of backbone cyclic peptide libraries (cycloscan) based on the sequence of the lead antagonist; and (iv) design and synthesis of a peptidomimetic prototype insecticide. The BBC-NBA approach was applied to PBAN and led to the discovery of a potent linear lead antagonist and a potent backbone cyclic antagonist devoid of agnoistic activity which inhibited sex pheromone biosynthesis inHeliothis peltigera female moths.