Considerable advances have been made in the study of opioid receptor structure and function, and in recent years evidence has accumulated that opioid receptors can interact with one another. Early studies demonstrated that antagonism of delta opioid receptors (DOR) prevents the development of tolerance and dependence to morphine. Opioid receptors, like most G-protein coupled receptors, can form dimers and higher order oligomers. Heterodimerization has been proposed to lead to altered signaling pathways, and heterodimerization of DOR with mu opioid receptors (MOR) has been proposed to be responsible for the modulation of morphine analgesia. Thus interacting MOR and DOR is an attractive target for drug development. Affinity labels, compounds that bind to their targets covalently, provide a unique approach to studying ligand-receptor interactions and offer advantages over reversible ligands for a number of studies. This competitive renewal application builds on our successes during the previous grant period in the design and identification of affinity label derivatives of opioid peptides for MOR and DOR. During the two-year grant period these studies will be expanded to include the use of these peptide-based affinity labels to study coexpressed MOR and DOR at the molecular level. The proposed research consists of two specific aims: 1) The synthesis of additional opioid peptide-based affinity labels for DOR that are needed for studies of coexpressed receptors, and the incorporation of additional functionalities, i.e., a fluorescent group, into selected peptides. Selected peptides will be evaluated for opioid receptor affinity and wash-resistant binding to opioid receptors in cells coexpressing MOR and DOR as well as in cells expressing a single opioid receptor. Affinity label derivatives will be used to examine the interaction of MOR and DOR in cellular studies;a novel multifunctional peptide will be used in receptor isolation studies to determine how the ligand binds to its receptor;and 2) The design of novel opioid peptide analogs containing an affinity label that are tailored to bind to interacting MOR and DOR;these peptides will undergo detailed pharmacological evaluation in cells coexpressing both receptors. As a result of this research we expect to identify affinity label derivatives of opioid peptides that retain high affinity and bind covalently to their receptors. By studying the interactions of these unique and novel pharmacological tools with opioid receptors in cells coexpressing MOR and DOR we will obtain valuable information on the interactions of these two receptors as well as on ligand-receptor interactions. Such information could be crucial in developing novel types of narcotic agents with improved pharmacological profiles and decreased side effects.
Opioid narcotic analgesics such as morphine have been a mainstay in the treatment of severe pain, but these agents have serious side effects such as dependence, tolerance and respiratory depression that limit their therapeutic use. Despite considerable effort in designing new compounds, there has been limited success to date in improving the therapeutic profiles of narcotic analgesics. Thus there is a need to develop new analgesics and other agents targeting opioid receptors with decreased side effects and improved therapeutic profiles. The proposed affinity label derivatives of opioid peptides represent unique pharmacological tools that can be used to study interacting mu and delta opioid receptors. The information on receptor-receptor interactions obtained from these studies could be crucial in developing new classes of narcotic agents with improved therapeutic profiles.
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