The long term objective of this research is to understand the structural basis by which Pro-Leu-Gly-NH2 (PLG) and its peptidomimetic analogues modulate CNS dopamine receptors and to elucidate the molecular mechanism underlying this modulation. In pursuing this objective, we plan to synthesize analogues of highly potent PLG peptidomimetics to test the hypothesis that the carbonyl groups in these molecules are important for their intrinsic activity. Analogues will also be made which are substituted with hydrophobic groups capable of mimicking the leucyl side chain found in PLG. Synthesis of analogues of the diketopiperazine lactam peptidomimetic will be made to test the hypothesis that it is the type II beta-turn conformation that is the important conformational feature of dopamine receptor modulating activity while the N-terminal C5 conformation is the structural feature responsible for the potency of the PLG peptidomimetics. Different sized ring analogues of the highly constrained spiro bicyclic lactam PLG peptidomimetic will be made so as to alter the phi2, psi2, and phi3 torsion angles and thereby determine the effect this has on dopamine receptor modulating activity. Finally, two general types of potential radioligands for the putative PLG receptor will be obtained and evaluated. One type will be a tritiated diketopiperazine lactam peptidomimetic,while in a second approach the 4- hydroxybenzyl group will be incorporated into the structures of the highly potent PLG analogues for the purpose of radiolabeling these compounds with 125I. The PLG peptidomimetics synthesized will be evaluated for their ability to modulate (3H]spiroperidol/N- propylnorapomorphine competitive binding to D2 receptors in the presence or absence of 5 prime-guanylylimidodiphosphate, a norhydrolyzable analog of GTP. They will also be evaluated for their ability to enhance the binding of dopamine receptor agonists to either the D1, D2, D3 D4 or D5 receptors using SH-SY5Y human neuroblastoma cell lines transfected with either the D1,D2, D3, D4 or D5 receptor genes. Selected PLG peptidomimetics will be tested for their activity in in vivo animal models of Parkinson's disease (6- hydroxydopamine-lesioned rat model and the MPTP mouse model) and dyskinesia disorders (haloperidol-induced D2 receptor supersensitivity model and the L-DOPA-induced dyskinesia model). Finally, studies will be conducted to further determine the functional interaction of PLG and its bioactive analogues with G-proteins in the striatum through the use of specific antisense phosphothioate oligonucleotides by examining: (1) the ability of Gs and Gi antisense oligonucleotides to modulate rotational behavior induced by dopamine agonists in 6-hydroxydopamine lesioned rats in the presence and absence of PLG analogues; (2) the G protein levels and mRNA expression for Gi alter treating rats with D2 receptor agonists in the absence and presence of PLG peptidomimetics, and (3) the effect of PLG peptidomimetics on GTPase activity of purified G-proteins uncoupled from the receptor. These studies will enhance our understanding of dopamine receptor modulation and provide groundwork for the development of new therapeutic agents for the treatment of such neurological disorders as Parkinson's disease and tardive dyskinesia.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
2R01NS020036-11
Application #
2263754
Study Section
Bio-Organic and Natural Products Chemistry Study Section (BNP)
Project Start
1983-12-01
Project End
1999-07-31
Budget Start
1995-09-30
Budget End
1996-07-31
Support Year
11
Fiscal Year
1995
Total Cost
Indirect Cost
Name
University of Minnesota Twin Cities
Department
Pharmacology
Type
Schools of Pharmacy
DUNS #
168559177
City
Minneapolis
State
MN
Country
United States
Zip Code
55455
Basu, Dipannita; Tian, Yuxin; Bhandari, Jayant et al. (2013) Effects of the dopamine D2 allosteric modulator, PAOPA, on the expression of GRK2, arrestin-3, ERK1/2, and on receptor internalization. PLoS One 8:e70736
Basu, Dipannita; Castellano, Jessica M; Thomas, Nancy et al. (2013) Cell-free protein synthesis and purification of human dopamine D2 receptor long isoform. Biotechnol Prog 29:601-8
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Bhagwanth, Swapna; Mishra, Ram K; Johnson, Rodney L (2013) Development of peptidomimetic ligands of Pro-Leu-Gly-NH(2) as allosteric modulators of the dopamine D(2) receptor. Beilstein J Org Chem 9:204-14
Bhagwanth, Swapna; Mishra, Satyendra; Daya, Ritesh et al. (2012) Transformation of Pro-Leu-Gly-NH2 peptidomimetic positive allosteric modulators of the dopamine D2 receptor into negative modulators. ACS Chem Neurosci 3:274-84
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Mann, Amandeep; Verma, Vaneeta; Basu, Dipannita et al. (2010) Specific binding of photoaffinity-labeling peptidomimetics of Pro-Leu-Gly-NH2 to the dopamine D2L receptor: evidence for the allosteric modulation of the dopamine receptor. Eur J Pharmacol 641:96-101
Wood, Richard L; Young-Dixon, Brendan J; Roy, Abhrajeet et al. (2010) Evaluation of Density Functionals, SCC-DFTB, Neglect of Diatomic Differential Overlap (NDDO) Models and Molecular Mechanics Methods for Prolyl-Leucyl-Glycinamide (PLG) and Structural Derivatives. Theochem 944:76-82
Raghavan, Bhooma; Skoblenick, Kevin J; Bhagwanth, Swapna et al. (2009) Allosteric modulation of the dopamine D2 receptor by Pro-Leu-Gly-NH2 peptidomimetics constrained in either a polyproline II helix or a type II beta-turn conformation. J Med Chem 52:2043-51

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