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Discovery of Small Peptides and Peptidomimetics Targeting the Substance P 1-7 Binding Site: Focus on Design, Synthesis, Structure-Activity Relationships and Drug-Like Properties
Uppsala University, Disciplinary Domain of Medicine and Pharmacy, Faculty of Pharmacy, Department of Medicinal Chemistry.
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Biologically active peptides are important for many physiological functions in the human body and therefore serve as interesting starting points in drug discovery processes. In this work the neuropeptide substance P 1–7 (SP1–7, H-Arg-Pro-Lys-Pro-Gln-Gln-Phe-OH), which has been demonstrated to reduce neuropathic pain and attenuate opioid withdrawal symptoms in animal models, has been addressed in a medicinal chemistry program with the overall aim of transforming this bioactive peptide into more drug-like compounds. Specific binding sites for this neuropeptide have been detected in the brain and the spinal cord. Interestingly, the smaller neuropeptide endomorphin-2 (EM-2, H-Tyr-Pro-Phe-Phe-NH2) also interacts with these binding sites, although 10-fold less efficient.

In this work the structure–activity relationship of SP1–7 and EM-2, regarding their affinity to the SP1–7 binding site was elucidated using alanine scans, truncation, and terminal modifications. The C-terminal part of both peptides, and especially the C-terminal phenylalanine, was crucial for binding affinity. Moreover, the C-terminal functional group should preferably be a primary amide. The truncation studies finally resulted in the remarkable discovery of H-Phe-Phe-NH2 as an equally good binder as the heptapeptide SP1–7. This dipeptide amide served as a lead compound for further studies. In order to improve the drug-like properties and to find a plausible bioactive conformation, a set of rigidified and methylated dipeptides of different stereochemistry, and analogs with reduced peptide character, were synthesized and evaluated regarding binding, metabolic stability and absorption. Small SP1–7 analogs with retained affinity and substantially improved permeability and metabolic stability were identified.

Beside peptide chemistry the synthetic work included the development of a fast and convenient microwave-assisted protocol for direct arylation of imidazoles. Furthermore, microwave-assisted aminocarbonylation using Mo(CO)6 as a solid carbon monoxide source was investigated in the synthesis of MAP amides and for coupling of imidazoles with amino acids.

In a future perspective the present findings, together with the fact that some of the SP1–7 analogs discovered herein have been shown to reproduce the biological effects of SP1-7 in animal studies related to neuropathic pain and opioid dependence, can ultimately have an impact on drug discovery in these two areas.

Place, publisher, year, edition, pages
Uppsala: Acta Universitatis Upsaliensis , 2011. , 88 p.
Series
Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy, ISSN 1651-6192 ; 143
Keyword [en]
substance P 1-7, peptidomimetics, structure-activity relationship, drug-like properties, phenylalanine, imidazole, MAP aryl amides, carbonylation
National Category
Medicinal Chemistry
Research subject
Medicinal Chemistry
Identifiers
URN: urn:nbn:se:uu:diva-149480ISBN: 978-91-554-8040-0OAI: oai:DiVA.org:uu-149480DiVA: diva2:406174
Public defence
2011-05-06, B42, Husargatan 3, Uppsala, 09:15 (English)
Opponent
Supervisors
Available from: 2011-04-15 Created: 2011-03-20 Last updated: 2011-05-05Bibliographically approved
List of papers
1. Small peptides mimicking substance P (1-7) and encompassing a C-terminal amide functionality
Open this publication in new window or tab >>Small peptides mimicking substance P (1-7) and encompassing a C-terminal amide functionality
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2008 (English)In: Neuropeptides, ISSN 0143-4179, E-ISSN 1532-2785, Vol. 42, no 1, 31-37 p.Article in journal (Refereed) Published
Abstract [en]

Some of the biological effects demonstrated after administration of substance P (SP) in vivo can indirectly be attributed to the fragmentation of the undecapeptide to its N-terminal bioactive fragment SP1–7. This heptapeptide (H-Arg-Pro-Lys-Pro-Gln-Gln-Phe-OH) is a major bioactive metabolite from SP that frequently exerts similar biological effects as the parent peptide but also, in several cases, completely opposite actions. Specific binding sites for the heptapeptide SP1–7 that are separate from the SP preferred NK receptors have been identified. In this study we demonstrate that (a) the C-terminal part of the SP metabolite SP1–7 is most important for binding as deduced from an Ala scan and that a replacement of Phe7 for Ala is deleterious, (b) truncation of the N-terminal amino acid residues of SP1–7 delivers peptides with retained binding activity, although with somewhat lower binding affinities than SP1–7 and (c) a C-terminal amide group as a replacement for the terminal carboxy group of SP1–7 and for all of the truncated ligands synthesized affords approximately 5–10-fold improvements of the binding affinities.

Keyword
Substance P (1–7), SP (1–7), SP1–7, Substance P, Structure–activity relationships (SAR), Ala scan, Peptidomimetics
National Category
Pharmaceutical Sciences
Identifiers
urn:nbn:se:uu:diva-97823 (URN)10.1016/j.npep.2007.11.002 (DOI)000254231100003 ()18093649 (PubMedID)
Available from: 2008-11-21 Created: 2008-11-21 Last updated: 2014-11-12Bibliographically approved
2. Discovery of Dipeptides with High Affinity to the Specific Binding Site for Substance P1-7
Open this publication in new window or tab >>Discovery of Dipeptides with High Affinity to the Specific Binding Site for Substance P1-7
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2010 (English)In: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 53, no 6, 2383-2389 p.Article in journal (Refereed) Published
Abstract [en]

Substance P 1-7 (SP1-7, H-Arg-Pro-Lys-Pro-Gln-Gln-Phe-OH) is the major bioactive metabolite of substance P. The interest in this heptapeptide originates from the observation that it modulates, and in certain cases opposes the effects of the parent peptide. e.g., the nociceptive effect. The p-opioid receptor agonist endomorphin-2 (EM-2, H-Tyr-Pro-Phe-Phe-NH2) has been found to also interact with the specific binding site of SP1-7 with only a 10-fold lower affinity compared to the native peptide. Considering the smaller size of EM-2 compared to the target heptapeptide, it was selected as a lead compound in the development of low-molecular-weight ligands to the SP1-7 binding site. An alanine scan and truncation study led to the unexpected discovery of the dipeptide H-Phe-Phe-NH2 (K-i = 1.5 nM), having equal affinity as the endogenous heptapeptide SP1-7. Moreover, the studies show that the C-terminal phenylalanine amide is crucial for the affinity of the dipeptide.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-136801 (URN)10.1021/jm901352b (DOI)000275711000006 ()20178322 (PubMedID)
Available from: 2010-12-14 Created: 2010-12-14 Last updated: 2012-10-03Bibliographically approved
3. Constrained H-Phe-Phe-NH2 Analogues With High Affinity to the Substance P 1-7 Binding Site and With Improved Metabolic Stability and Cell Permeability
Open this publication in new window or tab >>Constrained H-Phe-Phe-NH2 Analogues With High Affinity to the Substance P 1-7 Binding Site and With Improved Metabolic Stability and Cell Permeability
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2013 (English)In: Journal of Medicinal Chemistry, ISSN 0022-2623, E-ISSN 1520-4804, Vol. 56, no 12, 4953-4965 p.Article in journal (Refereed) Published
Abstract [en]

We recently reported the discovery of H-Phe-Phe-NH2 as a small and high affinity ligand for the substance P 1-7 (SP1-7, H-Arg-Pro-Lys-Pro-Gln-Gln-Phe-OH) specific binding site and its intriguing ability to reduce neuropathic pain. With the overall aim to develop stable and orally bioavailable SP1-7 mimetics, the dipeptide was chosen as a lead compound. Herein the structure-activity relationship (SAR) of a set of modified H-Phe-Phe-NH2 analogues is presented together with their potential active uptake by PEPT1 transporter, intestinal permeability, and metabolic stability. Local constraints via peptide backbone methylation or preparation of cyclized analogues based on pyrrolidine were evaluated and were shown to significantly improve the in vitro pharmacokinetic properties. The SAR was rationalized by deriving a plausible binding pose for the high affinity ligands. Rigidification using a 3-phenylpyrrolidine moiety in the C-terminal of H-Phe-Phe-NH2 resulted in high affinity and improved intrinsic clearance and intestinal epithelial permeability.

National Category
Medical and Health Sciences
Identifiers
urn:nbn:se:uu:diva-149482 (URN)10.1021/jm400209h (DOI)000321237100011 ()23735006 (PubMedID)
Available from: 2011-03-24 Created: 2011-03-20 Last updated: 2013-09-09Bibliographically approved
4. Discovery and Pharmacokinetic Profiling of Phenylalanine Based Carbamates as Novel Substance P 1-7 Analogues
Open this publication in new window or tab >>Discovery and Pharmacokinetic Profiling of Phenylalanine Based Carbamates as Novel Substance P 1-7 Analogues
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(English)Manuscript (preprint) (Other academic)
Identifiers
urn:nbn:se:uu:diva-149995 (URN)
Available from: 2011-03-24 Created: 2011-03-24 Last updated: 2011-05-03
5. Design and Synthesis of N-Terminal Imidazole Based H-Phe-Phe-NH2 Mimetics
Open this publication in new window or tab >>Design and Synthesis of N-Terminal Imidazole Based H-Phe-Phe-NH2 Mimetics
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(English)Manuscript (preprint) (Other academic)
Identifiers
urn:nbn:se:uu:diva-149996 (URN)
Available from: 2011-03-24 Created: 2011-03-24 Last updated: 2011-05-03
6. Microwave-Assisted Synthesis of Weinreb and MAP Aryl Amides via Pd-Catalyzed Heck Aminocarbonylation Using Mo(CO)(6) or W(CO)(6)
Open this publication in new window or tab >>Microwave-Assisted Synthesis of Weinreb and MAP Aryl Amides via Pd-Catalyzed Heck Aminocarbonylation Using Mo(CO)(6) or W(CO)(6)
2011 (English)In: Journal of Organic Chemistry, ISSN 0022-3263, E-ISSN 1520-6904, Vol. 76, no 3, 978-981 p.Article in journal (Refereed) Published
Abstract [en]

A simple and expedient process for the Heck aminocarbonylative synthesis of Weinreb and MAP amide acylating agents, from aryl halides, is reported. This methodology utilizes solid sources of CO making it readily accessible to chemists working in small-scale laboratory applications.

National Category
Pharmaceutical Sciences
Identifiers
urn:nbn:se:uu:diva-148925 (URN)10.1021/jo102151u (DOI)000286577400027 ()
Available from: 2011-03-15 Created: 2011-03-14 Last updated: 2013-07-15Bibliographically approved

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