scholarly journals In Silico, In Vitro and In Vivo Pharmacodynamic Characterization of Novel Analgesic Drug Candidate Somatostatin SST4 Receptor Agonists

2021 ◽  
Vol 11 ◽  
Author(s):  
Boglárka Kántás ◽  
Éva Szőke ◽  
Rita Börzsei ◽  
Péter Bánhegyi ◽  
Junaid Asghar ◽  
...  
Keyword(s):  
Oral Administration ◽  
G Protein ◽  
In Silico ◽  
The Novel ◽  
Pain Model ◽  
Protein Activation ◽  
Single Oral Administration ◽  
G Protein Activation

Background: Somatostatin released from the capsaicin-sensitive sensory nerves mediates analgesic and anti-inflammatory effects via its receptor subtype 4 (SST4) without influencing endocrine functions. Therefore, SST4 is considered to be a novel target for drug development in pain, especially chronic neuropathy which is a great unmet medical need.Purpose and Experimental Approach: Here, we examined the in silico binding, SST4-linked G protein activation and β-arrestin activation on stable SST4 expressing cells and the effects of our novel pyrrolo-pyrimidine molecules (20, 100, 500, 1,000, 2,000 µg·kg−1) on partial sciatic nerve ligation-induced traumatic mononeuropathic pain model in mice.Key Results: The novel compounds bind to the high affinity binding site of SST4 the receptor and activate the G protein. However, unlike the reference SST4 agonists NNC 26-9100 and J-2156, they do not induce β-arrestin activation responsible for receptor desensitization and internalization upon chronic use. They exert 65–80% maximal anti-hyperalgesic effects in the neuropathy model 1 h after a single oral administration of 100–500 µg·kg−1 doses.Conclusion and Implications: The novel orally active compounds show potent and effective SST4 receptor agonism in vitro and in vivo. All four novel ligands proved to be full agonists based on G protein activation, but failed to recruit β-arrestin. Based on their potent antinociceptive effect in the neuropathic pain model following a single oral administration, they are promising candidates for drug development.

scholarly journals Gelsolin Modulates Phospholipase C Activity In Vivo through Phospholipid Binding

1997 ◽  
Vol 138 (4) ◽  
pp. 811-820 ◽  
Author(s):  
Hui-qiao Sun ◽  
Keng-mean Lin ◽  
Helen L. Yin
Keyword(s):  
G Protein ◽  
Live Cells ◽  
Dependent Manner ◽  
Permeabilized Cells ◽  
Protein Activation ◽  
G Protein Activation ◽  
Effective Inhibition ◽  
Actin Regulatory Proteins

Gelsolin and CapG are actin regulatory proteins that remodel the cytoskeleton in response to phosphatidylinositol 4,5-bisphosphate (PIP2) and Ca2+ during agonist stimulation. A physiologically relevant rise in Ca2+ increases their affinity for PIP2 and can promote significant interactions with PIP2 in activated cells. This may impact divergent PIP2- dependent signaling processes at the level of substrate availability. We found that CapG overexpression enhances PDGF-stimulated phospholipase Cγ (PLCγ) activity (Sun, H.-q., K. Kwiatkowska, D.C. Wooten, and H.L. Yin. 1995. J. Cell Biol. 129:147–156). In this paper, we examined the ability of gelsolin and CapG to compete with another PLC for PIP2 in live cells, in semiintact cells, and in vitro. We found that CapG and gelsolin overexpression profoundly inhibited bradykinin-stimulated PLCβ. Inhibition occurred at or after the G protein activation step because overexpression also reduced the response to direct G protein activation with NaF. Bradykinin responsiveness was restored after cytosolic proteins, including gelsolin, leaked out of the overexpressing cells. Conversely, exogenous gelsolin added to permeabilized cells inhibited response in a dose-dependent manner. The washout and addback experiments clearly establish that excess gelsolin is the primary cause of PLC inhibition in cells. In vitro experiments showed that gelsolin and CapG stimulated as well as inhibited PLCβ, and only gelsolin domains containing PIP2-binding sites were effective. Inhibition was mitigated by increasing PIP2 concentration in a manner consistent with competition between gelsolin and PLCβ for PIP2. Gelsolin and CapG also had biphasic effects on tyrosine kinase– phosphorylated PLCγ, although they inhibited PLCγ less than PLCβ. Our findings indicate that as PIP2 level and availability change during signaling, cross talk between PIP2-regulated proteins provides a selective mechanism for positive as well as negative regulation of the signal transduction cascade.

scholarly journals A novel nucleoside rescue metabolic pathway may be responsible for therapeutic effect of orally administered cordycepin

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Jong Bong Lee ◽  
Masar Radhi ◽  
Elena Cipolla ◽  
Raj D. Gandhi ◽  
Sarir Sarmad ◽  
...  
Keyword(s):  
Oral Administration ◽  
Metabolic Pathway ◽  
Oral Absorption ◽  
Therapeutic Effects ◽  
The Novel ◽  
Drug Candidates ◽  
Biopharmaceutical Properties

Abstract Although adenosine and its analogues have been assessed in the past as potential drug candidates due to the important role of adenosine in physiology, only little is known about their absorption following oral administration. In this work, we have studied the oral absorption and disposition pathways of cordycepin, an adenosine analogue. In vitro biopharmaceutical properties and in vivo oral absorption and disposition of cordycepin were assessed in rats. Despite the fact that numerous studies showed efficacy following oral dosing of cordycepin, we found that intact cordycepin was not absorbed following oral administration to rats. However, 3′-deoxyinosine, a metabolite of cordycepin previously considered to be inactive, was absorbed into the systemic blood circulation. Further investigation was performed to study the conversion of 3′-deoxyinosine to cordycepin 5′-triphosphate in vitro using macrophage-like RAW264.7 cells. It demonstrated that cordycepin 5′-triphosphate, the active metabolite of cordycepin, can be formed not only from cordycepin, but also from 3′-deoxyinosine. The novel nucleoside rescue metabolic pathway proposed in this study could be responsible for therapeutic effects of adenosine and other analogues of adenosine following oral administration. These findings may have importance in understanding the physiology and pathophysiology associated with adenosine, as well as drug discovery and development utilising adenosine analogues.

scholarly journals Leishmania Proteomics: An in Silico Perspective

2020 ◽  
Author(s):  
Carlos A. Padilla ◽  
María J. Álvarez ◽  
Aldo F. Combariza
Keyword(s):  
Geographical Distribution ◽  
In Silico ◽  
Chemical Species ◽  
Biologically Active ◽  
Infectious Episode ◽  
Protein Activation ◽  
Enzymatic Function ◽  
Active Chemical

We report on the state of the art of scientific literature about proteins recognized as potential targets for the development of Leishmania treatments through the search of biologically active chemical species, either from experimental in vitro, in vivo, or in silico sources. We classify the gathered information, in several ways: vector taxonomy and geographical distribution, parasite taxonomic and geographical distribution and enzymatic function (oxidoreductases, transferases, hydrolases, lyases, isomerases, ligases and cytokines). Our aim is to provide a much needed reference layout for research efforts aimed to understand the underpinning physical interactions in ligand-protein activation/inactivation processes. In the specific case of Leishmania, we focus on enzymes known to be part of the biochemical molecular processes initiated following a Leishmania infectious episode.

A comprehensive pharmacokinetic/pharmacodynamics analysis of the novel IGF1R/INSR inhibitor BI 893923 applying in vitro, in vivo and in silico modeling techniques

2016 ◽  
Vol 77 (6) ◽  
pp. 1303-1314 ◽  
Author(s):  
Melanie I. Titze ◽  
Otmar Schaaf ◽  
Marco H. Hofmann ◽  
Michael P. Sanderson ◽  
Stephan K. Zahn ◽  
...  
Keyword(s):  
In Silico ◽  
The Novel ◽  
In Silico Modeling ◽  
Modeling Techniques

scholarly journals Leishmania Proteomics: An in Silico Perspective

2019 ◽  
Author(s):  
Carlos A. Padilla ◽  
Maria J. Alvarez ◽  
Aldo Combariza
Keyword(s):  
Geographical Distribution ◽  
In Silico ◽  
Chemical Species ◽  
Cascade Reactions ◽  
Biologically Active ◽  
Infectious Episode ◽  
Protein Activation ◽  
Enzymatic Function

We report on the state of the art of proteins recognized as potential targets for the development of leishmania treatments through the search of biologically active chemical species, either from experimental in vitro, in vivo, or in silico sources.  We classify the gathered information, in several ways: vector taxonomy and geographical distribution, leishmania parasite taxonomic and geographical distribution and enzymatic function (oxidoreductases, transferases, hydrolases, lyases, isomerases, ligases and cytokines). Our aim is to provide a much needed reference layout for research efforts aimed to understand the background of ligand-protein activation/inactivation processes, in this specific case, related with enzymes known to be part of biochemical cascade reactions initiated following a leishmania infectious episode.

scholarly journals Adhesion G protein-coupled receptors are activated by exposure of a cryptic tethered agonist

2015 ◽  
Vol 112 (19) ◽  
pp. 6194-6199 ◽  
Author(s):  
Hannah M. Stoveken ◽  
Alexander G. Hajduczok ◽  
Lei Xu ◽  
Gregory G. Tall
Keyword(s):  
G Protein ◽  
Clinical Utility ◽  
G Protein Coupled Receptors ◽  
Activation Mechanism ◽  
Inhibitory Influence ◽  
Neighboring Cell ◽  
Protein Activation ◽  
G Protein Activation ◽  
G Protein Coupled

The large class of adhesion G protein-coupled receptors (aGPCRs) bind extracellular matrix or neighboring cell-surface ligands to regulate organ and tissue development through an unknown activation mechanism. We examined aGPCR activation using two prototypical aGPCRs, GPR56 and GPR110. Active dissociation of the noncovalently bound GPR56 or GPR110 extracellular domains (ECDs) from the respective seven-transmembrane (7TM) domains relieved an inhibitory influence and permitted both receptors to activate defined G protein subtypes. After ECD displacement, the newly revealed short N-terminal stalk regions of the 7TM domains were found to be essential for G protein activation. Synthetic peptides comprising these stalks potently activated GPR56 or GPR110 in vitro or in cells, demonstrating that the stalks comprise a tethered agonist that was encrypted within the ECD. Establishment of an aGPCR activation mechanism provides a rational platform for the development of aGPCR synthetic modulators that could find clinical utility toward aGPCR-directed disease.

scholarly journals Specific inhibition of GPCR-independent G protein signaling by a rationally engineered protein

2017 ◽  
Vol 114 (48) ◽  
pp. E10319-E10328 ◽  
Author(s):  
Anthony Leyme ◽  
Arthur Marivin ◽  
Marcin Maziarz ◽  
Vincent DiGiacomo ◽  
Maria P. Papakonstantinou ◽  
...  
Keyword(s):  
G Protein ◽  
Rational Design ◽  
G Protein Signaling ◽  
Guanine Nucleotide ◽  
Protein Signaling ◽  
Nucleotide Exchange ◽  
Protein Activation ◽  
Conserved Sequence ◽  
G Protein Activation

Activation of heterotrimeric G proteins by cytoplasmic nonreceptor proteins is an alternative to the classical mechanism via G protein-coupled receptors (GPCRs). A subset of nonreceptor G protein activators is characterized by a conserved sequence named the Gα-binding and activating (GBA) motif, which confers guanine nucleotide exchange factor (GEF) activity in vitro and promotes G protein-dependent signaling in cells. GBA proteins have important roles in physiology and disease but remain greatly understudied. This is due, in part, to the lack of efficient tools that specifically disrupt GBA motif function in the context of the large multifunctional proteins in which they are embedded. This hindrance to the study of alternative mechanisms of G protein activation contrasts with the wealth of convenient chemical and genetic tools to manipulate GPCR-dependent activation. Here, we describe the rational design and implementation of a genetically encoded protein that specifically inhibits GBA motifs: GBA inhibitor (GBAi). GBAi was engineered by introducing modifications in Gαi that preclude coupling to every known major binding partner [GPCRs, Gβγ, effectors, guanine nucleotide dissociation inhibitors (GDIs), GTPase-activating proteins (GAPs), or the chaperone/GEF Ric-8A], while favoring high-affinity binding to all known GBA motifs. We demonstrate that GBAi does not interfere with canonical GPCR-G protein signaling but blocks GBA-dependent signaling in cancer cells. Furthermore, by implementing GBAi in vivo, we show that GBA-dependent signaling modulates phenotypes during Xenopus laevis embryonic development. In summary, GBAi is a selective, efficient, and convenient tool to dissect the biological processes controlled by a GPCR-independent mechanism of G protein activation mediated by cytoplasmic factors.

scholarly journals Novel Drug-Like Somatostatin Receptor 4 Agonists are Potential Analgesics for Neuropathic Pain

2019 ◽  
Vol 20 (24) ◽  
pp. 6245 ◽  
Author(s):  
Boglárka Kántás ◽  
Rita Börzsei ◽  
Éva Szőke ◽  
Péter Bánhegyi ◽  
Ádám Horváth ◽  
...  
Keyword(s):  
Neuropathic Pain ◽  
G Protein ◽  
Somatostatin Receptor ◽  
Sensory Nerves ◽  
Mechanical Hyperalgesia ◽  
Protein Activation ◽  
G Protein Activation ◽  
In Vivo Testing ◽  
Chronic Neuropathy

Somatostatin released from the capsaicin-sensitive sensory nerves mediates analgesic and anti-inflammatory effects via the somatostatin sst4 receptor without endocrine actions. Therefore, sst4 is considered to be a novel target for drug development in pain including chronic neuropathy, which is an emerging unmet medical need. Here, we examined the in silico binding, the sst4-linked G-protein activation on stable receptor expressing cells (1 nM to 10 μM), and the effects of our novel pyrrolo-pyrimidine molecules in mouse inflammatory and neuropathic pain models. All four of the tested compounds (C1–C4) bind to the same binding site of the sst4 receptor with similar interaction energy to high-affinity reference sst4 agonists, and they all induce G-protein activation. C1 is the more efficacious (γ-GTP-binding: 218.2% ± 36.5%) and most potent (EC50: 37 nM) ligand. In vivo testing of the actions of orally administered C1 and C2 (500 µg/kg) showed that only C1 decreased the resiniferatoxin-induced acute neurogenic inflammatory thermal allodynia and mechanical hyperalgesia significantly. Meanwhile, both of them remarkably reduced partial sciatic nerve ligation-induced chronic neuropathic mechanical hyperalgesia after a single oral administration of the 500 µg/kg dose. These orally active novel sst4 agonists exert potent anti-hyperalgesic effect in a chronic neuropathy model, and therefore, they can open promising drug developmental perspectives.

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