Teresa Tarragó

Baicalin, a prodrug able to reach the CNS, is a prolyl oligopeptidase inhibitor

Prolyl oligopeptidase is a cytosolic serine peptidase that hydrolyzes proline-containing peptides at the carboxy terminus of proline residues. It has been associated with schizophrenia, bipolar affective disorder, and related neuropsychiatric disorders and therefore may have important clinical implications. In a previous work, we used (19)F NMR to search for new prolyl oligopeptidase inhibitors from a library of traditional Chinese medicine plant extracts, and identified several extracts as powerful inhibitors of this peptidase. Here, the flavonoid baicalin was isolated as the active component of an extract of Scutellaria baicalensis roots having prolyl oligopeptidase inhibitory activity. Baicalin inhibited prolyl oligopeptidase in a dose-dependent manner. Inhibition experiments using baicalin analogs showed that the sugar moiety was not necessary for activity. The IC(50)s of baicalin and its aglycone derivative baicalein were rather similar, showing that the sugar moiety was not involved in the interaction of baicalin with POP. These results were confirmed by saturation transfer difference NMR experiments. To further understand the absorption and transport mechanisms of baicalin and baicalein, we evaluated their transport in vitro through the gastrointestinal tract and the blood-brain barrier using a Parallel Artificial Membrane Permeability Assay. The molecule which potentially crosses both barriers was identified as baicalein, the aglycone moiety of baicalin. Our results show that baicalin is a new prodrug able to inhibit prolyl oligopeptidase. As baicalin is a natural compound with a long history of safe administration to humans, it is a highly attractive base from which to develop new treatments for schizophrenia, bipolar affective disorder, and related neuropsychiatric diseases.

Comparative effects of β2-adrenergic agonists on muscle waste associated with tumour growth

The implantation of the Yoshida AH-130 ascites hepatoma (a fast growing tumour) to rats resulted in a dramatic loss of both white adipose tissue and muscle (skeletal and cardiac) mass. Administration of beta2-adrenergic agonists to tumour-bearing rats resulted in a partial recovery of skeletal muscle and heart mass. Treatment of the tumour-bearing animals with the different drugs (salbutamol, salmeterol and clenbuterol) did not influence tumour growth or food intake so it can be suggested that the effects were solely due to metabolic changes. In addition, while the three drugs had clear effects on gastrocnemius muscles, clenbuterol and salbutamol had also an effect on soleus, and salbutamol had a clear effect on cardiac muscle. It is suggested that any of the studied beta2-adrenergic agonists (but perhaps, particularly salmeterol) could be used clinically in the treatment of cancer cachexia.

A Signaling Mechanism Coupling Netrin-1/Deleted in Colorectal Cancer Chemoattraction to SNARE-Mediated Exocytosis in Axonal Growth Cones

Directed cell migration and axonal guidance are essential steps in neural development. Both processes are controlled by specific guidance cues that activate the signaling cascades that ultimately control cytoskeletal dynamics. Another essential step in migration and axonal guidance is the regulation of plasmalemma turnover and exocytosis in leading edges and growth cones. However, the cross talk mechanisms linking guidance receptors and membrane exocytosis are not understood. Netrin-1 is a chemoattractive cue required for the formation of commissural pathways. Here, we show that the Netrin-1 receptor deleted in colorectal cancer (DCC) forms a protein complex with the t-SNARE (target SNARE) protein Syntaxin-1 (Sytx1). This interaction is Netrin-1 dependent both in vitro and in vivo, and requires specific Sytx1 and DCC domains. Blockade of Sytx1 function by using botulinum toxins abolished Netrin-1-dependent chemoattraction of axons in mouse neuronal cultures. Similar loss-of-function experiments in the chicken spinal cord in vivo using dominant-negative Sytx1 constructs or RNAi led to defects in commissural axon pathfinding reminiscent to those described in Netrin-1 and DCC loss-of-function models. We also show that Netrin-1 elicits exocytosis at growth cones in a Sytx1-dependent manner. Moreover, we demonstrate that the Sytx1/DCC complex associates with the v-SNARE (vesicle SNARE) tetanus neurotoxin-insensitive vesicle-associated membrane protein (TI-VAMP) and that knockdown of TI-VAMP in the commissural pathway in the spinal cord results in aberrant axonal guidance phenotypes. Our data provide evidence of a new signaling mechanism that couples chemotropic Netrin-1/DCC axonal guidance and Sytx1/TI-VAMP SNARE proteins regulating membrane turnover and exocytosis.

all-d proline-rich cell-penetrating peptides : a preliminary in vivo internalization study

Proline-rich cell-penetrating peptides, particularly the SAP (sweet arrow peptide), (VRLPPP)(3), have been proposed to be useful intracellular delivery vectors, as a result of their lack of cytotoxicity combined with their capacity to be internalized by cells. A common limitation of the therapeutic use of peptides is metabolic instability. In general, peptides are quickly degraded by proteases upon entry into the bloodstream. The use of all-D-peptide derivatives is emerging as a fruitful strategy to circumvent this degradation problem. In this context, we report on the internalization behaviour, protease-resistance enhancement and self-assembly properties of an all-D version of SAP [(vrlppp)(3)]. The cellular uptake of (vrlppp)(3) was evaluated in an in vivo assay in mice. Both flow cytometry and confocal laser-scanning microscopy experiments showed that a carboxyfluoresceinated version of the molecule, carboxyfluorescein-(vrlppp)(3), is internalized rapidly in white blood cells and kidney cells. Significant fluorescence was also detected in other organs such as the spleen and the liver. Finally, the toxicity of (vrlppp)(3) was examined, and no significant differences in the main biochemical parameters nor in weight were detected compared with controls.

Identification by 19F NMR of traditional Chinese medicinal plants possessing prolyl oligopeptidase inhibitory activity.

Prolyl oligopeptidase is a cytosolic serine peptidase that hydrolyzes proline‐containing peptides at the carboxy termini of the proline residues. This peptidase has been associated with schizophrenia, bipolar affective disorder, and related neuropsychiatric disorders and might therefore have important clinical implications. Traditional Chinese medicinal (TCM) plants provide a rich source of unexplored compounds for strategies to find novel POP inhibitors, but the traditional methodologies used to identify POP inhibitors could have some limitations when working with natural products: interference with the colorimetric or fluorimetric detection methods commonly used to screen for POP inhibitors can result in the generation of false positives or false negatives. Since NMR screening is less prone to such interference, we decided to explore the use of 19F NMR to screen for POP inhibitors. We synthesized a new 19F‐labeled POP substrate—Z‐Gly‐Pro‐Phe‐4(CF3)‐NH2—and used it to search for new POP inhibitors in TCM plant extracts. We identified several plants with high POP‐inhibitory activity and show here that the combination of 19F NMR and TCM plant extracts is a useful tool for identifying new POP inhibitors.

The Natural Product Berberine is a Human Prolyl Oligopeptidase Inhibitor

Prolyl oligopeptidase is a cytosolic serine peptidase that hydrolyzes proline‐containing peptides at the carboxy terminus. This peptidase has been associated with schizophrenia, bipolar affective disorder, and related neuropsychiatric disorders, and therefore may have important clinical implications. Among the strategies used to find novel prolyl oligopeptidase inhibitors, traditional Chinese medicinal plants provide a rich source of unexplored compounds. We used 19F NMR spectroscopy to search for new prolyl oligopeptidase inhibitors in a library of traditional Chinese medicine plant extracts. Several extracts were identified as powerful inhibitors of this peptidase. The alkaloid berberine was the prolyl oligopeptidase inhibitory molecule isolated from Rhizoma coptidis extract. Berberine inhibited prolyl oligopeptidase in a dose‐dependent manner. As berberine is a natural compound that has been safely administered to humans, it opens up new perspectives for the treatment of neuropsychiatric diseases. The results described herein suggest that the initiation of clinical trials in patients with schizophrenia, bipolar affective disorder, or related diseases in which cognitive capabilities are affected should be undertaken with either the extract or pure BBR.

ENPDA: an evolutionary structure-based de novo peptide design algorithm

SummaryOne of the goals of computational chemists is to automate the de novo design of bioactive molecules. Despite significant advances in computational approaches to ligand design and binding energy evaluation, novel procedures for ligand design are required. Evolutionary computation provides a new approach to this design endeavor. We propose an evolutionary tool for de novo peptide design, based on the evaluation of energies for peptide binding to a user-defined protein surface patch. Special emphasis has been placed on the evaluation of the proposed peptides, leading to two different evaluation heuristics. The software developed was successfully tested on the design of ligands for the proteins prolyl oligopeptidase, p53, and DNA gyrase.

Low molecular weight inhibitors of Prolyl Oligopeptidase: a review of compounds patented from 2003 to 2010.

Introduction: Prolyl Oligopeptidase (POP) is a serine peptidase that cleaves post-proline bonds in short peptides. Besides the direct hydrolytic regulation function over peptides, neuropeptides and peptide hormones, POP is probably involved in the regulation of the inositol pathway and participates in protein-protein interactions. Experimental data show that POP inhibitors have neuroprotective, anti-amnesic and cognition-enhancing properties. These compounds are considered therapeutic agents of interest for the treatment of cognitive deficits related to neuropsychiatric and neurodegenerative diseases, such as Alzheimers disease and Parkinsons disease. Recent findings pointed to the involvement of POP in angiogenesis, although the exact mechanism is still under study. Areas covered: This review comprises patents and patent applications involving POP inhibitors patented between 2003 and 2010, classified as peptidomimetics, heteroaryl ketones and alkaloids. The binding processes and the mechanisms of inhibition of these inhibitors are also discussed, together with their in vivo effects. Expert opinion: The major part of the repertory of POP inhibitors derived from systematical modification of the canonical compound benzyloxycarbonyl-prolyl-prolinal (ZPP). Nevertheless, only two of them have progressed into the clinical trials. One possible reason for this failure is the lack of studies concerning pharmacodynamics, pharmacokinetics and toxicity, together with the absence of suitable animal models. Moreover, POP is still not a well-defined therapeutic target. Further studies are required for the elucidation of the biological role of POP and to validate the therapeutic action of inhibitors in cognitive processes. In contrast, the involvement of POP in protein-protein interactions together with the recent evidences in angiogenesis opens alternative approaches to the traditional active site-directed inhibitors, as well as new therapeutic applications.

Small Peptide Inhibitors Disrupt a High-Affinity Interaction between Cytoplasmic Dynein and a Viral Cargo Protein

ABSTRACT Several viruses target the microtubular motor system in early stages of the viral life cycle. African swine fever virus (ASFV) protein p54 hijacks the microtubule-dependent transport by interaction with a dynein light chain (DYNLL1/DLC8). This was shown to be a high-affinity interaction, and the residues gradually disappearing were mapped on DLC8 to define a putative p54 binding surface by nuclear magnetic resonance (NMR) spectroscopy. The potential of short peptides targeting the binding domain to disrupt this high-affinity protein-protein interaction was assayed, and a short peptide sequence was shown to bind and compete with viral protein binding to dynein. Given the complexity and number of proteins involved in cellular transport, the prevention of this viral-DLC8 interaction might not be relevant for successful viral infection. Thus, we tested the capacity of these peptides to interfere with viral infection by disrupting dynein interaction with viral p54. Using this approach, we report on short peptides that inhibit viral growth.

Recent patents of dipeptidyl peptidase IV inhibitors.

Introduction: The serine exopeptidase DPP IV is a dual protein able to work as an enzyme and an interacting protein. The incretin molecules glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP) are hydrolyzed by DPP IV into inactive forms, which are unable to promote insulin secretion. Therefore, DPP IV is a validated target for the treatment of type 2 diabetes mellitus (T2DM), and a number of inhibitors have been reported in the literature as antidiabetic drugs. Areas covered: DPP IV inhibitor patents from 2006 are included in this review. Documents are classified into chemical groups depending on the main claim. Groups are: i) pyrrolidines and thiazolidines; ii) cyclohexanes, piperidines, piperazines, pyridines and pyrimidines; iii) fused 5-carbon cycles; iv) pyridine, pyrimidine and pyrazine-based bicyclic structures; v) indoles, condensed-imidazoles and xanthines; vi) pyrido-pyrimidines, quinolones, isoquinolines, quinozalines, quinoxalines, naphthyridines, quinolones and quinazolinones; vii) benzoquinolizines, fused aminopiperidines and fused triazoles; viii) other heterocyclic structures and ix) peptidomimetics. Expert opinion: Research in finding new DPP IV inhibitors is intense, despite the number of reported molecules. This is mainly because marketed compounds have been approved in the last 5 years and long-term side effects have not been detected. The perfect inhibitor for the T2DM treatment would therefore be a molecule that inhibits GLP-1 and GIP degradation by DPP IV, but does not affect the activity of the protease in other substrates, nor disturbs the communication of DPP IV with other proteins.