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1.
Exp Parasitol ; 262: 108787, 2024 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-38759776

RESUMEN

New affordable drugs are needed for the treatment of infection with the protozoan parasite Trypanosoma cruzi responsible for the Chagas disease (CD). Only two old drugs are currently available, nifurtimox and benznidazole (Bz) but they exhibit unwanted side effects and display a weak activity in the late chronic phase of the disease. In this context, we evaluated the activity of a series of aryl-pyrazolone derivatives against T cruzi, using both bloodstream trypomastigote and intracellular amastigote forms of the parasite. The test compounds originate from a series of anticancer agents targeting the immune checkpoint ligand PD-L1 and bear an analogy with known anti-trypanosomal pyrazolones. A first group of 6 phenyl-pyrazolones was tested, revealing the activity of a single pyridyl-pyrazolone derivative. Then a second group of 8 compounds with a common pyridyl-pyrazolone core was evaluated. The in vitro testing process led to the identification of two non-cytotoxic and highly potent molecules against the intracellular form of T. cruzi, with an activity comparable to Bz. Moreover, one compound revealed an activity largely superior to that of Bz against bloodstream trypomastigotes, while being non-cytotoxic (selectivity index >1000). Unfortunately, the compound showed little activity in vivo, most likely due to its very limited plasma stability. However, the study opens novel perspectives for the design of new anti-trypanosomal products and the mechanism of action of the compounds is discussed.


Asunto(s)
Enfermedad de Chagas , Pirazolonas , Tripanocidas , Trypanosoma cruzi , Trypanosoma cruzi/efectos de los fármacos , Pirazolonas/farmacología , Pirazolonas/química , Tripanocidas/farmacología , Tripanocidas/química , Animales , Ratones , Enfermedad de Chagas/tratamiento farmacológico , Enfermedad de Chagas/parasitología , Piridinas/farmacología , Piridinas/química , Concentración 50 Inhibidora , Nitroimidazoles/farmacología , Nitroimidazoles/química
2.
Molecules ; 28(8)2023 Apr 15.
Artículo en Inglés | MEDLINE | ID: mdl-37110727

RESUMEN

Orally-active anticancer small molecules targeting the PD-1/PD-L1 immune checkpoint are actively searched. Phenyl-pyrazolone derivatives with a high affinity for PD-L1 have been designed and characterized. In addition, the phenyl-pyrazolone unit acts as a scavenger of oxygen free radicals, providing antioxidant effects. The mechanism is known for the drug edaravone (1) which is also an aldehyde-reactive molecule. The present study reports the synthesis and functional characterization of new molecules (2-5) with an improved anti-PD-L1 activity. The leading fluorinated molecule 5 emerges as a potent checkpoint inhibitor, avidly binding to PD-L1, inducing its dimerization, blocking PD-1/PD-L1 signaling mediated by phosphatase SHP-2 and reactivating the proliferation of CTLL-2 cells in the presence of PD-L1. In parallel, the compound maintains a significant antioxidant activity, characterized using electron paramagnetic resonance (EPR)-based free radical scavenging assays with the probes DPPH and DMPO. The aldehyde reactivity of the molecules was investigated using 4-hydroxynonenal (4-HNE), which is a major lipid peroxidation product. The formation of drug-HNE adducts, monitored by high resolution mass spectrometry (HRMS), was clearly identified and compared for each compound. The study leads to the selection of compound 5 and the dichlorophenyl-pyrazolone unit as a scaffold for the design of small molecule PD-L1 inhibitors endowed with antioxidant properties.


Asunto(s)
Antioxidantes , Receptor de Muerte Celular Programada 1 , Antioxidantes/farmacología , Antioxidantes/metabolismo , Receptor de Muerte Celular Programada 1/metabolismo , Dimerización , Transducción de Señal , Aldehídos
3.
J Cell Biochem ; 123(3): 543-556, 2022 03.
Artículo en Inglés | MEDLINE | ID: mdl-34927768

RESUMEN

Receptor-interacting protein kinase 3 (RIPK3) can induce necroptosis, apoptosis, or cell proliferation and is silenced in several hematological malignancies. We previously reported that RIPK3 activity independent of its kinase domain induces caspase-mediated p65/RelA cleavage, resulting in N-terminal 1-361 and C-terminal 362-549 fragments. We show here that a noncleavable p65/RelA D361E mutant expressed in DA1-3b leukemia cells decreases mouse survival times and that coexpression of p65/RelA fragments increases the tumorigenicity of B16F1 melanoma cells. This aggressiveness in vivo did not correlate with NF-κB activity measured in vitro. The fragments and p65/RelA D361E mutant induced different expression profiles in DA1-3b and B16F1 cells. Stemness markers were affected: p65/RelA D361E increased ALDH activity in DA1-3b cells, and fragment expression increased melanoma sphere formation in B16/F1 cells. p65/RelA fragments and the D361E noncleavable mutant decreased oxidative or glycolytic cell metabolism, with differences observed between models. Thus, p65/RelA cleavage initiated by kinase-independent RIPK3 activity in cancer cells is not neutral and induces pleiotropic effects in vitro and in vivo that may vary across tumor types.


Asunto(s)
Melanoma , FN-kappa B , Animales , Apoptosis , Caspasas/metabolismo , Ratones , FN-kappa B/metabolismo , Fosforilación , Proteína Serina-Treonina Quinasas de Interacción con Receptores/genética , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/farmacología , Factor de Transcripción ReIA/genética , Factor de Transcripción ReIA/metabolismo
4.
Mov Disord ; 37(8): 1761-1767, 2022 08.
Artículo en Inglés | MEDLINE | ID: mdl-35708213

RESUMEN

BACKGROUND: Pathogenic variants in the LRRK2 gene are a common monogenic cause of Parkinson's disease. However, only seven variants have been confirmed to be pathogenic. OBJECTIVES: We identified two novel LRRK2 variants (H230R and A1440P) and performed functional testing. METHODS: We transiently expressed wild-type, the two new variants, or two known pathogenic mutants (G2019S and R1441G) in HEK-293 T cells, with or without LRRK2 kinase inhibitor treatment. We characterized the phosphorylation and kinase activity of the mutants by western blotting. Thermal shift assays were performed to determine the folding and stability of the LRRK2 proteins. RESULTS: The two variants were found in two large families and segregate with the disease. They display altered LRRK2 phosphorylation and kinase activity. CONCLUSIONS: We identified two novel LRRK2 variants which segregate with the disease. The results of functional testing lead us to propose these two variants as novel causative mutations for familial Parkinson's disease. © 2022 The Authors. Movement Disorders published by Wiley Periodicals LLC on behalf of International Parkinson and Movement Disorder Society.


Asunto(s)
Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina , Enfermedad de Parkinson , Células HEK293 , Humanos , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina/genética , Mutación/genética , Enfermedad de Parkinson/genética , Enfermedad de Parkinson/patología , Proteínas Serina-Treonina Quinasas/genética
5.
Int J Mol Sci ; 23(14)2022 Jul 12.
Artículo en Inglés | MEDLINE | ID: mdl-35887019

RESUMEN

The review highlights how protein-protein interactions (PPIs) have determining roles in most life processes and how interactions between protein partners are involved in various human diseases. The study of PPIs and binding interactions as well as their understanding, quantification and pharmacological regulation are crucial for therapeutic purposes. Diverse computational and analytical methods, combined with high-throughput screening (HTS), have been extensively used to characterize multiple types of PPIs, but these procedures are generally laborious, long and expensive. Rapid, robust and efficient alternative methods are proposed, including the use of Microscale Thermophoresis (MST), which has emerged as the technology of choice in drug discovery programs in recent years. This review summarizes selected case studies pertaining to the use of MST to detect therapeutically pertinent proteins and highlights the biological importance of binding interactions, implicated in various human diseases. The benefits and limitations of MST to study PPIs and to identify regulators are discussed.


Asunto(s)
Ensayos Analíticos de Alto Rendimiento , Proteínas , Fenómenos Biofísicos , Descubrimiento de Drogas/métodos , Ensayos Analíticos de Alto Rendimiento/métodos , Humanos , Unión Proteica , Proteínas/química , Temperatura
6.
Molecules ; 27(10)2022 May 21.
Artículo en Inglés | MEDLINE | ID: mdl-35630791

RESUMEN

Small molecules targeting the PD-1/PD-L1 checkpoint are actively searched to complement the anticancer arsenal. Different molecular scaffolds have been reported, including phenyl-pyrazolone derivatives which potently inhibit binding of PD-L1 to PD-1. These molecules are structurally close to antioxidant drug edaravone (EDA) used to treat amyotrophic lateral sclerosis. For this reason, we investigated the capacity of five PD-L1-binding phenyl-pyrazolone compounds (1-5) to scavenge the formation of oxygen free radicals using electron spin resonance spectroscopy with DPPH/DMPO probes. In addition, the reactivity of the compounds toward the oxidized base 5-formyluracil (5fU) was assessed using chromatography coupled to mass spectrometry and photodiode array detectors. The data revealed that the phenyl-pyrazolone derivatives display antioxidant properties and exhibit a variable reactivity toward 5fU. Compound 2 with a N-dichlorophenyl-pyrazolone moiety cumulates the three properties, being a potent PD-L1 binder, a robust antioxidant and an aldehyde-reactive compound. On the opposite, the adamantane derivative 5 is a potent PD-L1 binding with a reduced antioxidant potential and no aldehyde reactivity. The nature of the substituent on the phenyl-pyrazolone core modulates the antioxidant capacity and reactivity toward aromatic aldehydes. The molecular signature of the compound can be adapted at will, to confer additional properties to these PD-L1 binders.


Asunto(s)
Antineoplásicos , Pirazolonas , Aldehídos , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Antioxidantes/farmacología , Antígeno B7-H1/metabolismo , Fluorouracilo , Receptor de Muerte Celular Programada 1
7.
Bioorg Med Chem ; 28(10): 115463, 2020 05 15.
Artículo en Inglés | MEDLINE | ID: mdl-32241621

RESUMEN

1-Phenyl-3-methyl-5-pyrazolone is a reagent, known as PMP, used to derivatize monosaccharides for the study of polysaccharides composition and structure, and for the dosage of carbohydrates in complex media. The same molecule is also known as edaravone, a drug approved for the treatment of stroke and amyotrophic lateral sclerosis. It is also a reactive molecule susceptible to form stable adducts with aromatic aldehydes, such as formylpterin and vanillin. In addition, the molecule serves as a scaffold to design of edaravone analogs and drug conjugates, with various pharmacological properties (antioxidant, anticancer, antiviral). We have analyzed the multiple usages of PMP/edaravone to highlight the reactivity of the molecule and its wide range of applications. This phenyl-pyrazolone compound, considered by many as a biochemical reagent and by other as a clinically useful drug, has not yet revealed the full extent of its capacities and benefits.


Asunto(s)
Esclerosis Amiotrófica Lateral/tratamiento farmacológico , Edaravona/uso terapéutico , Fármacos Neuroprotectores/uso terapéutico , Accidente Cerebrovascular/tratamiento farmacológico , Edaravona/síntesis química , Edaravona/química , Humanos , Estructura Molecular , Fármacos Neuroprotectores/síntesis química , Fármacos Neuroprotectores/química
8.
Bioorg Chem ; 96: 103643, 2020 03.
Artículo en Inglés | MEDLINE | ID: mdl-32035298

RESUMEN

The majority of cancers detected every year are treated with anti-cancer compounds. Unfortunately, many tumors become resistant to antineoplastic drugs. One option is to use cocktails of compounds acting on different targets to try to overcome the resistant cells. This type of approach can produce good results, but is often accompanied by a sharp increase of associated side effects. The strategy presented herein focuses on the use of a single compound acting on two different biological targets enhancing potency and lowering the toxicity of the chemotherapy. In this light, the approach presented in the current study involves the dual inhibition of human pyruvate dehydrogenase kinase-1 (PDHK1) and tubulin polymerization using mono-, di- and tri-chloroacetate-loaded benzophenones and benzothiophenones. Synthesized molecules were evaluated in vitro on tubulin polymerization and on pyruvate dehydrogenase kinase 1. The cell cycle distribution after treatment of DA1-3b leukemic cells with active compounds was tested. Twenty-two benzo(thio)phenones have been selected by the National Cancer Institute (USA) for evaluation of their anti-proliferative potential against NCI-60 cancer cell lines including multidrug-resistant tumor cell lines. Seventeen molecules proved to be very effective in combating the growth of tumor cells exhibiting inhibitory activities up to nanomolar range. The molecular docking of best antitumor molecules in the study was realized with GOLD in the tubulin and PDHK1 binding sites, and allowed to understand the positioning of active molecules. Chloroacetate-loaded benzo(thio)phenones are dual targeted tubulin- and pyruvate dehydrogenase kinase 1 (PDHK1)-binding antitumor agents and exhibited superior antitumor activity compared to non-chlorinated congeners particularly on leukemia, colon, melanoma and breast cancer cell lines.


Asunto(s)
Acetatos/farmacología , Antineoplásicos/farmacología , Benzofenonas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/antagonistas & inhibidores , Moduladores de Tubulina/farmacología , Acetatos/química , Antineoplásicos/química , Benzofenonas/química , Línea Celular Tumoral , Ensayos de Selección de Medicamentos Antitumorales , Humanos , Ligandos , Simulación del Acoplamiento Molecular , Neoplasias/tratamiento farmacológico , Neoplasias/metabolismo , Inhibidores de Proteínas Quinasas/química , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora/metabolismo , Tubulina (Proteína)/metabolismo , Moduladores de Tubulina/química
9.
Arch Pharm (Weinheim) ; 352(5): e1800227, 2019 May.
Artículo en Inglés | MEDLINE | ID: mdl-30947375

RESUMEN

Unprecedented triazinyl-isoxazoles were afforded via an effective cycloaddition reaction between nitrile oxides and the scarcely described 2-ethynyl-4,6-dimethoxy-1,3,5-triazine as dipolarophile. The biological evaluation of the newly synthesized compounds showed that the inhibition of human farnesyltransferase by zinc complexation could be improved with triazine-isoxazole moieties. The replacement of the isoxazole unit by a pyrrolidin-2-one was detrimental to the inhibitory activity while the pyrrolidin-2-thione derivatives conserved the biological potential. The potential of selected compounds to disrupt protein farnesylation in Chinese hamster ovary (CHO) cells transfected with pEGFP-CAAX was also evaluated.


Asunto(s)
Inhibidores Enzimáticos/farmacología , Farnesiltransferasa/antagonistas & inhibidores , Isoxazoles/farmacología , Pirrolidinonas/farmacología , Triazinas/farmacología , Animales , Células CHO , Cricetulus , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/síntesis química , Inhibidores Enzimáticos/química , Farnesiltransferasa/metabolismo , Humanos , Isoxazoles/química , Estructura Molecular , Pirrolidinonas/química , Relación Estructura-Actividad , Triazinas/química
10.
BMC Cancer ; 18(1): 1219, 2018 Dec 04.
Artículo en Inglés | MEDLINE | ID: mdl-30514258

RESUMEN

BACKGROUND: Metastatic melanoma is one of the most aggressive forms of cancer in humans. Among its types, mucosal melanomas represent one of the most highly metastatic and aggressive forms, with a very poor prognosis. Because they are rare in Caucasian individuals, unlike cutaneous melanomas, there has been fewer epidemiological, clinical and genetic evaluation of mucosal melanomas. Moreover, the lack of predictive models fully reproducing the pathogenesis and molecular alterations of mucosal melanoma makes its treatment challenging. Interestingly, dogs are frequently affected by melanomas of the oral cavity that are characterized, as their human counterparts, by focal infiltration, recurrence, and metastasis to regional lymph nodes, lungs and other organs. In dogs, some particular breeds are at high risk, suggesting a specific genetic background and strong genetic drivers. Altogether, the striking homologies in clinical presentation, histopathological features, and overall biology between human and canine mucosal melanomas make dogs invaluable natural models with which to investigate tumor development, including tumor ætiology, and develop tailored treatments. METHODS: We developed and characterized two canine oral melanoma cell lines from tumors isolated from dog patients with distinct clinical profiles; with and without lung metastases. The cells were characterized using immunohistochemistry, pharmacology and genetic studies. RESULTS: We have developed and immunohistochemically, genetically, and pharmacologically characterized. Two cell lines (Ocr_OCMM1X & Ocr_OCMM2X) were produced through mouse xenografts originating from two clinically contrasting melanomas of the oral cavity. Their exhaustive characterization showed two distinct biological and genetic profiles that are potentially linked to the stage of malignancy at the time of diagnosis and sample collection of each melanoma case. These cell lines thus constitute relevant tools with which to perform genetic and drug screening analyses for a better understanding of mucosal melanomas in dogs and humans. CONCLUSIONS: The aim of this study was to establish and characterize xenograft-derived canine melanoma cell lines with different morphologies, genetic features and pharmacological sensitivities that constitute good predictive models for comparative oncology. These cell lines are relevant tools to advance the use of canine mucosal melanomas as natural models for the benefit of both veterinary and human medicine.


Asunto(s)
Melanoma/diagnóstico por imagen , Melanoma/genética , Neoplasias de la Boca/diagnóstico por imagen , Neoplasias de la Boca/genética , Neoplasias Cutáneas/diagnóstico por imagen , Neoplasias Cutáneas/genética , Animales , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Línea Celular Tumoral , Perros , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Melanoma/tratamiento farmacológico , Ratones , Ratones Desnudos , Neoplasias de la Boca/tratamiento farmacológico , Neoplasias Cutáneas/tratamiento farmacológico , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto/métodos , Melanoma Cutáneo Maligno
11.
Nat Med ; 13(1): 35-7, 2007 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-17159985

RESUMEN

Abdominal pain is common in the general population and, in patients with irritable bowel syndrome, is attributed to visceral hypersensitivity. We found that oral administration of specific Lactobacillus strains induced the expression of mu-opioid and cannabinoid receptors in intestinal epithelial cells, and mediated analgesic functions in the gut-similar to the effects of morphine. These results suggest that the microbiology of the intestinal tract influences our visceral perception, and suggest new approaches for the treatment of abdominal pain and irritable bowel syndrome.


Asunto(s)
Dolor Abdominal/fisiopatología , Intestinos/fisiopatología , Lactobacillus acidophilus/fisiología , Receptores de Cannabinoides/fisiología , Receptores Opioides/fisiología , Dolor Abdominal/prevención & control , Administración Oral , Analgésicos Opioides/administración & dosificación , Analgésicos Opioides/farmacología , Animales , Antagonistas de Receptores de Cannabinoides , Colon/efectos de los fármacos , Colon/microbiología , Colon/fisiopatología , Relación Dosis-Respuesta a Droga , Células HT29 , Humanos , Indoles/administración & dosificación , Indoles/farmacología , Intestinos/efectos de los fármacos , Intestinos/microbiología , Masculino , Ratones , Ratones Endogámicos BALB C , Morfina/administración & dosificación , Morfina/farmacología , Naloxona/administración & dosificación , Naloxona/farmacología , Antagonistas de Narcóticos/administración & dosificación , Antagonistas de Narcóticos/farmacología , Probióticos/administración & dosificación , Probióticos/farmacología , Ratas , Ratas Sprague-Dawley , Receptor Cannabinoide CB2/antagonistas & inhibidores , Receptor Cannabinoide CB2/biosíntesis , Receptor Cannabinoide CB2/fisiología , Receptores de Cannabinoides/biosíntesis , Receptores Opioides/biosíntesis , Receptores Opioides mu/antagonistas & inhibidores , Receptores Opioides mu/biosíntesis , Receptores Opioides mu/fisiología , Recto/efectos de los fármacos , Recto/microbiología , Recto/fisiopatología
12.
iScience ; 27(4): 109417, 2024 Apr 19.
Artículo en Inglés | MEDLINE | ID: mdl-38510131

RESUMEN

Multiple myeloma (MM) is an incurable hematological malignancy in which MYC alterations contribute to the malignant phenotype. Nevertheless, MYC lacks therapeutic druggability. Here, we leveraged large-scale loss-of-function screens and conducted a small molecule screen to identify genes and pathways with enhanced essentiality correlated with MYC expression. We reported a specific gene dependency in glutaminase (GLS1), essential for the viability and proliferation of MYC overexpressing cells. Conversely, the analysis of isogenic models, as well as cell lines dataset (CCLE) and patient datasets, revealed GLS1 as a non-oncogenic dependency in MYC-driven cells. We functionally delineated the differential modulation of glutamine to maintain mitochondrial function and cellular biosynthesis in MYC overexpressing cells. Furthermore, we observed that pharmaceutical inhibition of NAMPT selectively affects MYC upregulated cells. We demonstrate the effectiveness of combining GLS1 and NAMPT inhibitors, suggesting that targeting glutaminolysis and NAD synthesis may be a promising strategy to target MYC-driven MM.

13.
Cancers (Basel) ; 15(7)2023 Apr 06.
Artículo en Inglés | MEDLINE | ID: mdl-37046846

RESUMEN

Tetraspanin CD81 plays major roles in cell-cell interactions and the regulation of cellular trafficking. This cholesterol-embarking transmembrane protein is a co-receptor for several viruses, including HCV, HIV-1 and Chikungunya virus, which exploits the large extracellular loop EC2 for cell entry. CD81 is also an anticancer target implicated in cancer cell proliferation and mobility, and in tumor metastasis. CD81 signaling contributes to the development of solid tumors (notably colorectal, liver and gastric cancers) and has been implicated in the aggressivity of B-cell lymphomas. A variety of protein partners can interact with CD81, either to regulate attachment and uptake of viruses (HCV E2, claudin-1, IFIM1) or to contribute to tumor growth and dissemination (CD19, CD44, EWI-2). CD81-protein interactions can be modulated with molecules targeting the extracellular domain of CD81, investigated as antiviral and/or anticancer agents. Several monoclonal antibodies anti-CD81 have been developed, notably mAb 5A6 active against invasion and metastasis of triple-negative breast cancer cells. CD81-EC2 can also be targeted with natural products (trachelogenin and harzianoic acids A-B) and synthetic compounds (such as benzothiazole-quinoline derivatives). They are weak CD81 binders but offer templates for the design of new compounds targeting the open EC2 loop. There is no anti-CD81 compound in clinical development at present, but this structurally well-characterized tetraspanin warrants more substantial considerations as a drug target.

14.
Biochem Pharmacol ; 209: 115445, 2023 03.
Artículo en Inglés | MEDLINE | ID: mdl-36739094

RESUMEN

Immune checkpoints inhibition is a privileged approach to combat cancers and other human diseases. The TIM-3 (T cell immunoglobulin and mucin-domain containing-3) inhibitory checkpoint expressed on different types of immune cells is actively investigated as an anticancer target, with a dozen of monoclonal antibodies in (pre)clinical development. A soluble form sTIM-3 can be found in the plasma of patients with cancer and other diseases. This active circulating protein originates from the proteolytic cleavage by two ADAM metalloproteases of the membrane receptor shared by tumor and non-tumor cells, and extracellular vesicles. In most cancers but not all, overexpression of mTIM-3 at the cell surface leads to high level of sTIM-3. Similarly, elevated levels of sTIM-3 have been reported in chronic autoimmune diseases, inflammatory gastro-intestinal diseases, certain viral and parasitic diseases, but also in cases of organ transplantation and in pregnancy-related pathologies. We have analyzed the origin of sTIM-3, its methods of dosage in blood or plasma, its presence in multiple diseases and its potential role as a biomarker to follow disease progression and/or the treatment response. In contrast to sPD-L1 generated by different classes of proteases and by alternative splicing, sTIM-3 is uniquely produced upon ADAM-dependent shedding, providing a more homogenous molecular entity and a possibly more reliable molecular marker. However, the biological functionality of sTIM-3 remains insufficiently characterized. The review shed light on pathologies associated with an altered expression of sTIM-3 in human plasma and the possibility to use sTIM-3 as a diagnostic or therapeutic marker.


Asunto(s)
Receptor 2 Celular del Virus de la Hepatitis A , Neoplasias , Humanos , Biomarcadores/metabolismo , Membrana Celular/metabolismo , Receptor 2 Celular del Virus de la Hepatitis A/metabolismo , Neoplasias/metabolismo
15.
Biomed Hub ; 8(1): 1-9, 2023.
Artículo en Inglés | MEDLINE | ID: mdl-36938364

RESUMEN

Introduction: The quest for small molecule inhibitors of the PD-1/PD-L1 checkpoint continues in parallel to the extensive development of monoclonal antibodies directed against this immune checkpoint. Drug screening strategies are being set up to identify novel PD-L1 inhibitors. Methods: A virtual screening based on molecular docking with the PD-L1 protein dimer has been performed to identify a new binder. Binding of the identified ligand to PD-L1 has been validated experimentally using a microscale thermophoresis (MST) assay. The cellular effect of the compound was evidenced using a fluorescence resonance energy transfer (FRET) assay based on activation of tyrosine phosphatase SHP-2. Results: We have identified the potent Wnt/ß-catenin inhibitor KYA1797K as a weak PD-L1 binder. Molecular docking suggested that the compound can bind to the interface of a PD-L1 dimer, with a geometry superimposable to that of the reference PD-L1 inhibitor BMS-202. The atypical 2-thioxo-4-thiazolidinone motif of KYA1797K, derived from the natural product rhodanine, plays a major role in the interaction with PD-L1. Binding of KYA1797K to recombinant hPD-L1 was validated experimentally, using MST. The drug was found to bind modestly but effectively to hPD-L1. The FRET assay confirmed the weak capacity of KYA1797K to interfere with the activation of SHP-2 upon its interaction with human PD-1. Discussion: Collectively, the data show that KYA1797K could function as a weak modulator of the PD-1/PD-L1 checkpoint. This effect may contribute, at least partially, to the reported capacity of the ß-catenin inhibitor to downregulate PD-L1 in cancer cells. The work also underlines the interest to further consider the rhodanine moiety as a chemical motif for the design of new PD-L1 binders.

16.
Cancers (Basel) ; 14(14)2022 Jul 11.
Artículo en Inglés | MEDLINE | ID: mdl-35884428

RESUMEN

Monoclonal antibodies targeting the PD-1/PD-L1 immune checkpoint have considerably improved the treatment of some cancers, but novel drugs, new combinations, and treatment modalities are needed to reinvigorate immunosurveillance in immune-refractory tumors. An option to elicit antitumor immunity against cancer consists of using approved and marketed drugs known for their capacity to modulate the expression and functioning of the PD-1/PD-L1 checkpoint. Here, we have reviewed several types of drugs known to alter the checkpoint, either directly via the blockade of PD-L1 or indirectly via an action on upstream effectors (such as STAT3) to suppress PD-L1 transcription or to induce its proteasomal degradation. Specifically, the repositioning of the approved drugs liothyronine, azelnidipine (and related dihydropyridine calcium channel blockers), niclosamide, albendazole/flubendazole, and a few other modulators of the PD-1/PD-L1 checkpoint (repaglinide, pimozide, fenofibrate, lonazolac, propranolol) is presented. Their capacity to bind to PD-L1 or to repress its expression and function offer novel perspectives for combination with PD-1 targeted biotherapeutics. These known and affordable drugs could be useful to improve the therapy of cancer.

17.
Eur J Med Chem ; 236: 114343, 2022 Jun 05.
Artículo en Inglés | MEDLINE | ID: mdl-35429911

RESUMEN

Immuno-therapy has become a leading strategy to fight cancer. Over the past few years, immuno-therapies using checkpoint inhibitor monoclonal antibodies (mAbs) against programmed death receptor 1 (PD-1) and programmed death ligand 1 (PD-L1) have demonstrated improved survival compared with chemotherapy. We describe the microwave-assisted synthesis and the characterization of an innovative series of synthetic compounds endowed with nanomolar activity against PD-L1. The properties of the compounds were characterized using several biophysical techniques including microscale thermophoresis (MST) and fluorescence resonance energy transfer (FRET) measurements. A few small molecules demonstrated a high affinity for human PD-L1, potently disrupted the PD-L1:PD-1 interaction and inhibited Src homology region 2 domain-containing phosphatase (SHP2) recruitment to PD-1. More than 30 molecules from the pyrazolone family have been synthesized and 5 highly potent "PD-L1 silencing compounds" have been identified, based on in vitro measurements. Structure-activity relationships have been defined and ADME properties were evaluated. The phenyl-pyrazolone unit offers novel perspectives to design PD-L1-targeting agents, potentially useful to combat cancer and other pathologies implicating the PD-1/PD-L1 checkpoint.


Asunto(s)
Inhibidores de Puntos de Control Inmunológico , Neoplasias , Pirazolonas , Antígeno B7-H1 , Humanos , Ligandos , Receptor de Muerte Celular Programada 1 , Pirazolonas/farmacología
18.
Cells ; 11(6)2022 03 17.
Artículo en Inglés | MEDLINE | ID: mdl-35326469

RESUMEN

The Leucine Rich Repeat Kinase 2 (LRRK2) gene is a major genetic determinant of Parkinson's disease (PD), encoding a homonymous multi-domain protein with two catalytic activities, GTPase and Kinase, involved in intracellular signaling and trafficking. LRRK2 is phosphorylated at multiple sites, including a cluster of autophosphorylation sites in the GTPase domain and a cluster of heterologous phosphorylation sites at residues 860 to 976. Phosphorylation at these latter sites is found to be modified in brains of PD patients, as well as for some disease mutant forms of LRRK2. The main aim of this study is to investigate the functional consequences of LRRK2 phosphorylation or dephosphorylation at LRRK2's heterologous phosphorylation sites. To this end, we generated LRRK2 phosphorylation site mutants and studied how these affected LRRK2 catalytic activity, neurite outgrowth and lysosomal physiology in cellular models. We show that phosphorylation of RAB8a and RAB10 substrates are reduced with phosphomimicking forms of LRRK2, while RAB29 induced activation of LRRK2 kinase activity is enhanced for phosphodead forms of LRRK2. Considering the hypothesis that PD pathology is associated to increased LRRK2 kinase activity, our results suggest that for its heterologous phosphorylation sites LRRK2 phosphorylation correlates to healthy phenotypes and LRRK2 dephosphorylation correlates to phenotypes associated to the PD pathological processes.


Asunto(s)
Enfermedad de Parkinson , Humanos , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina/genética , Proteína 2 Quinasa Serina-Treonina Rica en Repeticiones de Leucina/metabolismo , Lisosomas/metabolismo , Enfermedad de Parkinson/metabolismo , Fosforilación/fisiología , Transducción de Señal
19.
Sci Immunol ; 7(78): eade5686, 2022 12 09.
Artículo en Inglés | MEDLINE | ID: mdl-36459543

RESUMEN

Cytokines interact with their receptors in the extracellular space to control immune responses. How the physicochemical properties of the extracellular space influence cytokine signaling is incompletely elucidated. Here, we show that the activity of interleukin-2 (IL-2), a cytokine critical to T cell immunity, is profoundly affected by pH, limiting IL-2 signaling within the acidic environment of tumors. Generation of lactic acid by tumors limits STAT5 activation, effector differentiation, and antitumor immunity by CD8+ T cells and renders high-dose IL-2 therapy poorly effective. Directed evolution enabled selection of a pH-selective IL-2 mutein (Switch-2). Switch-2 binds the IL-2 receptor subunit IL-2Rα with higher affinity, triggers STAT5 activation, and drives CD8+ T cell effector function more potently at acidic pH than at neutral pH. Consequently, high-dose Switch-2 therapy induces potent immune activation and tumor rejection with reduced on-target toxicity in normal tissues. Last, we show that sensitivity to pH is a generalizable property of a diverse range of cytokines with broad relevance to immunity and immunotherapy in healthy and diseased tissues.


Asunto(s)
Interleucina-2 , Neoplasias , Humanos , Factor de Transcripción STAT5 , Linfocitos T CD8-positivos , Citocinas , Concentración de Iones de Hidrógeno
20.
J Exp Clin Cancer Res ; 41(1): 110, 2022 Mar 28.
Artículo en Inglés | MEDLINE | ID: mdl-35346305

RESUMEN

BACKGROUND: CD44 is a multifunctional membrane glycoprotein. Through its heparan sulfate chain, CD44 presents growth factors to their receptors. We have shown that CD44 and Tropomyosin kinase A (TrkA) form a complex following nerve growth factor (NGF) induction. Our study aimed to understand how CD44 and TrkA interact and the consequences of inhibiting this interaction regarding the pro-tumoral effect of NGF in breast cancer. METHODS: After determining which CD44 isoforms (variants) are involved in forming the TrkA/CD44 complex using proximity ligation assays, we investigated the molecular determinants of this interaction. By molecular modeling, we isolated the amino acids involved and confirmed their involvement using mutations. A CD44v3 mimetic peptide was then synthesized to block the TrkA/CD44v3 interaction. The effects of this peptide on the growth, migration and invasion of xenografted triple-negative breast cancer cells were assessed. Finally, we investigated the correlations between the expression of the TrkA/CD44v3 complex in tumors and histo-pronostic parameters. RESULTS: We demonstrated that isoform v3 (CD44v3), but not v6, binds to TrkA in response to NGF stimulation. The final 10 amino acids of exon v3 and the TrkA H112 residue are necessary for the association of CD44v3 with TrkA. Functionally, the CD44v3 mimetic peptide impairs not only NGF-induced RhoA activation, clonogenicity, and migration/invasion of breast cancer cells in vitro but also tumor growth and metastasis in a xenograft mouse model. We also detected TrkA/CD44v3 only in cancerous cells, not in normal adjacent tissues. CONCLUSION: Collectively, our results suggest that blocking the CD44v3/TrkA interaction can be a new therapeutic option for triple-negative breast cancers.


Asunto(s)
Neoplasias de la Mama , Receptores de Hialuranos , Factor de Crecimiento Nervioso , Receptor trkA , Animales , Neoplasias de la Mama/genética , Femenino , Humanos , Receptores de Hialuranos/metabolismo , Ratones , Factor de Crecimiento Nervioso/farmacología , Isoformas de Proteínas , Receptor trkA/metabolismo
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