ABSTRACT
SUMMARY: High-throughput screens (HTS) have been utilized to assess the efficacy of single drugs against patient tumor samples with the purpose of optimizing precision therapy, but testing the synergy of drug combinations can identify the ideal second drug to add. With novel sophisticated HTS, effective venetoclax combinations can be revealed that provide the cell state, phenotype, and molecular features of the susceptible and resistant cell populations. See related article by Eide, Kurtz et al., p. 452 (14) .
Subject(s)
Leukemia, Myeloid, Acute , Humans , Drug Evaluation, Preclinical , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/genetics , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Bridged Bicyclo Compounds, Heterocyclic/therapeutic use , Sulfonamides/pharmacology , Sulfonamides/therapeutic useABSTRACT
The continuous progression in the field of electrotherapies implies the development of multifunctional materials exhibiting excellent electrochemical performance and biocompatibility, promoting cell adhesion, and possessing antibacterial properties. Since the conditions favouring the adhesion of mammalian cells are similar to conditions favouring the adhesion of bacterial cells, it is necessary to engineer the surface to exhibit selective toxicity, i.e., to kill or inhibit the growth of bacteria without damaging mammalian tissues. The aim of this paper is to introduce a surface modification approach based on a subsequent deposition of silver and gold particles on the surface of a conducting polymer, poly(3,4-ethylenedioxythiophene) (PEDOT). The resulting PEDOT-Au/Ag surface is found to possess optimal wettability, roughness, and surface features making it an excellent platform for cell adhesion. By depositing Ag particles on PEDOT surface decorated with Au particles, it is possible to reduce toxic effects of Ag particles, while maintaining their antibacterial activity. Besides, electroactive and capacitive properties of PEDOT-Au/Ag account for its applicability in various electroceutical therapies.
Subject(s)
Gold , Silver , Animals , Silver/pharmacology , Silver/chemistry , Gold/chemistry , Polymers/chemistry , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Bridged Bicyclo Compounds, Heterocyclic/chemistry , Anti-Bacterial Agents/pharmacology , MammalsABSTRACT
AMP-activated protein kinase (AMPK) regulates the balance between cellular anabolism and catabolism dependent on energy resources to maintain proliferation and survival. Small-compound AMPK activators show anti-cancer activity in preclinical models. Using the direct AMPK activator GSK621, we show that the unfolded protein response (UPR) is activated by AMPK in acute myeloid leukemia (AML) cells. Mechanistically, the UPR effector protein kinase RNA-like ER kinase (PERK) represses oxidative phosphorylation, tricarboxylic acid (TCA) cycle, and pyrimidine biosynthesis and primes the mitochondrial membrane to apoptotic signals in an AMPK-dependent manner. Accordingly, in vitro and in vivo studies reveal synergy between the direct AMPK activator GSK621 and the Bcl-2 inhibitor venetoclax. Thus, selective AMPK-activating compounds kill AML cells by rewiring mitochondrial metabolism that primes mitochondria to apoptosis by BH3 mimetics, holding therapeutic promise in AML.
Subject(s)
AMP-Activated Protein Kinases/metabolism , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Imidazoles/pharmacology , Leukemia, Myeloid, Acute/drug therapy , Pyrimidinones/pharmacology , Sulfonamides/pharmacology , Unfolded Protein Response/physiology , eIF-2 Kinase/metabolism , Adolescent , Adult , Aged , Aged, 80 and over , Animals , Antineoplastic Agents/pharmacology , Apoptosis/physiology , Cell Line, Tumor , Citric Acid Cycle/drug effects , Drug Evaluation, Preclinical , Female , HEK293 Cells , HL-60 Cells , Humans , Leukemia, Myeloid, Acute/genetics , Leukemia, Myeloid, Acute/metabolism , Male , Mice , Middle Aged , Mitochondria/metabolism , Oxidative Phosphorylation/drug effects , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , THP-1 Cells , U937 Cells , Young AdultABSTRACT
Everolimus, an oral mammalian target of rapamycin complex 1 (mTORC1) inhibitor, presents a therapeutic option in metastatic renal cell carcinoma (RCC) patients who were intolerant to, or previously failed, immune- and vascular endothelial growth factor-targeted therapies. However, the onset of drug resistance limits its clinical use. One possible mechanism underpinning the resistance is that inhibiting mTORC1 by everolimus results in mTORC2-dependent activation of v-Akt murine thymoma viral oncogene (AKT) and upregulation of hypoxia-inducible transcription factors (HIF). Norcantharidin (NCTD) is a demethylated derivative of cantharidin with antitumor properties which is an active ingredient of the traditional Chinese medicine Mylabris. In this study, everolimus-resistant RCC cells (786-O-R) obtained by chronic everolimus treatment revealed higher level of HIF2α and over-activated mTORC2 pathway and NCTD inhibits cell proliferation in both everolimus-resistant and -sensitive RCC cells by arresting cell cycle in G0/G1 phase and reducing cell cycle-related proteins of C-Myc and cyclin D. Furthermore, NCTD shows synergistic anticancer effects combined with everolimus in everolimus-resistant 786-O-R cells. Mechanically, NCTD repressed both mTORC1 and mTORC2 signaling pathways as well as downstream molecular signaling pathways, such as p-4EBP1, p-AKT, HIF1α and HIF2α. Our findings provide sound evidence that combination of NCTD and everolimus is a potential therapeutic strategy for treating RCC and overcoming everolimus resistance by dual inhibition of mTORC1 and mTORC2.
Subject(s)
Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Carcinoma, Renal Cell/pathology , Drug Resistance, Neoplasm/drug effects , Everolimus/pharmacology , Kidney Neoplasms/pathology , Basic Helix-Loop-Helix Transcription Factors/drug effects , Cell Line, Tumor , Cell Proliferation/drug effects , Humans , Mechanistic Target of Rapamycin Complex 1/drug effects , Mechanistic Target of Rapamycin Complex 2/drug effects , Proto-Oncogene Proteins c-akt/drug effects , Signal Transduction/drug effectsABSTRACT
The coronavirus disease 2019 (COVID-19) global pandemic has turned into the largest public health and economic crisis in recent history impacting virtually all sectors of society. There is a need for effective therapeutics to battle the ongoing pandemic. Repurposing existing drugs with known pharmacological safety profiles is a fast and cost-effective approach to identify novel treatments. The COVID-19 etiologic agent is the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a single-stranded positive-sense RNA virus. Coronaviruses rely on the enzymatic activity of the replication-transcription complex (RTC) to multiply inside host cells. The RTC core catalytic component is the RNA-dependent RNA polymerase (RdRp) holoenzyme. The RdRp is one of the key druggable targets for CoVs due to its essential role in viral replication, high degree of sequence and structural conservation and the lack of homologues in human cells. Here, we have expressed, purified and biochemically characterised active SARS-CoV-2 RdRp complexes. We developed a novel fluorescence resonance energy transfer-based strand displacement assay for monitoring SARS-CoV-2 RdRp activity suitable for a high-throughput format. As part of a larger research project to identify inhibitors for all the enzymatic activities encoded by SARS-CoV-2, we used this assay to screen a custom chemical library of over 5000 approved and investigational compounds for novel SARS-CoV-2 RdRp inhibitors. We identified three novel compounds (GSK-650394, C646 and BH3I-1) and confirmed suramin and suramin-like compounds as in vitro SARS-CoV-2 RdRp activity inhibitors. We also characterised the antiviral efficacy of these drugs in cell-based assays that we developed to monitor SARS-CoV-2 growth.
Subject(s)
Antiviral Agents/chemistry , Antiviral Agents/pharmacology , Coronavirus RNA-Dependent RNA Polymerase/antagonists & inhibitors , Drug Evaluation, Preclinical , SARS-CoV-2/enzymology , Small Molecule Libraries/pharmacology , Animals , Benzoates/pharmacology , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Chlorocebus aethiops , Coronavirus RNA-Dependent RNA Polymerase/metabolism , Enzyme Assays , Fluorescence Resonance Energy Transfer , High-Throughput Screening Assays , Holoenzymes/metabolism , Reproducibility of Results , SARS-CoV-2/drug effects , Small Molecule Libraries/chemistry , Suramin/pharmacology , Vero Cells , Viral Nonstructural Proteins/antagonists & inhibitors , Viral Nonstructural Proteins/metabolismABSTRACT
The number and activity of Cav1.2 channels in the cardiomyocyte sarcolemma tunes the magnitude of Ca2+-induced Ca2+ release and myocardial contraction. ß-Adrenergic receptor (ßAR) activation stimulates sarcolemmal insertion of CaV1.2. This supplements the preexisting sarcolemmal CaV1.2 population, forming large "superclusters" wherein neighboring channels undergo enhanced cooperative-gating behavior, amplifying Ca2+ influx and myocardial contractility. Here, we determine this stimulated insertion is fueled by an internal reserve of early and recycling endosome-localized, presynthesized CaV1.2 channels. ßAR-activation decreased CaV1.2/endosome colocalization in ventricular myocytes, as it triggered "emptying" of endosomal CaV1.2 cargo into the t-tubule sarcolemma. We examined the rapid dynamics of this stimulated insertion process with live-myocyte imaging of channel trafficking, and discovered that CaV1.2 are often inserted into the sarcolemma as preformed, multichannel clusters. Similarly, entire clusters were removed from the sarcolemma during endocytosis, while in other cases, a more incremental process suggested removal of individual channels. The amplitude of the stimulated insertion response was doubled by coexpression of constitutively active Rab4a, halved by coexpression of dominant-negative Rab11a, and abolished by coexpression of dominant-negative mutant Rab4a. In ventricular myocytes, ßAR-stimulated recycling of CaV1.2 was diminished by both nocodazole and latrunculin-A, suggesting an essential role of the cytoskeleton in this process. Functionally, cytoskeletal disruptors prevented ßAR-activated Ca2+ current augmentation. Moreover, ßAR-regulation of CaV1.2 was abolished when recycling was halted by coapplication of nocodazole and latrunculin-A. These findings reveal that ßAR-stimulation triggers an on-demand boost in sarcolemmal CaV1.2 abundance via targeted Rab4a- and Rab11a-dependent insertion of channels that is essential for ßAR-regulation of cardiac CaV1.2.
Subject(s)
Calcium Channels, L-Type/metabolism , Myocytes, Cardiac/metabolism , Receptors, Adrenergic, beta/metabolism , Sarcolemma/metabolism , rab4 GTP-Binding Proteins/metabolism , Animals , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Cell Line , Cells, Cultured , Endosomes/metabolism , Female , Heart Ventricles/cytology , Humans , Mice , Mice, Inbred C57BL , Myocytes, Cardiac/drug effects , Myocytes, Cardiac/physiology , Nocodazole/pharmacology , Protein Transport , Thiazolidines/pharmacologyABSTRACT
BACKGROUND: Impetigo, a highly contagious bacterial skin infection commonly occurring in young children, but adults may also be affected. The superficial skin infection is mainly caused by Staphylococcus aureus (S. aureus) and less frequently by Streptococcus pyogenes (S. pyogenes). Antimicrobial resistance has become a worldwide concern and needs to be addressed when selecting treatment for impetigo patients. An evidence-based impetigo treatment algorithm was developed to address the treatment of impetigo for pediatric and adult populations. METHODS: An international panel of pediatric dermatologists, dermatologists, pediatricians, and pediatric infectious disease specialists employed a modified Delphi technique to develop the impetigo treatment algorithm. Treatment recommendations were evidence-based, taking into account antimicrobial stewardship and the increasing resistance to oral and topical antibiotics. RESULTS: The algorithm includes education and prevention of impetigo, diagnosis and classification, treatment measures, and follow-up and distinguishes between localized and widespread or epidemic outbreaks of impetigo. The panel adopted the definition of localized impetigo of fewer than ten lesions and smaller than 36 cm2 area affected in patients of two months and up with no compromised immune status. Resistance to oral and topical antibiotics prescribed for the treatment of impetigo such as mupirocin, retapamulin, fusidic acid, have been widely reported. CONCLUSIONS: When prescribing antibiotics, it is essential to know the local trends in antibiotic resistance. Ozenoxacin cream 1% is highly effective against S. pyogenes and S. aureus, including methycyllin-susceptible and resistant strains (MRSA), and may be a suitable option for localized impetigo.J Drugs Dermatol. 2021;20(2):134-142. doi:10.36849/JDD.5475 THIS ARTICLE HAD BEEN MADE AVAILABLE FREE OF CHARGE. PLEASE SCROLL DOWN TO ACCESS THE FULL TEXT OF THIS ARTICLE WITHOUT LOGGING IN. NO PURCHASE NECESSARY. PLEASE CONTACT THE PUBLISHER WITH ANY QUESTIONS.
Subject(s)
Anti-Bacterial Agents/therapeutic use , Critical Pathways/standards , Impetigo/drug therapy , Staphylococcus aureus/drug effects , Streptococcus pyogenes/drug effects , Aminopyridines/pharmacology , Aminopyridines/therapeutic use , Anti-Bacterial Agents/pharmacology , Antimicrobial Stewardship/standards , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Bridged Bicyclo Compounds, Heterocyclic/therapeutic use , Delphi Technique , Diterpenes/pharmacology , Diterpenes/therapeutic use , Drug Resistance, Bacterial , Evidence-Based Medicine/standards , Fusidic Acid/pharmacology , Fusidic Acid/therapeutic use , Humans , Impetigo/diagnosis , Impetigo/microbiology , Microbial Sensitivity Tests/standards , Mupirocin/pharmacology , Mupirocin/therapeutic use , Practice Guidelines as Topic , Quinolones/pharmacology , Quinolones/therapeutic use , Skin Cream/pharmacology , Skin Cream/therapeutic use , Staphylococcus aureus/isolation & purification , Streptococcus pyogenes/isolation & purification , Systematic Reviews as TopicABSTRACT
Vascular calcification may result from stimulation of osteogenic signalling with upregulation of the transcription factors CBFA1, MSX2 and SOX9, as well as alkaline phosphatase (ALPL), which degrades and thus inactivates the calcification inhibitor pyrophosphate. Osteogenic signalling further involves upregulation of the Ca2+-channel ORAI1. The channel is activated by STIM1 and then accomplishes store-operated Ca2+ entry. ORAI1 and STIM1 are upregulated by the serum & glucocorticoid inducible kinase 1 (SGK1) which is critically important for osteogenic signalling. Stimulators of vascular calcification include vasopressin. The present study explored whether exposure of human aortic smooth muscle cells (HAoSMCs) to vasopressin upregulates ORAI1 and/or STIM1 expression, store-operated Ca2+ entry and osteogenic signalling. To this end, HAoSMCs were exposed to vasopressin (100 nM, 24 h) without or with additional exposure to ORAI1 blocker MRS1845 (10 µM) or SGK1 inhibitor GSK-650394 (1 µM). Transcript levels were measured using q-RT-PCR, cytosolic Ca2+-concentration ([Ca2+]i) by Fura-2-fluorescence, and store-operated Ca2+ entry from increase of [Ca2+]i following re-addition of extracellular Ca2+ after store depletion with thapsigargin (1 µM). As a result, vasopressin enhanced the transcript levels of ORAI1 and STIM1, store-operated Ca2+ entry, as well as the transcript levels of CBFA1, MSX2, SOX9 and ALPL. The effect of vasopressin on store-operated Ca2+ entry as well as on transcript levels of CBFA1, MSX2, SOX9 and ALPL was virtually abrogated by MRS1845 and GSK-650394. In conclusion, vasopressin stimulates expression of ORAI1/STIM1, thus augmenting store-operated Ca2+ entry and osteogenic signalling. In HAoSMCs, vasopressin (VP) upregulates Ca2+ channel ORAI1 and its activator STIM1. VP upregulates store-operated Ca2+ entry (SOCE) and osteogenic signalling (OS). VP-induced SOCE, OS and Ca2+-deposition are disrupted by ORAI1 inhibitor MRS1845. VP-induced SOCE, OS and Ca2+-deposition are disrupted by SGK1 blocker GSK-650394. KEY MESSAGES: ⢠In HAoSMCs, vasopressin (VP) upregulates Ca2+ channel ORAI1 and its activator STIM1. ⢠VP upregulates store-operated Ca2+ entry (SOCE) and osteogenic signalling (OS). ⢠VP-induced SOCE, OS and Ca2+-deposition are disrupted by ORAI1 inhibitor MRS1845. ⢠VP-induced SOCE, OS and Ca2+-deposition are disrupted by SGK1 blocker GSK-650394.
Subject(s)
Calcium Signaling/drug effects , Muscle, Smooth, Vascular/cytology , Myocytes, Smooth Muscle/drug effects , ORAI1 Protein/biosynthesis , Vascular Calcification/metabolism , Vasopressins/pharmacology , Aorta/cytology , Benzoates/pharmacology , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Calcium Channel Blockers/pharmacology , Calcium Signaling/physiology , Cells, Cultured , Drug Evaluation, Preclinical , Humans , Immediate-Early Proteins/antagonists & inhibitors , Immediate-Early Proteins/physiology , Myocytes, Smooth Muscle/metabolism , Neoplasm Proteins/biosynthesis , Neoplasm Proteins/genetics , Neoplasm Proteins/physiology , Nitrendipine/analogs & derivatives , Nitrendipine/pharmacology , ORAI1 Protein/antagonists & inhibitors , ORAI1 Protein/genetics , Osteogenesis/drug effects , Protein Serine-Threonine Kinases/antagonists & inhibitors , Protein Serine-Threonine Kinases/physiology , Stromal Interaction Molecule 1/biosynthesis , Stromal Interaction Molecule 1/genetics , Stromal Interaction Molecule 1/physiology , Transcription, Genetic/drug effects , Up-Regulation/drug effects , Vascular Calcification/prevention & controlABSTRACT
Renal cell carcinoma (RCC) is generally acknowledged as the most resistant primary malignancy unresponsive to conventional radiotherapy and chemotherapy treatments. Norcantharidin (NCTD), a therapeutic compound derived from medicinal plants, has been shown to trigger apoptosis, as well as antimetastatic and antioxidant activities in several tumor cells. However, NCTD's mechanism of antitumor activity in the RCC cell line remains unclear. In this study, we report that NCTD led to a time- and dose-dependent inhibition of cell proliferation. It had also markedly induced apoptosis and G2/M phase cell cycle arrest in a dose-dependent manner by decreasing the expressions of pro-caspase-3, pro-caspase-9, cyclin B1, and pCDC25C while increasing active caspase-3, cleaved-PARP, P21, and pCDC2 levels. Interestingly, NCTD treatment provoked the phosphorylation of extracellular-regulated protein kinase (ERK) and c-Jun-N-terminal kinase (JNK), but not of p38 MAPK. Moreover, SCH772984 and SP600125, ERK and JNK inhibitors, respectively, could partially abolish NCTD-induced apoptosis and G2/M phase cell cycle arrest. Collectively, these findings suggest that NCTD might activate JNK and ERK signaling pathways, consequently inducing apoptosis and G2/M arrest through the modulation of related proteins. This study provided evidence that NCTD is a promising therapeutic drug for the treatment of RCC.
Subject(s)
Antineoplastic Agents/pharmacology , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Carcinoma, Renal Cell/drug therapy , Kidney Neoplasms/drug therapy , Antineoplastic Agents/administration & dosage , Apoptosis/drug effects , Bridged Bicyclo Compounds, Heterocyclic/administration & dosage , Carcinoma, Renal Cell/pathology , Cell Line, Tumor , Cell Proliferation/drug effects , Dose-Response Relationship, Drug , G2 Phase Cell Cycle Checkpoints/drug effects , Humans , Kidney Neoplasms/pathology , M Phase Cell Cycle Checkpoints/drug effects , MAP Kinase Signaling System/drug effects , Signal Transduction/drug effects , Time Factors , p38 Mitogen-Activated Protein Kinases/metabolismABSTRACT
Cantharidin (CTD) is the main bioactive component of Cantharides, which is called Banmao in Traditional Chinese Medicine (TCM). Norcantharidin (NCTD) is a structural modifier of CTD. To compare with CTD, NCTD has lighter side effects and stronger bioactivity in anti-cancer through inhibiting cell proliferation, causing apoptosis and autophagy, overwhelming migration and metastasis, affecting immunity as well as lymphangiogenesis. Examples of these effects include suppressing Protein Phosphatase 2A and modulating Wnt/beta catenin signal, with Caspase family proteins, AMPK pathway and c-Met/EGFR pathway involving respectively. Moreover, NCTD has the effects of immune enhancement, anti-platelet aggregation and inhibition of renal interstitial fibrosis with distinct signaling pathways. The immunological effects induced by NCTD are related to the regulation of macrophage polarization and LPS-mediated immune response. The antiplatelet activity that NCTD induced is relevant to the inhibition of platelet signaling and the downregulation of α2 integrin. Furthermore, some of novel derivatives designed and synthesized artificially show stronger biological activities (e.g., anticancer effect, enzyme inhibition effect, antioxidant effect) and lower toxicity than NCTD itself. Plenty of literatures have reported various pharmacological effects of NCTD, particularly the anticancer effect, which has been widely concerned in clinical application and laboratory research. In this review, the pharmaceutical activities and derivatives of NCTD are discussed, which can be reference for further study.
Subject(s)
Antineoplastic Agents/pharmacology , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Neoplasms/drug therapy , Animals , Apoptosis/drug effects , Autophagy/drug effects , Cantharidin/chemistry , Cell Proliferation/drug effects , Humans , Medicine, Chinese Traditional , Neoplasms/pathologyABSTRACT
Bcl-2 inhibitors display an effective activity in acute myeloid leukemia (AML), but its clinical efficacy as a monotherapy was limited in part owing to failure to target other antiapoptotic Bcl-2 family proteins, such as Mcl-1. In this context, the combination strategy may be a promising approach to overcome this barrier. Here, we report the preclinical efficacy of a novel strategy combining ABT-199 with triptolide (TPL), a natural product extracted from a traditional Chinese medicine, in AML. Combination treatment exhibited markedly increased cytotoxicity in leukemic cells irrespective of p53 status while largely sparing normal cells of the hematopoietic lineage. Moreover, co-administration of ABT-199 with TPL dramatically suppressed leukemia progression as well as prolonged animal survival in a xenograft AML model. The potentiated effect of ABT-199 and TPL against AML was associated with activation of the mitochondrum-related intrinsic apoptotic pathway through a mechanism reciprocally modulating Bcl-2 family proteins. In this case, TPL not only downregulated Mcl-1 but also upregulated proapoptotic BH3-only proteins, thereby overcoming the resistance toward ABT-199. Conversely, ABT-199 abrogated Bcl-2-mediated cytoprotection against TPL. Together, these findings suggest that the regimen combining TPL and ABT-199 might be active against AML by inducing robust apoptosis through reciprocal regulation of anti- and proapoptotic Bcl-2 family proteins, therefore providing a strong rationale for the clinical investigation of this combination regimen for the treatment of AML.
Subject(s)
Apoptosis , Bridged Bicyclo Compounds, Heterocyclic/therapeutic use , Diterpenes/therapeutic use , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/metabolism , Phenanthrenes/therapeutic use , Proto-Oncogene Proteins c-bcl-2/metabolism , Sulfonamides/therapeutic use , Adolescent , Adult , Aged , Animals , Apoptosis/drug effects , Blast Crisis/pathology , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Cell Line, Tumor , Child , Diterpenes/pharmacology , Drug Synergism , Epoxy Compounds/pharmacology , Epoxy Compounds/therapeutic use , Female , Humans , Male , Mice, Inbred BALB C , Mice, Nude , Middle Aged , Mitochondria/drug effects , Mitochondria/metabolism , Phenanthrenes/pharmacology , Sulfonamides/pharmacology , Tumor Suppressor Protein p53/metabolism , Xenograft Model Antitumor AssaysABSTRACT
Targeting neuroendocrine receptors can be considered as another interesting approach to treating fibrotic disorders. Previously, we could demonstrate that tropisetron, a classical serotonin receptor blocker, can modulate collagen synthesis and acts in vitro through the α7 nicotinic acetylcholine receptor (α7nAchR). Here, we used a pharmacologic approach with specific α7nAchR agonists to validate this hypothesis. PHA-543613, an α7nAchR-specific agonist, not only prevented but also reversed established skin fibrosis of mice injected with bleomycin. Interestingly, agonistic stimulation of α7nAchR also attenuated experimental skin fibrosis in the non-inflammation driven adenovirus coding for TGFß receptor Iact mouse model, indicating fibroblast-mediated and not only anti-inflammatory effects of such agents. The fibroblast-mediated effects were confirmed in vitro using human dermal fibroblasts, in which the α7nAchR-specific agonists strongly reduced the impact of TGFß1-mediated expression on collagen and myofibroblast marker expression. These actions were linked to modulation of the redox-sensitive transcription factor JunB and impairment of the mitochondrial respiratory system. Our results indicate that pharmacologic stimulation of the α7nAchR could be a promising target for treatment of patients with skin fibrotic diseases. Moreover, our results suggest a mechanistic axis of collagen synthesis regulation through the mitochondrial respiratory system.
Subject(s)
Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Quinuclidines/pharmacology , Scleroderma, Systemic/drug therapy , Skin/pathology , alpha7 Nicotinic Acetylcholine Receptor/agonists , Adenoviridae/genetics , Animals , Bleomycin/toxicity , Bridged Bicyclo Compounds, Heterocyclic/therapeutic use , Cells, Cultured , Collagen/metabolism , Disease Models, Animal , Drug Evaluation, Preclinical , Fibroblasts/drug effects , Fibroblasts/metabolism , Fibrosis , Genetic Vectors/administration & dosage , Genetic Vectors/genetics , Humans , Male , Mice , Primary Cell Culture , Quinuclidines/therapeutic use , Receptor, Transforming Growth Factor-beta Type I/genetics , Scleroderma, Systemic/chemically induced , Scleroderma, Systemic/genetics , Scleroderma, Systemic/pathology , Skin/cytology , Skin/drug effects , alpha7 Nicotinic Acetylcholine Receptor/metabolismABSTRACT
BCL-2 is an antiapoptotic protein that plays a critical role acute and chronic leukemias. Venetoclax is an orally selective BCL-2 inhibitor and BH3 mimetic approved in chronic lymphocytic leukemia and in combination with low dose cytarabine or hypomethylating agent in acute myeloid leukemia for the treatment of patients unfit for intensive chemotherapy. This article reviews the biology of BCL-2, focusing on its relationship to the myeloid microenvironment, and discusses the rationale for BCL-2 inhibition in myelodysplastic syndrome (MDS). Clinical trials testing venetoclax in MDS patients are under way. Potential biomarkers for clinical response to BCL-2 inhibition are discussed. Therapeutic opportunities for venetoclax in the therapeutic landscape of MDS are explored.
Subject(s)
Bridged Bicyclo Compounds, Heterocyclic/therapeutic use , Myelodysplastic Syndromes/drug therapy , Sulfonamides/therapeutic use , Animals , Antineoplastic Agents/pharmacology , Antineoplastic Agents/therapeutic use , Apoptosis/drug effects , Apoptosis/genetics , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Clinical Studies as Topic , Disease Management , Disease Models, Animal , Disease Susceptibility , Drug Evaluation, Preclinical , Humans , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/etiology , Mitochondria/drug effects , Mitochondria/genetics , Mitochondria/metabolism , Myelodysplastic Syndromes/diagnosis , Myelodysplastic Syndromes/etiology , Myelodysplastic Syndromes/metabolism , Proto-Oncogene Proteins c-bcl-2/genetics , Proto-Oncogene Proteins c-bcl-2/metabolism , Sulfonamides/pharmacology , Treatment OutcomeABSTRACT
Purpose/Aim of the Study: Wnt/ß-catenin signaling was reported to be activated in pulmonary fibrosis, and was focused on as a target for antifibrotic therapy. However, the mechanism how the inhibition of Wnt/ß-catenin signaling ameliorate pulmonary fibrosis has not been fully elucidated. The purpose of this study is to explore the target cells of Wnt/ß-catenin inhibition in pulmonary fibrosis and to examine the antifibrotic effect of the novel inhibitor PRI-724 specifically disrupting the interaction of ß-catenin and CBP. Materials and Methods: The effect of C-82, an active metabolite of PRI-724, on the expression of TGF-ß1 and α-smooth muscle actin (SMA) was examined on fibroblasts and macrophages. We also examined the effects of PRI-724 in mouse model of bleomycin-induced pulmonary fibrosis. Results: The activation and increased accumulation of ß-catenin in the canonical pathway were detected in lung fibroblasts as well as macrophages stimulated by Wnt3a using Western blotting. Treatment with C-82 reduced CBP protein and increased p300 protein binding to ß-catenin in the nucleus of lung fibroblasts. In addition, C-82 inhibited the expression of SMA in lung fibroblasts treated with TGF-ß, indicating the inhibition of myofibroblast differentiation. In the fibrotic lungs induced by bleomycin, ß-catenin was stained strongly in macrophages, but the staining of ß-catenin in alveolar epithelial cells and fibroblasts was weak. The administration of PRI-724 ameliorated pulmonary fibrosis induced by bleomycin in mice when administered with a late, but not an early, treatment schedule. Analysis of bronchoalveolar fluid (BALF) showed a decreased number of alveolar macrophages. In addition, the level of TGF-ß1 in BALF was decreased in mice treated with PRI-724. C-82 also inhibited the production of TGF-ß1 by alveolar macrophages. Conclusions: These results suggest that the ß-catenin/CBP inhibitor PRI-724 is a potent antifibrotic agent that acts by modulating the activity of macrophages in the lungs.
Subject(s)
Bridged Bicyclo Compounds, Heterocyclic/therapeutic use , Pulmonary Fibrosis/drug therapy , Pyrimidinones/therapeutic use , beta Catenin/antagonists & inhibitors , Animals , Bleomycin , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Drug Evaluation, Preclinical , Fibroblasts/metabolism , Macrophages/metabolism , Male , Mice, Inbred C57BL , Pulmonary Fibrosis/chemically induced , Pulmonary Fibrosis/metabolism , Pyrimidinones/pharmacology , Transforming Growth Factor beta1/metabolism , Wnt Signaling Pathway/drug effects , beta Catenin/metabolismABSTRACT
The amygdala plays a critical role in emotional-affective aspects of behaviors and pain modulation. The central nucleus of amygdala (CeA) serves major output functions, and neuroplasticity in the CeA is linked to pain-related behaviors in different models. Activation of Gi/o-coupled group II metabotropic glutamate receptors (mGluRs), which consist of mGluR2 and mGluR3, can decrease neurotransmitter release and regulate synaptic plasticity. Group II mGluRs have emerged as targets for neuropsychiatric disorders and can inhibit pain-related processing and behaviors. Surprisingly, site and mechanism of antinociceptive actions of systemically applied group II mGluR agonists are not clear. Our previous work showed that group II mGluR activation in the amygdala inhibits pain-related CeA activity, but behavioral and spinal consequences remain to be determined. Here we studied the contribution of group II mGluRs in the amygdala to the antinociceptive effects of a systemically applied group II mGluR agonist (LY379268) on behavior and spinal dorsal horn neuronal activity, using the kaolin/carrageenan-induced knee joint arthritis pain model. Audible and ultrasonic vocalizations (emotional responses) and mechanical reflex thresholds were measured in adult rats with and without arthritis (5-6â¯h postinduction). Extracellular single-unit recordings were made from spinal dorsal horn wide dynamic range neurons of anesthetized (isoflurane) rats with and without arthritis (5-6â¯h postinduction). Systemic (intraperitoneal) application of a group II mGluR agonist (LY379268) decreased behaviors and activity of spinal neurons in the arthritis pain model but not under normal conditions. Stereotaxic administration of LY379268 into the CeA mimicked the effects of systemic application. Conversely, stereotaxic administration of a group II mGluR antagonist (LY341495) into the CeA reversed the effects of systemic application of LY379268 on behaviors and dorsal horn neuronal activity in arthritic rats. The data show for the first time that the amygdala is the critical site of action for the antinociceptive behavioral and spinal neuronal effects of systemically applied group II mGluR agonists.
Subject(s)
Amino Acids/pharmacology , Arthritis, Experimental , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Central Amygdaloid Nucleus/drug effects , Excitatory Amino Acid Agonists/pharmacology , Nociception/drug effects , Posterior Horn Cells/drug effects , Receptors, Metabotropic Glutamate/agonists , Amygdala/drug effects , Animals , Arthralgia , Behavior, Animal/drug effects , Carrageenan , Excitatory Amino Acid Antagonists/pharmacology , Kaolin , Pain/metabolism , Rats , Receptors, Metabotropic Glutamate/metabolism , Spinal Cord/cytology , Spinal Cord/drug effects , Spinal Cord/metabolism , Vocalization, Animal/drug effects , Xanthenes/pharmacologyABSTRACT
BACKGROUND: Our previous research confirmed that electroacupuncture (EA) stimulus elicits neuroprotective effects against cerebral ischemic injury through α7 nicotinic acetylcholine receptor (α7nAChR)-mediated inhibition of high-mobility group box 1 release mechanism. This study investigated whether the signal transducer of α7nAChR and inhibition of NLRP3 inflammasome are involved in the neuroprotective effects of EA stimulus. METHODS: In adult male Sprague-Dawley rats, the focal cerebral ischemic injury was induced by middle cerebral artery occlusion (MCAO) models for 1.5 h. The expression of NLRP3 inflammasome in the penumbral tissue following reperfusion was assessed by western blotting and immunoflourescent staining. The infarct size, neurological deficit score, TUNEL staining and the expression of proinflammatory factors or anti-inflammatory cytokines were evaluated at 72 h after reperfusion in the presence or absence of either α7nAChR antagonist (α-BGT) or agonist (PHA-543,613). RESULTS: The contents of inflammasome proteins were gradually increased after cerebral ischemia/reperfusion (I/R). EA stimulus attenuated NLRP3 inflammasome mediated inflammatory reaction and regulated the balance between proinflammatory factors and anti-inflammatory cytokines. The agonist of α7nAChR induced similar neuroprotective effects as EA stimulus. In contrast, α7nAChR antagonist reversed not only the neuroprotective effects, but also the inhibitory effects of NLRP3 inflammasome and the regulatory effects on the balance between proinflammatory factors and anti-inflammatory cytokines. CONCLUSIONS: These results provided compelling evidence that α7nAChR played a pivotal role in regulating the activation and expression of NLRP3 inflammasome in neurons after cerebral I/R. These findings highlighted a novel anti-inflammatory mechanism of EA stimulus by α7nAChR modulating the inhibition of NLRP3 inflammasome, suggesting that α7nAChR-dependent cholinergic anti-inflammatory system and NLRP3 inflammasome in neurons might act as potential therapeutic targets in EA induced neuroprotection against cerebral ischemic injury.
Subject(s)
Brain Ischemia/metabolism , Brain Ischemia/therapy , Electroacupuncture/methods , Inflammasomes/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , alpha7 Nicotinic Acetylcholine Receptor/metabolism , Animals , Blotting, Western , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , In Situ Nick-End Labeling , Infarction, Middle Cerebral Artery/metabolism , Infarction, Middle Cerebral Artery/therapy , Inflammation/metabolism , Inflammation/therapy , Injections, Intraventricular , Male , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Quinuclidines/pharmacology , Rats , Rats, Sprague-Dawley , Signal Transduction/drug effects , alpha7 Nicotinic Acetylcholine Receptor/agonists , alpha7 Nicotinic Acetylcholine Receptor/antagonists & inhibitorsABSTRACT
In order to find the endogenous potential biomarkers of in vitro hepatic injury caused by NCTD-Na and elucidate the mechanism of hepatic injury of NCTD-Na,ultra-high performance liquid chromatography coupled quadrupole time-of-flight mass spectrometry(UPLC-Q-TOF-MS/MS) was used for lipidomics detection.Multivariate statistical analysis was used to study the endogenous lipid metabolic changes of human normal liver cells LO2 injury after the treatment with sodium norcantharidate(NCTD-Na).The results showed that the half maximal inhibitory concentration(IC50) of NCTD-Na was 0.034 mmol·L-1.A total of 280 differential metabolites were found between the control group and the low-dose group,with VIP > 2.0 and P<0.05.At the same time,a total of 273 differential metabolites were found between the control group and the high-dose group,with VIP > 2.0 and P<0.05.Cell metabolite profiles showed clear separation among control group,the low-dose group and the high-dose group,and 111 differential metabolites were found,with VIP > 2.0,P<0.05,RSD<30% and in a dose-dependent manner.It was found that most of the above differential metabolites were lipid metabolites after the analysis of simple preparnation methods and database search.A total of 32 potential biomarkers were identified,including 3 phosphatidylcholine(PC),5 lysophosphatidylcholine(Lyso PC),3 ceramide(Cer),1 sphingomyelin(SM),1 phosphatidylethanolamine(PE),10 lysophosphatidylethanolamine(LysoPE),4 diacylglycerol(DG),1 Phosphatidic acid(PA),1 lysophosphatidic acid(Lyso PA),1 phosphatidyl glycerol(PG),1 fatty acid hydroxy fatty acid(FAHFA) and 1 phosphatidylserine(PS).The changes of PCs,Cers,SM,PE and DGs were closely related liver protection,DNA methylation and self-repair in hepatocytes,apoptosis,methylation and detoxification of carcinogens,as well as lipid peroxides production process.Also,they had impact on the proliferation of hepatocytes,differentiation and gene transcription disorders.Cells stimulated by NCTD-Na could promote the production of PA as well as the synthesis and catabolism of FAHFA in a variety of ways.The levels of Lyso PCs,LysoPEs and Lyso PA were correlated with PCs,PE and PA;PE and PS might have valgus during apoptosis,triggering phagocytosis.
Subject(s)
Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Hepatocytes/drug effects , Lipid Metabolism , Cells, Cultured , Hepatocytes/metabolism , Humans , Lipids/analysis , Tandem Mass SpectrometryABSTRACT
Geranylgeranoic acid (GGA) has been reported to induce autophagic cell death via upregulation of lipid-induced unfolded protein response in several human hepatoma-derived cell lines, and its 4,5-didehydro derivative has been developed as a preventive agent against second primary hepatoma in clinical trials. We have previously reported that GGA is a natural diterpenoid synthesized in several medicinal herbs. Here, we provide unequivocal evidence for de novo GGA biosynthesis in mammals. First, with normal male Wistar rats, the levels of GGA in liver were found to be far greater than those in other organs analyzed. Second, we demonstrated the metabolic GGA labeling from the 13C-labeled mevalonolactone in the human hepatoma-derived cell line, HuH-7. Isotopomer spectral analysis revealed that approximately 80% of the cellular GGA was newly synthesized from mevalonate (MVA) in 12 h and the acid picked up preexisting farnesyl diphosphate (FPP) and geranylgeranyl diphosphate (GGPP), suggesting that GGA is derived from FPP and GGPP through the MVA pathway. Third, zaragozic acid A, a squalene synthase inhibitor, induced dose-dependent upregulation of endogenous GGA content in HuH-7 cells and their concomitant cell death. These results strongly suggest that a cancer-preventive GGA is biosynthesized via the MVA pathway in mammals.
Subject(s)
Diterpenes/metabolism , Mevalonic Acid/metabolism , Animals , Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Cell Death/drug effects , Cell Line, Tumor , Male , Rats , Rats, Wistar , Tricarboxylic Acids/pharmacologyABSTRACT
HYPOTHESIS: SENS-401 (R-azasetron besylate) is effective against severe acoustic trauma-induced hearing loss. BACKGROUND: SENS-401 has calcineurin inhibiting properties and attenuates cisplatin-induced hearing loss in a rat model. Cisplatin-induced and acoustic trauma-induced hearing loss share common apoptotic pathways. METHODS: The dose-response relationship of SENS-401 (6.6âmg/kg BID, 13.2âmg/kg BID, 26.4âmg/kg QD) and treatment time-window (13.2âmg/kg BID starting 24, 72, and 96âh posttrauma) versus placebo for 28 days were evaluated in a male rat model of severe acoustic trauma-induced hearing loss (120âdB SPL, 2âh) using auditory brainstem response (ABR) and distortion product otoacoustic emissions (DPOAE) measures followed by cochlear outer hair cell (OHC) counting with myosin-VIIa immunolabeling. RESULTS: All SENS-401 doses improved ABR threshold shift and recovery, reaching statistical significance (pâ<â0.05) for ABR threshold recoveries after 28-days treatment. DPOAE amplitude loss and recovery improved markedly for 13.2âmg/kg BID SENS-401, reaching significance after 14 days (pâ<â0.05). Significant improvements in ABR threshold shifts/recovery and DPOAE amplitude loss occurred with up to 96-hours delay in initiating SENS-401 (pâ<â0.05), and in DPOAE amplitude recovery with up to 72-hours delay (pâ<â0.05). Significantly more surviving OHCs were present after SENS-401 treatment compared with placebo after 24 to 96-hours delay posttrauma, with up to 5.3-fold more cells in the basal cochlea turn. CONCLUSIONS: In vivo data support the otoprotective potential of twice daily oral SENS-401. Improvements in hearing loss recovery make SENS-401 a promising clinical candidate for acoustic trauma-induced hearing loss, including when treatment is not initiated immediately.
Subject(s)
Bridged Bicyclo Compounds, Heterocyclic/pharmacology , Evoked Potentials, Auditory, Brain Stem/drug effects , Hearing Loss, Noise-Induced/drug therapy , Hearing Loss, Sensorineural/drug therapy , Oxazines/pharmacology , Acoustic Stimulation/adverse effects , Animals , Bridged Bicyclo Compounds, Heterocyclic/administration & dosage , Bridged Bicyclo Compounds, Heterocyclic/therapeutic use , Cisplatin/toxicity , Hair Cells, Auditory, Outer/drug effects , Hearing Loss, Sensorineural/chemically induced , Male , Otoacoustic Emissions, Spontaneous/drug effects , Oxazines/administration & dosage , Oxazines/therapeutic use , Rats , Rats, WistarABSTRACT
BH3 mimetics are novel targeted drugs with remarkable specificity, potency and enormous potential to improve cancer therapy. However, acquired resistance is an emerging problem. We report the rapid development of resistance in chronic lymphocytic leukemia cells isolated from patients exposed to increasing doses of navitoclax (ABT-263), a BH3 mimetic. To mimic such rapid development of chemoresistance, we developed simple resistance models to three different BH3 mimetics, targeting BCL-2 (ABT-199), BCL-XL (A-1331852) or MCL-1 (A-1210477), in relevant hematologic cancer cell lines. In these models, resistance could not be attributed to either consistent changes in expression levels of the anti-apoptotic proteins or interactions among different pro- and anti-apoptotic BCL-2 family members. Using genetic silencing, pharmacological inhibition and metabolic supplementation, we found that targeting glutamine uptake and its downstream signaling pathways, namely glutaminolysis, reductive carboxylation, lipogenesis, cholesterogenesis and mammalian target of rapamycin signaling resulted in marked sensitization of the chemoresistant cells to BH3 mimetic-mediated apoptosis. Furthermore, our findings highlight the possibility of repurposing widely used drugs, such as statins, to target intermediary metabolism and improve the efficacy of BH3 mimetic therapy.