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1.
Molecules ; 24(23)2019 Nov 22.
Article in English | MEDLINE | ID: mdl-31771098

ABSTRACT

The identification of signaling pathways that are involved in gliomagenesis is crucial for targeted therapy design. In this study we assessed the biological and therapeutic effect of ingenol-3-dodecanoate (IngC) on glioma. IngC exhibited dose-time-dependent cytotoxic effects on large panel of glioma cell lines (adult, pediatric cancer cells, and primary cultures), as well as, effectively reduced colonies formation. Nevertheless, it was not been able to attenuate cell migration, invasion, and promote apoptotic effects when administered alone. IngC exposure promoted S-phase arrest associated with p21CIP/WAF1 overexpression and regulated a broad range of signaling effectors related to survival and cell cycle regulation. Moreover, IngC led glioma cells to autophagy by LC3B-II accumulation and exhibited increased cytotoxic sensitivity when combined to a specific autophagic inhibitor, bafilomycin A1. In comparison with temozolomide, IngC showed a mean increase of 106-fold in efficacy, with no synergistic effect when they were both combined. When compared with a known compound of the same class, namely ingenol-3-angelate (I3A, Picato®), IngC showed a mean 9.46-fold higher efficacy. Furthermore, IngC acted as a potent inhibitor of protein kinase C (PKC) activity, an emerging therapeutic target in glioma cells, showing differential actions against various PKC isotypes. These findings identify IngC as a promising lead compound for the development of new cancer therapy and they may guide the search for additional PKC inhibitors.


Subject(s)
Antineoplastic Agents/pharmacology , Brain Neoplasms/enzymology , Diterpenes/pharmacology , Euphorbia/chemistry , Glioma/enzymology , Protein Kinase C/antagonists & inhibitors , Antineoplastic Agents/chemistry , Autophagy , Brain Neoplasms/drug therapy , Cell Cycle/drug effects , Cell Line, Tumor , Cell Movement/drug effects , Cell Proliferation/drug effects , Diterpenes/chemistry , Drug Screening Assays, Antitumor , Gene Expression Regulation, Neoplastic/drug effects , Glioma/drug therapy , Humans , Plant Extracts/chemistry , Plant Extracts/pharmacology , Signal Transduction/drug effects
2.
Invest New Drugs ; 37(5): 1029-1035, 2019 10.
Article in English | MEDLINE | ID: mdl-30706338

ABSTRACT

The latex from Euphorbia tirucalli is used in Brazil as a folk medicine for several diseases, including cancer. Recently, we showed a cytotoxic activity of E. tirucalli euphol in a wide range of cancer cell lines. Moreover, we showed that euphol inhibits proliferation, motility and colony formation in pancreatic cancer cells, induces autophagy and sensitizes glioblastoma cells to temozolomide cytotoxicity. Herein, we report in vitro activity of three semi-synthetic ingenol compounds derived from E. tirucalli, IngA (ingenol-3-trans-cinnamate), IngB (ingenol-3-hexanoate) and IngC (ingenol-3-dodecanoate), against a large panel of human cancer cell lines. Antineoplastic effects of the three semi-synthetic compounds were assessed using MTS assays on 70 cancer cell lines from a wide array of solid tumors. Additionally, their antitumor potential was compared with known compounds of the same class, namely ingenol-3-angelate (Picato®) and ingenol 3,20-dibenzoate and in combination with standard chemotherapeutic agents. We observed that IngA, B, and C exhibited dose-dependent cytotoxic effects. Amongst the semi-synthetic compounds, IngC displayed the best activity across the tumor cell lines. In comparison with ingenol-3-angelate and ingenol 3,20-dibenzoate, IngC showed a mean of 6.6 and 3.6-fold higher efficacy, respectively, against esophageal cancer cell lines. Besides, IngC sensitized esophageal cancer cells to paclitaxel treatment. In conclusion, the semi-synthetic ingenol compounds, in particular, IngC, demonstrated a potent antitumor activity on all cancer cell lines evaluated. Although the underlying mechanisms of action of IngC are not elucidated, our results provide insights for further studies suggesting IngC as a putative therapy for cancer treatment.


Subject(s)
Antineoplastic Agents, Phytogenic/pharmacology , Apoptosis/drug effects , Cell Proliferation/drug effects , Diterpenes/pharmacology , Euphorbia/chemistry , Neoplasms/drug therapy , Neoplasms/pathology , Antineoplastic Agents, Phytogenic/chemistry , Diterpenes/chemistry , Humans , Tumor Cells, Cultured
3.
Invest New Drugs ; 37(2): 223-237, 2019 04.
Article in English | MEDLINE | ID: mdl-29931585

ABSTRACT

Glioblastoma (GBM) is the most frequent and aggressive type of brain tumor. There are limited therapeutic options for GBM so that new and effective agents are urgently needed. Euphol is a tetracyclic triterpene alcohol, and it is the main constituent of the sap of the medicinal plant Euphorbia tirucalli. We previously identified anti-cancer activity in euphol based on the cytotoxicity screening of 73 human cancer cells. We now expand the toxicological screening of the inhibitory effect and bioactivity of euphol using two additional glioma primary cultures. Euphol exposure showed similar cytotoxicity against primary glioma cultures compared to commercial glioma cells. Euphol has concentration-dependent cytotoxic effects on cancer cell lines, with more than a five-fold difference in the IC50 values in some cell lines. Euphol treatment had a higher selective cytotoxicity index (0.64-3.36) than temozolomide (0.11-1.13) and reduced both proliferation and cell motility. However, no effect was found on cell cycle distribution, invasion and colony formation. Importantly, the expression of the autophagy-associated protein LC3-II and acidic vesicular organelle formation were markedly increased, with Bafilomycin A1 potentiating cytotoxicity. Finally, euphol also exhibited antitumoral and antiangiogenic activity in vivo, using the chicken chorioallantoic membrane assay, with synergistic temozolomide interactions in most cell lines. In conclusion, euphol exerted in vitro and in vivo cytotoxicity against glioma cells, through several cancer pathways, including the activation of autophagy-associated cell death. These findings provide experimental support for further development of euphol as a novel therapeutic agent for GBM, either alone or in combination chemotherapy.


Subject(s)
Autophagy , Drug Resistance, Neoplasm/drug effects , Drug Synergism , Euphorbia/chemistry , Glioblastoma/pathology , Lanosterol/analogs & derivatives , Temozolomide/pharmacology , Antineoplastic Agents, Alkylating/pharmacology , Apoptosis , Brain Neoplasms/drug therapy , Brain Neoplasms/pathology , Cell Movement , Cell Proliferation , Glioblastoma/drug therapy , Humans , Lanosterol/pharmacology , Tumor Cells, Cultured
4.
Exp Ther Med ; 16(2): 557-566, 2018 Aug.
Article in English | MEDLINE | ID: mdl-30112023

ABSTRACT

A large number of classic antineoplastic agents are derived from plants. Euphorbia tirucalli L. (Euphorbiaceae) is a subtropical and tropical plant, used in Brazilian folk medicine against many diseases, including cancer, yet little is known about its true anticancer properties. The present study evaluated the antitumor effect of the tetracyclic triterpene alcohol, euphol, the main constituent of E. tirucalli in a panel of 73 human cancer lines from 15 tumor types. The biological effect of euphol in pancreatic cells was also assessed. The combination index was further used to explore euphol interactions with standard drugs. Euphol showed a cytotoxicity effect against several cancer cell lines (IC50 range, 1.41-38.89 µM), particularly in esophageal squamous cell (11.08 µM) and pancreatic carcinoma cells (6.84 µM), followed by prostate, melanoma, and colon cancer. Cytotoxicity effects were seen in all cancer cell lines, with more than half deemed highly sensitive. Euphol inhibited proliferation, motility and colony formation in pancreatic cancer cells. Importantly, euphol exhibited synergistic interactions with gemcitabine and paclitaxel in pancreatic and esophageal cell lines, respectively. To the best of our knowledge, this study constitutes the largest in vitro screening of euphol efficacy on cancer cell lines and revealed its in vitro anti-cancer properties, particularly in pancreatic and esophageal cell lines, suggesting that euphol, either as a single agent or in combination with conventional chemotherapy, is a potential anti-cancer drug.

5.
J Diet Suppl ; 15(5): 673-683, 2018 Sep 03.
Article in English | MEDLINE | ID: mdl-29190155

ABSTRACT

Fatigue is frequent among oncologic patients. Unpurified Paullinia cupana dry extract showed encouraging results for chemotherapy-induced fatigue in our previous studies. We report two randomized, double-blind studies with a standardized dry purified Paullinia cupana extract named PC-18. For both studies, we recruited early breast cancer patients who had an increase in their fatigue scores after their first cycle of adjuvant chemotherapy. In the first study, we compared an oral dose of 37.5 mg of PC-18 twice daily with placebo. In the second study, we examined PC-18 at either 7.5 or 12.5 mg orally twice daily versus placebo. In both studies, PC-18 was not superior to placebo as assessed by both Chalder and Brief Fatigue Inventory (BFI) fatigue questionnaires, probably reflecting unexpectedly good placebo antifatigue activity. Since all capsules employed in both studies contained about 100 mg of magnesium silicate as an excipient, we retrospectively evaluated frozen serum samples from the second study and found a significant increase in magnesium levels after patients received placebo. By multivariate analysis, higher prerandomization magnesium levels and higher BFI scores together with the use of a 12.5 mg dose of PC-18 all correlated significantly with higher posttreatment BFI scores. We observed no significant toxicities in any of the trials. We conclude that the absence of differences between PC-18 and placebo may be due to the unexpectedly high antifatigue activity of the placebo in these studies. Further studies evaluating the role of magnesium supplementation for chemotherapy-induced fatigue are needed.


Subject(s)
Antineoplastic Agents/adverse effects , Fatigue/drug therapy , Paullinia , Phytotherapy , Plant Extracts/therapeutic use , Breast Neoplasms/drug therapy , Double-Blind Method , Fatigue/chemically induced , Female , Humans , Magnesium/administration & dosage , Magnesium/blood , Middle Aged , Placebos , Seeds/chemistry , Surveys and Questionnaires
6.
PLoS One ; 12(5): e0174516, 2017.
Article in English | MEDLINE | ID: mdl-28467486

ABSTRACT

BACKGROUND: Some latency-reversing agents (LRAs) inhibit HIV-specific CD8+ T cell responses. In a prior study of protein kinase C (PKC) agonists, we found that bryostatin-1 inhibited elite controller/suppressor (ES) CD8+ T cell suppressive activity whereas prostratin had no effect. Ingenol-B is another PKC agonist with potent LRA activity both by itself and in combination with the bromodomain inhibitor JQ1; however its effect on CD8+ T cell mediated control of HIV-1 replication is unknown. METHODS: CD8+ T cells were isolated from ES and treated with bryostatin-1, prostratin, ingenol-B, and JQ1 as well as a combination of each PKC-agonist with JQ1. The cells were then tested in the viral suppression assay. To assess possible mechanisms of inhibition, CD8+ T cells were treated with the LRAs and analyzed for the expression of various immune cell markers. RESULTS: Ingenol-B had no effect on the ability of ES CD8+ T cells to suppress viral replication, however, the combination of ingenol-B and JQ1 caused a modest, but significant decrease in this suppressive capacity. The mechanism of the inhibitory effect of the JQ1 and ingenol-B combination relative to ingenol-B alone was unclear but the effect appeared to be dose dependent. CONCLUSIONS: Ingenol-B does not inhibit HIV-specific CD8+ T cell responses in vitro. These responses are however modestly inhibited when 100 nMingenol-B is combined with JQ1. Since HIV-specific CD8+ T cell activity may be essential for the eradication of reactivated latently infected cells, the potency of latency-reversal activity of drug combinations must be balanced against the effects of the combinations on HIV-specific CD8+ T cell responses.


Subject(s)
CD8-Positive T-Lymphocytes/drug effects , Diterpenes/pharmacology , HIV-1/immunology , Azepines/administration & dosage , Azepines/pharmacology , CD8-Positive T-Lymphocytes/immunology , Cell Death/drug effects , Diterpenes/administration & dosage , Dose-Response Relationship, Drug , Flow Cytometry , Humans , Lymphocyte Activation/drug effects , Triazoles/administration & dosage , Triazoles/pharmacology
7.
AIDS ; 31(1): 5-14, 2017 01 02.
Article in English | MEDLINE | ID: mdl-27898590

ABSTRACT

OBJECTIVE: Resting CD4 T cells have been recognized as the major cell reservoir of latent HIV-1 during antiretroviral therapy (ART). Using an simian immunodeficiency virus (SIV)/macaque model for AIDS and HIV-related neurocognitive disorders we assessed the contribution of the brain to viral latency and reactivation. DESIGN: Pigtailed macaques were dual inoculated with SIVDeltaB670 and SIV17E-Fr and treated with an efficacious central nervous system-penetrant ART. After 500 days of viral suppression animals were treated with two cycles of latency reversing agents and increases in viral transcripts were examined. METHODS: Longitudinal plasma and cerebrospinal fluid (CSF) viral loads were analyzed by quantitative and digital droplet PCR. After necropsy, viral transcripts in organs were analyzed by PCR, in-situ hybridization, and phylogenetic genotyping based on env V1 loop sequences. Markers for neuronal damage and CSF activation were measured by ELISA. RESULTS: Increases in activation markers and plasma and CSF viral loads were observed in one animal treated with latency reversing agents, despite ongoing ART. SIV transcripts were identified in occipital cortex macrophages by in-situ hybridization and CD68 staining. The most abundant SIV genotype in CSF was unique and expanded independent from viruses found in the periphery. CONCLUSION: The central nervous system harbors latent SIV genomes after long-term viral suppression by ART, indicating that the brain represents a potential viral reservoir and should be seriously considered during AIDS cure strategies.


Subject(s)
Brain/virology , Neurocognitive Disorders/virology , Simian Acquired Immunodeficiency Syndrome/virology , Simian Immunodeficiency Virus/physiology , Virus Activation , Virus Latency , Animals , Anti-Retroviral Agents/therapeutic use , Cerebrospinal Fluid/virology , Disease Models, Animal , Enzyme-Linked Immunosorbent Assay , Gene Products, env/genetics , Genotyping Techniques , In Situ Hybridization , Macaca , Neurocognitive Disorders/drug therapy , Phylogeny , Plasma/virology , Polymerase Chain Reaction , Sequence Analysis, DNA , Simian Acquired Immunodeficiency Syndrome/complications , Simian Acquired Immunodeficiency Syndrome/drug therapy , Viral Load
8.
PLoS Pathog ; 11(7): e1005063, 2015 Jul.
Article in English | MEDLINE | ID: mdl-26225566

ABSTRACT

The persistence of latently infected cells in patients under combinatory antiretroviral therapy (cART) is a major hurdle to HIV-1 eradication. Strategies to purge these reservoirs are needed and activation of viral gene expression in latently infected cells is one promising strategy. Bromodomain and Extraterminal (BET) bromodomain inhibitors (BETi) are compounds able to reactivate latent proviruses in a positive transcription elongation factor b (P-TEFb)-dependent manner. In this study, we tested the reactivation potential of protein kinase C (PKC) agonists (prostratin, bryostatin-1 and ingenol-B), which are known to activate NF-κB signaling pathway as well as P-TEFb, used alone or in combination with P-TEFb-releasing agents (HMBA and BETi (JQ1, I-BET, I-BET151)). Using in vitro HIV-1 post-integration latency model cell lines of T-lymphoid and myeloid lineages, we demonstrated that PKC agonists and P-TEFb-releasing agents alone acted as potent latency-reversing agents (LRAs) and that their combinations led to synergistic activation of HIV-1 expression at the viral mRNA and protein levels. Mechanistically, combined treatments led to higher activations of P-TEFb and NF-κB than the corresponding individual drug treatments. Importantly, we observed in ex vivo cultures of CD8+-depleted PBMCs from 35 cART-treated HIV-1+ aviremic patients that the percentage of reactivated cultures following combinatory bryostatin-1+JQ1 treatment was identical to the percentage observed with anti-CD3+anti-CD28 antibodies positive control stimulation. Remarkably, in ex vivo cultures of resting CD4+ T cells isolated from 15 HIV-1+ cART-treated aviremic patients, the combinations bryostatin-1+JQ1 and ingenol-B+JQ1 released infectious viruses to levels similar to that obtained with the positive control stimulation. The potent effects of these two combination treatments were already detected 24 hours post-stimulation. These results constitute the first demonstration of LRA combinations exhibiting such a potent effect and represent a proof-of-concept for the co-administration of two different types of LRAs as a potential strategy to reduce the size of the latent HIV-1 reservoirs.


Subject(s)
Bryostatins/pharmacology , CD4-Positive T-Lymphocytes/virology , Gene Expression Regulation, Viral/drug effects , HIV-1/drug effects , CD4-Positive T-Lymphocytes/drug effects , Diterpenes/metabolism , HIV-1/physiology , Humans , Positive Transcriptional Elongation Factor B/metabolism , Virus Activation/drug effects , Virus Latency/drug effects
9.
Virology ; 462-463: 328-39, 2014 Aug.
Article in English | MEDLINE | ID: mdl-25014309

ABSTRACT

The ability of HIV to establish long-lived latent infection is mainly due to transcriptional silencing of viral genome in resting memory T lymphocytes. Here, we show that new semi-synthetic ingenol esters reactivate latent HIV reservoirs. Amongst the tested compounds, 3-caproyl-ingenol (ING B) was more potent in reactivating latent HIV than known activators such as SAHA, ingenol 3,20-dibenzoate, TNF-α, PMA and HMBA. ING B activated PKC isoforms followed by NF-κB nuclear translocation. As virus reactivation is dependent on intact NF-κB binding sites in the LTR promoter region ING B, we have shown that. ING B was able to reactivate virus transcription in primary HIV-infected resting cells up to 12 fold and up to 25 fold in combination with SAHA. Additionally, ING B promoted up-regulation of P-TEFb subunits CDK9/Cyclin T1. The role of ING B on promoting both transcription initiation and elongation makes this compound a strong candidate for an anti-HIV latency drug combined with suppressive HAART.


Subject(s)
Diterpenes/metabolism , HIV-1/drug effects , HIV-1/physiology , Virus Activation/drug effects , Virus Latency/drug effects , Cell Line , Esters/metabolism , Humans , Virus Replication/drug effects
10.
AIDS ; 28(11): 1555-66, 2014 Jul 17.
Article in English | MEDLINE | ID: mdl-24804860

ABSTRACT

OBJECTIVE: Although HAART effectively suppresses viral replication, it fails to eradicate latent viral reservoirs. The 'shock and kill' strategy involves the activation of HIV from latent reservoirs and targeting them for eradication. Our goal was to develop new approaches for activating HIV from latent reservoirs. DESIGN: We investigated capacity of Ingenol B (IngB), a newly modified derivative of Ingenol ester that was originally isolated from a Brazilian plant in Amazon, for its capacity and mechanisms of HIV reactivation. METHODS: Reactivation of HIV-1 by IngB was evaluated in J-Lat A1 cell culture model of HIV latency as well as in purified primary CD4 T cells from long-term HAART-treated virologically-suppressed HIV-infected individuals. The underlining molecular mechanisms of viral reactivation were investigated using flow cytometry, RT-qPCR and chromatin immunoprecipitation. RESULTS: IngB is highly effective in reactivating HIV in J-Lat A1 cells with relatively low cellular toxicity. It is also able to reactivate latent HIV in purified CD4 T cells from HAART-treated HIV-positive individuals ex vivo. Our data show that IngB may reactivate HIV expression by both activating protein kinase C (PKC)δ-nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) pathway and directly inducing NF-κB protein expression. Importantly, IngB has a synergistic effect with JQ1, a BET bromodomain inhibitor, in latent HIV reactivation. CONCLUSIONS: IngB is a new promising compound to activate latent HIV reservoirs. Our data suggest that formulating novel derivatives from Ingenol esters may be an innovative approach to develop new lead compounds to reactivate latent HIV.


Subject(s)
CD4-Positive T-Lymphocytes/drug effects , Diterpenes/metabolism , NF-kappa B/metabolism , Protein Kinase C/metabolism , Signal Transduction , Virus Activation/drug effects , Virus Latency/drug effects , Adult , Aged , CD4-Positive T-Lymphocytes/virology , Cells, Cultured , Chromatin Immunoprecipitation , Flow Cytometry , HIV-1/drug effects , Humans , Male , Middle Aged , Proviruses/drug effects , Real-Time Polymerase Chain Reaction
11.
PLoS One ; 9(5): e97257, 2014.
Article in English | MEDLINE | ID: mdl-24827152

ABSTRACT

HIV infection is not cleared by antiretroviral drugs due to the presence of latently infected cells that are not eliminated with current therapies and persist in the blood and organs of infected patients. New compounds to activate these latent reservoirs have been evaluated so that, along with HAART, they can be used to activate latent virus and eliminate the latently infected cells resulting in eradication of viral infection. Here we describe three novel diterpenes isolated from the sap of Euphorbia tirucalli, a tropical shrub. These molecules, identified as ingenols, were modified at carbon 3 and termed ingenol synthetic derivatives (ISD). They activated the HIV-LTR in reporter cell lines and human PBMCs with latent virus in concentrations as low as 10 nM. ISDs were also able to inhibit the replication of HIV-1 subtype B and C in MT-4 cells and human PBMCs at concentrations of EC50 0.02 and 0.09 µM respectively, which are comparable to the EC50 of some antiretroviral currently used in AIDS treatment. Control of viral replication may be caused by downregulation of surface CD4, CCR5 and CXCR4 observed after ISD treatment in vitro. These compounds appear to be less cytotoxic than other diterpenes such as PMA and prostratin, with effective dose versus toxic dose TI>400. Although the mechanisms of action of the three ISDs are primarily attributed to the PKC pathway, downregulation of surface receptors and stimulation of the viral LTR might be differentially modulated by different PKC isoforms.


Subject(s)
Anti-HIV Agents/pharmacology , Diterpenes/pharmacology , Euphorbia/metabolism , HIV-1/drug effects , Virus Latency/drug effects , Virus Replication/drug effects , Antiretroviral Therapy, Highly Active/methods , CD4 Antigens/metabolism , Cell Line , Down-Regulation/drug effects , HIV Infections/drug therapy , HIV Infections/virology , HIV Seropositivity/drug therapy , HIV Seropositivity/metabolism , HIV Seropositivity/virology , Humans , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/virology , Receptors, CCR5/metabolism , Receptors, CXCR4/metabolism , Transcription, Genetic/drug effects , Up-Regulation/drug effects
12.
J Diet Suppl ; 10(4): 325-34, 2013 Dec.
Article in English | MEDLINE | ID: mdl-24237188

ABSTRACT

PURPOSE: Paullinia cupana (guaraná) is an Amazonian plant that has been previously shown to be effective in treating chemotherapy-related fatigue (CRF) in patients with breast cancer. We aimed to evaluate the efficacy of a purified dry extract of P. cupana (PC-18) in patients with various solid tumors treated with chemotherapy. METHODS: We included 40 patients with solid tumors who showed increases in their Brief Fatigue Inventory (BFI) questionnaire scores after 1 week of systemic chemotherapy. PC-18 was administered at 37.5 mg by mouth two times per day (PO bid), starting after 1 week of chemotherapy, for 3 weeks (induction phase). Patients who had an improvement in or stabilization of their BFI scores were randomized to receive either PC-18 at the same dose or placebo for the following 3 weeks (maintenance phase). RESULTS: After PC-18 treatment, the BFI fatigue scores improved or stabilized in 36 out of the 40 patients (mean BFI score difference = 2.503; 95% confidence interval: 1.716-3.375, p = .0002). Three weeks after randomization (16 patients on PC-18 and 17 on placebo), we observed no significant differences in the BFI, Functional Assessment of Chronic Illness Therapy, Hospital Anxiety and Depression Scale, and Pittsburgh Sleep Quality Index scores between patients randomized to PC-18 versus placebo. CONCLUSIONS: We conclude that the PC-18 extract may be effective for the treatment of CRF in patients with a variety of solid tumors. A conditioning effect, which was observed in patients who had a beneficial effect of PC-18 on CRF, may explain the better than expected fatigue scores of the placebo-treated patients.


Subject(s)
Antineoplastic Agents/adverse effects , Fatigue/drug therapy , Neoplasms , Paullinia , Phytotherapy , Plant Extracts/therapeutic use , Adult , Aged , Fatigue/chemically induced , Female , Humans , Male , Middle Aged , Neoplasms/drug therapy , Plant Extracts/pharmacology , Quality of Life , Surveys and Questionnaires
13.
Eur J Pharmacol ; 698(1-3): 413-20, 2013 Jan 05.
Article in English | MEDLINE | ID: mdl-23099255

ABSTRACT

Inflammation underlies the development and progression of a number of skin disorders including psoriasis, atopic dermatitis and cancer. Therefore, novel antiinflammatory agents are of great clinical interest for prevention and treatment of these conditions. Herein, we demonstrated the underlying molecular mechanisms of the antiinflammatory activity of euphol, a tetracyclic triterpene isolated from the sap of Euphorbia tirucalli, in skin inflammation induced by 12-O-tetradecanoylphorbol-13-acetate (TPA) in mice. Topical application of euphol (100 µg/ear) significantly inhibited TPA-induced ear edema and leukocyte influx through the reduction of keratinocyte-derived chemokine (CXCL1/KC) and macrophage inflammatory protein (MIP)-2 levels. At the intracellular level, euphol reduced TPA-induced extracellular signal-regulated protein kinase (ERK) activation and cyclooxygenase-2 (COX-2) upregulation. These effects were associated with euphol's ability to prevent TPA-induced protein kinase C (PKC) activation, namely PKCα and PKCδ isozymes. Our data indicate that topical application of euphol markedly inhibits the inflammatory response induced by TPA. Thus, euphol represents a promising agent for the management of skin diseases with an inflammatory component.


Subject(s)
Extracellular Signal-Regulated MAP Kinases/metabolism , Lanosterol/analogs & derivatives , MAP Kinase Signaling System/drug effects , Protein Kinase C/metabolism , Skin Diseases/chemically induced , Skin Diseases/drug therapy , Tetradecanoylphorbol Acetate/adverse effects , Administration, Topical , Animals , Anti-Inflammatory Agents, Non-Steroidal/administration & dosage , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Cyclooxygenase 2/metabolism , Edema/chemically induced , Edema/drug therapy , Edema/immunology , Edema/pathology , Enzyme Activation/drug effects , Inflammation/chemically induced , Inflammation/drug therapy , Inflammation/immunology , Inflammation/pathology , Lanosterol/administration & dosage , Lanosterol/pharmacology , Lanosterol/therapeutic use , Leukocytes/drug effects , Leukocytes/immunology , Male , Mice , Mitogen-Activated Protein Kinase 1/metabolism , Mitogen-Activated Protein Kinase 3/metabolism , Skin Diseases/immunology , Skin Diseases/pathology , Up-Regulation/drug effects
14.
Neuropharmacology ; 63(4): 593-605, 2012 Sep.
Article in English | MEDLINE | ID: mdl-22613837

ABSTRACT

Persistent pains associated with inflammatory and neuropathic states are prevalent and debilitating diseases, which still remain without a safe and adequate treatment. Euphol, an alcohol tetracyclic triterpene, has a wide range of pharmacological properties and is considered to have anti-inflammatory action. Here, we assessed the effects and the underlying mechanisms of action of euphol in preventing inflammatory and neuropathic pain. Oral treatment with euphol (30 and 100 mg/kg) reduced carrageenan-induced mechanical hyperalgesia. Likewise, euphol given through the spinal and intracerebroventricular routes prevented mechanical hyperalgesia induced by carrageenan. Euphol consistently blocked the mechanical hyperalgesia induced by complete Freund's adjuvant, keratinocyte-derived chemokine, interleukin-1ß, interleukin-6 and tumor necrosis factor-alpha associated with the suppression of myeloperoxidase activity in the mouse paw. Oral treatment with euphol was also effective in preventing the mechanical nociceptive response induced by ligation of the sciatic nerve and also significantly reduced the levels and mRNA of cytokines/chemokines in both paw and spinal cord tissues following i.pl. injection of complete Freund's adjuvant. In addition, the pre-treatment with either CB1R or CB2R antagonists, as well as the knockdown gene of the CB1R and CB2R, significantly reversed the antinociceptive effect of euphol. Interestingly, even in higher doses, euphol did not cause any relevant action in the central nervous system. Considering that few drugs are currently available for the treatment of chronic pain states, the present results provided evidence that euphol constitutes a promising molecule for the management of inflammatory and neuropathic pain states.


Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Edema/prevention & control , Hyperalgesia/prevention & control , Lanosterol/analogs & derivatives , Neuralgia/prevention & control , Receptor, Cannabinoid, CB1/metabolism , Receptor, Cannabinoid, CB2/metabolism , Animals , Anti-Inflammatory Agents, Non-Steroidal/administration & dosage , Anti-Inflammatory Agents, Non-Steroidal/antagonists & inhibitors , Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Behavior, Animal/drug effects , Cytokines/genetics , Cytokines/metabolism , Dose-Response Relationship, Drug , Down-Regulation/drug effects , Edema/immunology , Edema/metabolism , Gene Knockdown Techniques , Hindlimb/drug effects , Hindlimb/metabolism , Hyperalgesia/immunology , Hyperalgesia/metabolism , Lanosterol/administration & dosage , Lanosterol/antagonists & inhibitors , Lanosterol/pharmacology , Lanosterol/therapeutic use , Male , Mice , Neuralgia/immunology , Neuralgia/metabolism , Neutrophil Infiltration/drug effects , Pain Measurement , RNA, Messenger/metabolism , Receptor, Cannabinoid, CB1/antagonists & inhibitors , Receptor, Cannabinoid, CB1/genetics , Receptor, Cannabinoid, CB2/antagonists & inhibitors , Receptor, Cannabinoid, CB2/genetics , Spinal Cord/drug effects , Spinal Cord/metabolism
15.
Biochem Pharmacol ; 83(4): 531-42, 2012 Feb 15.
Article in English | MEDLINE | ID: mdl-22155310

ABSTRACT

Multiple sclerosis (MS) is a severe chronic T cell-mediated autoimmune inflammatory disease of the central nervous system (CNS), the existing therapy of which is only partially effective and is associated with undesirable side effects. Euphol, an alcohol tetracyclic triterpene, has a wide range of pharmacological properties and is considered to have anti-inflammatory action. However there are no reports about the effects and mechanisms of euphol in experimental autoimmune encephalomyelitis (EAE), an established model of MS. Here we report the effects and the underlying mechanisms of action of euphol in EAE. Euphol (1-10mg/kg) was administered orally at different time-points of EAE. Immunological and inflammatory responses were evaluated by real-time PCR, Western blot and flow cytometry assays. We provide evidence that euphol significantly attenuates neurological signs of EAE. These beneficial effects of euphol seem to be associated with the down-regulation of mRNA and protein expression of some pro-inflammatory mediators such as TNF-α, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in the CNS. Furthermore, in vitro, euphol consistently inhibited the T cell-mediated immune response including the production of T(H)1 and T(H)17 cytokines in spleen cells of untreated EAE animals. Likewise, oral euphol treatment inhibited the infiltration of T(H)17 myelin-specific cells into the CNS through the adhesion molecule, lymphocyte function-associated antigen 1 (LFA-1). Our findings reveal that oral administration of euphol consistently reduces and limits the severity and development of EAE. Therefore, euphol might represent a potential molecule of interest for the treatment of MS and other T(H)17 cell-mediated inflammatory diseases.


Subject(s)
Encephalomyelitis, Autoimmune, Experimental/drug therapy , Encephalomyelitis, Autoimmune, Experimental/prevention & control , Lanosterol/analogs & derivatives , Animals , Central Nervous System/drug effects , Central Nervous System/metabolism , Cyclooxygenase 2/genetics , Cyclooxygenase 2/metabolism , Cytokines/genetics , Cytokines/metabolism , Disease Models, Animal , Dose-Response Relationship, Drug , Drinking/drug effects , Drug Administration Schedule , Eating/drug effects , Female , Gene Expression Regulation/drug effects , Lanosterol/chemistry , Lanosterol/therapeutic use , Mice , Mice, Inbred C57BL , Molecular Structure , RNA, Messenger/genetics , RNA, Messenger/metabolism , T-Lymphocytes/drug effects , T-Lymphocytes/metabolism , Time Factors , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/metabolism
16.
PLoS One ; 6(11): e27122, 2011.
Article in English | MEDLINE | ID: mdl-22073270

ABSTRACT

BACKGROUND: The tetracyclic triterpene euphol is the main constituent found in the sap of Euphorbia tirucalli. This plant is widely known in Brazilian traditional medicine for its use in the treatment of several kinds of cancer, including leukaemia, prostate and breast cancers. Here, we investigated the effect of euphol on experimental models of colitis and the underlying mechanisms involved in its action. METHODOLOGY/PRINCIPAL FINDINGS: Colitis was induced in mice either with dextran sulfate sodium (DSS) or with 2,4,6-trinitrobenzene sulfonic acid (TNBS), and the effect of euphol (3, 10 and 30 mg/kg) on colonic injury was assessed. Pro-inflammatory mediators and cytokines were measured by immunohistochemistry, enzyme-Linked immunoabsorbent assay (ELISA), real time-polymerase chain reaction (RT-PCR) and flow cytometry. Preventive and therapeutic oral administration of euphol attenuated both DSS- and TNBS-induced acute colitis as observed by a significant reduction of the disease activity index (DAI), histological/microscopic damage score and myeloperoxidase (MPO) activity in colonic tissue. Likewise, euphol treatment also inhibited colon tissue levels and expression of IL-1ß, CXCL1/KC, MCP-1, MIP-2, TNF-α and IL-6, while reducing NOS2, VEGF and Ki67 expression in colonic tissue. This action seems to be likely associated with inhibition of activation of nuclear factor-κB (NF-κB). In addition, euphol decreased LPS-induced MCP-1, TNF-α, IL-6 and IFN-γ, but increased IL-10 secretion from bone marrow-derived macrophages in vitro. Of note, euphol, at the same schedule of treatment, markedly inhibited both selectin (P- and E-selectin) and integrin (ICAM-1, VCAM-1 and LFA-1) expression in colonic tissue. CONCLUSIONS/SIGNIFICANCE: Together, these results clearly demonstrated that orally-administered euphol, both preventive or therapeutic treatment were effective in reducing the severity of colitis in two models of chemically-induced mouse colitis and suggest this plant-derived compound might be a potential molecule in the management of inflammatory bowel diseases.


Subject(s)
Colitis/drug therapy , Lanosterol/analogs & derivatives , Animals , Colitis/metabolism , Cytokines/metabolism , Enzyme-Linked Immunosorbent Assay , Lanosterol/pharmacology , Lanosterol/therapeutic use , Macrophages/drug effects , Macrophages/metabolism , Mice , NF-kappa B/metabolism , Real-Time Polymerase Chain Reaction
17.
Article in English | MEDLINE | ID: mdl-19815648

ABSTRACT

Trichilia catigua is a native plant of Brazil; its barks are used by some local pharmaceutical companies to prepare tonic drinks, such as Catuama. The present study was addressed to evaluate the effects of T. catigua hydroalcoholic extract in mouse nociception behavioral models, and to evaluate the possible mechanisms involved in its actions. Male Swiss mice were submitted to hot-plate, writhing and von Frey tests, after oral treatment with T. catigua extract (200 mg kg(-1), p.o.). The extract displayed antinociceptive effect in all three models. For characterization of the mechanisms involved in the antinociceptive action of the extract, the following pharmacological treatments were done: naloxone (2.5 mg kg(-1), s.c.), SR141716A (10 mg kg(-1), i.p.), SCH23390 (15 µg kg(-1), i.p.), sulpiride (50 mg kg(-1), i.p.), prazosin (1 mg kg(-1), i.p.), bicuculline (1 mg kg(-1), i.p.) or dl-p-chlorophenylalanine methyl ester (PCPA, 100 mg kg(-1), i.p.). In these experiments, the action of T. catigua extract was evaluated in the hot-plate test. The treatment with SCH23390 completely prevented the antinociceptive effect, while naloxone partially prevented it. The possible involvement of the dopaminergic system in the actions of T. catigua extract was substantiated by data showing the potentiation of apomorphine-induced hypothermia and by the prevention of haloperidol-induced catalepsy. In conclusion, the antinociceptive effects of T. catigua extract seem to be mainly associated with the activation of dopaminergic system and, to a lesser extent, through interaction with opioid pathway.

18.
Eur J Pharmacol ; 569(3): 228-36, 2007 Aug 27.
Article in English | MEDLINE | ID: mdl-17559833

ABSTRACT

This study evaluated the anti-inflammatory properties of two sesquiterpenes isolated from Cordia verbenacea's essential oil, alpha-humulene and (-)-trans-caryophyllene. Our results revealed that oral treatment with both compounds displayed marked inhibitory effects in different inflammatory experimental models in mice and rats. alpha-humulene and (-)-trans-caryophyllene were effective in reducing platelet activating factor-, bradykinin- and ovoalbumin-induced mouse paw oedema, while only alpha-humulene was able to diminish the oedema formation caused by histamine injection. Also, both compounds had important inhibitory effects on the mouse and rat carrageenan-induced paw oedema. Systemic treatment with alpha-humulene largely prevented both tumor necrosis factor-alpha (TNFalpha) and interleukin-1beta (IL-1beta) generation in carrageenan-injected rats, whereas (-)-trans-caryophyllene diminished only TNFalpha release. Furthermore, both compounds reduced the production of prostaglandin E(2) (PGE(2)), as well as inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX-2) expression, induced by the intraplantar injection of carrageenan in rats. The anti-inflammatory effects of alpha-humulene and (-)-trans-caryophyllene were comparable to those observed in dexamethasone-treated animals, used as positive control drug. All these findings indicate that alpha-humulene and (-)-trans-caryophyllene, derived from the essential oil of C. verbenacea, might represent important tools for the management and/or treatment of inflammatory diseases.


Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/pharmacology , Cordia/chemistry , Sesquiterpenes/pharmacology , Administration, Oral , Animals , Anti-Inflammatory Agents/pharmacology , Anti-Inflammatory Agents, Non-Steroidal/isolation & purification , Brazil , Cyclooxygenase 2/drug effects , Cyclooxygenase 2/metabolism , Dexamethasone/pharmacology , Dinoprostone/biosynthesis , Edema/chemically induced , Edema/drug therapy , Interleukin-1beta/drug effects , Interleukin-1beta/metabolism , Male , Medicine, Traditional , Mice , Monocyclic Sesquiterpenes , Nitric Oxide Synthase Type II/drug effects , Nitric Oxide Synthase Type II/metabolism , Oils, Volatile/chemistry , Plant Components, Aerial , Plants, Medicinal , Polycyclic Sesquiterpenes , Rats , Rats, Wistar , Sesquiterpenes/isolation & purification , Tumor Necrosis Factor-alpha/drug effects , Tumor Necrosis Factor-alpha/metabolism
19.
J Ethnopharmacol ; 110(2): 323-33, 2007 Mar 21.
Article in English | MEDLINE | ID: mdl-17084568

ABSTRACT

The anti-inflammatory and anti-allergic effects of the essential oil of Cordia verbenacea (Boraginaceae) and some of its active compounds were evaluated. Systemic treatment with the essential oil of Cordia verbenacea (300-600mg/kg, p.o.) reduced carrageenan-induced rat paw oedema, myeloperoxidase activity and the mouse oedema elicited by carrageenan, bradykinin, substance P, histamine and platelet-activating factor. It also prevented carrageenan-evoked exudation and the neutrophil influx to the rat pleura and the neutrophil migration into carrageenan-stimulated mouse air pouches. Moreover, Cordia verbenacea oil inhibited the oedema caused by Apis mellifera venom or ovalbumin in sensitized rats and ovalbumin-evoked allergic pleurisy. The essential oil significantly decreased TNFalpha, without affecting IL-1beta production, in carrageenan-injected rat paws. Neither the PGE(2) formation after intrapleural injection of carrageenan nor the COX-1 or COX-2 activities in vitro were affected by the essential oil. Of high interest, the paw edema induced by carrageenan in mice was markedly inhibited by both sesquiterpenic compounds obtained from the essential oil: alpha-humulene and trans-caryophyllene (50mg/kg, p.o.). Collectively, the present results showed marked anti-inflammatory effects for the essential oil of Cordia verbenacea and some active compounds, probably by interfering with TNFalpha production. Cordia verbenacea essential oil or its constituents might represent new therapeutic options for the treatment of inflammatory diseases.


Subject(s)
Anti-Allergic Agents/pharmacology , Anti-Inflammatory Agents/pharmacology , Cordia/chemistry , Oils, Volatile/pharmacology , Plant Extracts/pharmacology , Animals , Carrageenan , Cyclooxygenase 1/drug effects , Cyclooxygenase 2/drug effects , Dinoprostone , Edema/chemically induced , Edema/drug therapy , Interleukin-1beta/drug effects , Mice , Monocyclic Sesquiterpenes , Neutrophils/drug effects , Peroxidase/drug effects , Peroxidase/metabolism , Phytotherapy , Plant Leaves , Plants, Medicinal , Polycyclic Sesquiterpenes , Rats , Rats, Wistar , Sesquiterpenes/pharmacology , Tumor Necrosis Factor-alpha/drug effects
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