Half-maximal Effective Dose Research Articles

The electrophysiological effects of Tongyang Huoxue granules on the ignition phase during hypoxia/reoxygenation injury in sinoatrial node cells.

This study was undertaken to explore the potential therapeutic effects of Tongyang Huoxue Granules (TYHX) on sinoatrial node (SAN) dysfunction, a cardiac disorder characterized by impaired impulse generation or conduction. The research question addressed whether TYHX could positively influence SAN ion channel function, specifically targeting the sodium-calcium exchanger (I NCX) and L-type calcium channel (I CaL) of the SAN. Sinoatrial node cells (SANCs) were isolated and cultured from neonatal Japanese big-eared white rabbits within 24h of birth. The study encompassed five groups: Control, H/R (hypoxia/reoxygenation), H/R+100μg/mL TYHX, H/R+200μg/mL TYHX, and H/R+400μg/mL TYHX. The H/R model, simulating hypoxia/reoxygenation stress, was induced within 5days of culture. Whole-cell patch clamp technique was employed to record currents following a 3-min perfusion and stabilization period with TYHX. TYHX administration demonstrated improvements in the ignition phase of impaired SANCs. The half-maximal effective dose of TYHX, as determined by SANC beating frequency, was found to be 323.63μg/mL. Inward current density of I NCX increased in response to TYHX (200 and 400μg/mL), while TYHX enhanced I CaL current density in H/R SANCs, with 400μg/mL exhibiting greater efficacy. Additionally, TYHX regulated the gating mechanisms of I CaL by right-shifting the steady-state inactivation curve and accelerating recovery from inactivation. Notably, TYHX increased the activation time constant under 200 and 400μg/mL, prolonged the fast inactivation time constant τ1 with 400μg/mL, and extended the slow inactivation time constant τ2 with 100 and 400μg/mL. The findings suggest that TYHX may hold promise as a therapeutic intervention for sinus node dysfunction, offering potential avenues for drug development aimed at safeguarding SAN function.

Analysis of the Stimulative Effect of Tryptophan on Hepatic Protein Synthesis in Rats.

Tryptophan is an essential amino acid important as a protein building block, but it also serves as substrate for the generation of several bioactive compounds with important physiological roles. Furthermore, tryptophan has been reported to have a unique role as a nutritional signaling molecule that regulates protein synthesis in mouse and rat liver. In the present study, the acute effects of tryptophan on protein synthesis were confirmed and compared with those of leucine in rats. Eighteen hours fasted rats were orally administered of tryptophan or leucine at a dose of 135 mg/100 g body weight by gavage and then sacrificed 1 h after administration. The effects of tryptophan and leucine on the rate of protein synthesis were evaluated by the surface sensing of translation (SUnSET) method. We also examined the ability of tryptophan to induce activation of the mTOR pathway by measuring phosphorylation of 4E-BP1 and S6K1. Oral administration of tryptophan led to a stimulation of the rate of protein synthesis concomitant with activation of mTOR pathway in the liver, but not in skeletal muscle. We also investigated the sensitivity of liver protein synthesis to tryptophan administration. The half-maximal effective doses (ED50) of tryptophan in stimulating 4E-BP1 and S6K1 phosphorylation were both about 60% of daily intake. The effect of tryptophan on hepatic protein synthesis was similar to that of leucine on muscle protein synthesis, and the sensitivity of liver protein synthesis to tryptophan administration appeared to be almost the same or slightly lower than that of muscle protein synthesis to leucine administration.

Effect of gender factor on the effective dose of oxycodone for inhibiting responses to laryngeal mask airway insertion in elderly patients undergoing ophthalmic surgery

Objective: To evaluate the effect of gender factor on the effective dose of oxycodone for inhibiting responses to laryngeal mask airway (LMA) insertion in elderly patients undergoing ophthalmic surgery. Methods: From June to October 2021, 56 elderly patients, including 26 females and 30 males, aged from 65 to 80 (72±5) years, with a body mass index (BMI) of 18.5-24.9 kg/m2 and American Society of Anesthesiologists (ASA) physical status of Ⅰ or Ⅱ, who underwent elective ophthalmic surgery requiring LMA insertion in the Beijing Tongren Hospital affiliated to Capital Medical University were selected. Patients were divided into two groups according to gender: elderly female group and elderly male group. The modified Dixon sequential method was used. Oxycodone 0.10 mg/kg was injected intravenously in the first patient. Etomidate 0.2 mg/kg and cisatracurium 0.1 mg/kg were administrated 5 min later. After 3 min, the laryngeal mask was placed when the bispectral index (BIS) ≤60. The positive response to LMA insertion was defined as an increase in the maximum mean arterial pressure or the maximum heart rate>20% of the baseline value within 2 min after insertion. When the response to LMA insertion was positive, the dose of oxycodone was increased in the next patient, otherwise the dose was decreased, and the ratio of adjacent dose was 1.1. This process was repeated until the 7th turning point occurred. The half-maximal effective doses (ED50) and 95% confidence intervals (CI) of oxycodone for inhibiting laryngeal mask insertion were calculated and compared between the two groups. Meanwhile, the adverse reactions during anesthesia induction were observed. Results: All 26 patients in the elderly female group completed the test, but one patient in the elderly male group withdrew due to poor alignment of laryngeal mask, and finally 29 patients completed the study. There were 13 cases and 14 cases who had positive response to LMA insertion in the elderly female and male groups, respectively. The ED50 (95%CI) of oxycodone for inhibiting laryngeal mask insertion in the elderly male group was 0.096 (0.083-0.112) mg/kg, which was higher than that in the elderly female group [0.081 (0.073-0.098) mg/kg, P=0.008]. No adverse reactions such as choking cough, muscle tremor, hypoxemia, nausea, vomiting, reflux and aspiration occurred in 55 patients, except that one patient in the elderly female group had transient hypotension after induction, which improved after symptomatic treatment. Conclusion: The ED50 of oxycodone for inhibiting laryngeal mask insertion reaction in ophthalmic surgery is different in patients with different genders, which is 0.096 (0.083-0.112) mg/kg in the elderly male group and 0.081 (0.073-0.098) mg/kg in the elderly female group.

Effects of Mitomycin-C and 5-Fluorouracil on Ocular Adnexal Sebaceous Carcinoma Cells

To investigate the effects of mitomycin-C (MMC) and 5-fluorouracil (5-FU) on the viability, proliferation, and migratory capacity of cultured ocular adnexal sebaceous carcinoma (SC) cells. Laboratory investigation. Human SC cell lines (Bascom Palmer 50 and 52 [BP50 and BP52]) and human limbal stem cells (LSCs) were treated with various concentrations of MMC and 5-FU. Cytotoxicity was assessed with the tetrazolium MTT colorimetric viability assay on normal corneal vs tumor cells. Growth curves and scratch assays were performed to characterize the effects of these chemotherapeutic agents on SC proliferation and migration, respectively. MMC decreased BP52 cell viability in a dose-dependent manner with a half-maximal effective dose (EC50) of 11.8 μM after 72 hours. SC viability decreased >50% at 80 mM 5-FU after 72 hours. MMC reduced LSC viability in a dose-dependent manner with an EC50 value of 3.24 μM, and 5-FU decreased LSC viability >50% at 160 μM. MMC decreased SC cell proliferation and migration in a dose-dependent manner. 5-FU displayed antiproliferative effects but did not affect cell migration at concentrations below 1000 μM. Our in vitro data corroborate clinical observations that MMC is efficacious for treating ocular adnexal SC, albeit at the expense of LSC viability. Our findings also demonstrate that topical 5-FU exhibits antiproliferative effects that supersede its cancer-killing and antimigratory effects on cultured SC cells.

The First-in-Class UBA1 Inhibitor, TAK-243, in Combination With Radiotherapy for YAP1 and BEND3 Biomarker-Defined Small Cell Lung Cancer

The First-in-Class UBA1 Inhibitor, TAK-243, in Combination With Radiotherapy for YAP1 and BEND3 Biomarker-Defined Small Cell Lung Cancer

MP40-09 MOLECULAR ASPECTS OF RESISTANCE TO CISPLATIN AND OSIMERTINIB IN PENILE CANCER

You have accessJournal of UrologyPenile & Testicular Cancer I (MP40)1 Sep 2021MP40-09 MOLECULAR ASPECTS OF RESISTANCE TO CISPLATIN AND OSIMERTINIB IN PENILE CANCER Anita Thomas, Olesya Vakhrusheva, Martin MIchaelis, Jindrich Cinatl, Florian Rothweiler, Maarten Albersen, Axel Haferkamp, Eva Jüngel, and Igor Tsaur Anita ThomasAnita Thomas More articles by this author , Olesya VakhrushevaOlesya Vakhrusheva More articles by this author , Martin MIchaelisMartin MIchaelis More articles by this author , Jindrich CinatlJindrich Cinatl More articles by this author , Florian RothweilerFlorian Rothweiler More articles by this author , Maarten AlbersenMaarten Albersen More articles by this author , Axel HaferkampAxel Haferkamp More articles by this author , Eva JüngelEva Jüngel More articles by this author , and Igor TsaurIgor Tsaur More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000002055.09AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Metastatic penile cancer (PeCa) is hallmarked by rapidly evolving resistance to chemotherapy worsening prognosis. Since PeCa is a rare disease, translational research aimed at identifying novel options of counteracting resistance is limited by a scarcity of biomaterials, tumor models as well as funding. In the current project, PeCa cell lines resistant to cisplatin and osimertinib were successfully established. Moreover, molecular mechanisms of resistance as well as efficacy of drug treatment were analyzed. METHODS: Therapy-naïve UKF-PeC3 cells were adapted to growth in the presence of 2 μg/ml cisplatin (UKF-PeC3rCIS2) or 2 μM osimertinib (UKF-PeC3rOSI2). Tumor cell growth, proliferation, apoptosis, cell cycle phases and regulating proteins as well as akt/mTOR signaling pathway proteins were investigated in therapy-naïve and cisplatin- and in osimertinib resistant cell lines by the MTT dye reduction assay, BrdU assay, flow cytometry and western blot. Finally, effects of the drug treatment with the mTOR-inhibitor everolimus on PeCa cancer growth were determined in both cell lines. RESULTS: The IC50 (half-maximal effective dose) value for cisplatin (5.48 μg/ml vs. 61.76 μg/ml) and osimertinib (5.04 μM vs. 17.02 μM) was increased significantly in resistant cells compared to treatment-naïve cells confirming acquired resistance in UKF-PeC3rCIS2 and UKF-PeC3rOSI2. Accordingly, both resistant cell lines showed a significantly weaker response in viability and proliferation assays. Flow cytometry detected a modulation of cell cycle phases in the state of resistance. Furthermore, protein expression profiles of selected proteins of the akt/mTOR signaling pathway demonstrated an upregulation of pAKT, p4EBP1 and pmTOR in both resistant cell lines compared to therapy-naïve cells. Therapeutic potential of this pathway was further corroborated by efficacy testing of everolimus. CONCLUSIONS: Resistance patterns and mechanisms were identified, emphasizing the involvement of the AKT/mTOR signaling pathway and paving the way for development of novel treatment strategies in PeCa. Targeting this pathway might prolong efficacy of therapeutic regimens or induce a re-sensitization of treatment-resistant PeCa cells. Source of Funding: Brigitte- und Dr. Konstanze Wegener-Stiftung © 2021 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 206Issue Supplement 3September 2021Page: e717-e718 Advertisement Copyright & Permissions© 2021 by American Urological Association Education and Research, Inc.MetricsAuthor Information Anita Thomas More articles by this author Olesya Vakhrusheva More articles by this author Martin MIchaelis More articles by this author Jindrich Cinatl More articles by this author Florian Rothweiler More articles by this author Maarten Albersen More articles by this author Axel Haferkamp More articles by this author Eva Jüngel More articles by this author Igor Tsaur More articles by this author Expand All Advertisement Loading ...

A network meta-analysis of the dose\u2013response effects of lurasidone on acute schizophrenia

We compared the efficacy, safety, and acceptability of lurasidone at different doses to establish the dose–response relationships of lurasidone therapeutic and adverse effects in acute schizophrenia. Included trials were 4- to 16-week, fixed-dose, randomized controlled trials of lurasidone in adults with acute schizophrenia. Different doses of lurasidone, other antipsychotics, and placebo were considered as independent treatments. Apart from all-cause dropout rates, four therapeutic and four adverse outcomes were included in the frequentist network meta-analysis (NMA). Lurasidone 160, 120, 80, 40, and 20 mg/day were studied in ten trials of 3,366 adults with schizophrenia exacerbation. Lurasidone 160 mg/day reduced Positive and Negative Syndrome Scale (PANSS) total scores significantly more than lurasidone 120, 80, 40, and 20 mg/day (mean differences = − 7.63, − 7.04, − 8.83, and − 12.25, respectively). All-cause dropout rates were significantly lower in participants receiving lurasidone 160 mg/day and 80 mg/day compared with those taking placebo. The half-maximal effective doses of lurasidone for PANSS total, PANSS positive, and MADRS score reductions were higher than 80 mg/day. The confidence of all NMA estimates was low or very low. Lurasidone 160 mg/day is currently the most efficacious and acceptable dose for acute schizophrenia. Its maximal effective doses may be higher than 160 mg/day.

Consciousness, Anesthesia, and Acetylcholine.

Consciousness, Anesthesia, and Acetylcholine.

TLR9 expression in chronic lymphocytic leukemia identifies a promigratory subpopulation and novel therapeutic target

Chronic lymphocytic leukemia (CLL) remains incurable despite B-cell receptor–targeted inhibitors revolutionizing treatment. This suggests that other signaling molecules are involved in disease escape mechanisms and resistance. Toll-like receptor 9 (TLR9) is a promising candidate that is activated by unmethylated cytosine guanine dinucleotide–DNA. Here, we show that plasma from patients with CLL contains significantly more unmethylated DNA than plasma from healthy control subjects (P < .0001) and that cell-free DNA levels correlate with the prognostic markers CD38, β2-microglobulin, and lymphocyte doubling time. Furthermore, elevated cell-free DNA was associated with shorter time to first treatment (hazard ratio, 4.0; P = .003). We also show that TLR9 expression was associated with in vitro CLL cell migration (P < .001), and intracellular endosomal TLR9 strongly correlated with aberrant surface expression (sTLR9; r = 0.9). In addition, lymph node–derived CLL cells exhibited increased sTLR9 (P = .016), and RNA-sequencing of paired sTLR9hi and sTLR9lo CLL cells revealed differential transcription of genes involved in TLR signaling, adhesion, motility, and inflammation in sTLR9hi cells. Mechanistically, a TLR9 agonist, ODN2006, promoted CLL cell migration (P < .001) that was mediated by p65 NF-κB and STAT3 transcription factor activation. Importantly, autologous plasma induced the same effects, which were reversed by a TLR9 antagonist. Furthermore, high TLR9 expression promoted engraftment and rapid disease progression in a NOD/Shi-scid/IL-2Rγnull mouse xenograft model. Finally, we showed that dual targeting of TLR9 and Bruton's tyrosine kinase (BTK) was strongly synergistic (median combination index, 0.2 at half maximal effective dose), which highlights the distinct role for TLR9 signaling in CLL and the potential for combined targeting of TLR9 and BTK as a more effective treatment strategy in this incurable disease.

Anti-inflammatory and antioxidant activities of the root and bark extracts of Vitex grandifolia (Verbanaceae)

Medicinal plants form the basis for many medical treatments and a precursor for most drugs. This study examined the anti-inflammatory and antioxidant properties of the root and bark extracts of Vitex grandifolia. Soxhlet extraction was used to obtain the crude extracts. The 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging, hydrogen peroxide and phosphomolybdenum (PM) assays were used to evaluate antioxidant properties. Total phenolic content determination was by the Folin Ciocalteu method. The carrageenan-induced paw edema model in day-old chicks was used to evaluate anti-inflammatory activity. The root and bark ethanolic extracts displayed similar antioxidant activities in the DPPH and PM assays. The bark extract was a better anti-oxidant than the root extract in the hydrogen peroxide assay with an inhibition concentration of 218.90 µg/mL. Both extracts showed similar levels of phenolic compounds, with values of 5.72 ± 1.96 and 5.09 ± 2.25 g/100g gallic acid equivalent (GAE) for root and bark extracts respectively. For anti-inflammatory action, the bark extract, with a half maximal effective dose (ED50) of 34.48 ± 6.00 mg/kg, proved superior to root extract (ED50 of 76.90 ± 7.76 mg/kg). The bark extract which gave the best anti-inflammatory activity was fractionated using butanol, ethyl acetate and hexane to identify the fraction in which the anti-inflammatory principles were concentrated. The ED50 for the fractions were in the order ethyl acetate < hexane < butanol. The results obtained showed that both root and bark extracts were potent anti-inflammatory and anti-oxidant agents and validates their use in folkloric medicine for treating inflammation-related diseases.

Pharmacokinetic and Pharmacodynamic Properties of Cemdisiran, an RNAi Therapeutic Targeting Complement Component 5, in Healthy Subjects and Patients with Paroxysmal Nocturnal Hemoglobinuria

BackgroundCemdisiran, an N-acetylgalactosamine (GalNAc) conjugated RNA interference (RNAi) therapeutic, is currently under development for the treatment of complement-mediated diseases by suppressing liver production of complement 5 (C5) protein. This study was designed to evaluate the safety, tolerability, pharmacokinetics (PK), and pharmacodynamics (PD) of cemdisiran in healthy subjects and in patients with paroxysmal nocturnal hemoglobinuria (PNH) in order to support dose selection for late-stage clinical trials.MethodsHealthy volunteers (HVs; n = 32, including 12 Japanese subjects) were randomized (3:1) to receive single doses of subcutaneous cemdisiran (50–900 mg) or placebo, or repeat doses of subcutaneous cemdisiran (100–600 mg) or placebo weekly, biweekly, weekly/biweekly, or weekly/monthly for 5, 8, or 13 weeks (n = 24). Cemdisiran 200 or 400 mg was administered weekly in an open-label manner, for varying durations, as monotherapy in three eculizumab-naïve PNH patients or in combination with eculizumab in three PNH patients who were receiving stable label doses of eculizumab (900 or 1200 mg biweekly) before the start of the study. After the last dose of cemdisiran, patients were followed for safety and ongoing pharmacologic effects with the eculizumab regimen (600 or 900 mg every month).ResultsIn HVs, cemdisiran was rapidly converted to a major active metabolite, AS(N-2)3′-cemdisiran, both declining below the lower limit of quantification (LLOQ) in plasma within 48 h, and showing minimal renal excretion. AS(N-2)3′-cemdisiran exhibited more than dose-proportional PK. The C5 protein reductions were dose-dependent, with > 90% reduction of C5 protein beginning on days 21–28 and maintained for 10–13 months following single and biweekly doses of 600 mg. The dose–response relationship, described by an inhibitory sigmoid maximum effect (Emax) model, estimated half-maximal effective dose (ED50) of 14.0 mg and maximum C5 reduction of 99% at 600 mg. The PK and PD were similar between Japanese and non-Japanese subjects, and PNH patients and HVs. One of 48 subjects tested transiently positive for antidrug antibody with low titer, with no impact on PK or PD. In PNH patients, C5 suppression by cemdisiran enabled effective inhibition of residual C5 levels with lower dose and/or dosing frequency of eculizumab, which was maintained for 6–10 months after the last dose of cemdisiran.ConclusionsConsistent with the PK/PD properties of liver targeting GalNac conjugates, cemdisiran and AS(N-2)3′-cemdisiran plasma concentrations declined rapidly while showing rapid and robust C5 suppression maintained up to 13 months following single and multiple doses, which indicates long residence times of cemdisiran within hepatocytes. The long PD duration of action in liver, low immunogenicity and acceptable safety profiles enables low, infrequent SC dosing and support further evaluation of cemdisiran in complement-mediated diseases as monotherapy or in combination with a C5 inhibitor antibody.Clinical Trial Registration NoNCT02352493.Electronic supplementary materialThe online version of this article (10.1007/s40262-020-00940-9) contains supplementary material, which is available to authorized users.

Glycolaldehyde-modified proteins cause adverse functional and structural aortic remodeling leading to cardiac pressure overload

Growing evidence supports the role of advanced glycation end products (AGEs) in the development of diabetic vascular complications and cardiovascular diseases (CVDs). We have shown that high-molecular-weight AGEs (HMW-AGEs), present in our Western diet, impair cardiac function. Whether HMW-AGEs affect vascular function remains unknown. In this study, we aimed to investigate the impact of chronic HMW-AGEs exposure on vascular function and structure. Adult male Sprague Dawley rats were daily injected with HMW-AGEs or control solution for 6 weeks. HMW-AGEs animals showed intracardiac pressure overload, characterized by increased systolic and mean pressures. The contraction response to PE was increased in aortic rings from the HMW-AGEs group. Relaxation in response to ACh, but not SNP, was impaired by HMW-AGEs. This was associated with reduced plasma cyclic GMP levels. SOD restored ACh-induced relaxation of HMW-AGEs animals to control levels, accompanied by a reduced half-maximal effective dose (EC50). Finally, collagen deposition and intima-media thickness of the aortic vessel wall were increased with HMW-AGEs. Our data demonstrate that chronic HMW-AGEs exposure causes adverse vascular remodelling. This is characterised by disturbed vasomotor function due to increased oxidative stress and structural changes in the aorta, suggesting an important contribution of HMW-AGEs in the development of CVDs.

Inhibition of Opisthorchis felineus glutathione-dependent prostaglandin synthase by resveratrol correlates with attenuation of cholangiocyte neoplasia in a hamster model of opisthorchiasis

Food-borne trematodiases represent major neglected parasitic diseases. Trematodes of the family Opisthorchiidae including Opisthorchis felineus, Opisthorchis viverrini and Clonorchis sinensis are ranked eight on the global list of the 24 most prevalent food-borne parasites. Chronic O. felineus infection symptoms include precancerous lesions with the potential for malignancy. In recent decades, liver flukes of the family Opisthorchiidae have been extensively scientifically explored, however despite this the molecular mechanisms of O. felineus pathogenicity and its carcinogenic potential have not been studied. Opisthorchis felineus glutathione-dependent prostaglandin synthase (GST σ) is the major component of the excretory–secretory product of this liver fluke. We hypothesised that the activity of this enzyme is involved in the infection pathogenesis, including the formation of precancerous lesions. To test this hypothesis and to gain insights into the mechanisms of precancerous lesion formation, we (i) investigated whether excretory parasitic GST σ retains its enzymatic activity, (ii) tested resveratrol (RSV) as a possible inhibitor of this enzyme, and (iii) assessed biliary neoplasia and oxidative DNA damage as well as the expression of neoplasia and fibrogenesis marker genes after prolonged administration of RSV in a hamster model. RSV was found to inhibit GST σ enzymatic activity in a dose-dependent manner (R = 0.85, P < 0.001; half-maximal effective dose (ED50) = 48.6 μM). Prolonged administration of RSV significantly suppressed high-grade biliary neoplasia (P = 0.008), attenuated upregulation of hyperplasia and fibrogenesis-related genes (Tgfb, α-SMA and CK7), and decreased the elevated oxidative DNA damage. Taking into account that RSV can influence a wide range of pathways, further research is needed to confirm the role of GST σ in O. felineus pathogenicity. Nevertheless, the chemopreventive effect of RSV targeting biliary neoplasia formation might be useful for improving the outcomes in infected populations and represents a compelling rationale for RSV testing in combination chemotherapy of opisthorchiasis.

Inhibition of experimental visceral pain in rodents by cebranopadol.

The aim of this study was to investigate the efficacy of cebranopadol in two rodent models of visceral pain. Cebranopadol is a first-in-class analgesic with agonist activity at the nociceptin/orphanin FQ opioid peptide receptor and classical µ-, δ- and κ-opioid peptide receptors. Colitis was induced in Naval Medical Research Institute mice by intra-rectal infusion of mustard oil. The effects of intravenous cebranopadol pretreatment on spontaneous pain behaviours and referred allodynia and hyperalgesia were assessed. Pancreatitis was induced in Sprague-Dawley rats by intravenous administration of dibutyltin dichloride. After 6 days, the effects of intravenous cebranopadol on withdrawal reactions to mechanical abdominal stimulation with von Frey filaments were assessed. In mice with experimental colitis, cebranopadol dose-dependently inhibited spontaneous pain behaviours and allodynic and hyperalgesic withdrawal reactions, with half-maximal effective dose values of 4.6 µg/kg [95% confidence interval (CI): 2.9-7.9] for inhibition of spontaneous pain behaviours, 2.2 µg/kg (95% CI: 1.3-3.4) for inhibition of referred allodynia and 2.4 µg/kg (95% CI: 1.4-3.6) for inhibition of referred hyperalgesia in mice with colitis. In rats with experimental pancreatitis, cebranopadol dose-dependently inhibited abdominal tactile allodynia (half-maximal effective dose, 0.13 µg/kg; 95% CI: 0.03-0.49). Behavioural manifestations of visceral pain were almost completely abolished at the highest doses tested in mice (17.2 µg/kg, intravenous) and rats (2.4 µg/kg, intravenous). We conclude that cebranopadol is a potent and effective antiallodynic and antihyperalgesic agent in rodent models of visceral pain.

Investigational RNAi Therapeutic Targeting C5 Is Efficacious in Pre-clinical Models of Myasthenia Gravis

Complement-mediated damage to the neuromuscular junction (NMJ) is a key mechanism of pathology in myasthenia gravis (MG), and therapeutics inhibiting complement have shown evidence of efficacy in the treatment of MG. In this study, we describe the development of a subcutaneously administered N-acetylgalactosamine (GalNAc)-conjugated small interfering RNA (siRNA) targeting the C5 component of complement that silences C5 expression in the liver (ALN-CC5). Treatment of wild-type rodents with ALN-CC5 resulted in robust and durable suppression of liver C5 expression. Dose-dependent serum C5 suppression was observed in non-human primates, with a lowering of serum C5 of up to 97.5% and the concomitant inhibition of serum complement activity. C5 silencing was efficacious in ameliorating disease symptoms in two standard rat models of MG, demonstrating the key role of circulating C5 in pathology at the NMJ. Improvement in disease activity scores and NMJ pathology was observed at intermediate levels of complement activity inhibition, suggesting that complete ablation of complement activity may not be required for efficacy in MG. The pre-clinical studies of ALN-CC5 and efficacy of C5 silencing in rat models of MG support further clinical development of ALN-CC5 as a potential therapeutic for the treatment of MG and other complement-mediated disorders.

Determining RBE for development of lung fibrosis induced by fractionated irradiation with carbon ions utilizing fibrosis index and high-LET BED model

Background and purposesCarbon ion radiotherapy (CIRT) with raster scanning technology is a promising treatment for lung cancer and thoracic malignancies. Determining normal tissue tolerance of organs at risk is of utmost importance for the success of CIRT. Here we report the relative biological effectiveness (RBE) of CIRT as a function of dose and fractionation for development of pulmonary fibrosis using well established fibrosis index (FI) model. Materials and MethodsDose series of fractionated clinical quality CIRT versus conventional photon irradiation to the whole thorax were compared in C57BL6 mice. Quantitative assessment of pulmonary fibrosis was performed by applying the FI to computed tomography (CT) data acquired 24-weeks post irradiation. RBE was calculated as the ratio of photon to CIRT dose required for the same level of FI. Further RBE predictions were performed using the derived equation from high-linear energy transfer biologically effective dose (high-LET BED) model. ResultsThe averaged lung fibrosis RBE of 5-fraction CIRT schedule was determined as 2.75 ± 0.55. The RBE estimate at the half maximum effective dose (RBEED50) was estimated at 2.82 for clinically relevant fractional sizes of 1–6 Gy. At the same dose range, an RBE value of 2.81 ± 0.40 was predicted by the high-LET BED model. The converted biologically effective dose (BED) of CIRT for induction of half maximum FI (BEDED50) was identified to be 58.12 Gy3.95. In accordance, an estimated RBE of 2.88 was obtained at the BEDED50 level. The LQ model radiosensitivity parameters for 5-fraction was obtained as αH = 0.3030 ± 0.0037 Gy−1 and βH = 0.0056 ± 0.0007 Gy−2. ConclusionThis is the first report of RBE estimation for CIRT with the endpoint of pulmonary fibrosis in-vivo. We proposed in present study a novel way to mathematically modeling RBE by integrating RBEmax and α/βL based on conventional high-LET BED conception. This model well predicted RBE in the clinically relevant dose range but is sensitive to the uncertainties of α/β estimates from the reference photon irradiation (α/βL). These findings will assist to eliminate current uncertainties in prediction of CIRT induced normal tissue complications and builds a solid foundation for development of more accurate in-vivo data driven RBE estimates.

Single, very low rituximab doses in healthy volunteers - a pilot and a randomized trial: implications for dosing and biosimilarity testing

There are no dose-finding trials available for rituximab that could guide dosing in non-malignant diseases. We hypothesized that currently used doses (≥375 mg/m2) exceed several hundred-fold the half-maximal effective dose, which is most sensitive for detecting putative differences between biosimilars and important for dose finding. In an open label, exploratory trial healthy volunteers received single infusions of rituximab at doses of 0.1, 0.3 or 1.0 mg/m2. Subsequently, in a double-blind, randomized trial healthy volunteers received single infusions of two rituximab products at doses of 0.1 and 0.3 mg/m2. In the exploratory trial rituximab transiently depleted CD20+ cells by a mean 68% (range: 57–95%), 74% (55–82%) and 97% (94–100%) immediately after the infusion of 0.1 (n = 4), 0.3 (n = 4) and 1 mg/m2 (n = 8), respectively. In the randomized trial CD20+ cells decreased by a mean 48% (25–84%) − 55% (26–85%) and 81 (67–89%) – 87% (77–96%) after infusion of 0.1 mg/m2 (n = 12) or 0.3 mg/m2 (n = 8 proposed biosimilar, n = 4 reference product) of the proposed biosimilar or the reference product, respectively. It is important to understand that in healthy volunteers <1% of the authorized rituximab doses depletes almost all circulating B lymphocytes. Thus, for non-malignant diseases alternative, more cost-effective dosing regimens seem plausible, but require clinical testing. (EudraCT-No. 2010–023781–45; EudraCT-No. 2013–001077–24).

Gastroprotective activity of the methanol extract from peels of Plinia edulis (Vell.) Sobral fruits and its isolated triterpenes: maslinic and ursolic acids.

According to the Brazilian folk medicine, the leaves of Plinia edulis (Vell.) Sobral (Myrtaceae), known as cambuca, are indicated in the treatment of gastric disorders. Infusions of P. edulis leaves were previously demonstrated to contain both maslinic (MA) and ursolic acids (UA). Both triterpenes have also been identified in the methanolic extract of peels from P. edulis fruit (MEPPE); however, the antiulcer effects of MEPPE have not yet been studied. This study therefore evaluates the gastroprotective potential of MEPPE, MA, and UA using ethanol/HCl- and indomethacin-induced gastric ulcers in mice. In addition, the in vitro effects of these compounds on the H+, K+-ATPase activity and on the free radical DPPH were measured. When used at concentration of 100μg/mL, both MEPPE and UA were found to reduce the DPPH radical levels by 78.66 and 60.14%, respectively. However, MA did not reduce DPPH radical levels. Our results illustrated the antiulcer effects of MEPPE, MA, and UA against experimental ulcer models when administered by either the oral or the intraperitoneal routes. In addition, MEPPE reduced the size of ethanol/HCl-induced ulcers in a dose-dependent manner (log half-maximal effective oral dose, LogED50 = 1.09). Interestingly, UA promoted gastroprotection at lower doses than MA by increasing the production of mucin levels at 692%; however, it does not alter the activity of H+, K+-ATPase. In contrast, both MEPPE and MA, when incubated at concentrations of 10 and 100μg/mL, inhibited H+, K+-ATPase activity in 61.81, 68.37, 54.04, and 70.45%, respectively. These results confirm that MEPPE, MA, and UA display gastroprotective activity through different modes of action; MA inhibits H+, K+-ATPase activity, whereas UA favour the mucus barrier.

A Novel High-Mannose Specific Lectin from the Green Alga Halimeda renschii Exhibits a Potent Anti-Influenza Virus Activity through High-Affinity Binding to the Viral Hemagglutinin.

We have isolated a novel lectin, named HRL40 from the green alga Halimeda renschii. In hemagglutination-inhibition test and oligosaccharide-binding experiment with 29 pyridylaminated oligosaccharides, HRL40 exhibited a strict binding specificity for high-mannose N-glycans having an exposed (α1-3) mannose residue in the D2 arm of branched mannosides, and did not have an affinity for monosaccharides and other oligosaccharides examined, including complex N-glycans, an N-glycan core pentasaccharide, and oligosaccharides from glycolipids. The carbohydrate binding profile of HRL40 resembled those of Type I high-mannose specific antiviral algal lectins, or the Oscillatoria agardhii agglutinin (OAA) family, which were previously isolated from red algae and a blue-green alga (cyanobacterium). HRL40 potently inhibited the infection of influenza virus (A/H3N2/Udorn/72) into NCI-H292 cells with half-maximal effective dose (ED50) of 2.45 nM through high-affinity binding to a viral envelope hemagglutinin (KD, 3.69 × 10−11 M). HRL40 consisted of two isolectins (HRL40-1 and HRL40-2), which could be separated by reverse-phase HPLC. Both isolectins had the same molecular weight of 46,564 Da and were a disulfide -linked tetrameric protein of a 11,641 Da polypeptide containing at least 13 half-cystines. Thus, HRL40, which is the first Type I high-mannose specific antiviral lectin from the green alga, had the same carbohydrate binding specificity as the OAA family, but a molecular structure distinct from the family.

The Pharmacodynamic Effects of Rituximab at Very Low Doses

No dose-finding trials are available for rituximab that could guide dosing in non-malignant diseases. We hypothesized that currently used doses (≥375mg/m2) exceed several hundred-fold the half-maximal effective dose, which is most sensitive for detecting putative differences between biosimilars and important for dose-finding. In an exploratory, dose-finding, trial healthy volunteers received single infusions of 0.1 (n=4), 0.3 (n=4) and 1 mg/m2 (n=8) rituximab. Subsequently, in a randomized, double-blind trial, healthy volunteers received single infusions of 0.1 (n=24) or 0.3 mg/m2 (n=12) of two rituximab products. CD19/20+ cell counts were measured, pharmacokinetics and, immunogenicity were assessed. Single infusions of 0.1, 0.3 and 1 mg/m2 rituximab transiently depleted CD20+ cells by a mean 68% (95%CI: 24-100%), 74% (95%CI: 59-84%) and 97% (95%CI: 95-99%), respectively. In the randomized trial infusion of 0.1mg/m2 or 0.3mg/m2 proposed biosimilar or reference rituximab decreased CD20+cells by 45% (95%CI: 32-58%) - 55% (95%CI: 45-66%) and 81 (95%CI: 73-87%) – 87% (95%CI: 74-100%), respectively. In the randomized trial, 26 of 36 patients developed human anti-chimeric antibodies and 9 of 36 patients developed neutralizing anti-drug antibodies. Pharmacokinetic analyses were limited by the assay sensitivity and the very low rituximab doses. However, there was a clear pharmacokinetic signal during the first 24 hours in the 1mg/m2 group. It is important to understand that <1% of the authorized rituximab doses depletes all circulating B lymphocytes in healthy volunteers. This will help particularly countries struggling to meet the financial burden of therapy with biologics.