Over‐activation of microglia cells in the brain contributes to neurodegenerative processes promoted by the production of various neurotoxic factors including pro‐inflammatory cytokines and nitric ...oxide. Recently, accumulating evidence has suggested that mitochondrial dynamics are an important constituent of cellular quality control and function. However, the role of mitochondrial dynamics in microglial activation is still largely unknown. In this study, we determined whether mitochondrial dynamics are associated with the production of pro‐inflammatory mediators in lipopolysaccharide (LPS)‐stimulated immortalization of murine microglial cells (BV‐2) by a v‐raf/v‐myc carrying retrovirus (J2). Excessive mitochondrial fission was observed in lentivirus‐transfected BV‐2 cells stably expressing DsRed2‐mito following LPS stimulation. Furthermore, mitochondrial localization of dynamin‐related protein 1 (Drp1) (a key regulator of mitochondrial fission) was increased and accompanied by de‐phosphorylation of Ser637 in Drp1. Interestingly, inhibition of LPS‐induced mitochondrial fission and reactive oxygen species (ROS) generation by Mdivi‐1 and Drp1 knock‐down attenuated the production of pro‐inflammatory mediators via reduced nuclear factor kappa‐light‐chain‐enhancer of activated B cells (NF‐κB) and mitogen‐activated protein kinase (MAPK) signaling. Our results demonstrated for the first time that mitochondrial fission regulates mitochondrial ROS production in activated microglial cells and influences the expression of pro‐inflammatory mediators through the activation of NF‐κB and MAPK. We therefore suggest that mitochondrial dynamics may be essential for understanding pro‐inflammatory mediator expression in activated microglial cells. This could represent a new therapeutic approach for preventing neurodegenerative diseases.
LPS induced excessive mitochondrial fission through mitochondrial localization of de‐phosphorylation of Ser637 Drp1. Interestingly, inhibition of LPS‐induced mitochondrial fission and mitochondrial ROS generation by Mdivi‐1 and Drp1 shRNA attenuate the production of pro‐inflammatory mediators via reduced NF‐κB and MAPK signaling. Our results suggest that mitochondrial dynamics may be essential for understanding pro‐inflammatory mediator expression in activated microglial cells.
LPS induced excessive mitochondrial fission through mitochondrial localization of de‐phosphorylation of Ser637 Drp1. Interestingly, inhibition of LPS‐induced mitochondrial fission and mitochondrial ROS generation by Mdivi‐1 and Drp1 shRNA attenuate the production of pro‐inflammatory mediators via reduced NF‐κB and MAPK signaling. Our results suggest that mitochondrial dynamics may be essential for understanding pro‐inflammatory mediator expression in activated microglial cells.
Dynamin‐related protein 1 (DRP1) is a mitochondrial membrane GTPase and regulates mitochondrial fission. In this study, we found that the cytokine RANKL increased the expression of DRP1 and its ...receptor proteins, Fis1, Mid49, and Mid 51, during osteoclast formation in mouse bone marrow‐derived macrophages. Inactivation of the kinase GSK3β appeared to induce DRP1 expression. DRP1 knockdown or the DRP1 inhibitor Mdivi1 suppressed osteoclast differentiation via downregulation of c‐Fos and NFATc1, the key transcription factor for osteoclast formation. Finally, the DRP1 inhibitor suppressed lipopolysaccharide‐induced osteoclast formation in a calvarial model and ovariectomy‐induced bone loss in vivo. Taken together, our data demonstrate that DRP1 positively contributes to RANKL‐induced osteoclast differentiation by regulating the c‐Fos–NFATc1 axis, suggesting the importance of mitochondrial DRP1 in osteoclastogenesis.
In drug discovery or preclinical stages of development, potency parameters such as IC
,
, or
have been routinely used to predict the parameters of efficacious exposure (AUC,
, etc.) in humans. ...However, to our knowledge, the fundamental assumption that the potency
is correlated with the efficacious concentration
in humans has not been investigated extensively. Thus, the present review examined this assumption by comparing a wide range of published pharmacokinetic (PK) and potency data. If the drug potency
and its
effectiveness in humans are well correlated, the steady-state average unbound concentrations in humans
=
·F·Dose/(
·τ) =
·AUCss/τ after treatment with approved dosage regimens should be higher than, or at least comparable to, the potency parameters assessed
. We reviewed the ratios of
/potency
for a total of 54 drug entities (13 major therapeutic classes) using the dosage, PK, and
potency reported in the published literature. For 54 drugs, the
/
potency ratios were < 1 for 38 (69%) and < 0.1 for 22 (34%) drugs. When the ratios were plotted against
(unbound fraction), "ratio < 1" was predominant for drugs with high protein binding (90% of drugs with
≤ 5%; i.e., 28 of 31 drugs). Thus, predicting the
efficacious unbound concentrations in humans using only
potency data and
should be avoided, especially for molecules with high protein binding.
We aimed to determine the role of extracellular peroxiredoxin 1 (Prdx1) in the pathogenesis of bacterial infections and inflammatory bone disease.
We first investigated the role of Prdx1 using ...knockout mice. Next, we determined the role of extracellular Prdx1 in bacterial infections by using a neutralizing antibody against Prdx1. We finally investigated whether blockade of extracellular Prdx1 affected high- or low-grade inflammatory bone diseases using calvarial osteolysis, collagen-induced arthritis (CIA), and microgravity-induced bone loss in mouse models.
The lack of Prdx1 increased susceptibility to infections by Listeria monocytogenes or Escherichia coli. Prdx1 is released into the serum upon E. coli infection, and blockade of extracellular Prdx1 confers significant protection against bacterial infections. Our data suggested that circulating Prdx1 is increased by the development of osteolytic disease, and that blockade of extracellular Prdx1 exerts therapeutic effects against high- and low-grade inflammatory bone loss. In addition, the release of Prdx1 under inflammatory osteolytic conditions partly depends on non-canonical TIR-domain-containing adapter-inducing interferon-β (TRIF)-caspase-11-gasdemin D (GSDMD) inflammasome pathways.
Extracellular Prdx1 is involved in the development of bacterial infections and inflammatory bone disease. Thus, extracellular Prdx1 may represent a novel therapeutic target for bacterial infections or inflammatory osteolytic diseases.
•The bias caused by early information loss in the time-to-event modeling has never been reported.•Using the PIEM-algorithm, the biased modeling results (Kaplan-Meier plot) caused by early information ...loss were corrected.•This method may help researchers obtain unbiased time-to-event models when the early information loss is severe.
Typical clinical data can suffer routine information loss when event times are rounded to the nearest day and right-censored at the end of follow-up. Because of the daily basis recording system, for the first 24 h, there are no events, which can damage the estimation of the Weibull survival model. Its estimation bias is inevitable since, for this short period, massive events might have occurred, the data is missing, and the fitted Weibull model is to show a steep slope. This phenomenon of estimation bias caused by the information loss caused by the problem of measurement resolution has not been properly discussed so far.
We propose a partial imputation Expectation Maximization (PIEM)-algorithm to estimate missing lifetimes only for day 1 at the mode among the whole clinical follow-up days. Based on various Weibull distributions, we simulated clinical sets after rounding and censoring raw event times and prepared chimera sets by partially substituting the imputed lifetimes only for the 24 h at the mode among the entire clinical sets.
For shape parameter ≤ 1, almost all the 95% prediction intervals (PIs) of both parameters in the chimera sets include their true values, while those in the clinical sets miss most of the true shape parameters and some of the true scale parameters. Estimating a small proportion of missing data only for the 24 h period, while keeping the rest as they are, greatly reduces biases of both scale and shape parameters. For shape parameter >1, the chimera sets consistently outperform the clinical sets.
The PIEM-algorithm may be applied as an intuitive tool for time-to-event modeling of survival data with this kind of information loss.
In drug discovery or preclinical stages of development, potency parameters such as $IC_{50}$, $K_i$, or $K_d$ in vitro have been routinely used to predict the parameters of efficacious exposure (AUC, ...$C_{min}$, etc.) in humans. However, to our knowledge, the fundamental assumption that the potency in vitro is correlated with the efficacious concentration in vivo in humans has not been investigated extensively. Thus, the present review examined this assumption by comparing a wide range of published pharmacokinetic (PK) and potency data. If the drug potency in vitro and its in vivo effectiveness in humans are well correlated, the steady-state average unbound concentrations in humans $C_{u_-ss.avg}=f_u{\cdot}F{\cdot}Dose/(CL{\cdot}{\tau})=f_u{\cdot}AUCss/{\tau}$ after treatment with approved dosage regimens should be higher than, or at least comparable to, the potency parameters assessed in vitro. We reviewed the ratios of $C_{u_-ss.avg}$/potency in vitro for a total of 54 drug entities (13 major therapeutic classes) using the dosage, PK, and in vitro potency reported in the published literature. For 54 drugs, the $C_{u_-ss.avg}$/in vitro potency ratios were < 1 for 38 (69%) and < 0.1 for 22 (34%) drugs. When the ratios were plotted against $f_u$ (unbound fraction), "ratio < 1" was predominant for drugs with high protein binding (90% of drugs with $f_u{\leq}5%$; i.e., 28 of 31 drugs). Thus, predicting the in vivo efficacious unbound concentrations in humans using only in vitro potency data and $f_u$ should be avoided, especially for molecules with high protein binding.
•In silico experiments and simulations using physiologically based pharmacokinetic (PBPK) and allometric approaches have played an important role in pharmaceutical research and drug ...development.•Commercial PBPK platforms or relevant services are costly and generally demand a large amount of input data, but their prediction methods or key references are not fully known to users.•We developed DallphinAtoM, a user-friendly GUI tool predicting human PK through assessment of various references on PBPK and allometric methods.•DallphinAtoM is a free, straightforward and transparent tool enabling fast and reliable human PK simulation.
In silico experiments and simulations using physiologically based pharmacokinetic (PBPK) and allometric approaches have played an important role in pharmaceutical research and drug development. These methods integrate diverse data from preclinical and clinical development, and have been widely applied to in vitro-in vivo extrapolation (IVIVE) of absorption, distribution, metabolism, and excretion (ADME).
To develop a user-friendly open tool predicting human PK, we assessed various references on PBPK and allometric methods published so far. They were integrated into a software system named “DallphinAtoM” (Drugs with ALLometry and PHysiology Inside-Animal to huMan), which has a user-friendly platform that can handle complex PBPK models and allometric models with a relatively small amount of essential information of the drug. The models of DallphinAtoM support the integration of data gained during the nonclinical development phase, enable translation from animal to human, and allow the prediction of concentration-time profiles with predicted PK parameters.
We presented two illustrative applications using DallphinAtoM: (1) human PK simulation of an orally administered drug using PBPK method; and (2) simulation of intravenous infusion following a two-compartment model using the allometric scaling method.
We conclude that this is a straightforward and transparent tool allowing fast and reliable human PK simulation based on the latest knowledge on biochemical processes and physiology and provides valuable information for decision making during the early-phase drug development.
Osteoclasts (OCs) are resorptive cells responsible for bone erosion in diseases, including osteoporosis, periodontitis and rheumatoid arthritis. Montelukast is a cysteinyl leukotriene receptor 1 ...(CysLTR1) antagonist clinically used for the treatment of asthma. In the present study, the role of CysLTR1 on OC formation and bone loss was investigated using montelukast. Montelukast inhibited receptor activator of nuclear factor‑κB ligand (RANKL)‑induced OC formation in cultures of mouse bone marrow macrophages. Additionally, montelukast suppressed actin ring formation and bone resorption activity of differentiated OCs. The inhibitory effect of montelukast was associated with impaired activation of extracellular signal‑regulated kinase, AKT serine/threonine kinase, and/or phospholipase Cγ2 signaling pathways downstream of RANK, followed by decreased expression of nuclear factor of activated T cells c1. Notably, OC formation was efficiently restored by addition of adenosine diphosphate, a P2Y12 agonist, as well as by addition of CysLT. Furthermore, similar to montelukast, P2Y12 blockade by a pharmacological inhibitor or siRNAs suppressed OC differentiation. These data indicate the involvement of the P2Y12 receptor in the inhibitory effect of montelukast on osteoclastogenesis. In vivo, montelukast significantly inhibited inflammation‑induced osteoclastogenesis in the calvarial model. Montelukast also served a protective role in a murine ovariectomy (OVX)‑ and unloading‑induced bone loss model. Altogether, these results confirmed that the CysLTR1 antagonist exerted an inhibitory effect on OC formation in vitro and in vivo. It may be useful for the treatment of bone diseases associated with excessive bone resorption.
Preclinical models suggest anticancer activity of IM156, a novel biguanide mitochondrial protein complex 1 inhibitor of oxidative phosphorylation (OXPHOS). This first-in-human dose-escalation study ...enrolled patients with refractory advanced solid tumors to determine the maximum tolerated dose (MTD) or recommended phase 2 dose (RP2D). Eligible patients received oral IM156 every other day (QOD) or daily (QD) and were assessed for safety, dose-limiting toxicities (DLTs), pharmacokinetics, and preliminary signals of efficacy. 22 patients with advanced cancers (gastric, n = 8; colorectal, n = 3; ovarian, n = 3; other, n = 8) received IM156 100 to 1,200 mg either QOD or QD. There were no DLTs. However, 1,200 mg QD was not well tolerated due to nausea; 800 mg QD was determined as the RP2D. The most frequent treatment-related AEs (TRAEs) were nausea (n = 15; 68%), diarrhea (n = 10; 46%), emesis (n = 9; 41%), fatigue (n = 4; 18%) and abdominal pain, constipation, and blood lactate increased (n = 2 each; 9%). Grade 3 nausea (n = 3; 14%) was the only grade ≥ 3 TRAE. Plasma exposures increased dose proportionally; mean Day 27 area under the curve (AUC
0-24
) values were higher following QD administration compared to the respective QOD regimen. Stable disease (SD), observed in 7 (32%) patients (confirmed in 2 9%), was the best response. To our knowledge, this is the first phase 1 study of an OXPHOS inhibitor that established a RP2D for further clinical development in cancer. Observed AEs of IM156 were manageable and SD was the best response.
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