The impact of URMC-099 (URMC099) on the in vitro proliferation of "brain homing" MDA-MB-231 BR cells expressing eGFP (eGFP8.4) and their parental cell line, MDA-MB-231, is assessed. Both cell groups are exposed to either 200 nM URMC-099 or vehicle alone. Cells treated with URMC-099 exhibit comparable growth rates to those treated with the vehicle. Cell viability exceeds 99% in all instances[2].
体内研究
URMC-099 exhibits a moderate terminal elimination half-life (t1/2=1.92 h, 2.14 h and 2.72 h for C57 BL/6 mice (10 mg/kg, oral dosing), C57 BL/6 mice (2.5 mg/kg, iv), C57 BL/6 mice (10 mg/kg, iv))[1].
URMC-099 (URMC099) is evaluated for its impact on tumor formation in vivo using a well-established mouse xenograft model of breast cancer brain metastasis. In this study, eGFP8.4 cells are injected into the left ventricle of immunodeficient nu/nu mice, followed by treatment with either URMC-099 (10 mg/kg) or vehicle alone every 12 hours for 20 days. This dosage is selected based on its demonstrated efficacy in inhibiting MLK3 in mice, penetrating the blood-brain barrier effectively, and potently inhibiting phosphorylation of Jun N-terminal kinase (JNK) in brain tissue. Then, mice are euthanized, and the number of brain metastases (BM) is quantified. Each treatment group comprises fifteen mice. Brain metastases are detected in 60% of mice, consistent with previous findings using this xenograft model. Notably, URMC-099 treatment significantly increases the total number of brain metastases in mice (p<0.05, two-tailed t-test). A similar trend is observed for micrometastases, while the number of macrometastases remains statistically comparable between mice treated with URMC-099 and those given vehicle alone[2].
体外研究
The impact of URMC-099 (URMC099) on the in vitro proliferation of "brain homing" MDA-MB-231 BR cells expressing eGFP (eGFP8.4) and their parental cell line, MDA-MB-231, is assessed. Both cell groups are exposed to either 200 nM URMC-099 or vehicle alone. Cells treated with URMC-099 exhibit comparable growth rates to those treated with the vehicle. Cell viability exceeds 99% in all instances[2].
作用机制
URMC-099 inhibits MLK3 by interacting with MLK3 ATP binding site.
URMC-099 细胞实验
Cell Line
Concentration
Treated Time
Description
References
HeLa cells
200 nM
48 hours
URMC-099 enhanced ZIKV infection
J Virol. 2019 Aug 28;93(18):e00758-19.
Huh7 cells
200 nM
48 hours
URMC-099 enhanced ZIKV infection
J Virol. 2019 Aug 28;93(18):e00758-19.
BEnd.3 cells
200 nM
1 hour
To evaluate the preventive effect of URMC-099 on barrier integrity and inflammation in bEnd.3 monolayers. Results showed that URMC-099 pretreatment significantly reduced barrier damage and inflammatory responses induced by COVID-QDs or Spike protein.
ACS Appl Nano Mater. 2023 Aug 7;6(16):15094-15107.
Human monocyte-derived macrophages (MDMs)
400 ng/ml
1, 3, 5 days
Evaluate the ability of URMC-099 to affect the anti-HIV activity of nanoformulated ARVs, results showed that URMC-099 co-administered with 1 μM nanoATV reduced virion production by 4-fold
J Clin Invest. 2017 Mar 1;127(3):857-873.
BV-2 cells
100 nM
12 hours
URMC-099 significantly reduced the phagocytic activity of Tat-activated BV-2 cells.
J Neurosci. 2013 Jun 12;33(24):9998-10010.
Human monocyte-derived macrophages
1 µM
14 days
To evaluate the effect of URMC-099 on autophagy markers such as LC3B and p62, results showed that URMC-099 significantly increased autophagosome formation and drug retention in autophagosomes.
Nanomedicine (Lond). 2018 Sep;13(17):2139-2154.
Human monocyte-derived macrophages (MDM)
10 ng/mL
16 hours
To evaluate the effect of URMC-099 on HIV-1 infection, results showed URMC-099 enhanced the antiviral activity of nanoATV
Nanomedicine. 2016 Jan;12(1):109-22.
Primary rat hippocampal neurons
100 nM
18 hours
URMC-099 protected neuronal axons from destruction by Tat-activated microglia.
J Neurosci. 2013 Jun 12;33(24):9998-10010.
HEK293 cells
5 µM and 10 µM
24 hours
To evaluate the effect of MLK3 overexpression on AP-1 activity, results showed that MLK3 overexpression significantly increased AP-1-mediated luciferase activity.
Int J Mol Sci. 2022 Sep 17;23(18):10874.
Mouse N9 microglia cells
0.20 µM (EC50)
24 hours
To assess the inhibitory effect of URMC-099 on TNFα secretion in LPS-stimulated microglia, results showed URMC-099 significantly and dose-dependently decreased TNFα secretion
Cell Chem Biol. 2019 Dec 19;26(12):1703-1715.e37.
Bend.3 cells
300 nM
30 min pretreatment followed by 16 hours stimulation
To evaluate the inhibitory effect of URMC-099 on IL-1β-induced brain endothelial cell activation. Results showed URMC-099 pretreatment virtually abolished the IL-1β-induced increase in VCAM-1 immunoreactivity and reduced basal PECAM-1 immunoreactivity.
FASEB J. 2022 Jun;36(6):e22343.
Murine microglia
100 nM
30 minutes
URMC-099 inhibited phosphorylation of MKK3/MKK4 and p38/JNK in Aβ42-stimulated microglia, reduced the expression of pro-inflammatory cytokines IL-1β, IL-6, and TNF-α, and induced the expression of anti-inflammatory cytokines IL-4 and IL-13.
J Neuroinflammation. 2016 Jul 11;13(1):184.
BV-2 cells
100 nM
30 minutes
URMC-099 significantly reduced Tat-induced JNK phosphorylation but had minimal effect on p38 MAPK phosphorylation.
J Neurosci. 2013 Jun 12;33(24):9998-10010.
BV-2 cells
100 nM
4, 8, 12 hours
URMC-099 significantly decreased the production of TNFα, IL-6, and MCP-1 but had no effect on IL-10 expression.
J Neurosci. 2013 Jun 12;33(24):9998-10010.
U-118 MG
3 µM
48 hours
Evaluate the inhibitory effect of URMC-099 on glioblastoma cell proliferation, results showed that URMC-099 significantly inhibited cell proliferation
Cancer Cell Int. 2021 Jan 6;21(1):24.
U-87 MG
3 µM
48 hours
Evaluate the inhibitory effect of URMC-099 on glioblastoma cell proliferation, results showed that URMC-099 significantly inhibited cell proliferation
Cancer Cell Int. 2021 Jan 6;21(1):24.
SNB-19 cells
200 nM
48 hours
URMC-099 significantly enhanced ZIKV infection in a dose-dependent manner
J Virol. 2019 Aug 28;93(18):e00758-19.
RAW264.7 cells
10 µM
5-60 minutes
To evaluate the effect of LPS stimulation on inflammatory gene expression in RAW264.7 cells, results showed that LPS significantly increased the mRNA levels of COX-2 and CCL-12.
Int J Mol Sci. 2022 Sep 17;23(18):10874.
Human motor neurons
0.6 µM
72 hours
To evaluate the neuroprotective effects of URMC-099 against ER-stress-mediated neurodegeneration, results showed URMC-099 restored survival to 90% at 0.6 μM
Cell Chem Biol. 2019 Dec 19;26(12):1703-1715.e37.
Primary human monocytes
100, 300, 1000 nM
8 hours
Inhibition of HIV-1 Tat-induced cytokine release
J Med Chem. 2013 Oct 24;56(20):8032-48.
Mouse microglial BV-2 cells
100, 300, 1000 nM
8 hours
Inhibition of LPS-induced TNF-α release
J Med Chem. 2013 Oct 24;56(20):8032-48.
URMC-099 动物实验
Species
Animal Model
Administration
Dosage
Frequency
Description
References
Mice
BALB/cJ mice and humanized NSG mice
Intraperitoneal injection
10 mg/kg
Twice daily for 21 days
Assess the effect of URMC-099 on autophagy induction and pharmacokinetics of nanoformulated ARVs in vivo, results showed URMC-099 treatment increased plasma DTG levels by 52.3-fold
J Clin Invest. 2017 Mar 1;127(3):857-873.
Mice
HIV-1 Tat injection model
Intraperitoneal injection
10 mg/kg
Every 12 hours, total of 3 doses before Tat injection and 2 doses post injection
Inhibition of HIV-1 Tat-induced JNK signaling pathway activation
J Med Chem. 2013 Oct 24;56(20):8032-48.
APPSwDI/mNos2−/− AD mice
Orthopedic surgery model (tibial fracture/fixation)
Intraperitoneal injection
10 mg/kg
Three doses at 12-hour intervals before surgery, with the last dose immediately prior to surgery.
To evaluate the prophylactic effect of URMC-099 on hippocampal vascular vulnerability and synaptic damage following orthopedic surgery. Results showed URMC-099 prophylaxis reduced cerebrovascular VCAM-1 expression, prevented BBB breakdown and synapse loss, and inhibited microglial activation.
FASEB J. 2022 Jun;36(6):e22343.
Mice
HIV-1 Tat exposure model
Intraperitoneal injection
10 mg/kg
Every 12 hours throughout the experiment
URMC-099 reduced Tat-induced inflammatory cytokine production, protected neuronal architecture, and altered the morphologic and ultrastructural response of microglia to HIV-1 Tat exposure.
J Neurosci. 2013 Jun 12;33(24):9998-10010.
NOD/SCID/IL2R γc−/− mice
HIV-1-infected humanized mouse model
Intraperitoneal injection
10 mg/kg
Daily for three weeks
To evaluate the antiviral effect of URMC-099 in combination with nanoATV/r, results showed combined treatment significantly reduced viral load and infected cell numbers
Three injections, 12 hours apart, last injection one hour before surgery
Prevent surgery-induced neuroinflammation and cognitive impairment
J Neuroinflammation. 2019 Oct 28;16(1):193
Mice
MLK3 knockout mice
Intraperitoneal injection
10 mg/kg
Twice daily for 14 days
To evaluate the effect of MLK3 kinase inhibition on left ventricular function
JCI Insight. 2021 Sep 22;6(18):e149075
Balb/C nude mice
Subcutaneous glioblastoma xenograft model
Intraperitoneal injection
3 mg/kg
Once daily for 7 days
Evaluate the inhibitory effect of URMC-099 on glioblastoma xenograft growth, results showed that URMC-099 combined with AZD6482 significantly inhibited tumor growth
Cancer Cell Int. 2021 Jan 6;21(1):24.
BALB/c mice
ZIKV infection model
Intracerebral injection
5 mg/kg
Single dose, observed for 3 days
URMC-099 significantly increased ZIKV E protein levels and viral RNA load in mouse brains
J Virol. 2019 Aug 28;93(18):e00758-19.
Mice
Breast cancer model
Intraperitoneal injection
7.5 mg/kg
Once daily for 3 weeks
Evaluate the effect of MLK3 inhibition on T cell activation and cytotoxicity, results showed increased CD8+ GZMB+ T cell population
Proc Natl Acad Sci U S A. 2020 Apr 7;117(14):7961-7970
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