MEK1 (IC50 = 2-7 μM); MEK2 (IC50 = 50 μM); ERK1; ERK2; Autophagy

PD98059 inhibits either basal MEK1 or a partially activated MEK created by changing serine at residues 218 and 222 to glutamate (MEK-2E) with an IC50 of 2 M. The MAPK homologues JNK and P38 are not inhibited by PD98059 in any way. Raf kinase, cAMP-dependent kinase, protein kinase C, v-Src, epidermal growth factor (EGF) receptor kinase, insulin receptor kinase, PDGF receptor kinase, and phosphatidylinositol 3-kinase are just a few of the additional kinases that PD98059 does not inhibit. With an IC50 of ~10 μM and ~7 μM, respectively, PD98059 blocks PDGF-stimulated MAPK activation and thymidine incorporation into 3T3 cells.[1] PD98059 has an IC50 of 4 μM, which makes it potently inhibit MEK1 activation by Raf or MEK kinase, and an IC50 of 50 μM, which makes it weakly inhibit MEK2 activation by Raf. In KB and PC12 cells as well as in Swiss 3T3 cells, PD98059 does not prevent the activation of the MEK homologues MKK4 and RK kinase, which take part in the stress and interleukin-1-stimulated kinase cascades.[2] PD98059 completely prevents the differentiation of PC12 cells brought on by nerve growth factor (NGF) without affecting cell viability.[3] A dose-dependent inhibition of RAW264.7 cell proliferation in the presence of RANKL by PD98059 causes a visible reduction in TRAP-positive cells in the culture.[4]

Mice are treated A reduction in infarct volume occurs when PD98059 is administered 30 minutes before focal cerebral ischemia to prevent damage.[5] Based on pancreatic wet weight and histology, PD98059 pretreatment (10 mg/kg per i.v. injection) given 30 minutes prior to hourly cerulein injections for three hours significantly reduces the severity of cerulein-induced acute pancreatitis.[6] A reduction in all the measured parameters of inflammation is brought on by giving mice PD98059 (10 mg/kg) an hour after carrageenan.[7]

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The aim of the present study is to evaluate the contribution of mitogen-activated protein kinase 1-3 MAPK3/MAPK1) in a model of acute lung inflammation in mice. Injection of carrageenan into the pleural cavity of mice elicited an acute inflammatory response characterized by: accumulation of fluid containing a large number of neutrophils (PMNs) in the pleural cavity, infiltration of PMNs in lung tissues and subsequent adhesion molecule expression (I-CAM and P-selectin), lipid peroxidation, and increased production of tumour necrosis factor-alpha, (TNF-alpha) and interleukin-1beta (IL-1beta). Furthermore, carrageenan induced lung apoptosis (Bax and Bcl-2 expression) as well as nitrotyrosine formation, NF-kB activation, and pJNK expression, as determined by immunohistochemical analysis of lung tissues and the degree of lung inflammation and tissue injury (histological score). Administration of PD98059, an inhibitor of MAPK3/MAPK1 (10 mg/kg) 1 h after carrageenan caused a reduction in all the parameters of inflammation measured. Thus, based on these findings we propose that inhibitors of the MAPK3/MAPK1 signaling pathways, such as PD98059, may be useful in the treatment of various inflammatory diseases [7].

In vitro suppression of MEK and ERK activities [12]
Interest in the use of PD98059 as a tool in signal transduction research is derived from the specificity with which it inhibits MEK but no other kinases (Alessi et al., 1995; Dudley et al., 1995). A kinase cascade assay was used to quantify inhibition of MEK by PD98059 (Fig. 2). This assay uses a mutated, constitutively activated form of MEK to phosphorylate and activate ERK1, which in turn phosphorylates MBP. Hence, the activity of MEK can be monitored by measuring either ERK1 or MBP...

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PD98059 [2-(2'-amino-3'-methoxyphenyl)-oxanaphthalen-4-one] is a flavonoid and a potent inhibitor of mitogen-activated protein kinase kinase (MEK). Concentrations of PD98059 of /=10 microM. In vivo exposure of cultures to 95%) of the dually phosphorylated forms of extracellular signal-regulated kinase (IC50 = 1 muM). Treatment of cultures with PD98059 of >/=1 muM either at the time of addition or up to 48 hr before the addition of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) suppressed in a concentration-dependent manner the accumulation of induced steady state CYP1A1, CYP1B1, and NQO1 mRNAs. The addition of PD98059 to rat liver cytosol just before the addition of TCDD suppressed TCDD binding (IC50 = 4 muM) and aryl hydrocarbon receptor (AHR) transformation (IC50 = 1 muM), as measured by sucrose gradient centrifugation and electrophoretic mobility shift assays. Flavone and flavanone, two closely related structural analogs of PD98059, inhibited AHR transformation by TCDD with IC50 values similar to that obtained with PD98059. However, neither analog was as potent as PD98059 in inhibiting MEK (IC50 approximately 190 muM for both). These results suggest that PD98059 is a ligand for the AHR and functions as an AHR antagonist at concentrations commonly used to inhibit MEK and signaling processes that entail MEK activation. [12]

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In order to measure the incorporation of 32P into myelin basic protein (MBP), glutathione S-transferase (GST) fusion proteins containing either the 44-kDa MAPK (GST-MAPK) or the 45-kDa MEK (GST-MEK1) are used. Assays are carried out in 50 μL of a solution containing 10 μg of GST-MEK1, 0.5 μg of GST-MAPK, and 40 μg of MBP, along with 50 mM Tris, pH 7.4, 10 mM MgCl2, 2 mM EGTA, and 10 μM [γ-32P]ATP. The addition of Laemmli SDS sample buffer stops reactions after 15 minutes of incubation at 30°C. By using SDS/10% PAGE, phosphorylated MBP is resolved.

Cells are plated into multi-well plates at a density of 10,000–20,000/mL for monolayer growth. The cell growth medium is then supplemented with various concentrations of PD98059 48 hours later, and incubation is then carried out for an additional 3 days. Following trypsin incubation, cells are then extracted from the wells and counted using a Coulter Counter. Cells are seeded into 35-mm dishes at a density of 5,000–10,000 cells per dish with growth medium containing the desired concentrations of PD98059 and 0.3% agar for growth in soft agar. Visible colonies are manually counted using a dissecting microscope after 7–10 days of growth.

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Cell lines and cell cultures. [11]
Three human AML cell lines were cultured in RPMI 1640 containing 10% heat-inactivated FCS. OCI-AML-3 and MOLM-13 cells have wild-type p53, whereas p53 is disabled in HL-60 by large deletion of the TP53. Cell lines were harvested in log-phase growth, seeded at a density of 2 × 105 cells/mL and exposed to the Nutlin-3a and/or PD98059. In experiments involving combinations of Nutlin-3a and PD98059, OCI-AML-3 and HL-60 cells were treated with Nutlin-3a at 0, 1, 2.5, 5, and 10 μmol/L and MOLM-13 cells at 0, 0.4, 1, 2, and 4 μmol/L, in the absence or presence of 20 μmol/L PD98059. The two agents were added simultaneously to cells, and they were cultured for 24 h. Cell viability was evaluated by triplicate counts of trypan blue dye–excluding cells. Experiments were done at least in duplicate.

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Real-time quantitative PCR. [11]
OCI-AML-3 cells were treated with 20 μmol/L PD98059 and/or 5 μmol/L Nutlin-3a for 3 h. RNA was prepared from cells using RNeasy Mini Kit, and first-strand cDNA was generated using random hexamers (SuperScript III First-Strand Synthesis SuperMix) from 1 μg total RNA. The mRNA expression levels of p21, Mdm2, Noxa, ABL, and 18S were quantified using TaqMan gene expression assays. Quantitative real-time PCR was done using an ABI Prism 7500 Sequence Detection System and TaqMan Universal Master Mix. The reaction was initiated by a hold for 10 min at 95°C followed by 40 cycles 15 s at 95°C and 1 min at 60°C. Two probes for 18S (Hs99999901_s1) and p21 (Hs00355782_m1) were used. The specificity of the PCR products was confirmed by melting curve analysis with ABI SDS 2.0 software. Relative expression levels were calculated based on the difference in CT values between the test samples and control untreated cells. This was normalized with expression levels of 18S using the equation Etarget(CTtesttarget − CTcontroltarget)/Eref(CTtestref − CTcontrolref). The real-time PCR experiments were carried out in triplicate.

Male Sprague–Dawley rats with acute pancreatitis
10 mg/kg
Injection i.v.

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PD98059 (MEK1 Inhibitor) has been shown to act in vivo as a highly selective inhibitor of MEK1 activation and the MAP kinase cascade. In the present study, we have investigated the effects of PD98059, on the development of non-septic shock caused by zymosan in mice. Mice received either intraperitoneally zymosan (500mg/kg, administered i.p. as a suspension in saline) or vehicle (0.25ml/mouse saline). PD98059 (10mg/kg) was administered 1 and 6h after zymosan administration i.p. Organ failure and systemic inflammation in mice was assessed 18h after administration of zymosan and/or PD98059. Treatment of mice with PD98059 attenuated the peritoneal exudation and the migration of polymorphonuclear cells caused by zymosan. PD98059 also attenuated the lung, liver and pancreatic injury and renal dysfunction caused by zymosan as well as the increase of TNF-alpha and IL-1beta plasma levels caused by zymosan. Immunohistochemical analysis for inducible nitric oxide synthase (iNOS), nitrotyrosine, poly(ADP-ribose) (PAR), ICAM-1, P-selectin, Bax, Bcl-2 and FAS-ligand revealed positive staining in pancreatic and intestinal tissue obtained from zymosan-injected mice. The degree of staining for nitrotyrosine, iNOS, PAR, ICAM-1, P-selectin, Bax, Bcl-2 and FAS-ligand were markedly reduced in tissue sections obtained from zymosan-injected mice, which had received PD98059. Moreover treatment of mice with PD98059 (10mg/kg) attenuated the NF-kappaB activation and mitogen-activated protein kinases (MAPK) expression induced by zymosan injection. In addition, administration of zymosan caused a severe illness in the mice characterized by a systemic toxicity, significant loss of body weight and a 60% of mortality at the end of observation period. Treatment with PD98059 significantly reduced the development of systemic toxicity, the loss in body weight and the mortality (20%) caused by zymosan. This study provides evidence that PD98059 attenuates the degree of zymosan-induced non-septic shock in mice. Pharmacol Res. 2010 Feb;61(2):175-87.

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PD98059 (2-(2-Amino-3-methoxyphenyl)-4H-1-benzopyran-4-one) (Sigma-Aldrich, USA) was dissolved in 75% DMSO. The PD98059 (2.5 mcg/5 mcl, i.t.) was single or repeated preemptively administered 16 h and 1 h before CCI and then once daily for 7 days (the administration was according to our previous paper [17], [25]). The Vehicle-treated CCI-exposed rats received 75% DMSO according to the same schedule. There was no significant difference in pain behavior between no-treated and V(DMSO)-treated CCI-exposed rats. This method of PD98059 or vehicle administration was used throughout the study and is referred to in the text as “repeated administration”. At day 7th after CCI 30 min after PD98059 administration tactile allodynia was measured using von Frey test and thermal hyperalgesia was conducted using cold plate test. Additionally, at day 7th after CCI the vehicle-treated and PD98059-treated rats received a single i.t. vehicle, morphine (2.5 mcg/5 mcl) or buprenorphine (2.5 mcg/5 mcl) injection 30 min after PD98059, and then 30 min later the von Frey and/or cold plate tests were repeated. Since the dose of morphine 2.5 mcg/5 mcl in naïve rats produced maximal analgesic effect in tail-flick test. We have used lower dose of morphine for co-administration experiments, so that we would be able to observe the possible enhancement of opioid effectiveness. The vehicle-treated and PD98059-treated naïve rats (uninjured rats) received a single i.t. vehicle, morphine (0.5 mcg/5 mcl) or buprenorphine (2.5 mcg/5 mcl) injection 30 min after PD98059, and then 30 min later the tail flick test was performed (S1 Table and S1 Fig). https://pmc.ncbi.nlm.nih.gov/articles/PMC4591269/#sec002

[1]. Proc Natl Acad Sci U S A . 1995 Aug 15;92(17):7686-9.

[2]. J Biol Chem . 1995 Nov 17;270(46):27489-94.

[3]. J Biol Chem . 1995 Jun 9;270(23):13585-8.

[4]. J Biol Chem . 2002 Dec 6;277(49):47366-72.

[5]. Proc Natl Acad Sci U S A . 1999 Oct 26;96(22):12866-9.

[6]. Pancreas . 2002 Oct;25(3):251-9.

[7]. Int J Immunopathol Pharmacol . 2009 Oct-Dec;22(4):937-50.

[8]. Melanoma Res . 2001 Feb;11(1):11-9.

[9]. Int Immunopharmacol . 2007 Jan;7(1):36-45.

[10]. Int J Cancer . 2003 Nov 10;107(3):478-85.

[11]. Cancer Res . 2007 Apr 1;67(7):3210-9.

[12]. Mol Pharmacol . 1998 Mar;53(3):438-45.

2-(2-amino-3-methoxyphenyl)chromen-4-one is a member of the class of monomethoxyflavones that is 3'-methoxyflavone bearing an additional amino substituent at position 2'. It has a role as an EC 2.7.11.24 (mitogen-activated protein kinase) inhibitor and a geroprotector. It is a monomethoxyflavone and an aromatic amine.
PD-98059 is an inhibitor of MAP-kinase kinase activation.
2-(2-Amino-3-methoxyphenyl)-4H-1-benzopyran-4-one has been reported in Pestalotiopsis neglecta with data available.
MEK Inhibitor PD-98059 is a cell-permeable, selective mitogen-activated protein (MAP) kinase inhibitor which exhibits activity through the inhibition of the phosphorylation and activation of MAP kinase.

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Treatment of cells with a variety of growth factors triggers a phosphorylation cascade that leads to activation of mitogen-activated protein kinases (MAPKs, also called extracellular signal-regulated kinases, or ERKs). We have identified a synthetic inhibitor of the MAPK pathway. PD 098059 [2-(2'-amino-3'-methoxyphenyl)-oxanaphthalen-4-one] selectively inhibited the MAPK-activating enzyme, MAPK/ERK kinase (MEK), without significant inhibitory activity of MAPK itself. Inhibition of MEK by PD 098059 prevented activation of MAPK and subsequent phosphorylation of MAPK substrates both in vitro and in intact cells. Moreover, PD 098059 inhibited stimulation of cell growth and reversed the phenotype of ras-transformed BALB 3T3 mouse fibroblasts and rat kidney cells. These results indicate that the MAPK pathway is essential for growth and maintenance of the ras-transformed phenotype. Further, PD 098059 is an invaluable tool that will help elucidate the role of the MAPK cascade in a variety of biological settings.[1]

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PD 098059 has been shown previously to inhibit the dephosphorylated form of mitogen-activated protein kinase kinase-1 (MAPKK1) and a mutant MAPKK1(S217E,S221E), which has low levels of constitutive activity (Dudley, D. T., Pang, L., Decker, S. J., Bridges, A. J., and Saltiel, A. R. (1995) Proc. Natl. Acad. Sci. U.S.A. 92, 7686-7689). Here we report that PD 098059 does not inhibit Raf-activated MAPKK1 but that it prevents the activation of MAPKK1 by Raf or MEK kinase in vitro at concentrations (IC50 = 2-7 microM) similar to those concentrations that inhibit dephosphorylated MAPKK1 or MAPKK1(S217E,S221E). PD 098059 inhibited the activation of MAPKK2 by Raf with a much higher IC50 value (50 microM) and did not inhibit the phosphorylation of other Raf or MEK kinase substrates, indicating that it exerts its effect by binding to the inactive form of MAPKK1. PD 098059 also acts as a specific inhibitor of the activation of MAPKK in Swiss 3T3 cells, suppressing by 80-90% its activation by a variety of agonists. The high degree of specificity of PD 098059 in vitro and in vivo is indicated by its failure to inhibit 18 protein Ser/Thr kinases (including two other MAPKK homologues) in vitro by its failure to inhibit the in vivo activation of MAPKK and MAP kinase homologues that participate in stress and interleukin-1-stimulated kinase cascades in KB and PC12 cells, and by lack of inhibition of the activation of p70 S6 kinase by insulin or epidermal growth factor in Swiss 3T3 cells. PD 098059 (50 microM) inhibited the activation of p42MAPK and isoforms of MAP kinase-activated protein kinase-1 in Swiss 3T3 cells, but the extent of inhibition depended on how potently c-Raf and MAPKK were activated by any particular agonist and demonstrated the enormous amplification potential of this kinase cascade. PD 098059 not only failed to inhibit the activation of Raf by platelet-derived growth factor, serum, insulin, and phorbol esters in Swiss 3T3 cells but actually enhanced Raf activity. The rate of activation of Raf by platelet-derived growth factor was increased 3-fold, and the subsequent inactivation that occurred after 10 min was prevented. These results indicate that the activation of Raf is suppressed and that its inactivation is accelerated by a downstream component(s) of the MAP kinase pathway.[2]

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The mitogen-activated protein kinase (MAP kinase) pathway is thought to play an important role in the actions of neurotrophins. A small molecule inhibitor of the upstream kinase activator of MAP kinase, MAP kinase kinase (MEK) was examined for its effect on the cellular action of nerve growth factor (NGF) in PC-12 pheochromocytoma cells. PD98059 selectively blocks the activity of MEK, inhibiting both the phosphorylation and activation of MAP kinases in vitro. Pretreatment of PC-12 cells with the compound completely blocked the 4-fold increase in MAP kinase activity produced by NGF. Half-maximal inhibition was observed at 2 microM PD98059, with maximal effects at 10-100 microM. The tyrosine phosphorylation of immunoprecipitated MAP kinase was also completely blocked by the compound. In contrast, the compound was without effect on NGF-dependent tyrosine phosphorylation of the pp140trk receptor or its substrate Shc and did not block NGF-dependent activation of phosphatidylinositol 3'-kinase. However, PD98059 completely blocked NGF-induced neurite formation in these cells without altering cell viability. These data indicate that the MAP kinase pathway is absolutely required for NGF-induced neuronal differentiation in PC-12 cells.[3]

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Activation of the Raf/MEK/ERK pathway and inactivation of wild-type p53 by Mdm2 overexpression are frequent molecular events in acute myelogenous leukemia (AML). We investigated the interaction of Raf/MEK/ERK and p53 pathways after their simultaneous blockades using a selective small-molecule antagonist of Mdm2, Nutlin-3a, and a pharmacologic MEK-specific inhibitor, PD98059. We found that PD98059, which itself has minimal apoptogenic activity, acts synergistically with Nutlin-3a to induce apoptosis in wild-type p53 AML cell lines OCI-AML-3 and MOLM-13. Interestingly, PD98059 enhanced nuclear proapototic function of p53 in these cells. In accordance with the activation of transcription-dependent apoptosis, PD98059 treatment promoted the translocation of p53 from the cytoplasm to the nucleus in OCI-AML-3 cells, in which p53 primarily initiates transcription-independent apoptosis when cells are treated with Nutlin-3a alone. The critical role of p53 localization in cells with increased p53 levels was supported by enhanced apoptosis induction in cells cotreated with Nutlin-3a and the nuclear export inhibitor leptomycin B. PD98059 prevented p53-mediated induction of p21 at the transcriptional level. The repressed expression of antiapototic p21 also seemed to contribute to synergism between PD98059 and Nutlin-3a because (a) the synergistic apoptogenic effect was preserved in G(1) cells, (b) p53-mediated induction of p21 was preferentially seen in G(1) cells, (c) PD98059 strongly antagonized p21 induction by Nutlin-3a, and (d) cells with high p21 levels were resistant to apoptosis. This is the first report showing that the Raf/MEK/ERK pathway regulates the subcellular localization of p53 and the relative contribution of transcription-dependent and transcription-independent pathways in p53-mediated apoptosis.[10]

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Anticancer drugs docetaxel and vinorelbine suppress cell growth by altering microtubule assembly and activating the proapoptotic signal pathway. Vinorelbine and docetaxel have been approved for treating several advanced cancers. However, their efficacy in the management of advanced hormone-refractory prostate cancer remains to be clarified. Microtubule damage by some anticancer drugs can activate the ERK survival pathway, which conversely compromises chemotherapeutic efficacy. We analyzed the effect of ERK inhibitors PD98059 and U0126 on vinorelbine- and docetaxel-induced cell growth suppression of androgen-independent prostate cancer cells. In androgen-independent C-81 LNCaP cells, inhibition of ERK by PD98059, but not U0126, plus docetaxel resulted in enhanced growth suppression by an additional 20% compared to the sum of each agent alone (p < 0.02). The combination treatment of docetaxel plus PD98059 also increased cellular apoptosis, which was in part due to the inactivation of Bcl-2 by increasing phosphorylated Bcl-2 by more than 6-fold and Bax expression by 3-fold over each agent alone. At these dosages, docetaxel alone caused only marginal phosphorylation of Bcl-2 (10%). Docetaxel plus U0126 had only 20% added effect on Bcl-2 phosphorylation compared to docetaxel alone. Nevertheless, both U0126 and PD98059 exhibited an enhanced effect on docetaxel-induced growth suppression in PC-3 cells. No enhanced effect was observed for vinorelbine plus PD98059 or U0126. Thus, the combination therapy of docetaxel plus PD98059 may represent a new anticancer strategy, requiring lower drug dosages compared to docetaxel monotherapy. This may lower the cytotoxicity and enhance tumor suppression in vivo. This finding of a combination effect could be of potential clinical importance in treating hormone-refractory prostate cancer.[9]

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Neuropathic pain treatment remains challenging due to ineffective therapy and resistance to opioid analgesia. Mitogen-activated protein kinase kinase (MAPKK) have been identified as the crucial regulators of pro- and antinociceptive factors. We used PD98059, an inhibitor of the MAPKK family members MEK1/2. The aim of study was to examine the influence of single and/or repeated PD98059 on nociception and opioid effectiveness in neuropathy. Moreover, we examined how PD98059 influences selected members of cellular pathways and cytokines. The PD98059 (2.5 mcg) was intrathecally preemptively administered before chronic constriction injury (CCI), and then once daily for 7 days. Additionally, at day 7 after CCI the PD98059-treated rats received a single injection of opioids. Using Western blot and qRT-PCR techniques in PD98059-treated rats we analyzed the mRNA and/or protein level of p38, ERK1/2, JNK, NF-kappaB, IL-1beta, IL-6, iNOS and IL-10 in the lumbar spinal cord. Our results indicate that PD98059 has an analgesic effects and potentiates morphine and/or buprenorphine analgesia. Parallel we observed that PD98059 inhibit upregulation of the CCI-elevated p38, ERK1/2, JNK and NF-kappaB protein levels. Moreover, PD98059 also prevented increase of pro- (IL-1beta, IL-6, and iNOS) but enhances anti-nociceptive (IL-10) factors. Summing up, PD98059 diminished pain and increased the effectiveness of opioids in neuropathy. The inhibition of MEKs might inactivate a variety of cell signaling pathways that are implicated in nociception. PLoS One. 2015 Oct 1;10(10):e0138583. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4591269/

C16H13NO3

267.2793

267.089

C, 71.90; H, 4.90; N, 5.24; O, 17.96

167869-21-8

167869-21-8

4713

Light yellow to yellow solid powder

1.3±0.1 g/cm3

453.1±45.0 °C at 760 mmHg

170 °C

221.9±25.0 °C

0.0±1.1 mmHg at 25°C

1.652

2.43

1

4

2

20

407

0

O1C2=C([H])C([H])=C([H])C([H])=C2C(C([H])=C1C1C([H])=C([H])C([H])=C(C=1N([H])[H])OC([H])([H])[H])=O

QFWCYNPOPKQOKV-UHFFFAOYSA-N

nChI=1S/C16H13NO3/c1-19-14-8-4-6-11(16(14)17)15-9-12(18)10-5-2-3-7-13(10)20-15/h2-9H,17H2,1H3

2-(2-amino-3-methoxyphenyl)chromen-4-one

PD98059; PD 98059; PD098059; 2-(2-amino-3-methoxyphenyl)-4H-chromen-4-one; PD-98059; 2-(2-Amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; PD 98,059; 2-(2-amino-3-methoxyphenyl)chromen-4-one; PD-98059

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: This product requires protection from light (avoid light exposure) during transportation and storage.

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO: 33.3~46 mg/mL (124.7~172.1 mM)
Ethanol: Insoluble (<1 mg/mL)
Water: Insoluble (<1 mg/mL)

Solubility in Formulation 1: 2.08 mg/mL (7.78 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 2: 4% DMSO+30% PEG 300+5% Tween 80+ddH2O: 1mg/mL

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Solubility in Formulation 3: 10 mg/mL (37.41 mM) in 50% PEG300 50% Saline (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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 (Please use freshly prepared in vivo formulations for optimal results.)