ZSTK474

Catalog No.S1072

For research use only.

ZSTK474 inhibits class I PI3K isoforms with IC50 of 37 nM in a cell-free assay, mostly PI3Kδ. Phase1/2.

ZSTK474 Chemical Structure

CAS No. 475110-96-4

Selleck's ZSTK474 has been cited by 67 publications

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Biological Activity

Description ZSTK474 inhibits class I PI3K isoforms with IC50 of 37 nM in a cell-free assay, mostly PI3Kδ. Phase1/2.
Features First orally administered PI3K inhibitor used in vivo.
Targets
PI3Kδ [2]
(Cell-free assay)
PI3Kα [2]
(Cell-free assay)
PI3K [1]
(Cell-free assay)
PI3Kβ [2]
(Cell-free assay)
PI3Kγ [2]
(Cell-free assay)
4.6 nM 16 nM 37 nM 44 nM 49 nM
In vitro

ZSTK474 at 1 μM potently reduces PI3K activity to 4.7% of the control level, whereas LY2194002 only reduces the activity to 44.6% of the control. ZSTK474 inhibits the activities of recombinant p110β, -γ, and -δ with IC50 of 17 nM, 53 nM, and 6 nM, respectively. ZSTK474 shows potent antiproliferative activity against a panel of 39 human cancer cell lines with mean GI50 of 0.32 μM, more effectively than that of LY294002 or wortmannin with mean GI50 of 7.4 μM or 10 μM, respectively. ZSTK474 treatment at 1 μM blocks membrane ruffling and generation of PIP3 induced by platelet-derived growth factor in murine embryonic fibroblasts (MEFs). ZSTK474 at 10 μM induces apoptosis in OVCAR3 cells, and induces complete G1-phase arrest but not apoptosis in A549 cells. ZSTK474 treatment at 0.5 μM significantly decreases the level of phosphorylated Akt and GSK-3β, as well as the cyclin D1 protein expression. ZSTK474 also inhibits the phosphorylation of other downstream signaling components that are involved in regulating cell proliferation including FKHRL1, FKHR, TSC-2, mTOR, and p70S6K in a dose-dependent manner. [1] ZSTK474 does not inhibit mTOR at 0.1 μM, and even at a concentration of 100 μM, ZSTK474 inhibits mTOR activity less than 40%. [2] ZSTK474 blocks VEGF-induced cell migration and the tube formation in human umbilical vein endothelial cells (HUVECs), and inhibits the expression of HIF-1α and secretion of VEGF in RXF-631L cells, exhibiting potent in vitro antiangiogenic activity. [3] ZSTK474 treatment inhibits the production of IFNγ and IL-17 in concanavalin A-activated T cells, and inhibits the proliferation and PGE(2) production by fibroblast-like synovial cells (FLS). [6]

Cell Data
Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
Sf21 insect cells NHHsTpJHfW6ldHnvckBie3OjeR?= NEW2cIcyKGh? Mly1TY5pcWKrdHnvckBw\iCkb4\pcoUhemWlb33ibY5idnRiUFmzT{BxOTFyZHXseIEh\XiycnXzd4VlKGmwIGPmNlEhcW6|ZXP0JINmdGy|IIXzbY5oKHCqb4PwbIF1cWS7bHnuc5NqfG:uIHHzJJN2[nO2cnH0[UBi\nSncjCxJIhzKGK7IIDoc5NxcG:rbXHnbY5oNCCLQ{WwQVAvPyCwTR?= NIP4e28zOTh6MkizNi=>
human HCT116 cells NWfJPYMyTnWwY4Tpc44h[XO|YYm= NXrCcmhsOTVibXnudy=> NFP6SZhKdmirYnn0bY9vKG:oIGDJT|NESSCKMUC0O3IhdXW2YX70MY1m\GmjdHXkJINmdGxic3nncoFtcW6pIHnuJIh2dWGwIFjDWFEyPiClZXzsd{BmgHC{ZYPzbY5oKFCWRV6gZZN{\XO|ZXSgZZMhcW6qaXLpeIlwdiCxZjDpcpN2dGmwLXnu[JVk\WRicFHreE9RU0JicHjvd5Bpd3K7bHH0bY9vKGG2IGTodlMxQCC2cnXheIVlKG[xcjCxOUBucW6|IHLl[o9z\SCrboP1cIlvKGOqYXzs[Y5o\SCvZXHzeZJm\CCjZoTldkA2KG2rboOgZpkhcW2vdX7vZoxwfHSrbnesJGlEPTB;N{igcm0> M37yZ|IyQDh{OEOy
human LNCAP cells NGjucXRRem:uaX\ldoF1cW:wIHHzd4F6 NWfkOGRlOyCmYYnz M{njRWFvfGmycn;sbYZmemG2aY\lJIFkfGm4aYT5JIFo[Wmwc4SgbJVu[W5iTF7DRXAh[2WubIOgZYZ1\XJiMzDkZZl{KGK7IF3UV{Bie3OjeTygTWM2OD1yLkKxJO69VQ>? NV[zdVc4OjB{Mke4PFE>
human NZB5 cells MW\Qdo9tcW[ncnH0bY9vKGG|c3H5 M2Xxb|Uh\GG7cx?= Mn71RY51cXC{b3zp[oVz[XSrdnWgZYN1cX[rdImgZYdicW6|dDDoeY1idiCQWlK1JINmdGy|IHX4dJJme3Orbnege4lt\CC2eYDlJJAyOTCjbIDoZUBie3Onc4Pl[EBieyCrbnPvdpBwemG2aX;uJI9nKFt|SG30bJlucWSrbnWgZYZ1\XJiNTDkZZl{NCCLQ{WwQVAvOjJizszN Ml3KNlE5QDJ6M{K=
human NZOV9 cells M13CPXBzd2yrZnXyZZRqd25iYYPzZZk> MWe1JIRigXN? MUTBcpRqeHKxbHnm[ZJifGm4ZTDhZ5Rqfmm2eTDh[4FqdnO2IHj1cYFvKE6cT2[5JINmdGy|IHX4dJJme3OrbnegdFEyOGGucHjhJItqdmG|ZTDZNVAzOUNibYX0ZY51KGG|c3Xzd4VlKGG|IHnuZ49zeG:{YYTpc44hd2ZiW{PIYZRpgW2rZHnu[UBi\nSncjC1JIRigXNuIFnDOVA:OC5{OTFOwG0> M4fkd|IyQDh{OEOy
human MDA-MB-468 cells NHHpOm1EgXSxdH;4bYPDqGG|c3H5 NUn1VphxPDhiaB?= MlfiR5l1d3SxeHnjbZR6KGGpYXnud5QhWFSHTj3k[YZq[2mnboSgbJVu[W5iTVTBMW1DNTR4ODDj[YxteyCjc4Pld5Nm\CCjczDpcohq[mm2aX;uJI9nKGOnbHyg[5Jwf3SqIHHmeIVzKDR6IHjyd{BjgSCFZXzsJHRqfGW{IEm2JIF{e2G7 M2n6NVI{Pzl3MkO5
human A549 cells MnrFSpVv[3Srb36gZZN{[Xl? M1[5NlExKM7:TR?= Mn[wNUBp M3\KbmlvcGmkaYTpc44hd2ZiUFmzT{BqdiCqdX3hckBCPTR7IHPlcIx{KGG|c3Xzd4VlKGG|IILl[JVkfGmxbjDpckBxSWu2IHzleoVtKGG2IEGwJJVOKGGodHXyJFEhcHJiYomgW4V{fGW{bjDicI91fGmwZzDhcoFtgXOrcx?= MnPUNlU4PjZ4M{O=
human DMS114 cells MoXvS5Jwf3SqIHnubIljcXSrb36gZZN{[Xl? NUW2VFNQT3Kxd4ToJIlvcGmkaYTpc44hd2ZiaIXtZY4hTE2VMUG0JINmdGy| MYqyNlM{PjJ2Nh?=
human MKN74 cells NHP5S3JIem:5dHigbY5pcWKrdHnvckBie3OjeR?= M4KxNWdzd3e2aDDpcohq[mm2aX;uJI9nKGi3bXHuJG1MVjd2IHPlcIx{ NFPjcG8zOjN|NkK0Oi=>
human SNB78 cells NXrwdIJpT3Kxd4ToJIlvcGmkaYTpc44h[XO|YYm= MnLrS5Jwf3SqIHnubIljcXSrb36gc4YhcHWvYX6gV25DPzhiY3XscJM> NGLMeIYzOjN|NkK0Oi=>
human St-4 cells Mnq2S5Jwf3SqIHnubIljcXSrb36gZZN{[Xl? MoHFS5Jwf3SqIHnubIljcXSrb36gc4YhcHWvYX6gV5QuPCClZXzsdy=> NYDVe3huOjJ|M{[yOFY>
human DU145 cells NHf6WYpIem:5dHigbY5pcWKrdHnvckBie3OjeR?= MnjES5Jwf3SqIHnubIljcXSrb36gc4YhcHWvYX6gSHUyPDViY3XscJM> NVXmOIJbOjJ|M{[yOFY>
human LOXIMVI cells NGDHS4xIem:5dHigbY5pcWKrdHnvckBie3OjeR?= NFrudFJIem:5dHigbY5pcWKrdHnvckBw\iCqdX3hckBNV1iLTW\JJINmdGy| NIfHWHQzOjN|NkK0Oi=>
human PC3 cells NWjxeWhYT3Kxd4ToJIlvcGmkaYTpc44h[XO|YYm= M4HQPGdzd3e2aDDpcohq[mm2aX;uJI9nKGi3bXHuJHBEOyClZXzsdy=> MXiyNlM{PjJ2Nh?=
human LOXIMVI cells M3i1NWdzd3e2aDDpcohq[mm2aX;uJIF{e2G7 NH62fYVIem:5dHigbY5pcWKrdHnvckBw\iCqdX3hckBNV1iLTW\JJINmdGy| MW[yNlM{PjJ2Nh?=
human OVCAR3 cells M{fQdGdzd3e2aDDpcohq[mm2aX;uJIF{e2G7 NHjVU3RIem:5dHigbY5pcWKrdHnvckBw\iCqdX3hckBQXkODUkOgZ4VtdHN? NEL0XHEzOjN|NkK0Oi=>
human SKOV3 cells MmSwS5Jwf3SqIHnubIljcXSrb36gZZN{[Xl? NYTPRXplT3Kxd4ToJIlvcGmkaYTpc44hd2ZiaIXtZY4hW0uRVkOgZ4VtdHN? M1O5[|IzOzN4MkS2
human KM12 cells MUfHdo94fGhiaX7obYJqfGmxbjDhd5NigQ>? MnvhS5Jwf3SqIHnubIljcXSrb36gc4YhcHWvYX6gT20yOiClZXzsdy=> MmjCNlI{OzZ{NE[=
human HT-29 cells NXTlWmRYT3Kxd4ToJIlvcGmkaYTpc44h[XO|YYm= NXThcHJwT3Kxd4ToJIlvcGmkaYTpc44hd2ZiaIXtZY4hUFRvMkmgZ4VtdHN? MUOyNlM{PjJ2Nh?=
human HCT15 cells M{LsOmdzd3e2aDDpcohq[mm2aX;uJIF{e2G7 MVvHdo94fGhiaX7obYJqfGmxbjDv[kBpfW2jbjDIR3QyPSClZXzsdy=> MmTXNlI{OzZ{NE[=
human NCI-H226 cells NYO3NVFoT3Kxd4ToJIlvcGmkaYTpc44h[XO|YYm= MYnHdo94fGhiaX7obYJqfGmxbjDv[kBpfW2jbjDOR2kuUDJ{NjDj[Yxtew>? MnfUNlI{OzZ{NE[=
human NCI-H522 cells  NH3qWHRIem:5dHigbY5pcWKrdHnvckBie3OjeR?= NGXZN49Iem:5dHigbY5pcWKrdHnvckBw\iCqdX3hckBPS0lvSEWyNkBk\Wyuc9Mg M1zSeVIzOzN4MkS2
human A549 cells NYPPWo46T3Kxd4ToJIlvcGmkaYTpc44h[XO|YYm= M4Lme2dzd3e2aDDpcohq[mm2aX;uJI9nKGi3bXHuJGE2PDliY3XscJM> MlvKNlI{OzZ{NE[=
human HCC2998 cells MnjLS5Jwf3SqIHnubIljcXSrb36gZZN{[Xl? M{W3emdzd3e2aDDpcohq[mm2aX;uJI9nKGi3bXHuJGhESzJ7OUigZ4VtdHN? M3TTNFIzOzN4MkS2
human SNB75 cells Mm\YS5Jwf3SqIHnubIljcXSrb36gZZN{[Xl? NWDDOYtXT3Kxd4ToJIlvcGmkaYTpc44hd2ZiaIXtZY4hW06EN{WgZ4VtdHN? M4HqWVIzOzN4MkS2
human OVCAR4 cells NWXse4tjT3Kxd4ToJIlvcGmkaYTpc44h[XO|YYm= MkTlS5Jwf3SqIHnubIljcXSrb36gc4YhcHWvYX6gU3ZESVJ2IHPlcIx{ NFjNemUzOjN|NkK0Oi=>
human OVCAR5 cells M36yc2dzd3e2aDDpcohq[mm2aX;uJIF{e2G7 M2LaeWdzd3e2aDDpcohq[mm2aX;uJI9nKGi3bXHuJG9XS0GUNTDj[Yxtew>? NEj5V3UzOjN|NkK0Oi=>
human OVCAR8 cells NHHYPJZIem:5dHigbY5pcWKrdHnvckBie3OjeR?= MmjRS5Jwf3SqIHnubIljcXSrb36gc4YhcHWvYX6gU3ZESVJ6IHPlcIx{ M1fyfFIzOzN4MkS2
human SKOV3 cells M2PxZ2dzd3e2aDDpcohq[mm2aX;uJIF{e2G7 MUTHdo94fGhiaX7obYJqfGmxbjDv[kBpfW2jbjDTT29XOyClZXzsdy=> M3L4TVIzOzN4MkS2
human ACHN cells NGO2SmVIem:5dHigbY5pcWKrdHnvckBie3OjeR?= MVrHdo94fGhiaX7obYJqfGmxbjDv[kBpfW2jbjDBR2hPKGOnbHzz NWHENHg{OjJ|M{[yOFY>
Assay
Methods Test Index PMID
Western blot p-PDK1 / p-GSK3β / p-AKT / AKT ; P-gp / MRP1 ; p-Rb / p27 / Cyclin D1 ; HIF-1α / HIF-1β 28388564 26918351 23812078
Growth inhibition assay Cell viability 28388564
In vivo Oral administration of ZSTK474 inhibits the growth of subcutaneously implanted mouse B16F10 melanoma tumors in a dose-dependent manner, producing tumor regression of 28.5%, 7.1%, or 4.9% on day 14 at 100, 200, or 400 mg/kg, respectively, which is superior to that of the four major anticancer drugs irinotecan, cisplatin, doxorubicin, and 5-fluorouracil at their respective maximum tolerable doses with tumor regression of 96%, 35.7%, 24%, or 68.3%, respectively. ZSTK474 treatment at 400 mg/kg completely inhibits the growth of A549, PC-3, and WiDr xenografts in mice, and induces the regression of A549 xenograft tumors. [1] ZSTK474 significantly inhibits tumor growth in the RXF-631L xenograft model, correlated with a significantly reduced number of microvessels in the ZSTK474-treated mice. [3] Oral administration of ZSTK474 ameliorates the progression of adjuvant-induced arthritis (AIA) in rats. [6]

Protocol (from reference)

Kinase Assay:[1]
  • Inhibition of PI3K activity:

    A549 cells are lysed in a buffer containing 20 mM Tris-HCl (pH 7.5), 150 mM NaCl, 5 mM EDTA, and 1% Igepal CA-630, the lysates are centrifuged at 20,000 g and 4 °C for 10 minutes, and the supernatants are used as cell lysate (protein = 2-4 mg/mL). To immunoprecipitate PI3K, 200 μL of cell lysate are incubated with anti-p85 polyclonal antibody and protein G-agarose (5 μL). PI3Kα, PI3Kβ, and PI3Kδ can be immunoprecipitated by the anti-p85 polyclonal antibody. Agarose beads containing immunoprecipitates are washed twice with buffer A (20 mM Tris-HCl at pH 7.5, 150 mM NaCl, 5 mM EDTA, and 1% Igepal CA-630), once with buffer B (500 mM LiCl and 100 mM Tris-HCl at pH 7.5), once with distilled water, and once with buffer C (100 mM NaCl and 20 mM Tris-HCl at pH 7.5). Immunoprecipitates are suspended in 20 μL of buffer C containing phosphatidylinositol of 200 μg/mL. The mixture is preincubated with increasing concentrations of ZSTK474 at 25 °C for 5 minutes. [γ-32P]ATP (2 μCi per assay mixture; final concentration, 20 μM) and MgCl2 (final concentration, 20 mM) are added to start the reaction. The reaction mixture is incubated at 25 °C for 20 minutes. Phosphorylated products of phosphatidylinositol are separated by thin-layer chromatography and visualized by autoradiography. The phosphatidylinositol-3-phosphate region is scraped from the plate, and radioactivity is also measured with liquid scintillation spectroscopy. The level of inhibition for ZSTK474 is determined as the percentage of 32P counts per minute obtained without ZSTK474.

Cell Research:[1]
  • Cell lines: MCF-7, HT-29, HCT-116, OVCAR3, A549, et al.
  • Concentrations: Dissolved in DMSO, final concentrations ~10 μM
  • Incubation Time: 48 hours
  • Method: Cells are exposed to increasing concentrations of ZSTK474 for 48 hours. The inhibition of cell proliferation is assessed by measuring changes in total cellular protein by use of a sulforhodamine B assay. Apoptosis is assessed by chromatin condensation or by flow cytometry. For chromatin condensation assay, cells are stained with Hoechst 33342 and examined by fluorescence microscopy. Morphologic changes induced by ZSTK474, such as the condensation of chromatin, are indicative of apoptosis. For flow cytometry analysis, cells are harvested, washed with ice-cold PBS, and fixed in 70% ethanol. Cells are then washed twice with ice-cold PBS again, treated with RNase A (500 μg/mL) at 37 °C for 1 hour, and stained with propidium iodide (25 μg/mL). The DNA content of the cells is analyzed with a flow cytometer.
Animal Research:[1]
  • Animal Models: Male BDF1 mice injected subcutaneously with B16F10 cells, and female BALB/c nude mice inoculated subcutaneously with A549, PC-3, or WiDr cells
  • Dosages: ~400 mg/kg/day
  • Administration: Orally

Solubility (25°C)

In vitro

In vivo

Add solvents to the product individually and in order
(Data is from Selleck tests instead of citations):
0.5% hydroxyethyl cellulose
For best results, use promptly after mixing.

30mg/mL

Chemical Information

Molecular Weight 417.41
Formula

C19H21F2N7O2

CAS No. 475110-96-4
Storage 3 years -20°C powder
2 years -80°C in solvent
Smiles C1COCCN1C2=NC(=NC(=N2)N3C4=CC=CC=C4N=C3C(F)F)N5CCOCC5

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Answers to questions you may have can be found in the inhibitor handling instructions. Topics include how to prepare stock solutions, how to store inhibitors, and issues that need special attention for cell-based assays and animal experiments.

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