Necrostatin-1

Synonyms: Nec-1

Necrostatin-1 (Nec-1) is a specific RIP1 (RIPK1) inhibitor and inhibits TNF-α-induced necroptosis with EC50 of 490 nM in 293T cells. Necrostatin-1 also blocks IDO and suppresses autophagy and apoptosis.

Necrostatin-1 Chemical Structure

Necrostatin-1 Chemical Structure

CAS: 4311-88-0

Selleck's Necrostatin-1 has been cited by 229 publications

Purity & Quality Control

Batch: Purity: 99.97%
99.97

Necrostatin-1 Related Products

Choose Selective RIP kinase Inhibitors

Cell Data

Cell Lines Assay Type Concentration Incubation Time Formulation Activity Description PMID
HT-22 Cytotoxicity Assay 10 μM 12 h protects against cell death induced by 5 mmol/L glutamate  17760869
Jurkat  Function Assay 200 μm 30 min reduces Naegleria fowleri-induced reactive oxygen species (ROS) generation 21535020
Jurkat  Cytotoxicity Assay 50/ 100/200 μm 1/3 h reduces Naegleria fowleri-induced cytotoxicity 21535020
SW13 Cell Viability Assay 100 μM 24 h DMSO increases cellular survival 22136818
8505c Cell Viability Assay 100 μM 24 h DMSO increases cellular survival 22136818
TPC-1 Cell Viability Assay 100 μM 24 h DMSO increases cellular survival 22136818
L929sA Apoptosis Assay 10 μM 1 h abrogates the interaction of caspase-8 with FADD 22362767
L929sA Apoptosis Assay 10 μM 1 h rescues cells expressing RIPK1ΔID from TNF-induced apoptosis 22362767
L929sA Apoptosis Assay 10 μM 1 h inhibits the apoptotic response to TNF 22362767
K562/Adr  Function Assay 60 μM 12 h increases the activity of caspases, caspase 8 and 9 22837689
K562 Function Assay 60 μM 12 h increases the activity of caspases, caspase 8 and 9 22837689
HL60/Adr Function Assay 60 μM 12 h increases the activity of caspases, caspase 8 and 9 22837689
HL60 Function Assay 60 μM 12 h increases the activity of caspases, caspase 8 and 9 22837689
K562/Adr  Function Assay 60 μM 12 h augments the caspase-3 activity 22837689
K562 Function Assay 60 μM 12 h augments the caspase-3 activity 22837689
HL60/Adr Function Assay 60 μM 12 h augments the caspase-3 activity 22837689
HL60 Function Assay 60 μM 12 h augments the caspase-3 activity 22837689
K562/Adr  Apoptosis Assay 60 μM 12 h enhances shikonin-induced apoptosis 22837689
K562 Apoptosis Assay 60 μM 12 h enhances shikonin-induced apoptosis 22837689
HL60/Adr Apoptosis Assay 60 μM 12 h enhances shikonin-induced apoptosis 22837689
HL60 Apoptosis Assay 60 μM 12 h enhances shikonin-induced apoptosis 22837689
SH-EP Apoptosis Assay 10 μM  72 h  inhibits IAP inhibitor- and Lexatumumab-induced apoptosis 22890322
NIH3T3  Function Assay 10/50 μM 1/3 h ameliorates TNFα-driven complex formation 23261677
HT-22  Function Assay 25 μM 0–30 min DMSO inhibits ERK Activation induced by glutamate 23307752
HT-22  Cell Viability Assay 10 μM 12 h DMSO protects against glutamate-induced cell death 23307752
KMS-12-BM Cytotoxicity Assay 90 µM 1 h blocks BAY 11-7082 induced rapid cell swelling 23527154
MM.1S  Cytotoxicity Assay 90 µM 1 h blocks BAY 11-7082 induced rapid cell swelling 23527154
ΔN-Karpas 299  Cytotoxicity Assay 20 μM 16 h inhibits CD30-induced cell death 23545938
MEFs Function Assay 20 μM 1/2/4 h suppresses TNFα-induced RIPK1 phosphorylation 23727581
MEFs Cytotoxicity Assay 2/6/20 μM 18 h inhibits TNFα-induced cell death in RelA KO MEFs 23727581
RMS13 Cell Viability Assay 40 μg/ml  24 h rescues GX15-070-induced loss of cell viability 23744296
TE671 Cell Viability Assay 40 μg/ml  24 h rescues GX15-070-induced loss of cell viability 23744296
U87 Function Assay 1 mmol/L 1.5-3 h suppresses the expression of RIP-1 caused by shikonin 23840441
C6 Function Assay 1 mmol/L 1.5-3 h suppresses the expression of RIP-1 caused by shikonin 23840441
U87 Cytotoxicity Assay 1 mmol/L 3 h blocks shikonin induced necrosis 23840441
C6 Cytotoxicity Assay 1 mmol/L 3 h blocks shikonin induced necrosis 23840441
U87 Cell Viability Assay 1 mmol/L 3 h attenuates Shikonin induced glioma cell death 23840441
C6 Cell Viability Assay 1 mmol/L 3 h attenuates Shikonin induced glioma cell death 23840441
L929 Function Assay 5 μg/ml 24 h blocks zVAD induced necroptosis and autophagy 23941769
L929 Function Assay 2 μg/ml  24 h promots caspase-6 (p20) activity and procaspase-6 cleavage 23941769
L929 Growth Inhibition Assay 2/5 μg/ml  24 h reverses the cell growth inhibition and cell death induced by TNFα alone as well as TNFα + zVAD 23941769
L929 Function Assay 2/5 μg/ml  24 h reversed the autophagy induced by TNFα alone as well as TNFα + zVAD 23941769
NRK-52E  Cell Viability Assay 20 μM 24 h inhibits increased Drp1 protein expression after TNF-α Stimulation and ATP Depletion 24351845
NRK-52E  Cell Viability Assay 20 μM 24 h increases cell viability after TNF-α Stimulation and ATP Depletion 24351845
NRK-52E  Cell Viability Assay 20 μM 24 h protects cells from cell death caused by ischemia injury 24351845
AGS Cell Viability Assay 60 μm 1 h prevents shikonin-induced cell death 24463199
L-540  Cell Viability Assay 60 μm 1 h reduces the Givinostat/Sorafenib-induced cell death 24561519
L-540  Function Assay 60 μm 1 h prevents the mitochondrial membrane depolarization 24561519
L-540  Function Assay 60 μm 1 h prevents the generation of ROS 24561519
SK-Hep1 Function Assay 60 μM  18 h blocks β-lapachone-mediated PAR accumulation and AIF translocation to the cytosol  24832602
SK-Hep1 Function Assay 60 μM  18 h inhibits β-Lapachone-induced leakage of HMGB-1  24832602
SK-Hep1 Function Assay 60 μM  18 h blocks β-lapachone-induced morphological change, cell death and PI uptake 24832602
OHC Function Assay 300 μM DMSO increases the number of apoptotic OHCs without altering the levels of CC8 after noise exposure 24874734
OHC Function Assay 300 μM DMSO diminishes noise-induced AMPK activation 24874734
OHC Function Assay 300 μM DMSO results in a reduction of noise-induced RIP1 and RIP3 immunofluorescence 24874734
OHC Function Assay 300 μM DMSO decreases noise-induced swollen nuclei  24874734
OHC Function Assay 300 μM DMSO increases noise-induced condensed nuclei 24874734
Huh7 Cell Viability Assay 50 µM 24/48 h DMSO prevents cell death of rAdHCV co-infected Huh7 cells 24973240
L929 Cell Viability Assay 30 μM 1 h inhibits TNF-α-induced cleavage of Topo I 25095742
L929 Cell Viability Assay 30 μM 1 h inhibits TNF-α-induced loss of cell viability 25095742
L929-A Function Assay 50 μM  12 h inhibits the TNFα-induced loss of mitochondrial membrane permeability 25398540
L929 Function Assay 50 μM  12 h inhibits TNFα-induced Bid cleavage 25398540
L929-N  Function Assay 50 μM  12 h blocks the cleavage of Caspase-3 and PARP 25398540
L929-A  Function Assay 50 μM  12 h blocks the cleavage of Caspase-3 and PARP 25398540
L929-N  Cell Viability Assay 50 μM  24 h blocks TNFα-induced cell death 25398540
L929-A  Cell Viability Assay 50 μM  24 h blocks TNFα-induced cell death 25398540
KMS-12-PE  Cell Viability Assay 60 μM 5 h inhibits SHK-induced cell death 25530098
SGC-7901 Cell Viability Assay 30 μM 1 h suppresses oxaliplatin-mediated cell death 25767076
BxPC-3 Function Assay 20 μM 24 h decreases the early necrotic cells 26000607
MiaPaCa-2 Function Assay 20 μM 24 h decreases the early necrotic cells 26000607
NCI-H28 Cell Viability Assay 10 μM 24 h prevents DAPE-induced reduction of NCI-H28 cell viability  26004138
BMDM  Function Assay 10 μM 30 min protects cells from TAKI-induced LDH release 26381601
MEFs  Cell Viability Assay 10 μM 48 h DMSO inhibits zVAD-promoted death of CNOT3-depleted MEFs 26437789
A549 Cell Viability Assay 50 μM 24 h inhibits MMS-induced cell death 26472723
Jurkat T Necroptosis assay Inhibition of TNF-alpha-induced necroptosis in FADD-deficient human Jurkat T cells, EC50 = 0.05 μM. 18467094
Jurkat Function assay Inhibition of endogenous RIP1 autophosphorylation in human Jurkat cells, EC50 = 0.182 μM. 18408713
Sf9 Function assay 30 mins Inhibition of recombinant human GST-fused RIPK1 (1 to 497 residues) expressed in baculovirus infected insect Sf9 cells in presence of 32P-gamma-ATP after 30 mins by autoradiogram-based Western blot method, IC50 = 0.182 μM. 28280261
Jurkat T Necroptosis assay Effective concentration required for inhibition of necroptosis in FADD deficient Jurkat T cells treated with TNF-alpha, EC50 = 0.49 μM. 16153840
Jurkat Necroptosis assay Inhibition of cellular necroptosis in TNFalpha treated FADD deficient human Jurkat cells, EC50 = 0.49 μM. 18408713
Jurkat T Necroptosis assay 30 uM 24 hrs Inhibition of necroptosis in TNF-alpha-induced human Jurkat T cells assessed as cell viability at 30 uM after 24 hrs 18467094
L929 Necroptosis assay 30 uM 24 hrs Inhibition of necroptosis in zVAD-induced mouse L929 cells assessed as cell viability at 30 uM after 24 hrs 18467094
L929 Necroptosis assay 30 uM 24 hrs Inhibition of necroptosis in TNF-alpha-induced mouse L929 cells assessed as cell viability at 30 uM after 24 hrs 18467094
3T3 Cell death assay 24 hrs Inhibition of death receptor signaling mediated necrotic cell death in mouse 3T3 cells assessed as cell viability after 24 hrs by ATP based viability assay in presence of TNFalpha and zVAD.fmk 16408008
3T3 Cell death assay 24 hrs Inhibition of death receptor signaling mediated necrotic cell death in mouse 3T3 cells assessed as cell viability after 24 hrs by ATP based viability assay in presence of FasL and zVAD.fmk 16408008
MEF Cell death assay 16 hrs Inhibition of death receptor signaling mediated necrotic cell death in SV40 transformed mouse MEF cells assessed as cell viability after 16 hrs by ATP based viability assay in presence of TNFalpha, CHX and zVAD.fmk 16408008
IEC18 Cell death assay Inhibition of death receptor signaling mediated necrotic cell death in rat IEC18 cells assessed as cell viability in presence of TNFalpha and zVAD.fmk 16408008
HL60 Cell death assay Inhibition of death receptor signaling mediated necrotic cell death in human HL60 cells assessed as cell viability in presence of TNFalpha and zVAD.fmk 16408008
L929 Cell death assay 24 hrs Inhibition of death receptor signaling mediated necrotic cell death in mouse L929 cells assessed as cell viability after 24 hrs by ATP based viability assay in presence of TNFalpha 16408008
Jurkat Necrosis assay Inhibition of necrosis in human Jurkat cells assessed as nuclear condensation by bright field microscopy in presence of FasL, CHX and zVAD-fmk 16408008
Jurkat Necrosis assay Inhibition of necrosis in human Jurkat cells assessed as organelle swelling by bright field microscopy in presence of FasL, CHX and zVAD-fmk 16408008
Jurkat Necrosis assay Inhibition of necrosis in human Jurkat cells assessed as early loss of plasma membrane integrity by bright field microscopy in presence of FasL, CHX and zVAD-fmk 16408008
Jurkat Necrosis assay Inhibition of necrosis in human Jurkat cells assessed as appearance of translucent cytosol in presence of FasL, CHX and zVAD-fmk 16408008
Jurkat Necrosis assay Inhibition of necrosis in human Jurkat cells deficient in FADD assessed as inhibition of nuclear condensation by bright field microscopy in presence of TNFalpha 16408008
Jurkat Necrosis assay Inhibition of necrosis in human Jurkat cells deficient in FADD assessed as inhibition of organelle swelling by bright field microscopy in presence of TNFalpha 16408008
Jurkat Necrosis assay Inhibition of necrosis in human Jurkat cells deficient in FADD assessed as inhibition of early loss of plasma membrane integrity by bright field microscopy in presence of TNFalpha 16408008
Jurkat Necrosis assay Inhibition of necrosis in human Jurkat cells deficient in FADD assessed as inhibition of appearance of translucent cytosol in presence of TNFalpha 16408008
U937 Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human U937 cells assessed as cell viability after 48 hrs by ATP based viability assay in presence of TNFalpha and zVAD-fmk 16408008
3T3 Cell death assay 24 hrs Inhibition of death receptor signaling mediated necroptotic cell death in mouse 3T3 cells assessed as cell viability after 24 hrs by ATP based viability assay in presence of TNFalpha and zVAD-fmk 16408008
Jurkat Cell death assay 24 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD assessed as decreased levels of PE-conjugated LC3-II (autophagy marker) after 24 hrs by Western blot method in presence of TNFalpha 16408008
L929 Cell death assay 24 hrs Inhibition of death receptor signaling mediated necroptotic cell death in mouse L929 cells assessed as decreased levels of PE-conjugated autophagy marker LC3-II after 24 hrs by Western blot method in presence of TNFalpha 16408008
3T3 Cell death assay 24 hrs Inhibition of death receptor signaling mediated necroptotic cell death in mouse 3T3 cells assessed as decreased levels of PE-conjugated autophagy marker LC3-II after 24 hrs by Western blot method in presence of TNFalpha and zVAD-fmk 16408008
3T3 Cell death assay 24 hrs Inhibition of death receptor signaling mediated necroptotic cell death in mouse 3T3 cells assessed as decreased levels of PE-conjugated autophagy marker LC3-II after 24 hrs by Western blot method in presence of FasL and zVAD-fmk 16408008
3T3 Cell death assay 24 hrs Inhibition of death receptor signaling mediated necroptotic cell death in mouse 3T3 cells assessed as decreased levels of PE-conjugated autophagy marker LC3-II after 24 hrs by Western blot method in presence of rapamycin 16408008
Jurkat Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD and expressing FKBP12-based dimerization domain assessed as cell viability after 48 hrs by FACS in presence of AP1510, zVAD-fmk 16408008
Jurkat Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD and expressing RIP kinase assessed as cell viability after 48 hrs by FACS in presence of AP1510, zVAD-fmk 16408008
Jurkat Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD and expressing RIP K45M mutant assessed as cell viability after 48 hrs by FACS in presence of AP1510, zVAD-fmk 16408008
Jurkat Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD and expressing RIP kinase domain assessed as cell viability after 48 hrs by FACS in presence of AP1510, zVAD-fmk 16408008
Jurkat Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD and expressing FKBP12-based dimerization domain assessed as cell viability after 48 hrs by FACS in presence of AP1510 16408008
Jurkat Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD and expressing RIP kinase assessed as cell viability after 48 hrs by FACS in presence of AP1510 16408008
Jurkat Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD and expressing RIP K45M mutant assessed as cell viability after 48 hrs by FACS in presence of AP1510 16408008
Jurkat Cell death assay 48 hrs Inhibition of death receptor signaling mediated necroptotic cell death in human Jurkat cells deficient in FADD and expressing RIP kinase domain assessed as cell viability after 48 hrs by FACS in presence of AP1510 16408008
Sf9 Function assay Inhibition of human RIP1 K45M mutant autophosphorylation expressed in Sf9 cells 18408713
Jurkat Cytoprotective assay 30 uM 1 hr Cytoprotective activity against FasL-induced necroptosis in human Jurkat cells assessed as increase in cell viability at 30 uM incubated for 1 hr followed by FasL stimulation measured after 20 hrs by Alamar blue assay 29541357
Jurkat Cytoprotective assay 30 uM 1 hr Cytoprotective activity against CHX-induced necroptosis in human Jurkat cells assessed as increase in cell viability at 30 uM incubated for 1 hr followed by CHX stimulation by Alamar blue assay 29541357
Jurkat Cytoprotective assay 30 uM 1 hr Cytoprotective activity against Z-VAD-induced necroptosis in human Jurkat cells assessed as increase in cell viability at 30 uM incubated for 1 hr followed by Z-VAD stimulation by Alamar blue assay 29541357
Jurkat Cytoprotective assay 30 uM 1 hr Cytoprotective activity against FasL-induced necroptosis in human Jurkat cells assessed as increase in cell viability at 30 uM incubated for 1 hr followed by FasL stimulation measured after 20 hrs by phase contrast microscopy 29541357
Jurkat Cytoprotective assay 30 uM 1 hr Cytoprotective activity against CHX-induced necroptosis in human Jurkat cells assessed as increase in cell viability at 30 uM incubated for 1 hr followed by CHX stimulation by phase contrast microscopy 29541357
Jurkat Cytoprotective assay 30 uM 1 hr Cytoprotective activity against Z-VAD-induced necroptosis in human Jurkat cells assessed as increase in cell viability at 30 uM incubated for 1 hr followed by Z-VAD stimulation by phase contrast microscopy 29541357
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Biological Activity

Description Necrostatin-1 (Nec-1) is a specific RIP1 (RIPK1) inhibitor and inhibits TNF-α-induced necroptosis with EC50 of 490 nM in 293T cells. Necrostatin-1 also blocks IDO and suppresses autophagy and apoptosis.
Features A powerful tool for characterizing the role of necroptosis with characterized primary target.
Targets
RIP1 [1]
(293T cells)
490 nM(EC50)
In vitro
In vitro

Necrostatin-1 (1-100 μM) inhibits the autophosphorylation of overexpressed and endogenous RIP1.It is found RIP1 is the primary cellular target responsible for the antinecroptosis activity of Necrostatin-1. [1]

Necrostatin-1 efficiently suppresses necroptotic cell death triggered by an array of stimuli in a variety of cell types. Necrostatin-1, previously identified as small-molecule inhibitor of necroptosis, inhibits RIP kinase-induced necroptosis and inhibits TNF-α-induced necroptosis in jurkat cells with EC50 of 490 nM. [2]

Kinase Assay RIP1 kinase assay
Phosphorylation of RIP1 requires its kinase activity. Expression constructs of FLAGtagged wild-type (WT) or a kinase-inactive pointmutant of RIP1 (K45M) are are transfected into 293T cells and RIP1 kinase assay is performed as described in the Methods in the presence of [γ-32P]ATP for 30 min at 30℃. Samples are subjected to SDS-PAGE and RIP1 band is visualized by autoradiography. Relative intensities of radioactive bands are quantified and are shown (ratio) in this and all other autoradiographs. In parallel to kinase reactions, a sample of beads is subjected to western blot analysis using anti-RIP1 antibody to ensure equal protein amounts in kinase reactions.
Cell Research Cell lines Jurkat, BALB/c 3T3, SV40-transformed MEF, L929
Concentrations 0.01-100 μM
Incubation Time --
Method

Cells are seeded in 96-well plates (white plates for luminescent assays; black plates for fluorescent assays; clear plates for MTT assay) at the density of 5,000-10,000 cells per well for adherent cells or 20,000-50,000 cells per well for suspension cells in 100 μl of the appropriate phenol red-free media. After incubation, we determined cell viability using one of the following methods. For the ATP assay, we used luminescence-based commercial kits and analyzed luminescence using a Wallac Victor II plate reader. For Sytox assay, we incubated cells with 1 μM Sytox Green reagent for 30 min at 37℃, and then performed fluorescent reading. Subsequently, we added 5 μl of 20% Triton X-100 solution into each well to produce maximal lysis and incubated cells for 1 h at 37℃, then performed the second reading. We calculated the ratio of values before and after Triton treatment and normalized it to the relevant controls not subjected to cytotoxic stimuli, as indicated in figure legends. For the MTT assay, we used the CellTiter 96 AQueous Non-Radioactive Cell Proliferation Assay kit. For PI exclusion assays, we added 2 μg/ml PI into the medium and immediately analyzed samples using FACSCalibur. For PI-annexin V assay we used the ApoAlert Annexin V-EGFP Apoptosis Kit. For DioC6 staining, we incubated cells with 40 nM DiOC6 for 30 min at 37 ℃, washed once and analyzed in FACSCalibur. For ROS analysis, we incubated cells with 5 μM dihydroethidium for 30 min at 37 ℃, washed once and analyzed in FACSCalibur. EM analyses are performed at the Harvard Medical School EM facility. We acquired bright-field images of the cells using an Axiovert 200 microscope.

Experimental Result Images Methods Biomarkers Images PMID
Immunofluorescence RIP1 / RIP3 30462730
In Vivo
In vivo

Necrostatin-1 (Nec-1) is a specific small molecule inhibitor of receptor-interacting protein kinase 1 (RIPK1) that specifically inhibits phosphorylation of RIPK1.

Animal Research Animal Models Male C57BL/6 mice
Dosages 0.0468 mg/Kg
Administration i.a.

Chemical Information & Solubility

Molecular Weight 259.33 Formula

C13H13N3OS

CAS No. 4311-88-0 SDF Download Necrostatin-1 SDF
Smiles CN1C(=O)C(NC1=S)CC2=CNC3=CC=CC=C32
Storage (From the date of receipt)

In vitro
Batch:

DMSO : 57 mg/mL ( (219.79 mM); Moisture-absorbing DMSO reduces solubility. Please use fresh DMSO.)

Water : Insoluble

Ethanol : Insoluble


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