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| Targets |
CDK4 RIPK3 RIPK1
RIPK3 (Receptor-interacting serine/threonine-protein kinase 3) and MLKL (Mixed lineage kinase domain-like pseudokinase). NecroIr1 activates these necroptosis proteins and also regulates CDK4 expression. It can activate RIPK1, RIPK3, and MLKL. |
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| ln Vitro |
In both CFSE-labeled L02 cells and non-labeled A549R cells, NecroIr1 (2 μM; 1-2 d) exhibits subcellular distribution with over 90% accumulation in the mitochondria[1]. The effects of NecroIr1 (1.5 μM and 3 μM; 24 h) include an increase in ROS production and a decrease in mitochondrial membrane potential[1]. NecroIr1 (1.5 μM and 3 μM; 24 h) enhances RIPK1 and RIPK3 phosphorylation and activates necroptosis proteins[1]. NecroIr1 (0.75 μM and 1.5 μM; 24 h) arrests the cell cycle in G0/G1 to cause necroptosis[1]. NecroIr1 (0.75-2.0 μM; 24 h) suppresses the growth of A549R cells[1].
In A549R cells, NecroIr1 (1.5 microM; 24 h) increases ROS generation and MMP loss, and increases phosphorylation of RIPK1 and RIPK3. It (0.75 microM; 24 h) arrests the cell cycle at G0/G1 to induce necroptosis and inhibits cell proliferation. At 2 microM, it accumulates over 90% in mitochondria. |
| ln Vivo |
NecroIr1 is expected to show anti-tumor efficacy in murine xenograft models bearing cisplatin-resistant lung tumors. It would likely be administered intravenously or intraperitoneally. Efficacy would be measured as tumor growth inhibition (TGI), with pharmacodynamic studies confirming the induction of necroptosis and modulation of CDK4 expression in tumor tissue.
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| Enzyme Assay |
Non-cellular assays would assess the direct binding or inhibition of purified MLKL or RIPK3 by NecroIr1. A typical experimental setup involves incubating the compound with the purified kinase domain of RIPK3 in a reaction buffer containing ATP and a specific substrate peptide. The reaction is stopped, and the level of phosphorylated substrate is measured using a time-resolved fluorescence resonance energy transfer (TR-FRET) or luminescence-based detection system to calculate the IC50.
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| Cell Assay |
Western Blot Analysis[1]
Cell Types: A549R cells Tested Concentrations: 1.5 μM and 3 μM Incubation Duration: 24 hrs (hours) Experimental Results: Increased phospho-RIPK1(p-PIPK1), total RIPK3, and phospho-RIPK3(p-PIPK3) level. Cell Cycle Analysis[1] Cell Types: A549R cells Tested Concentrations: 0 μM, 0.75 μM and 1.5 μM Incubation Duration: 24 hrs (hours) Experimental Results: Arrested cell cycle at G0/G1 phase in a dose-dependent manner. Cell Proliferation Assay[1] Cell Types: A549R cells Tested Concentrations: 0 μM, 0.75 μM, 1.5 μM and 3.0 μM Incubation Duration: 24 hrs (hours) Experimental Results: Inhibits cell proliferation in a dose-dependent manner. In vitro cell-based assays are performed using A549R (Cisplatin-resistant) cells. Cells are treated with NecroIr1 (0.75-3 microM; 24 h). Cell viability is assessed using MTT or CellTiter-Glo assays. To evaluate subcellular localization, cells are stained with MitoTracker dye and NecroIr1 fluorescence is visualized via confocal microscopy. Flow cytometry with JC-1 dye measures MMP loss. Western blotting is used to detect necroptosis markers (p-RIPK1, p-RIPK3, MLKL oligomerization) and cell cycle regulators. |
| Animal Protocol |
In vivo animal studies typically use a subcutaneous xenograft model in nude mice. A549R cells are injected into the flank to establish tumors. Once tumors reach a suitable size (e.g., ~100 mm3), animals are randomized and treated intravenously or intraperitoneally with NecroIr1. Tumor dimensions are measured with calipers every 2-3 days. At the study endpoint, tumors are harvested for ex vivo analysis, including histology (H&E), IHC (for CDK4, RIPK3, MLKL), and Western blotting.
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| ADME/Pharmacokinetics |
Standard pharmacokinetic parameters (half-life, clearance, volume of distribution) for this iridium(III) complex are not extensively cataloged. Following intravenous administration, metal-based complexes like NecroIr1 may exhibit a biphasic clearance profile, with an initial distribution phase and a prolonged terminal elimination phase due to strong protein or tissue binding.
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| Toxicity/Toxicokinetics |
Detailed preclinical toxicity data, such as the maximum tolerated dose (MTD) or no-observed-adverse-effect level (NOAEL), are not available in standard public databases. As an iridium-based complex that induces oxidative stress, standard safety precautions for handling potential cytotoxic compounds should be observed.
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| References | |
| Additional Infomation |
NecroIr1 is specifically designed to induce necroptosis in drug-resistant cancer cells, providing a research tool to bypass apoptosis resistance. Its ability to activate RIPK1/RIPK3/MLKL while regulating CDK4 places it at the intersection of cell death and cell cycle control, making it useful for studying the molecular crosstalk between these pathways. Along with NecroIr2, it represents a novel class of metal-based necroptosis inducers.
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| Molecular Formula |
C40H29CLIRN5O-
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| Molecular Weight |
823.36
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| Appearance |
Orange to red solid powder
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment, avoid exposure to moisture. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
DMSO :~100 mg/mL (~121.45 mM)
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 1.2145 mL | 6.0727 mL | 12.1454 mL | |
| 5 mM | 0.2429 mL | 1.2145 mL | 2.4291 mL | |
| 10 mM | 0.1215 mL | 0.6073 mL | 1.2145 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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