| Size | Price | Stock | Qty |
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| 100mg |
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| 500mg |
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| 1g |
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| Targets |
METTL16 (Methyltransferase-like protein 16). METTL16 is an RNA methyltransferase that methylates adenosine residues in RNA molecules, particularly U6 snRNA at position 47. METTL16-IN-1 inhibits the activity of METTL16.
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| ln Vitro |
In cell-free assays, METTL16-IN-1 inhibits METTL16 methyltransferase activity with an IC₅0 of 1.7 microM and a Kd of 1.35 microM. It also suppresses the binding interaction between U6 snRNA deletion and the METTL16 methyltransferase domain (MTD) with an IC₅0 of 2.5 microM. METTL16-IN-1 has antitumor activity, indicating its potential as a research tool for studying the role of METTL16 in cancer.
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| Enzyme Assay |
A general cell-free protocol for assessing METTL16 methyltransferase activity: A radiometric or fluorescence-based methyltransferase assay is used. Recombinant METTL16 protein (100 nM) is incubated with a 3′-biotinylated U6 snRNA substrate (1 uM) in assay buffer (50 mM Tris-HCl, pH 8.0, 150 mM NaCl, 1 mM DTT, 0.01% Tween-20) containing 1 uM S-adenosyl-L-methionine (SAM). Varying concentrations of METTL16-IN-1 (0.01 nM to 100 uM) are added. The reaction is incubated at 30degC for 60 minutes. The reaction is stopped by heating to 65degC for 5 minutes. The biotinylated RNA substrate is captured on streptavidin-coated plates. The methylated RNA is detected using an anti-methylated adenosine (m⁶A) antibody conjugated to HRP, followed by the addition of a chemiluminescent or colorimetric substrate. The IC₅0 is calculated from the inhibition curve. For Kd determination, a surface plasmon resonance (SPR) assay is used. METTL16 protein is immobilized on a sensor chip, and varying concentrations of METTL16-IN-1 are flowed over the chip. The association and dissociation rates are measured to calculate the Kd. For the U6 snRNA binding inhibition assay, a fluorescence polarization (FP) assay is used. The METTL16 MTD (methyltransferase domain) protein is incubated with a fluorescently labeled U6 snRNA probe (U6 snRNA deletion mutant). The increase in FP signal upon binding is measured. Varying concentrations of METTL16-IN-1 are added to compete for binding, and the IC₅0 is calculated.
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| Cell Assay |
A general cellular protocol for assessing METTL16 inhibition and m⁶A levels: Cancer cells (e.g., HeLa, HEK293T, or other METTL16-expressing cancer cell lines) are seeded in 6-well plates at 2×10⁵ cells/well. After 24 hours, the cells are treated with various concentrations of METTL16-IN-1 (0.1, 0.5, 1, 2.5, 5, 10, 25, 50 uM) for 48 hours. Total RNA is extracted from the treated cells using TRIzol reagent. For m⁶A quantification, the RNA is denatured and added to the wells of an m⁶A ELISA kit. The m⁶A level in the RNA samples is measured according to the manufacturer's instructions. For qRT-PCR analysis of METTL16 target genes, RNA is reverse transcribed to cDNA, and the expression of METTL16 target genes (e.g., MAT2A) is measured using TaqMan or SYBR Green qPCR. GAPDH or beta-actin is used as a loading control. For cell viability assays, cells are seeded in 96-well plates and treated with METTL16-IN-1 for 72 hours. Cell viability is assessed using the MTT or CellTiter-Glo assay. Apoptosis is assessed by flow cytometry using Annexin V-FITC/PI staining.
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| Animal Protocol |
A general animal protocol for evaluating the antitumor activity of METTL16-IN-1 in a xenograft model: Female BALB/c nude mice are subcutaneously injected with 5×10⁶ METTL16-expressing cancer cells (e.g., HeLa, HCT116, or A549 cells) in 0.1 mL of PBS mixed with Matrigel (1:1). When the average tumor volume reaches 100-150 mm3, the mice are randomized into treatment groups (n=8-10 per group). METTL16-IN-1 is formulated in a suitable vehicle (e.g., 10% DMSO, 40% PEG300, 5% Tween-80, 45% saline) and administered via oral gavage or intraperitoneal injection at doses of 10, 30, and 100 mg/kg once daily for 14-21 days. A vehicle control group is included. Tumor volume is measured twice weekly using a caliper, and body weight is recorded. At the end of the study, the mice are euthanized, and tumors are excised and weighed. Total RNA is extracted from the tumor tissues, and m⁶A levels are measured by ELISA. The expression of METTL16 target genes is measured by qRT-PCR. Tumor tissues are also processed for immunohistochemistry (Ki-67 and cleaved caspase-3).
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| ADME/Pharmacokinetics |
General pharmacokinetic protocol for METTL16-IN-1: Male Sprague-Dawley rats are administered METTL16-IN-1 via oral gavage (PO, 10 mg/kg) and intravenous (IV, 2 mg/kg) injection. The compound is formulated in a vehicle such as 10% DMSO, 40% PEG300, 5% Tween-80, 45% saline. Blood samples are collected at 0.083, 0.25, 0.5, 1, 2, 4, 8, 12, and 24 hours post-dose. Plasma concentrations of METTL16-IN-1 are quantified by LC-MS/MS. PK parameters (Cmax, Tmax, AUC, t½, clearance, Vd, and oral bioavailability) are calculated using non-compartmental analysis. METTL16-IN-1 has a molecular formula of C1₉H12BrN3O₆S2 and a molecular weight that can be calculated from this formula.
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| Toxicity/Toxicokinetics |
General toxicity protocol for METTL16-IN-1: A 14-day repeated-dose oral toxicity study is performed in ICR mice. METTL16-IN-1 is administered via oral gavage at doses of 30, 100, and 300 mg/kg/day for 14 consecutive days. Clinical signs, body weight, and food consumption are monitored daily. At the end of the study, blood samples are collected for hematology (complete blood count with differential) and serum chemistry (ALT, AST, ALP, BUN, creatinine, total protein, albumin, glucose). Gross necropsy is performed, and the weights of major organs (liver, kidney, spleen, heart, lung, brain, testes) are recorded. Histopathological examination of these organs is conducted. Because METTL16 is involved in RNA methylation, special attention is given to tissues with high proliferative activity, such as the bone marrow and gastrointestinal tract, for potential myelosuppression or gastrointestinal toxicity.
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| References | |
| Additional Infomation |
METTL16-IN-1 (Compound 45) is a potent inhibitor of the RNA methyltransferase METTL16. The molecular formula is C1₉H12BrN3O₆S2, which includes a bromine atom. The compound has an IC₅0 of 1.7 microM for METTL16 inhibition, a Kd of 1.35 microM, and inhibits the U6 snRNA-METTL16 interaction with an IC₅0 of 2.5 microM. METTL16-IN-1 has antitumor activity and is a valuable research tool for studying the role of METTL16 and RNA methylation (m⁶A) in cancer biology. METTL16 is emerging as a potential therapeutic target in various malignancies, including acute myeloid leukemia (AML) and hepatocellular carcinoma (HCC).
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| Molecular Formula |
C19H12BRN3O6S2
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| Molecular Weight |
522.35
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| Appearance |
Brown to reddish brown 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 |
| 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) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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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 | 1.9144 mL | 9.5721 mL | 19.1443 mL | |
| 5 mM | 0.3829 mL | 1.9144 mL | 3.8289 mL | |
| 10 mM | 0.1914 mL | 0.9572 mL | 1.9144 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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