3-Methyladenine (3-MA; Autophagy Inhibitor)

Alias: 3-Methyladenine, NSC-66389; NSC66389; NSC 66389
Cat No.:V0117 Purity: ≥98%
3-Methyladenine (also called 3-MA; Autophagy Inhibitor) is a novel, potent,cell-permeable and selective PI3K inhibitor.
3-Methyladenine (3-MA; Autophagy Inhibitor) Chemical Structure CAS No.: 5142-23-4
Product category: PI3K
This product is for research use only, not for human use. We do not sell to patients.
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Other Forms of 3-Methyladenine (3-MA; Autophagy Inhibitor):

  • 3-Methyladenine-d3 (3-methyladenine-d3)
Official Supplier of:
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Purity & Quality Control Documentation

Purity: ≥98%

Purity: ≥98%

Product Description

3-Methyladenine (also called 3-MA; Autophagy Inhibitor) is a novel, potent, cell-permeable and selective PI3K inhibitor. In HeLa cells, it has an inhibitory concentration (IC50) of 25 μM for Vps34 and 60 μM for PI3Kγ . It is also a class I PI3K inhibitor, which prevents cerebellar granule cells from apoptosis after serum/potassium deprivation, and thus blocks autophagic sequestration.

Biological Activity I Assay Protocols (From Reference)
Targets
PtdIns3Kγ (IC50 = 60 μM); Vps34 (IC50 = 25 μM); Autophagy; Mitophagy; Human Endogenous Metabolite
ln Vitro
The slight preference for Vps34 prevention by 3-Methyladenine probably arises from a hydrophobic ring specific to Vps34, which encircles the 3-methyl group of 3-Methyladenine. [1] Under both normal and starvation conditions, 3-Methyladenine has been shown to kill cancer cells. In addition to being able to inhibit autophagy, 3-methyladedine has the ability to inhibit cell invasion and migration, suggesting that it serves purposes aside from just inhibiting autophagy. Independent of the inhibition of autophagy, 3-methylladenine causes caspase-dependent cell death. When given 5 mM 3-Methyladenine, only 23% of glucose-starved HeLa cells exhibit GFP-LC3 puncta. In cells that have received 3-Methyladenine treatment, LC3-I levels rise while LC3-II levels fall between 12 and 48 hours. 3-Methyladenine blocks the transformation of LC3-I into LC3-II. The viability of HeLa cells treated for a day with 3-Methyladenine at 2.5 mM or 5 mM is unaffected, whereas the viability of the cells is reduced by 25.0% when treated for a day with 3-Methyladenine at 10 mM. Cell viability decreases by 11.5%, 38.0%, and 79.4%, respectively, after 2.5, 5 or 10 mM 3-Methyladenine has been added to the cells for two days. In a dose- and time-dependent manner, 3-Methyladenine reduces cell viability. 3-Methyladenine decreases cell viability in a time- and dose-dependent manner. 3-Methyladenine significantly shortens the duration of nocodazole-induced-prometaphase arrest. [2] 3-Methyladenine prevents SU11274-induced cell death by suppressing autophagy. [3] Significant LC3 I to II conversion is induced in wild type MEFs by prolonged treatment with 3-Methyladenine (up to 9 hours). But not wortmannin, prolonged 3-Methyladenine treatment significantly increases GFP-LC3 punctation/aggregation. 3-Methyladenine-induced LC3 conversion and free GFP liberation are ATG7-dependent. 3-Methyladenine treatment leads to evident increase of p62 protein level. 3-Methyladenine increases the p62 level even in Atg5−/− MEFs as well as in cells with DOX-mediated deletion of ATG5. 3-Methyladenine inhibits class I and class III PI3K in different temporal patterns. 3-Methyladenine-induced LC3 I to LC3 II conversion is dramatically compromised in Tsc2−/− cells compared with wild type cells. The mTOR complex 1's ability to inhibit autophagy is compromised by 3-methyladenine.[4]
ln Vivo
3-Methyladenine blocks autophagy through its effect on class III phosphatidylinositol 3-kinase (PI3K). 3-Methyladenine treatment does not alter the degree of hemorrhage compared with the subarachnoid hemorrhage (SAH) group. 3-Methyladenine pretreatment significantly aggravates neurological symptoms when compared with the SAH + vehicle group. When 3-Methyladenine is administered, autophagy is decreased. On the other hand, the SAH + 3-Methyladenine group significantly upregulates cleaved caspase-3. The number of TUNEL-positive cells in the right cortex is significantly higher in the SAH + 3-Methyladenine group than the SAH + vehicle group, which is consistent with the upregulation of cleaved caspase-3 expression.[5]
Enzyme Assay
HeLa cells are radiolabeled for 24 hours with 0.05 mCi/mL l-[U- 14C]valine. Cells are three times rinsed with PBS after the labeling period is over. With or without the addition of 10 mM 3-Methyladenine, cells are incubated for the specified periods of time in either full medium or EBSS.
Cell Assay
Cell (such as HeLa cell) viability is determined by a trypan blue exclusion assay. Briefly, adherent and floating cells are gathered and suspended in phosphate buffered saline (PBS, pH 7.4) at a final density of 1-2 × 106/mL after being treated with 3-Methyladenine. The cell suspension is added to an equal volume of 0.4% trypan blue solution (w/v, in PBS), and everything is thoroughly mixed. A hemacytometer is used to count the cells after 3 minutes of incubation at room temperature.
Animal Protocol
Adult male Sprague–Dawley rats weighing 300-350 g
400 nM
Intracerebral ventricular
References

[1]. Autophagy. 2010 Aug;6(6):805-7.

[2]. PLoS One. 2012;7(4):e35665.

[3]. J Pharmacol Sci. 2012;118(4):423-32.

These protocols are for reference only. InvivoChem does not independently validate these methods.
Physicochemical Properties
Molecular Formula
C6H7N5
Molecular Weight
149.1533
Exact Mass
149.0701
Elemental Analysis
C, 48.32; H, 4.73; N, 46.95
CAS #
5142-23-4
Appearance
Solid powder
SMILES
CN1C=NC(=N)C2=C1N=CN2
InChi Key
ZPBYVFQJHWLTFB-UHFFFAOYSA-N
InChi Code
InChI=1S/C6H7N5/c1-11-3-10-5(7)4-6(11)9-2-8-4/h2-3,7H,1H3,(H,8,9)
Chemical Name
3-methyl-7H-purin-6-imine
Synonyms
3-Methyladenine, NSC-66389; NSC66389; NSC 66389
HS Tariff Code
2934.99.9001
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)
Solubility Data
Solubility (In Vitro)
DMSO: ~3 mg/mL warming (~20.1 mM)
Water: ~20 mg/mL warming (~134.1 mM)
Ethanol: ~4 mg/mL (~26.8 mM)
Solubility (In Vivo)
5%DMSO+40%PEG300+5%Tween80+50%ddH2O: 10mg/ml (Please use freshly prepared in vivo formulations for optimal results.)
Preparing Stock Solutions 1 mg 5 mg 10 mg
1 mM 6.7047 mL 33.5233 mL 67.0466 mL
5 mM 1.3409 mL 6.7047 mL 13.4093 mL
10 mM 0.6705 mL 3.3523 mL 6.7047 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.

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Note: Chemical formula is case sensitive: C12H18N3O4  c12h18n3o4
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In vivo Formulation Calculator (Clear solution)
Step 1: Enter information below (Recommended: An additional animal to make allowance for loss during the experiment)
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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.
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Biological Data
  • 3-Methyladenine (3-MA)

    Prolonged treatment with 3-MA in full medium leads to accumulation of autophagic markers.J Biol Chem.2010 Apr 2;285(14):10850-61.

  • 3-Methyladenine (3-MA)

    3-MA increases autophagic flux.J Biol Chem.2010 Apr 2;285(14):10850-61.

  • 3-Methyladenine (3-MA)

    3-MA does not affect lysosomal functions.J Biol Chem.2010 Apr 2;285(14):10850-61.

  • 3-Methyladenine (3-MA)


    3-MA inhibits class I and class III PI3K in different temporal patterns.J Biol Chem.2010 Apr 2;285(14):10850-61.


  • 3-Methyladenine (3-MA)

    3-MA disrupts the function of mTOR complex I.J Biol Chem.2010 Apr 2;285(14):10850-61.

  • 3-Methyladenine (3-MA)

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