| Size | Price | Stock | Qty |
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| 1mg |
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| Other Sizes |
| Targets |
Auristatin
MMAD targets tubulin, specifically binding to the vinca domain on the beta-subunit of tubulin. This binding prevents the formation of microtubules and disrupts the mitotic spindle apparatus, leading to cell cycle arrest at the G2/M phase. The mc linker is stable in systemic circulation but is designed to be cleavable (cathepsin-sensitive) after ADC internalization into target cancer cells, releasing the active auristatin payload to exert its cytotoxic effects. |
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| ln Vitro |
The unbound MMAD payload demonstrates in vitro cytotoxic activity against a wide range of human cancer cell lines with IC50 values in the low nanomolar to picomolar range (e.g., 0.1-1 nM for ovarian, breast, and lung cancer cells). It disrupts tubulin polymerization and induces G2/M cell cycle arrest, leading to apoptosis. When conjugated via mc linker, the activity is masked until released, minimizing off-target toxicity.
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| ln Vivo |
The unlabeled MMAD (Monomethyl Auristatin D) is a potent inhibitor of xenograft tumor growth in mice when delivered via an ADC platform. When conjugated to a tumor-targeting antibody via the mc linker, Cys-mcMMAD ADCs specifically deliver the payload to tumor cells, resulting in significant tumor regression, often with durable responses. The efficacy is dose-dependent and typically correlates with target antigen expression on cancer cells.
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| Enzyme Assay |
The mechanism of action of the MMAD payload is evaluated using in vitro tubulin polymerization assays. Tubulin (purified from porcine brain, 3 mg/mL) is incubated in GTP-supplemented buffer at 37degC in the presence or absence of MMAD (0.1-100 nM). Assembly of tubulin into microtubules is monitored spectrophotometrically by measuring absorbance at 340 nm (turbidity) over time (30-60 minutes). IC50 values are derived from the inhibition of the maximal polymerization rate. Alternatively, a competition binding assay with [3H]-colchicine or [3H]-vinblastine can determine binding site specificity.
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| Cell Assay |
Cellular cytotoxicity of MMAD or its ADCs is assessed using standard proliferation assays. Target cancer cells (e.g., SK-BR-3 for HER2, NCI-N87 for gastric cancer) are seeded in 96-well plates (5,000 cells/well) and incubated overnight. Cells are treated with serially diluted ADC (0.0001-100 nM) for 72-96 hours. Viability is then quantified using the CellTiter-Glo luminescent assay (measuring ATP) or MTT colorimetric assay. IC50 values are calculated using non-linear regression. Apoptosis is confirmed by flow cytometry using Annexin V/PI double staining after 24-48 hours of treatment.
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| Animal Protocol |
For in vivo efficacy studies, female athymic nude mice bearing subcutaneous xenograft tumors (e.g., HER2-positive breast cancer xenografts) are randomized into treatment groups (n=5-10 per group) once tumors reach a mean volume of 100-200 mm3. ADC prepared using Cys-mcMMAD is administered intravenously via tail vein at various dose levels (e.g., 1, 3, 10 mg/kg) on days 0, 4, and 8 (Q4D x 3). Tumor volume (length × width2/2) and body weight are measured twice weekly. Efficacy endpoints include tumor growth inhibition percentage (%TGI), tumor regression (partial or complete response), and event-free survival. At study termination, tumors and organs are collected for histopathological analysis and payload quantitation.
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| ADME/Pharmacokinetics |
The pharmacokinetic profile of MMAD itself is not favorable (rapid clearance, short half-life). However, when conjugated in the ADC construct via the mc linker, the Cys-mcMMAD-containing ADC exhibits extended circulation typical of antibodies: half-life of 5-10 days in mice and 1-3 weeks in cynomolgus monkeys. The linker is stable in plasma, with less than 5% payload release over 14 days in vitro. Cleavage occurs primarily within lysosomes of target cells. The major elimination pathways are proteolytic degradation of the antibody scaffold and biliary/fecal excretion.
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| Toxicity/Toxicokinetics |
The toxicity of Cys-mcMMAD is primarily attributed to the mechanism-based toxicity of the free MMAD payload released from the ADC. MMAD inhibits tubulin, leading to mitotic arrest, which is toxic to rapidly dividing cells. On-target off-tumor toxicity (e.g., thrombocytopenia, neutropenia) is the dose-limiting toxicity of auristatin-based ADCs. Ocular toxicity (keratopathy) and peripheral neuropathy are also potential adverse effects. With the Cys-mc linker conjugation, systemic exposure to free payload is minimal, but hematological toxicity remains a class effect.
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| References | |
| Additional Infomation |
Cys-mcMMAD is the TFA salt of the drug-linker construct combining Monomethyl Auristatin D (MMAD), a potent microtubule inhibitor derived from dolastatin 10, with a maleimidocaproyl (mc) linker via a cysteine residue. This linker is specifically designed for conjugation to reduced interchain disulfide bond cysteines on monoclonal antibodies. MMAD is a substrate for the drug efflux pump P-glycoprotein (MDR1), which is a potential resistance mechanism in certain cancer cells. Cys-mcMMAD is for research use only and not approved for clinical use as a standalone agent.
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| Molecular Formula |
C54H84N8O11S2
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|---|---|
| Molecular Weight |
1085.42
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| Appearance |
White to off-white 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: (1). This product requires protection from light (avoid light exposure) during transportation and storage. (2). This product is not stable in solution, please use freshly prepared working solution for optimal results. |
| 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 :~200 mg/mL (~184.26 mM)
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| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 5 mg/mL (4.61 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 50.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 5 mg/mL (4.61 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 50.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. View More
Solubility in Formulation 3: ≥ 5 mg/mL (4.61 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution. |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 0.9213 mL | 4.6065 mL | 9.2130 mL | |
| 5 mM | 0.1843 mL | 0.9213 mL | 1.8426 mL | |
| 10 mM | 0.0921 mL | 0.4607 mL | 0.9213 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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