Cell lines

MCF7-R1 cells

Preparation Method

100,000 MCF7-R1 cells were seeded into each well of a 12-well culture plate, allowed to adhere overnight, and treated with 10 nM Monomethyl Auristatin E (MMAE) or conjugates in the presence of 10 U/mL heparin for 72 h. Then the cells were harvested with a TrypLE Express solution, stained by Annexin V-FITC and propidium iodide, and analyzed by flow cytometry using a NovoCyte 2060R flow cytometer.

Reaction Conditions

10 nM; for 72 h

Applications

After 72 h of treatment of MCF7-R1 cells with 10 nM conjugates, the cells were mainly in the early stage of apoptosis.

Animal models

Mice

Preparation Method

The breast tumor models were prepared via subcutaneous injection of 1 × 106 4T1 cells. When the tumor volumes were approximately 100 mm3, mice were divided into three groups: (i) saline; (ii) Monomethyl Auristatin E (MMAE) (0.2 mg/kg); and (iii) FRRG-MMAE nanoparticles (equivalent dose of 0.2 mg/kg based on MMAE contents). The mice were treated once every three days, and tumor volumes were calculated as the smallest diameter 2 × largest diameter × 0.53. The mice with a tumor volume of 2000 mm3 or larger were counted as dead. The toxicity study of FRRG-MMAE nanoparticles was assessed via histology.

Dosage form

0.2 mg/kg

Applications

The results demonstrated that free MMAE-treated mice showed rapid tumor growth compared to FRRG-MMAE-treated mice during monitoring for 15 days.Finally, mice treated with free MMAE showed significant body weight loss owing to the severe toxicity; eventually, all the mice were dead within five days of treatment.These results show the significantly reduced MMAE-related toxicity in the FRRG-MMAE nanoparticle group.

Monomethyl Auristatin E (MMAE), as a synthetic derivative of dolastatin 10, a linear pentapeptide originally isolated from the extracts of the sea hare Dolabella auriculari. Monomethyl Auristatin E, as a synthetic derivative of dolastatin 10, a linear pentapeptide originally isolated from the extracts of the sea hare Dolabella auriculari. Monomethyl Auristatin E can inhibit tubulin polymerization, thus blocking mitosis.^[1]

In vitro, Monomethyl Auristatin E and Monomethyl Auristatin E-phosphate shown the inhibition with IC50 of 2 and 48 nM, respectively, in PC-3 and C4-2B cell lines.^[1]In vitro experiment it shown that 5 nM MMAE resulted in 50% of HCT-116 cells blocked in G2/M and 2 nM in PANC-1 cells.^[2]MMAE shown the inhibition of cell growth with IC50 of 1.7 nM in HCT-116, 0.6 nM in PANC-1, and 5.6 nM in 779E cells, respectively.^[2]In addition, MMAE also showed markedly polarized transport in both MDCK-WT and MDCK-MDR1 cells with ERs at 13.6 and 44.5, respectively, resulting ratio of ratios of 3.3. MMAE shown dose-dependent cytotoxicity in HepG2, Hep3B2, H226, N87 or OVCAR3 cells.^[3]

In vivo, treatment with 30 mg/kg cAC10-vcMMAE in mice had no signs of toxicity.^[4]In vivo efficacy test it shown that treatment with 6.5 mg/kg DD1-MMAE (the bivalent DARPin dimer) or DFc-MMAE (a DARPin-Fc) twice weekly did not cause any sequela in mice.^[5]Moreover, nude mice were injected intravenously 2.5, 5, and 10 mg/kg hertuzumab-vcMMAE on day 0 resulted in obvious and sustained antitumor effects.^[6]

Abawi A, et al. Monomethyl Auristatin E Grafted-Liposomes to Target Prostate Tumor Cell Lines. Int J Mol Sci. 2021 Apr 15;22(8):4103.

Buckel L, et al. Tumor radiosensitization by monomethyl auristatin E: mechanism of action and targeted delivery. Cancer Res. 2015 Apr 1;75(7):1376-1387.

Liu-Kreyche P, et al. Lysosomal P-gp-MDR1 Confers Drug Resistance of Brentuximab Vedotin and Its Cytotoxic Payload Monomethyl Auristatin E in Tumor Cells. Front Pharmacol. 2019 Jul 17;10:749.

Francisco JA, et al. cAC10-vcMMAE, an anti-CD30-monomethyl auristatin E conjugate with potent and selective antitumor activity. Blood. 2003 Aug 15;102(4):1458-65.

Karsten L, et al. Bivalent EGFR-Targeting DARPin-MMAE Conjugates. Int J Mol Sci. 2022 Feb 23;23(5):2468.

Li H, et al. An anti-HER2 antibody conjugated with monomethyl auristatin E is highly effective in HER2-positive human gastric cancer. Cancer Biol Ther. 2016 Apr 2;17(4):346-54.

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