Figure 3.

In vitro cytotoxicity of anti-HER2 ADCs against HER2-expressing cancer cell lines. (a) For high copy number HER2-expressing cells, both anti-HER2 DAR = 8 MMAU ADC and DAR = 2 duostatin-3 ADC showed high cytotoxic activity, while both anti-HER2 naked antibody and an isotype IgG control antibody-MMAU conjugate showed only no or low cytotoxicity, respectively. All experiments were performed in triplicate except for anti-HER2 naked antibody; (b) For low copy number HER2-expressing cells, the anti-HER2 DAR = 8 MMAU ADC had superior activity compared with DAR = 2 duostatin-3 ADC. Both experiments were performed in triplicate; (c) To demonstrate bystander kill activity of MMAU ADCs, non-HER2-expressing Jurkat cells were co-cultured with HCC1954 cells that express HER2, and viability of Jurkat cells was measured after incubation with fluorescence-assisted cell sorting (FACS)-based assay. The anti-HER2 DAR = 8 MMAU ADC showed bystander kill activity, while neither the anti-HER2 DAR = 2 duostatin-3 ADC nor the free MMAU payload had cytotoxic activity towards Jurkat cells. The results are from a single representative experiment. IC₅₀ values are marked in the figure panels and they were determined using curve fitting by nonlinear regression as the concentration of the drug that causes 50% inhibition of cell viability compared to maximum inhibition. Error bars show the standard error of the mean, where applicable.