Fig 2: Generation of free drug in cells treated with anti-CD30-MCC-DM1. Intracellular (a) and extracellular (b) concentrations of Lys-MCC-DM1 from 0 to 72 h of exposure with anti-CD30-MCC-DM1 small molecule were determined with an LC-MS/MS method in CD30-positive (Karpas 299, HH, and L428) and CD30-negative (Ramos) cell cultures. Results for each test are plotted as the mean (± SEM).

Fig 3: Bystander activities of ADCs. CD30-positive and CD30-negative cells were co-cultured in dual chamber Transwell dishes. The CD30-positive cells (Karpas 299 cells, HH cells and L428 cells) were plated into the upper Transwell chamber, and CD30-negative Ramos cells were plated into the lower Transwell chamber. Cocultures were either untreated or treated with anti-CD30-MCC-DM1 or ADCETRIS. Counting represents the number of surviving CD30-positive and CD30-negative cells following 72, 96, and 120 h treatment. Results for each test are plotted as the mean (± SEM).

Fig 4: Binding and internalization of anti-CD30-MCC-DM1. a, Competition binding of anti-CD30-MCC-DM1 to CD30-positive cells. Karpas 299, HH and L428 cells were combined with biotinylated anti-CD30 mAb and serial dilutions of either anti-CD30 mAb or anti-CD30-MCC-DM1. The normalized fluorescence intensities were plotted versus mAb concentration as described in the “Materials and methods.” b, internalization of anti-CD30-MCC-DM1 into CD30-positive cells. pHAb dye-conjugated anti-CD30 mAb and anti-CD30-MCC-DM1 were added to the Karpas 299, HH, and L428 cells and incubated for various time durations. Mean and standard deviations from triplicate readings are plotted.

Fig 5: Pharmacokinetics of anti-CD30-MCC-DM1 in cynomolgus monkeys. Groups of cynomolgus monkeys (n = 8) were treated with a single dose of 1, 4, or 12 mg/kg anti-CD30-MCC-DM1 or 4 mg/kg anti-CD30 mAb mixed with 0.07 mg/kg DM1. Concentrations of total and conjugated antibody (a) as well as of DM1 (b) in plasma were analyzed by ELISA or LC-MS/MS and plotted as the mean (± SD).