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Preclinical Evaluation of 111 In-Labeled PEGylated Maytansine Nimotuzumab Drug Conjugates in EGFR-Positive Cancer Models

2019; Society of Nuclear Medicine and Molecular Imaging; Volume: 60; Issue: 8 Linguagem: Inglês

10.2967/jnumed.118.220095

1535-5667

Siddesh V. Hartimath, Elahe Alizadeh, V. Raja Solomon, Rufael Chekol, Wendy Bernhard, Wayne Hill, Ángel Casacó Parada, Kris Barreto, C. Ronald Geyer, Humphrey Fonge,

Monoclonal and Polyclonal Antibodies Research

Epidermal growth factor receptor I (EGFR) is overexpressed in most cancers of epithelial origin. Antibody drug conjugates (ADCs) with PEGylated-maytansine (PEG-DM1) show promise in vitro and in vivo. However, in vivo biodistribution data for ADCs with PEG-DM1 have not been reported. Development of methods to understand the real-time in vivo behavior of these ADCs is needed to move these compounds to the clinic. Methods: Here we have used noninvasive small-animal SPECT/CT imaging and ex vivo biodistribution to understand the in vivo behavior of PEG 6 -DM1 ADCs. We developed nimotuzumab ADCs conjugated to PEG 6 -DM1. We generated immunoconjugates with low (nimotuzumab-PEG 6 -DM1-Low) and high (nimotuzumab-PEG 6 -DM1-High) drug-to-antibody ratios. The drug-to-antibody of nimotuzumab-PEG 6 -DM1-Low and nimotuzumab-PEG 6 -DM1-High was 3.5 and 7.3, respectively. Quality control was performed using ultraviolet spectrophotometry, size-exclusion high-performance liquid chromatography, bioanalyzer, biolayer interferometry, and flow cytometry in EGFR-positive DLD-1 cells. These immunoconjugates were conjugated with DOTA and radiolabeled with 111 In. The in vitro binding and internalization rates of 111 In-nimotuzumab, 111 In-nimotuzumab-PEG 6 -DM1-Low, and 111 In-nimotuzumab-PEG 6 -DM1-High were characterized. Furthermore, the pharmacokinetics, biodistribution, and imaging characteristics were evaluated in normal and DLD-1 tumor–bearing mice. Results: Flow cytometry and biolayer interferometry showed a trend toward decreasing EGFR affinity with increasing number of PEG 6 -DM1 on the antibody. Despite the lower overall cellular binding of the PEG 6 -DM1 radioimmunoconjugates, internalization was higher for PEG 6 -DM1 ADCs than for the non-PEGylated ADC in the following order: 111 In-nimotuzumab-PEG 6 -DM1-High > 111 In-nimotuzumab-PEG 6 -DM1-Low > 111 In-nimotuzumab. Nuclear uptake of 111 In-nimotuzumab-PEG 6 -DM1-High was 4.4-fold higher than 111 In-nimotuzumab. Pharmacokinetics and biodistribution showed that 111 In-nimotuzumab-PEG 6 -DM1-High had the slowest blood and whole-body clearance rate. Uptake in DLD-1 tumors of 111 In-nimotuzumab was similar to 111 In-nimotuzumab-PEG 6 -DM1-Low but was significantly higher than for 111 In-nimotuzumab-PEG 6 -DM1-High. Tumor-to-background ratios for 111 In-nimotuzumab and 111 In-nimotuzumab-PEG 6 -DM1-Low were higher than for 111 In-nimotuzumab-PEG 6 -DM1-High. Conclusion: The results show that conjugation of multiple PEG 6 -DM1 reduces the affinity for EGFR in vitro. However, the reduced affinity is counteracted by the high internalization rate of constructs with PEG 6 -DM1 ADCs in vitro. The decreased affinity resulted in low tumor uptake of 111 In-nimotuzumab-PEG 6 -DM1-High, with a slow overall whole-body clearance rate. These data provide insights for evaluating the pharmacokinetics and normal -tissue toxicity and in determining dosing rate of PEGylated ADCs.