Abstract 2769: PET imaging shows dose-dependent pharmacokinetics of a 89Zr-labeled mesothelin/CD3 half-life extended bispecific T-cell engager molecule in a syngeneic mouse model
Authors: Suurs Frans V. , Lorenczewski Grit , Bailis Julie M. , Stienen Sabine , Friedrich Matthias , de Vries Elisabeth G.E. , de Groot Derk Jan A. , Lub-de Hooge Marjolijn N. 
Published: September 2020

BiTE® (bispecific T cell engager) immune therapy consists of two connected single-chain variable fragments, one targeting T-cells via CD3, and one targeting the tumor via a tumor-associated antigen. Given their short half-life, these molecules are administered by continuous intravenous (iv) infusion to ensure tumor accumulation. To extend plasma half-life, they are conjugated to an Fc domain, resulting in a molecule with an extended pharmacokinetic profile. To evaluate their biodistribution in a syngeneic mouse model, a molecule targeting murine mesothelin and murine CD3 (muMSLN HLE BiTE®), and a non-targeting control HLE BiTE® molecule were radiolabeled with the positron emission tomography (PET) isotope zirconium-89 (89Zr).

METHODS: The control and muMSLN HLE BiTE® were labeled with 89Zr. Immunocompetent BALB/c mice were engrafted in the lower mammary fat pad with mesothelin-positive murine mammary carcinoma cell line 4T1. PET imaging was performed at 1, 3, 5, 7 and 9 days after 50 µg 89Zr-muMSLN HLE BiTE® was administered by iv infusion (n = 8). Next, biodistribution of 10 µg (n = 6), 50 µg (n = 6) and 200 µg 89Zr-muMSLN HLE BiTE® (n = 5) and 50 µg 89Zr-control HLE BiTE® (n = 6) was compared via PET imaging on day 1, 3 and 5. PET-scans were quantified by mean standardized uptake value (SUVmean). Tracer injections were 4 MBq. To compare uptake in multiple groups, an analysis of variance followed by a post-hoc Tukey's multiple comparison test was performed. Values are expressed as mean ± standard deviation.

RESULTS: PET imaging following 50 µg 89Zr-muMSLN HLE BiTE® dosed iv revealed a blood elimination half-life of 63.4 hours. Uptake in tumor, spleen, thymus and liver were visible from 3 days post injection (pi). On day 5, tumor uptake was highest (SUVmean = 1.50 ± 0.2) with a tumor to blood ratio of 1.9 ± 0.3 and a spleen to blood of 1.7 ± 0.2. Heart SUVmean 5 days pi revealed that 10 μg 89Zr-muMSLN HLE cleared faster from the blood (0.5 ± 0.1) than the 50 μg (0.8 ± 0.1, P &lt; 0.01) and the 200 μg dose group (0.8 ± 0.1, P &lt; 0.01). Tumor SUVmean was higher in 50 µg (1.5 ± 0.2) than the 10 µg dose group (1.2 ± 0.1, P &lt; 0.01), but similar to the 200 μg dose group (1.3 ± 0.1, P = 0.28). Spleen uptake 5 days pi was dose-dependent (SUVmean: 10 µg = 1.6 ± 0.2; 50 µg = 1.3 ± 0.1; 200 µg = 0.8 ± 0.1, P &lt; 0.01). Spleen and tumor SUVmean 5 days pi of 50 µg 89Zr-muMSLN HLE BiTE® were higher than 50 µg 89Zr-control HLE BiTE® (spleen = 0.5 ± 0.1, P &lt; 0.01; tumor = 0.8 ± 0.1, P &lt; 0.01), while blood SUVmean of 50 µg 89Zr-control HLE BiTE® was similar (0.6 ± 0.1, P = 0.12).

CONCLUSION: PET imaging with a 89Zr-labeled muMSLN HLE BiTE® in a tumor-bearing syngeneic mouse model revealed a long blood half-life, specific uptake in tumor and spleen, and dose-dependent pharmacokinetics. Taken together, the HLE BiTE® molecule demonstrates an extended pharmacokinetic profile over previously evaluated small, canonical, BiTE® molecules.

Citation Format: Frans V. Suurs, Grit Lorenczewski, Julie M. Bailis, Sabine Stienen, Matthias Friedrich, Elisabeth G.E. de Vries, Derk Jan A. de Groot, Marjolijn N. Lub-de Hooge. PET imaging shows dose-dependent pharmacokinetics of a 89Zr-labeled mesothelin/CD3 half-life extended bispecific T-cell engager molecule in a syngeneic mouse model [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2769.