Labeling internalizing anti-epidermal growth factor receptor variant III monoclonal antibody with ¹⁷⁷Lu: in vitro comparison of acyclic and macrocyclic ligands☆

Abstract 

Introduction

The monoclonal antibody (mAb) L8A4, reactive with the epidermal growth factor receptor variant III (EGFRvIII), internalizes rapidly in glioma cells after receptor binding. Combining this tumor-specific mAb with the low-energy β-emitter ¹⁷⁷Lu would be an attractive approach for brain tumor radioimmunotherapy, provided that trapping of the radionuclide in tumor cells after mAb intracellular processing could be maximized.

Materials and Methods

L8A4 mAb was labeled with ¹⁷⁷Lu using the acyclic ligands [(R)-2-amino-3-(4-isothiocyanatophenyl)propyl]-trans-(S,S)-cyclohexane-1,2-diamine-pentaacetic acid (CHX-A″-DTPA), 2-(4-isothiocyanatobenzyl)-diethylenetriaminepenta-acetic acid (pSCN-Bz-DTPA) and 2-(4-isothiocyanatobenzyl)-6-methyldiethylenetriaminepentaacetic acid (1B4M-DTPA), and the macrocyclic ligands S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-tetraacetic acid (C-DOTA) and α-(5-isothiocyanato-2-methoxyphenyl)-1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (MeO-DOTA). Paired-label internalization and cellular processing assays were performed on EGFRvIII-expressing U87.ΔEGFR glioma cells over 24 h to directly compare ¹⁷⁷Lu-labeled L8A4 to L8A4 labeled with ¹²⁵I using either iodogen or N-succinimidyl 4-guanidinomethyl-3-[¹²⁵I]iodobenzoate ([¹²⁵I]SGMIB). In order to facilitate comparison of labeling methods, the primary parameter evaluated was the ratio of ¹⁷⁷Lu to ¹²⁵I activity retained in U87.ΔEGFR cells.

Results

All chelates demonstrated higher retention of internalized activity compared with mAb labeled using iodogen, with ¹⁷⁷Lu/¹²⁵I ratios of >20 observed for the three DTPA chelates at 24 h. When compared to L8A4 labeled using SGMIB, except for MeO-DOTA, internalized activity for ¹²⁵I was higher than ¹⁷⁷Lu from 1-8 h with the opposite behavior observed thereafter. At 24 h, ¹⁷⁷Lu/¹²⁵I ratios were between 1.5 and 3, with higher values observed for the three DTPA chelates.

Conclusions

The nature of the chelate used to label this internalizing mAb with ¹⁷⁷Lu influenced intracellular retention in vitro, although at early time points, only MeO-DOTA provided more favorable results than radioiodination of the mAb via SGMIB.

Keywords: Lu-177, Radioimmunotherapy, Epidermal growth factor receptor, DOTA, DTPA