Imaging xCT with Immuno-PET in Therapy-Resistant Non–Small Cell Lung Cancer

Therapy resistance presents a major clinical challenge in non–small cell lung cancer (NSCLC), contributing to its high mortality rate. Overexpression of the amino acid transporter xCT frequently mediates treatment resistance in NSCLC and can be therapeutically targeted with antibody–drug conjugates. Here, we developed a corresponding immuno-PET radiotracer to noninvasively evaluate xCT expression in NSCLC and visualize the pharmacokinetics of xCT-targeting antibodies. <b>Methods:</b> The specificity of xCT-targeting antibodies, HM30 and HM34, was assessed by Western blot, immunocytochemistry, and internalization studies. Next, these antibodies were conjugated to deferoxamine via their lysine residues and radiolabeled with ⁸⁹Zr. Immunoreactivity was determined by bead- and cell-based binding assays. Preclinical in&nbsp;vivo pharmacokinetics of the ⁸⁹Zr-labeled antibodies were evaluated through longitudinal imaging studies and ex vivo biodistribution using subcutaneous and orthotopic models of NSCLC. <b>Results:</b> Through multiple biochemical methods, we demonstrated excellent target specificity and rapid antibody internalization. Immuno-PET agents [⁸⁹Zr]Zr-DFO-HM30 and [⁸⁹Zr]Zr-DFO-HM34 were produced with good yield and radiochemical purity, with no impact on immunoreactivity. Both agents clearly delineated high xCT–expressing H460 tumors in&nbsp;vivo. Conversely, [⁸⁹Zr]Zr-DFO-HM30 binding was low in low xCT–expressing H1299 tumors. [⁸⁹Zr]Zr-DFO-HM30 imaging and autoradiography of H460 orthotopic lung tumors revealed lesion-specific binding. <b>Conclusion:</b> Our findings provide robust evidence for the use of immuno-PET as a research tool to image elevated xCT expression and visualize tumor penetration of xCT-directed antibodies. This tool can inform the development of enhanced antibody-based therapeutic candidates against treatment-resistant phenotypes to ultimately improve survival outcomes in patients with NSCLC.