Author: Andrew M Baschnagel, Jonathan W. Engle, Jessica E. Griffin, Gihan Gunaratne, Reinier Hernandez, Randall J Kimple, Aaron M. LeBeau, Natalie Y. Luo, Rachel Minne, Caroline M. Mork, Kwangok P Nickel, Jayden West, Madalynn R. Wopat 👨🔬
Affiliation: University of Wisconsin, University of Wisconsin, Department of Pathology and Laboratory Medicine, Department of Medical Physics, School of Medicine and Public Health, University of Wisconsin–Madison, University of Wisconsin - Department of Human Oncology, UW Madison, University of Wisconsin, Department of Medical Physics 🌍
Purpose: The Mesenchymal-Epithelial Transition gene (MET), a targetable cell surface receptor responsible for stimulating cell growth and proliferation, is altered in approximately 10% of non-small cell lung cancer (NSCLC) cases. Dysregulation of MET commonly occurs through amplification (METamp) or exon 14 skipping mutation (METex14). While FDA-approved inhibitors exist as first-line treatments, variable drug responses and resistance development necessitate new therapeutic approaches. We developed a shark-derived antibody (VNAR 2H4) targeting MET for therapeutic payload delivery.
Methods: After identifying 2H4's cross-species reactivity and high MET affinity, we assessed its targeting capabilities in MET-altered cell lines using flow cytometry and immunofluorescence. Cell-based assays evaluated 2H4-Fc's effects on signaling and viability. In vivo studies using [89Zr]Zr-2H4-Fc assessed tumor uptake via PET/CT imaging and biodistribution analysis. Time-activity curve integration with three compartmental modeling was performed to identify optimal activity of [177Lu]Lu-2H4-Fc to deliver effective dose. Finally, we evaluated [177Lu]Lu-2H4-Fc for tumor targeting and therapeutic efficacy using SPECT imaging, biodistribution analysis and tumor measurements in xenograft models.
Results: As a bivalent human Fc fusion protein, 2H4-Fc was found to selectively bind to and is internalized by both METamp and METex14 NSCLC cell lines. Notably, it did not inhibit downstream signaling nor affect cell viability. [89Zr]Zr-2H4-Fc demonstrated rapid localization and high tumor uptake in both xenografts with a measured %IA/g in the tumor to be 29.9% and 20.0% for METex14 and METamp respectively at 96 hr post injection. [177Lu]Lu-2H4-Fc demonstrated similar pharmacokinetic behavior with rapid attenuation in METamp tumors within 24 hours post injection. Tumor uptake was measured to be approximately 15 %IA/g at 120 hr post injection and delay in tumor growth was observed up to 21 days post injection.
Conclusion: 2H4-Fc, demonstrated high binding affinity and accumulation in MET-altered tumors and minimal surrounding tissue noise and cytotoxicity, highlighting its success as a theranostic agent.