Biodistribution-Driven Discovery Identifies a Glycosidase-Cleavable Linker to Reprogram Radiotheranostics

While radiopharmaceutical therapy (RPT) has become part of the standard-of-care for patients with advanced prostate cancers and neuroendocrine tumors (NETs), cures are elusive and normal tissue toxicity remain a challenge. Chemical groups susceptible to cleavage by enzymes present in tumors, tumor microenvironment or in normal tissues, have the potential to improve the therapeutic index for RPT. Using DOTA-TATE as an example and drawing from strategies used to develop antibody-drug conjugates, we designed, and synthesized, a chemically diverse series of linkers between the chelator (DOTA) and the targeting vector (TATE). Of the 10 agents we tested, two with cleavable linker domains reduced kidney retention compared to DOTA-TATE: the previously reported DOTA-MVK(ε)-TATE, and a novel agent bearing cleavable beta-galactose (β-Gal) unit, DOTA-β-Gal-TATE. In murine models of NETs, positron emission tomography (PET) was used to image yttrium-86 ( ⁸⁶ Y)-labeled variants and show that, while the ⁸⁶ Y-DOTA-MVK(ε)-TATE exhibits similar tumor uptake to the parent non-cleavable ⁸⁶ Y-DOTA-TATE, ⁸⁶ Y-DOTA-β-Gal-TATE shows enhanced tumor uptake, resulting in up to 10-fold improvement in the tumor-to-kidney ratios compared to ⁸⁶ Y-DOTA-TATE. In vitro and in vivo studies confirm high efficiency, enzyme-specific cleavage of ⁸⁶ Y-DOTA-MVK(ε)-TATE and ⁸⁶ Y-DOTA-β-Gal-TATE, supporting a key role for cleavable linker chemistry in the observed outcomes. RPT studies using actinium-225 ( ²²⁵ Ac)-labeled variants confirm that all agents are therapeutically effective and well tolerated. While both cleavable variants exhibit superior local control, overall survival, and more favorable toxicity profile when compared with ²²⁵ Ac-DOTA-TATE, ²²⁵ Ac-DOTA-β-Gal-TATE demonstrated lower nephrotoxicity. Our findings suggest a potentially generalizable strategy for improving the pharmacokinetics of radiopharmaceutical therapy agents.