• Imaging investigations examined the uptake and functionality of TAK-164, revealing positive associations between tumor uptake and GCC expression, which were consistent with the observed antitumor effects.• TAK-164 exhibits significant efficacy across various GCC-positive tumors, even in cases where TAK-264, a GCC-targeted auristatin ADC, has proven ineffective.• A robust correlation exists among the uptake of 89Zr-labeled TAK-164, the extent of GCC expression, and notably, the response to TAK-164 treatment in both GCC-expressing xenografts and PHTX models. This evidence concerns the gene GUCY2C and neoplasm.