SPECT and optical multimodality imaging of HIF-active tumors by using an oxygen-dependent degradation protein probe with interchangeable labeling system

Objectives The visualization of hypoxia-inducible factor (HIF)-active tumors is clinically important. We designed a novel in vivo imaging probe for it that consists of protein transduction domain, oxygen-dependent degradation domain, and HaloTag^® protein (POH). It can be used as a probe for various imaging modality by differently labeled HaloTag^® ligands (HL). We have already succeeded in monitoring HIF-active region in cancers with the near-infrared fluorescently labeled POH probe (NIRF-POH). In this study, we labeled POH with ¹¹¹In and evaluated their biodistribution in the tumor-bearing mice and their in vivo SPECT images, comparing with NIRF-POH.

Methods HL-DOTA and HL-BnDOTA were synthesized by the reaction of HL-NH₂ and NHS-DOTA or p-SCN-BnDOTA, and purified on ODS. They were labeled with ¹¹¹In, and then conjugated to POH. These radioactive probes were injected i.v. to mice with FM3A or Suit-2 tumors.

Results Both HL-DOTAs were labeled with ¹¹¹In showing good radiochemical yield (~80%). Conjugation of ¹¹¹In-HL-DOTAs to POH (¹¹¹In-POH) was confirmed as a single band around 46 kDa by SDS-PAGE. Their radiochemical purity was over 95%. The biodistribution of ¹¹¹In-POH was similar to that of NIRF-POH and ¹¹¹In-POH well accumulated in both tumors (0.8-2.6 % administered dose/g) at 24 h after injection. On in vivo SPECT images, POH was distributed strongly in the liver and kidney, but tumors could be identified at 1h after i.v.. And, as the activities of background areas as well as liver and kidneys gradually decreased, tumors were more clearly visualized at 24 h.

Conclusions ¹¹¹In-POH showed similar biodistribution to NIRF-POH. And, they could provide quantitative biodistribution data. Moreover, they were clearly imaged on in vivo SPECT for the first time. The SPECT and optical dual imaging system studied here would promote the clinical translation of our HIF imaging strategy established by animal experiments using NIRF-POH