Synthesis and Biological Evaluation of Novel ^99mTc-Labeled Palbociclib Derivatives Targeting Cyclin-Dependent Kinase 4/6 (CDK4/6) as Potential Cancer Imaging Agents

Cancer results from cell proliferation that exceeds normal growth control. There are various specific proteins that control and regulate the cell cycle, such as cyclin-dependent kinases (CDKs), cyclins, and retinoblastoma protein (pRb). The aberration of the cyclin D-CDK4/6-INK4-pRb pathway occurs frequently in cancers; thus, CDK4/6 is an attractive target for the development of radiopharmaceuticals for tumor imaging. In this study, we chose palbociclib, which was approved by the FDA for treating ER+/HER2– advanced breast cancer as the target vector and the isonitrile group, which can coordinate strongly with the [^99mTc­(CO)₃]⁺ core as the bifunctional chelator, to develop four novel ^99mTc-labeled radiotracers for tumor imaging. The ligands (L2, L3, L4, and L5) were synthesized by reacting palbociclib with isocyanide-containing active esters and then radiolabeling with a [^99mTc­(CO)₃]⁺ core to produce radiotracers (^99mTc–L2, ^99mTc–L3, ^99mTc–L4, and ^99mTc-L5) with high radiochemical purity (>95%) and good stability in vitro. The structures of the ^99mTc complexes were identified by preparation and characterization of the corresponding stable rhenium complexes. Partition coefficient results indicated that these complexes were lipophilic. A kinase inhibition assay demonstrated the high affinity of the stable Re complexes for CDK4. A cell study showed that all four complexes had substantial uptake by MCF-7 cells and could be significantly inhibited by palbociclib and nonradiolabeled ligand, indicating a CDK4/6-specific uptake mechanism. Biodistribution studies in nude mice bearing MCF-7 tumors showed that the complexes had obvious accumulation in tumors at 2 h postinjection. ^99mTc–L2 exhibited the highest tumor uptake and tumor/blood ratio, whereas ^99mTc–L4 showed the highest tumor/muscle ratio. The micro-SPECT/CT study showed that complex ^99mTc–L4 had visible uptake at the tumor site, and the accumulation was clearly reduced in the image after pretreatment with palbociclib, further indicating CDK4/6 specificity. All the results showed that the ^99mTc-labeled complexes in this work have the potential for tumor imaging.