Abstract
Purpose
[18F]FHBG has been used as a positron emission tomography (PET) imaging tracer for the monitoring of herpes simplex virus type 1 thymidine kinase (HSV1-tk), a reporter gene for cell and gene therapy in humans. However, this tracer shows inadequate blood-brain barrier (BBB) penetration and, therefore, would be limited for accurate quantification of reporter gene expression in the brain. Here, we report the synthesis and evaluation of 9-(4-[18F]fluoro-3-(hydroxymethyl)butyl)-2(phenylthio)-6-oxopurine ([18F]FHBT) as a new PET tracer for imaging reporter gene expression of HSV1-tk and its mutant HSV1-sr39tk, with the aim of improved BBB penetration.
Procedures
[18F]FHBT was prepared by using a tosylate precursor and [18F]KF. The cellular uptake of [18F]FHBT was performed in HSV1-sr39tk-positive (+) or HSV1-sr39tk-negative (−) MDA-MB-231 breast cancer cells. The specificity of [18F]FHBT to assess HSV1-sr39tk expression was evaluated by in vitro blocking studies using 1 mM of ganciclovir (GCV). Penetration of [18F]FHBT and [18F]FHBG across the BBB was assessed by dynamic PET imaging studies in normal mice.
Results
The tosylate precursor reacted with [18F]KF using Kryptofix2.2.2 followed by deprotection to give [18F]FHBT in 10 % radiochemical yield (decay-corrected). The uptake of [18F]FHBT in HSV1-sr39tk (+) cells was significantly higher than that of HSV1-sr39tk (−) cells. In the presence of GCV (1 mM), the uptake of [18F]FHBT was significantly decreased, indicating that [18F]FHBT serves as a selective substrate of HSV1-sr39TK. PET images and time-activity curves of [18F]FHBT in the brain regions showed similar initial brain uptakes (~ 12.75 min) as [18F]FHBG (P > 0.855). Slower washout of [18F]FHBT was observed at the later time points (17.75 − 57.75 min, P > 0.207).
Conclusions
Although [18F]FHBT showed no statistically significant improvement of BBB permeability compared with [18F]FHBG, we have demonstrated that the 2-(phenylthio)-6-oxopurine backbone can serve as a novel scaffold for developing HSV1-tk/HSV1-sr39tk reporter gene imaging agents for additional research in the future.
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Acknowledgments
We thank the Radiochemistry Facility at Stanford University for 18F-production. We would also like to thank Dr. Timothy Doyle at the Stanford Center for In Vivo Imaging (SCI3) for his technical assistance. Additionally, we extend our thanks to Dr. Michael B. Bolger and John A. DiBella at Simulations Plus, Inc. for lending us access to the ADMET Predictor software tool.
Funding
Financial support was provided by the Ben and Catherine Ivy Foundation (S.S.G.). This work was also supported in part by Grants-in-Aid for Scientific Research (Fund for the Promotion of Joint International Research) and the Mochida Memorial Foundation for Medical and Pharmaceutical Research (T.F.).
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Fuchigami, T., Haywood, T., Gowrishankar, G. et al. Synthesis and Characterization of 9-(4-[18F]Fluoro-3-(hydroxymethyl)butyl)-2-(phenylthio)-6-oxopurine as a Novel PET Agent for Mutant Herpes Simplex Virus Type 1 Thymidine Kinase Reporter Gene Imaging. Mol Imaging Biol 22, 1151–1160 (2020). https://doi.org/10.1007/s11307-020-01517-5
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DOI: https://doi.org/10.1007/s11307-020-01517-5