Abstract
1854
Objectives: The protein kinase mammalian target of rapamycin (mTOR) is a key regulator of cell growth and proliferation that is up-regulated in a number of cancers. Rapamycin, a 31-membered macrolide is a potent inhibitor of mTOR and three of its analogs are currently in clinical trials for cancer treatment in humans. The high affinity of rapamycin for mTOR complexed with the cellular protein FKBP12 (Kd = 2 nM) makes it possible to use positron emitter labeled rapamycin analogs as probes to image tumors for their drug sensitivity using Positron Emission Tomography (PET). The principal aim of this investigation was to prepare 11C, 18F and 124I labeled rapamycin analogs for that.
Methods: Rapamycin modified at 42-position with O-methyl, O-fluoroethyl, O-acetyl, O-fluoroacetyl, O-2-fluorobenzoyl, O-4-fluorobenzoyl, O-2-iodobenzoyl, O-3-iodobenzoyl, and O-4-iodobenzoyl groups, as well as modified at the 7-position with 2-fluorobenzyloxy, 3-fluorobenzyloxy, 4-fluorobenzyloxy, 2-iodobenzyloxy, 3-iodobenzyloxy, and 4-iodobenzyloxy groups were synthesized via alkylation, acylation, and substitution reactions on rapamycin with appropriate F- or I-substituted tagging moieties. The products were purified and characterized by 600 MHz 1H, 13C, 19F, HMBC and HMQC 2D NMR experiments as well as high resolution mass spectroscopy. The inhibitory activity of these analogs against mTOR was assessed in vitro by S6 phosphorylation assay in U87 glioblastoma cells. The feasibility of radiolabeling these complex rapamycin derivatives with 18F or 124I isotope was explored with a selected member of the group.
Results: The IC50 values of the newly synthesized analogs of rapamycin were found to be 2-65 fold less than unaltered rapamycin. The synthesis of 42-O-(4-[18F]fluorobenzoyl)rapamycin, as a model analog, was achieved by a two-step labeling procedure involving the preparation of no-carrier-added 4-[18F]fluorobenzoic acid and its condensation with rapamycin. The pure 18F-labeled rapamycin derivative was isolated by semi-preparative HPLC in 4±2 % (n=10) radiochemical yield. In vitro assays with this 18F-labeled analog showed that the probe was taken up by the U87 glioblastoma cells as well as by U87 cells over expressing EGFRvIII and restored with PTEN. Utilization of this analog in PET imaging of SCID mice bearing the same human tumor xenografts is currently under way.
Conclusions: A number of potent fluorinated and iodinated rapamycin based inhibitors of mTOR function has been synthesized. A member of this group has also been 18F-labeled for in vivo PET imaging purposes.
Research Support (if any): This investigation is supported by the DOE
- Society of Nuclear Medicine, Inc.