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Synthesis and evaluation of (18)F-labeled ATP competitive inhibitors of topoisomerase II as probes for imaging topoisomerase II expression.

ABSTRACT: Type II topoisomerase (Topo-II) is an ATP-dependent enzyme that is essential in the transcription, replication, and chromosome segregation processes and, as such, represents an attractive target for cancer therapy. Numerous studies indicate that the response to treatment with Topo-II inhibitors is highly dependent on both the levels and the activity of the enzyme. Consequently, a non-invasive assay to measure tumoral Topo-II levels has the potential to differentiate responders from non-responders. With the ultimate goal of developing a radiofluorinated tracer for positron emission tomography (PET) imaging, we have designed, synthesized, and evaluated a set of fluorinated compounds based on the structure of the ATP-competitive Topo-II inhibitor QAP1. Compounds 18 and 19b showed inhibition of Topo-II in in vitro assays and exhibited moderate, Topo-II level dependent cytotoxicity in SK-BR-3 and MCF-7 cell lines. Based on these results, (18)F-labeled analogs of these two compounds were synthesized and evaluated as PET probes for imaging Topo-II overexpression in mice bearing SK-BR-3 xenografts. [(18)F]-18 and [(18)F]-19b were synthesized from their corresponding protected tosylated derivatives by fluorination and subsequent deprotection. Small animal PET imaging studies indicated that both compounds do not accumulate in tumors and exhibit poor pharmacokinetics, clearing from the blood pool very rapidly and getting metabolized over. The insights gained from the current study will surely aid in the design and construction of future generations of PET agents for the non-invasive delineation of Topo-II expression.

SUBMITTER: Daumar P

PROVIDER: S-EPMC4406421 | biostudies-literature | 2014 Oct

REPOSITORIES: biostudies-literature

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Synthesis and evaluation of (18)F-labeled ATP competitive inhibitors of topoisomerase II as probes for imaging topoisomerase II expression.

Daumar Pierre P Zeglis Brian M BM Ramos Nicholas N Divilov Vadim V Sevak Kuntal Kumar KK Pillarsetty NagaVaraKishore N Lewis Jason S JS

European journal of medicinal chemistry 20140908

Type II topoisomerase (Topo-II) is an ATP-dependent enzyme that is essential in the transcription, replication, and chromosome segregation processes and, as such, represents an attractive target for cancer therapy. Numerous studies indicate that the response to treatment with Topo-II inhibitors is highly dependent on both the levels and the activity of the enzyme. Consequently, a non-invasive assay to measure tumoral Topo-II levels has the potential to differentiate responders from non-responder ...[more]

PMID: 25240701

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Project description:IntroductionMalignant gliomas frequently harbor mutations in the isocitrate dehydrogenase 1 (IDH1) gene. Studies suggest that IDH mutation contributes to tumor pathogenesis through mechanisms that are mediated by the neomorphic metabolite of the mutant IDH1 enzyme, 2-hydroxyglutarate (2-HG). The aim of this work was to synthesize and evaluate radiolabeled compounds that bind to the mutant IDH1 enzyme with the goal of enabling noninvasive imaging of mutant IDH1 expression in gliomas by positron emission tomography (PET).MethodsA small library of nonradioactive analogs were designed and synthesized based on the chemical structure of reported butyl-phenyl sulfonamide inhibitors of mutant IDH1. Enzyme inhibition assays were conducted using purified mutant IDH1 enzyme, IDH1-R132H, to determine the IC50 and the maximal inhibitory efficiency of the synthesized compounds. Selected compounds, 1 and 4, were labeled with radioiodine ((125)I) and/or (18)F using bromo- and phenol precursors, respectively. In vivo behavior of the labeled inhibitors was studied by conducting tissue distribution studies with [(125)I]1 in normal mice. Cell uptake studies were conducted using an isogenic astrocytoma cell line that carried a native IDH1-R132H mutation to evaluate the potential uptake of the labeled inhibitors in IDH1-mutated tumor cells.ResultsEnzyme inhibition assays showed good inhibitory potency for compounds that have iodine or a fluoroethoxy substituent at the ortho position of the phenyl ring in compounds 1 and 4 with IC50 values of 1.7 μM and 2.3 μM, respectively. Compounds 1 and 4 inhibited mutant IDH1 activity and decreased the production of 2-HG in an IDH1-mutated astrocytoma cell line. Radiolabeling of 1 and 4 was achieved with an average radiochemical yield of 56.6 ± 20.1% for [(125)I]1 (n = 4) and 67.5 ± 6.6% for [(18)F]4 (n = 3). [(125)I]1 exhibited favorable biodistribution characteristics in normal mice, with rapid clearance from the blood and elimination via the hepatobiliary system by 4 h after injection. The uptake of [(125)I]1 in tumor cells positive for IDH1-R132H was significantly higher compared to isogenic WT-IDH1 controls, with a maximal uptake ratio of 1.67 at 3 h post injection. Co-incubation of the labeled inhibitors with the corresponding nonradioactive analogs, and decreasing the normal concentrations of FBS (10%) in the incubation media substantially increased the uptake of the labeled inhibitors in both the IDH1-mutant and WT-IDH1 tumor cell lines, suggesting significant non-specific binding of the synthesized labeled butyl-phenyl sulfonamide inhibitors.ConclusionsThese data demonstrate the feasibility of developing radiolabeled probes for the mutant IDH1 enzyme based on enzyme inhibitors. Further optimization of the labeled inhibitors by modifying the chemical structure to decrease the lipophilicity and to increase potency may yield compounds with improved characteristics as probes for imaging mutant IDH1 expression in tumors.

Project description:IntroductionEquilibrium single-photon radionuclide imaging methods for assessing cardiac function and the integrity of the vascular system have long been in use for both clinical and research purposes. However, positron-emitting blood pool agents that could provide PET equivalents to these (and other) clinical procedures have not yet been adopted despite technical imaging advantages offered by PET. Our goal was to develop a PET blood pool tracer that not only meets necessary in vivo biological requirements but can be produced with an uncomplicated and rapid synthesis method which would facilitate clinical translation. Herein, albumin labeled with fluorine-18 was synthesized using a one-pot method and evaluated in vitro and in vivo in rats.MethodsA ligand (NODA-Bz-TFPE), containing NODA attached to a tetrafluorophenylester (TFPE) via a phenyl linker (Bz), was labeled with aluminum fluoride (Al[18F]F). Conjugation of the serum albumin with the ligand (Al[18F]F-NODA-Bz-TFPE), followed by purification (size exclusion chromatography), yielded the final product (Al[18F]F-NODA-Bz-RSA/HSA). In vitro stability was evaluated in human serum albumin by HPLC. Rat biodistributions and whole-body PET imaging over a 4 h time course were used for the in vivo evaluation.ResultsThis synthesis exhibited an overall radiochemical yield of 45 ± 10% (n = 30), a 50-min radiolabeling time, a radiochemical purity >99% and apparent stability up to 4 h in human serum. Blood had the highest retention of Al[18F]F-NODA-Bz-RSA at all times with a blood half-life of 5.2 h in rats. Al[18F]F-NODA-Bz-RSA distribution in most rat tissues remained relatively constant for up to 1 h, indicating that the tissue radioactivity content represents the respective tissue plasma volume. Dynamic whole-body PET images were in agreement with these findings.ConclusionsA new ligand has been developed and radiolabeled with Al[18F]F that allows rapid (50-min) preparation of fluorine-18 serum albumin in one-pot. In addition to increased synthetic efficiency, the construct appears to be metabolically stable in rats. This method could encourage wider use of PET to quantify cardiac function and tissue vascular integrity in both research and clinical settings.

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Synthesis and evaluation of (18)F-labeled ATP competitive inhibitors of topoisomerase II as probes for imaging topoisomerase II expression.

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Synthesis and evaluation of (18)F-labeled ATP competitive inhibitors of topoisomerase II as probes for imaging topoisomerase II expression.

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