Noninvasive quantification of SIRT1 expression-activity and pharmacologic inhibition in a rat model of intracerebral glioma using 2-[18F]BzAHA PET/CT/MRI

Abstract

Background: Several studies demonstrated that glioblastoma multiforme progression and recurrence is linked to epigenetic regulatory mechanisms. Sirtuin 1 (SIRT1) plays an important role in glioma progression, invasion, and treatment response and is a potential therapeutic target. The aim of this study is to test the feasibility of 2-[¹⁸F]BzAHA for quantitative imaging of SIRT1 expression-activity and monitoring pharmacologic inhibition in a rat model of intracerebral glioma.

Methods: Sprague Dawley rats bearing 9L (N = 12) intracerebral gliomas were injected with 2-[¹⁸F]BzAHA (300-500 µCi/animal i.v.) and dynamic positron-emission tomography (PET) imaging was performed for 60 min. Then, SIRT1 expression in 9L tumors (N = 6) was studied by immunofluorescence microscopy (IF). Two days later, rats with 9L gliomas were treated either with SIRT1 specific inhibitor EX-527 (5 mg/kg, i.p.; N = 3) or with histone deacetylases class IIa specific inhibitor MC1568 (30 mg/kg, i.p.; N = 3) and 30 min later were injected i.v. with 2-[¹⁸F]BzAHA. PET-computerized tomography-magnetic resonance (PET/CT/MR) images acquired after EX-527 and MC1568 treatments were co-registered with baseline images.

Results: Standard uptake values (SUVs) of 2-[¹⁸F]BzAHA in 9L tumors measured at 20 min post-radiotracer administration were 1.11 ± 0.058 and had a tumor-to-brainstem SUV ratio of 2.73 ± 0.141. IF of 9L gliomas revealed heterogeneous upregulation of SIRT1, especially in hypoxic and peri-necrotic regions. Significant reduction in 2-[¹⁸F]BzAHA SUV and distribution volume in 9L tumors was observed after administration of EX-527, but not MC1568.

Conclusions: PET/CT/MRI with 2-[¹⁸F]BzAHA can facilitate studies to elucidate the roles of SIRT1 in gliomagenesis and progression, as well as to optimize therapeutic doses of novel SIRT1 inhibitors.

Keywords: EX-527; SIRT1; epigenetics; glioma; molecular imaging.

Fig. 1

PET/CT/MRI of SIRT1 expression–activity in 9L intracerebral gliomas in rats. (A) Representative series of coronal images of the rat brain bearing intracerebral 9L glioma. The position of sections relative to bregma is indicated in millimeters on T2-weighted MR images. 2-[¹⁸F]BzAHA PET/CT images were obtained at 20 min post-injection of radiotracer and co-registered with T2-weighted MR images. The levels of 2-[¹⁸F]BzAHA accumulation in tumors and different structures of the brain were measured in standard uptake value (SUV). (B) SUV for 9L gliomas (N = 6) as compared to other structures within the brain. (C) SUV ratio normalized by the SUV of the brainstem region for 9L gliomas. PET/CT images are color coded to SUV. Data—mean ± SEM. Statistical significance was determined via one-way ANOVA, *P < .05, **P < .01, ***P < .001, ****P < .0001.

Fig. 2

Validation of 2-[¹⁸F]BzAHA PET/CT/MRI using histopathologic and immunofluorescence (IF) analyses of 9L glioma. (A) 2-[¹⁸F]BzAHA axial PET/CT image (top), (B) axial 2-[¹⁸F]BzAHA PET/MRI fusion image (bottom), PET/CT images are color-coded to standard uptake values (SUVs). (C) Macroscopic and (D) microscopic images (10× objective, scale bar—200 µm), of axial brain sections stained with H&E to visualize tumor structure. Microscopic images of SIRT1 IF and DAPI (E) 10x objective, scale bar—200 µm; (F) 20x objective, scale bar—100 µm.

Fig. 3

Monitoring pharmacologic inhibition of SIRT1 with 2-[¹⁸F]BzAHA PET/CT/MRI. (A) Representative coronal T2-weighted MRI and 2-[¹⁸F]BzAHA PET/CT/MR images depicting different regions of the rat brain with intracerebral 9L tumor (shown by a dotted line on a side-view of a 3D rendered image): through the area of the hippocampus (Hippo, top), 9L tumor (9L, middle), nucleus accumbens (NA, bottom). 2-[¹⁸F]BzAHA PET/CT images were obtained at 20 min post-radiotracer administration before (baseline) and after treatment with either EX-527 (SIRT1 selective inhibitor) or MC1568 (HDAC class IIa selective inhibitor). PET/CT images are color-coded to standard uptake values (SUVs). The levels of 2-[¹⁸F]BzAHA-derived radioactivity are expressed as (B) SUVs or (C) distribution volumes (DVs) in 9L tumors and different brain structures at baseline (N = 4) and after therapy with EX-527 (N = 3) or MC1568 (N = 3). Data—mean ± SEM. Statistical significance was determined using ANOVA for repeated measures; ****P < .0001.