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Attenuated Accumulation of Novel Fluorine (19F)-Labeled Bile Acid Analogues in Gallbladders of Fibroblast Growth Factor-15 (FGF15)-Deficient Mice.

ABSTRACT: Our work has focused on defining the utility of fluorine (19F)-labeled bile acid analogues and magnetic resonance imaging (MRI) to identify altered bile acid transport in vivo. In the current study, we explored the ability of this approach to differentiate fibroblast growth factor-15 (FGF15)-deficient from wild-type (WT) mice, a potential diagnostic test for bile acid diarrhea, a commonly misdiagnosed disorder. FGF15 is the murine homologue of human FGF19, an intestinal hormone whose deficiency is an underappreciated cause of bile acid diarrhea. In a pilot and three subsequent pharmacokinetic studies, we treated mice with two 19F-labeled bile acid analogues, CA-lys-TFA and CA-sar-TFMA. After oral dosing, we quantified 19F-labeled bile acid analogue levels in the gallbladder, liver, small and large intestine, and plasma using liquid chromatography mass spectrometry (LC-MS/MS). Both 19F bile acid analogues concentrated in the gallbladders of FGF15-deficient and WT mice, attaining peak concentrations at approximately 8.5 h after oral dosing. However, analogue levels in gallbladders of FGF15-deficient mice were several-fold less compared to those in WT mice. Live-animal 19F MRI provided agreement with our LC-MS/MS-based measures; we detected robust CA-lys-TFA 19F signals in gallbladders of WT mice but no signals in FGF15-deficient mice. Our finding that 19F MRI differentiates FGF15-deficient from WT mice provides additional proof-of-concept for the development of 19F bile acid analogues and 19F MRI as a clinical test to diagnose bile acid diarrhea due to FGF19 deficiency and other disorders.

SUBMITTER: Metry M

PROVIDER: S-EPMC6446233 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Attenuated Accumulation of Novel Fluorine (<sup>19</sup>F)-Labeled Bile Acid Analogues in Gallbladders of Fibroblast Growth Factor-15 (FGF15)-Deficient Mice.

Metry Melissa M Felton Jessica J Cheng Kunrong K Xu Su S Ai Yong Y Xue Fengtian F Raufman Jean-Pierre JP Polli James E JE

Molecular pharmaceutics 20181003 11

Our work has focused on defining the utility of fluorine (<sup>19</sup>F)-labeled bile acid analogues and magnetic resonance imaging (MRI) to identify altered bile acid transport in vivo. In the current study, we explored the ability of this approach to differentiate fibroblast growth factor-15 (FGF15)-deficient from wild-type (WT) mice, a potential diagnostic test for bile acid diarrhea, a commonly misdiagnosed disorder. FGF15 is the murine homologue of human FGF19, an intestinal hormone whose ...[more]

PMID: 30247920

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Project description:BackgroundCancer-associated fibroblasts are found in the stroma of epithelial tumors. They are characterized by overexpression of the fibroblast activation protein (FAP), a serine protease which was already proven as attractive target for chelator-based theranostics. Unfortunately, the value of gallium-68 labeled tracers is limited by their batch size and the short nuclide half-life. To overcome this drawback, radiolabeling with aluminum fluoride complexes and 6-fluoronicotinamide derivatives of the longer-lived nuclide fluorine-18 was established. The novel compounds were tested for their FAP-specific binding affinity. Uptake and binding competition were studied in vitro using FAP expressing HT-1080 cells. HEK cells transfected with the closely related dipeptidyl peptidase-4 (HEK-CD26) were used as negative control. Small animal positron emission tomography imaging and biodistribution experiments were performed in HT-1080-FAP xenografted nude mice. [18F]AlF-FAPI-74 was selected for PET/CT imaging in a non-small cell lung cancer (NSCLC) patient.ResultsIn vitro, 18F-labeled FAPI-derivatives demonstrated high affinity (EC50 = < 1 nm to 4.2 nm) and binding of up to 80% to the FAP-expressing HT1080 cells while no binding to HEK-CD26 cells was observed. While small animal PET imaging revealed unfavorable biliary excretion of most of the 18F-labeled compounds, the NOTA bearing compounds [18F]AlF-FAPI-74 and -75 achieved good tumor-to-background ratios, as a result of their preferred renal excretion. These two compounds showed the highest tumor accumulation in PET imaging. The organ distribution values of [18F]AlF-FAPI-74 were in accordance with the small animal PET imaging results. Due to its less complex synthesis, fast clearance and low background values, [18F]AlF-FAPI-74 was chosen for clinical imaging. PET/CT of a patient with metastasized non-small cell lung cancer (NSCLC), enabled visualization of the primary tumor and its metastases at the hepatic portal and in several bones. This was accompanied by a rapid clearance from the blood pool and low background in healthy organs.Conclusion[18F]AlF-labeled FAPI derivatives represent powerful tracers for PET. Owing to an excellent performance in PET imaging, FAPI-74 can be regarded as a promising precursor for [18F]AlF-based FAP-imaging.

Project description:The accumulation of α-synuclein aggregates (α-syn) in the human brain is an occurrence common to all α-synucleinopathies. Non-invasive detection of these aggregates in a living brain with a target-specific radiotracer is not yet possible. We have recently discovered that the inclusion of a methylenedioxy group in the structure of diarylbisthiazole (DABTA)-based tracers improves binding affinity and selectivity to α-syn. Subsequently, complementary in silico modeling and machine learning (ML) of tracer-protein interactions were employed to predict surface sites and structure-property relations for the binding of the ligands. Based on this observation, we developed a small focused library of DABTAs from which 4-(benzo[d][1,3]dioxol-5-yl)-4'-(3-[18F]fluoro-4-methoxyphenyl)-2,2'-bithiazole [ 18 F]d 2, 6-(4'-(3-[18F]fluoro-4-methoxyphenyl)-[2,2'-bithiazol]-4-yl)-[1,3]dioxolo[4,5-b]pyridine [ 18 F]d 4, 4-(benzo [d][1,3]dioxol-5-yl)-4'-(6-[18F]fluoropyridin-3-yl)-2,2'-bithiazole [ 18 F]d 6, and 6-(4'-(6-[18F]fluoropyridin-3-yl)-[2,2'-bithiazol]-4-yl)-[1,3]dioxolo[4,5-b]pyridine [ 18 F]d 8 were selected based on their high binding affinity to α-syn and were further evaluated. Binding assay experiments carried out with the non-radioactive versions of the above tracers d 2, d 4, d 6, and d 8 showed high binding affinity of the ligands to α-syn: 1.22, 0.66, 1.21, and 0.10 nM, respectively, as well as excellent selectivity over β-amyloid plaques (Aβ) and microtubular tau aggregates (>200-fold selectivity). To obtain the tracers, their precursors were radiolabeled either via an innovative ruthenium-mediated (SNAr) reaction ([ 18 F]d 2 and [ 18 F]d 4) or typical SNAr reaction ([ 18 F]d 6 and [ 18 F]d 8) with moderate-to-high radiochemical yields (13% - 40%), and high molar activity > 60 GBq/μmol. Biodistribution experiments carried out with the tracers in healthy mice revealed that [ 18 F]d 2 and [ 18 F]d 4 showed suboptimal brain pharmacokinetics: 1.58 and 4.63 %ID/g at 5 min post-injection (p.i.), and 1.93 and 3.86 %ID/g at 60 min p.i., respectively. However, [ 18 F]d 6 and [ 18 F]d 8 showed improved brain pharmacokinetics: 5.79 and 5.13 %ID/g at 5 min p.i.; 1.75 and 1.07 %ID/g at 60 min p.i.; and 1.04 and 0.58 %ID/g at 120 min p.i., respectively. The brain uptake kinetics of [ 18 F]d 6 and [ 18 F]d 8 were confirmed in a dynamic PET study. Both tracers also showed no brain radiometabolites at 20 min p.i. in initial in vivo stability experiments carried out in healthy mice. [ 18 F]d 8 seems very promising based on its binding properties and in vivo stability, thus encouraging further validation of its usefulness as a radiotracer for the in vivo visualization of α-syn in preclinical and clinical settings. Additionally, in silico and ML-predicted values correlated with the experimental binding affinity of the ligands.

Project description:Heart failure remains a major source of late morbidity and mortality after myocardial infarction (MI). The temporospatial presence of activated fibroblasts in the injured myocardium predicts the quality of cardiac remodeling after MI. Therefore, monitoring of activated fibroblasts is of great interest for studying cardiac remodeling after MI. Fibroblast activation protein (FAP) expression is upregulated in activated fibroblasts. This study investigated the feasibility of imaging activated fibroblasts with a new 68Ga-labeled FAP inhibitor (68Ga-FAPI-04) for PET imaging of fibroblast activation in a preclinical model of MI. Methods: MI and sham-operated rats were scanned with 68Ga-FAPI-04 PET/CT (1, 3, 6, 14, 23, and 30 d after MI) and with 18F-FDG (3 d after MI). Dynamic 68Ga-FAPI-04 PET and blocking studies were performed on MI rats 7 d after coronary ligation. After in vivo scans, the animals were euthanized and their hearts harvested for ex vivo analyses. Cryosections were prepared for autoradiography, hematoxylin and eosin (H&E), and immunofluorescence staining. Results: 68Ga-FAPI-04 uptake in the injured myocardium peaked on day 6 after coronary ligation. The tracer accumulated intensely in the MI territory, as identified by decreased 18F-FDG uptake and confirmed by PET/MR and H&E staining. Autoradiography and H&E staining of cross-sections revealed that 68Ga-FAPI-04 accumulated mainly at the border zone of the infarcted myocardium. In contrast, there was only minimal uptake in the infarct of the blocked rats, comparable to the uptake in the remote noninfarcted myocardium (PET image-derived ratio of infarct uptake to remote uptake: 6 ± 2). Immunofluorescence staining confirmed the presence of FAP-positive myofibroblasts in the injured myocardium. Morphometric analysis of the whole-heart sections demonstrated 3- and 8-fold higher FAP-positive fibroblast density in the border zone than in the infarct center and remote area, respectively. Conclusion: 68Ga-FAPI-04 represents a promising radiotracer for in vivo imaging of post-MI fibroblast activation. Noninvasive imaging of activated fibroblasts may have significant diagnostic and prognostic value, which could aid clinical management of patients after MI.

Project description:PurposeA series of radiotracers targeting fibroblast activation protein (FAP) with great pharmacokinetics have been developed for cancer diagnosis and therapy. Nevertheless, the use of dominant PET tracers, gallium-68-labeled FAPI derivatives, was limited by the short nuclide half-life and production scale, and the therapeutic tracers exhibited rapid clearance and insufficient tumor retention. In this study, we developed a FAP targeting ligand, LuFL, containing organosilicon-based fluoride acceptor (SiFA) and DOTAGA chelator, capable of labeling fluorine-18 and lutetium-177 in one molecular with simple and highly efficient labeling procedure, to achieve cancer theranostics.MethodsThe precursor LuFL (20) and [natLu]Lu-LuFL (21) were successfully synthesized and labeled with fluorine-18 and lutetium-177 using a simple procedure. A series of cellular assays were performed to characterize the binding affinity and FAP specificity. PET imaging, SPECT imaging, and biodistribution studies were conducted to evaluate pharmacokinetics in HT-1080-FAP tumor-bearing nude mice. A comparison study of [177Lu]Lu-LuFL ([177Lu]21) and [177Lu]Lu-FAPI-04 was carried out in HT-1080-FAP xenografts to determine the cancer therapeutic efficacy.ResultsLuFL (20) and [natLu]Lu-LuFL (21) demonstrated excellent binding affinity towards FAP (IC50: 2.29 ± 1.12 nM and 2.53 ± 1.87 nM), compared to that of FAPI-04 (IC50: 6.69 ± 0.88 nM). In vitro cellular studies showed that 18F-/177Lu-labeled 21 displayed high specific uptake and internalization in HT-1080-FAP cells. Micro-PET, SPECT imaging and biodistribution studies with [18F]/[177Lu]21 revealed higher tumor uptake and longer tumor retention than those of [68 Ga]/[177Lu]Ga/Lu-FAPI-04. The radionuclide therapy studies showed significantly greater inhibition of tumor growth for the [177Lu]21 group, than for the control group and the [177Lu]Lu-FAPI-04 group.ConclusionThe novel FAPI-based radiotracer containing SiFA and DOTAGA was developed as a theranostics radiopharmaceutical with simple and short labeling process, and showed promising properties including higher cellular uptake, better FAP binding affinity, higher tumor uptake and prolong retention compared to FAPI-04. Preliminary experiments with 18F- and 177Lu-labeled 21 showed promising tumor imaging properties and favorable anti-tumor efficacy.

Dataset Information

Attenuated Accumulation of Novel Fluorine (19F)-Labeled Bile Acid Analogues in Gallbladders of Fibroblast Growth Factor-15 (FGF15)-Deficient Mice.

Publications

Attenuated Accumulation of Novel Fluorine (<sup>19</sup>F)-Labeled Bile Acid Analogues in Gallbladders of Fibroblast Growth Factor-15 (FGF15)-Deficient Mice.

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