Total-body imaging of mu-opioid receptors with [11C]carfentanil in non-human primates.

PURPOSE: Mu-opioid receptors (MORs) are widely expressed in the central nervous system (CNS), peripheral organs, and immune system. This study measured the whole body distribution of MORs in rhesus macaques using the MOR selective radioligand [11C]carfentanil ([11C]CFN) on the PennPET Explorer. Both baseline and blocking studies were conducted using either naloxone or GSK1521498 to measure the effect of the antagonists on MOR binding in both CNS and peripheral organs. METHODS: The PennPET Explorer was used for MOR total-body PET imaging in four rhesus macaques using [11C]CFN under baseline, naloxone pretreatment, and naloxone or GSK1521498 displacement conditions. Logan distribution volume ratio (DVR) was calculated by using a reference model to quantitate brain regions, and the standard uptake value ratios (SUVRs) were calculated for peripheral organs. The percent receptor occupancy (%RO) was calculated to establish the blocking effect of 0.14 mg/kg naloxone or GSK1521498. RESULTS: The %RO in MOR-abundant brain regions was 75-90% for naloxone and 72-84% for GSK1521498 in blocking studies. A higher than 90% of %RO were observed in cervical spinal cord for both naloxone and GSK1521498. It took approximately 4-6 min for naloxone or GSK1521498 to distribute to CNS and displace [11C]CFN from the MOR. A smaller effect was observed in heart wall in the naloxone and GSK1521498 blocking studies. CONCLUSION: [11C]CFN total-body PET scans could be a useful approach for studying mechanism of action of MOR drugs used in the treatment of acute and chronic opioid use disorder and their effect on the biodistribution of synthetic opioids such as CFN. GSK1521498 could be a potential naloxone alternative to reverse opioid overdose.

Positron Emission Tomography with [18F]ROStrace Reveals Progressive Elevations in Oxidative Stress in a Mouse Model of Alpha-Synucleinopathy.

The synucleinopathies are a diverse group of neurodegenerative disorders characterized by the accumulation of aggregated alpha-synuclein (aSyn) in vulnerable populations of brain cells. Oxidative stress is both a cause and a consequence of aSyn aggregation in the synucleinopathies; however, noninvasive methods for detecting oxidative stress in living animals have proven elusive. In this study, we used the reactive oxygen species (ROS)-sensitive positron emission tomography (PET) radiotracer [18F]ROStrace to detect increases in oxidative stress in the widely-used A53T mouse model of synucleinopathy. A53T-specific elevations in [18F]ROStrace signal emerged at a relatively early age (6-8 months) and became more widespread within the brain over time, a pattern which paralleled the progressive development of aSyn pathology and oxidative damage in A53T brain tissue. Systemic administration of lipopolysaccharide (LPS) also caused rapid and long-lasting elevations in [18F]ROStrace signal in A53T mice, suggesting that chronic, aSyn-associated oxidative stress may render these animals more vulnerable to further inflammatory insult. Collectively, these results provide novel evidence that oxidative stress is an early and chronic process during the development of synucleinopathy and suggest that PET imaging with [18F]ROStrace holds promise as a means of detecting aSyn-associated oxidative stress noninvasively.

Imaging CAR T Cell Trafficking with eDHFR as a PET Reporter Gene.

Cell-based therapeutics have considerable promise across diverse medical specialties; however, reliable human imaging of the distribution and trafficking of genetically engineered cells remains a challenge. We developed positron emission tomography (PET) probes based on the small-molecule antibiotic trimethoprim (TMP) that can be used to image the expression of the Escherichia coli dihydrofolate reductase enzyme (eDHFR) and tested the ability of [18F]-TMP, a fluorine-18 probe, to image primary human chimeric antigen receptor (CAR) T cells expressing the PET reporter gene eDHFR, yellow fluorescent protein (YFP), and Renilla luciferase (rLuc). Engineered T cells showed an approximately 50-fold increased bioluminescent imaging signal and 10-fold increased [18F]-TMP uptake compared to controls in vitro. eDHFR-expressing anti-GD2 CAR T cells were then injected into mice bearing control GD2- and GD2+ tumors. PET/computed tomography (CT) images acquired on days 7 and 13 demonstrated early residency of CAR T cells in the spleen followed by on-target redistribution to the GD2+ tumors. This was corroborated by autoradiography and anti-human CD8 immunohistochemistry. We found a high sensitivity of detection for identifying tumor-infiltrating CD8 CAR T cells, ∼11,000 cells per mm3. These data suggest that the [18F]-TMP/eDHFR PET pair offers important advantages that could better allow investigators to monitor immune cell trafficking to tumors in patients.

Bacterial infection imaging with [18F]fluoropropyl-trimethoprim.

There is often overlap in the diagnostic features of common pathologic processes such as infection, sterile inflammation, and cancer both clinically and using conventional imaging techniques. Here, we report the development of a positron emission tomography probe for live bacterial infection based on the small-molecule antibiotic trimethoprim (TMP). [18F]fluoropropyl-trimethoprim, or [18F]FPTMP, shows a greater than 100-fold increased uptake in vitro in live bacteria (Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa) relative to controls. In a rodent myositis model, [18F]FPTMP identified live bacterial infection without demonstrating confounding increased signal in the same animal from other etiologies including chemical inflammation (turpentine) and cancer (breast carcinoma). Additionally, the biodistribution of [18F]FPTMP in a nonhuman primate shows low background in many important tissues that may be sites of infection such as the lungs and soft tissues. These results suggest that [18F]FPTMP could be a broadly useful agent for the sensitive and specific imaging of bacterial infection with strong translational potential.

Correlation analysis of [18F]ROStrace using ex vivo autoradiography and dihydroethidium fluorescent imaging in lipopolysaccharide-treated animals.

Reactive oxygen species (ROS) are believed to play an important role in the proinflammatory form of neuroinflammation. Therefore, the availability of a radiotracer labeled with a positron-emitting radionuclide that can measure levels of ROS in tissue could provide a valuable method for imaging neuroinflammation in vivo with the functional imaging technique positron emission tomography (PET). We previously reported the synthesis and in vivo evaluation of [18F]ROStrace, a radiotracer for imaging ROS in vivo with PET, in an LPS model of neuroinflammation. In the current study, we conducted additional validation studies aimed at determining the cellular localization of this radiotracer in the same model. Our results indicate that [18F]ROStrace is primarily localized in microglia/macrophages and neurons in LPS-treated animals, and provide further support in the use of this radiotracer as a PET-based probe for imaging the proinflammatory form of neuroinflammation.

Synthesis and evaluation of an AZD2461 [18F]PET probe in non-human primates reveals the PARP-1 inhibitor to be non-blood-brain barrier penetrant.

Poly(ADP-ribose)polymerase-1 inhibitor (PARPi) AZD2461 was designed to be a weak P-glycoprotein (P-gp) analogue of FDA approved olaparib. With this chemical property in mind, we utilized the AZD2461 ligand architecture to develop a CNS penetrant and PARP-1 selective imaging probe, in order to investigate PARP-1 mediated neuroinflammation and neurodegenerative diseases, such as Alzheimer's and Parkinson's. Our work led to the identification of several high-affinity PARPi, including AZD2461 congener 9e (PARP-1 IC50 = 3.9 ±â€¯1.2 nM), which was further evaluated as a potential 18F-PET brain imaging probe. However, despite the similar molecular scaffolds of 9e and AZD2461, our studies revealed non-appreciable brain-uptake of [18F]9e in non-human primates, suggesting AZD2461 to be non-CNS penetrant.

The nuclear receptor Rev-erbα controls circadian thermogenic plasticity.

Circadian oscillation of body temperature is a basic, evolutionarily conserved feature of mammalian biology. In addition, homeostatic pathways allow organisms to protect their core temperatures in response to cold exposure. However, the mechanism responsible for coordinating daily body temperature rhythm and adaptability to environmental challenges is unknown. Here we show that the nuclear receptor Rev-erbα (also known as Nr1d1), a powerful transcriptional repressor, links circadian and thermogenic networks through the regulation of brown adipose tissue (BAT) function. Mice exposed to cold fare considerably better at 05:00 (Zeitgeber time 22) when Rev-erbα is barely expressed than at 17:00 (Zeitgeber time 10) when Rev-erbα is abundant. Deletion of Rev-erbα markedly improves cold tolerance at 17:00, indicating that overcoming Rev-erbα-dependent repression is a fundamental feature of the thermogenic response to cold. Physiological induction of uncoupling protein 1 (Ucp1) by cold temperatures is preceded by rapid downregulation of Rev-erbα in BAT. Rev-erbα represses Ucp1 in a brown-adipose-cell-autonomous manner and BAT Ucp1 levels are high in Rev-erbα-null mice, even at thermoneutrality. Genetic loss of Rev-erbα also abolishes normal rhythms of body temperature and BAT activity. Thus, Rev-erbα acts as a thermogenic focal point required for establishing and maintaining body temperature rhythm in a manner that is adaptable to environmental demands.

Quantitative PET Reporter Gene Imaging with [11C]Trimethoprim.

There is a need for improved methods to image genetically engineered cells, including immune cells used for cell-based therapy. Given the genetic manipulation inherent to gene therapy, the use of a reporter protein is a logical solution and positron emission tomography (PET) can provide the desired sensitivity and spatial localization. We developed a broadly applicable PET imaging strategy based on the small bacterial protein E. coli dihydrofolate reductase (Ec dhfr) and its highly specific small molecule inhibitor, trimethoprim (TMP). The difference in TMP affinity for bacterial compared to mammalian DHFR suggests that a TMP radioligand would have a low background in unmodified mammalian tissues and high retention in Ec dhfr engineered cells, providing high contrast imaging. Here, we describe the in vitro properties of [11C]TMP and show over 10-fold increased signal in transgenic Ec dhfr cells compared to control. In a mouse xenograft model, [11C]TMP rapidly accumulated in Ec dhfr carrying cells within minutes of intravenous administration. Moreover, [11C]TMP can identify less than a million xenografted cells in a small volume in tissues other than the abdominal compartment. This limit of detection is a clinically relevant number and bodes well for clinical translation especially given that [11C]TMP is an isotopologue of clinically approved antibiotic.

The sigma-2 receptor as a therapeutic target for drug delivery in triple negative breast cancer.

BACKGROUND: Triple-negative breast cancer (TNBC) is associated with high relapse rates and increased mortality when compared with other breast cancer subtypes. In contrast to receptor positive breast cancers, there are no approved targeted therapies for TNBC. Identifying biomarkers for TNBC is of high importance for the advancement of patient care. The sigma-2 receptor has been shown to be overexpressed in triple negative breast cancer in vivo and has been characterized as a marker of proliferation. The aim of the present study was to define the sigma-2 receptor as a target for therapeutic drug delivery and biomarker in TNBC. METHODS: Three TNBC cell lines were evaluated: MDA-MB-231, HCC1937 and HCC1806. Sigma-2 compounds were tested for pharmacological properties specific to the sigma-2 receptor through competitive inhibition assays. Sigma-2 receptor expression was measured through radioligand receptor saturation studies. Drug sensitivity for taxol was compared to a sigma-2 targeting compound conjugated to a cytotoxic payload, SW IV-134. Cell viability was assessed after treatments for 2 or 48 h. Sigma-2 blockade was assessed to define sigma-2 mediated cytotoxicity of SW IV-134. Caspase 3/7 activation induced by SW IV-134 was measured at corresponding treatment time points. RESULTS: SW IV-134 was the most potent compound tested in two of the three cell lines and was similarly effective in all three. MDA-MB-231 displayed a statistically significant higher sigma-2 receptor expression and also was the most sensitive cell line evaluated to SW IV-134. CONCLUSION: Targeting the sigma-2 receptor with a cytotoxic payload was effective in all the three cell lines evaluated and provides the proof of concept for future development of a therapeutic platform for the treatment of TNBC.

Identification of a Putative α-synuclein Radioligand Using an in silico Similarity Search.

PURPOSE: Previous studies from our lab utilized an ultra-high throughput screening method to identify compound 1 as a small molecule that binds to alpha-synuclein (α-synuclein) fibrils. The goal of the current study was to conduct a similarity search of 1 to identify structural analogs having improved in vitro binding properties for this target that could be labeled with radionuclides for both in vitro and in vivo studies for measuring α-synuclein aggregates. METHODS: Using 1 as a lead compound in a similarity search, isoxazole derivative 15 was identified to bind to α-synuclein fibrils with high affinity in competition binding assays. A photocrosslinkable version was used to confirm binding site preference. Derivative 21, the iodo-analog of 15, was synthesized, and subsequently radiolabeled isotopologs [125I]21 and [11C]21 were successfully synthesized for use in in vitro and in vivo studies, respectively. [125I]21 was used in radioligand binding studies in post-mortem Parkinson's disease (PD) and Alzheimer's disease (AD) brain homogenates. In vivo imaging of an α-synuclein mouse model and non-human primates was performed with [11C]21. RESULTS: In silico molecular docking and molecular dynamic simulation studies for a panel of compounds identified through a similarity search, were shown to correlate with Ki values obtained from in vitro binding studies. Improved affinity of isoxazole derivative 15 for α-synuclein binding site 9 was indicated by photocrosslinking studies with CLX10. Design and successful (radio)synthesis of iodo-analog 21 of isoxazole derivative 15 enabled further in vitro and in vivo evaluation. Kd values obtained in vitro with [125I]21 for α-synuclein and Aß42 fibrils were 0.48 ± 0.08 nM and 2.47 ± 1.30 nM, respectively. [125I]21 showed higher binding in human postmortem PD brain tissue compared with AD tissue, and low binding in control brain tissue. Lastly, in vivo preclinical PET imaging showed elevated retention of [11C]21 in PFF-injected mouse brain. However, in PBS-injected control mouse brain, slow washout of the tracer indicates high non-specific binding. [11C]21 showed high initial brain uptake in a healthy non-human primate, followed by fast washout that may be caused by rapid metabolic rate (21% intact [11C]21 in blood at 5 min p.i.). CONCLUSION: Through a relatively simple ligand-based similarity search, we identified a new radioligand that binds with high affinity (<10 nM) to α-synuclein fibrils and PD tissue. Although the radioligand has suboptimal selectivity for α-synuclein towards Aß and high non-specific binding, we show here that a simple in silico approach is a promising strategy to identify novel ligands for target proteins in the CNS with the potential to be radiolabeled for PET neuroimaging studies.

A Novel Brain PET Radiotracer for Imaging Alpha Synuclein Fibrils in Multiple System Atrophy.

Abnormal α-synuclein (α-syn) aggregation characterizes α-synucleinopathies, including Parkinson's disease (PD) and multiple system atrophy (MSA). However, no suitable positron emission tomography (PET) radiotracer for imaging α-syn in PD and MSA exists currently. Our structure-activity relationship studies identified 4-methoxy-N-(4-(3-(pyridin-2-yl)-3,8-diazabicyclo[3.2.1]octan-8-yl)phenyl)benzamide (4i) as a PET radiotracer candidate for imaging α-syn. In vitro assays revealed high binding of 4i to recombinant α-syn fibrils (inhibition constant (Ki) = 6.1 nM) and low affinity for amyloid beta (Aß) fibrils in Alzheimer's disease (AD) homogenates. However, [3H]4i also exhibited high specific binding to AD, progressive supranuclear palsy, and corticobasal degeneration tissues as well as PD and MSA tissues, suggesting notable affinity to tau. Nevertheless, the specific binding to pathologic α-syn aggregates in MSA post-mortem brain tissues was significantly higher than in PD tissues. This finding demonstrated the potential use of [11C]4i as a PET tracer for imaging α-syn in MSA patients. Nonhuman primate PET studies confirmed good brain uptake and rapid washout for [11C]4i.

[18F]ROStrace detects oxidative stress in vivo and predicts progression of Alzheimer's disease pathology in APP/PS1 mice.

BACKGROUND: Oxidative stress is implicated in the pathogenesis of the most common neurodegenerative diseases, such as Alzheimer's disease (AD). However, tracking oxidative stress in the brain has proven difficult and impeded its use as a biomarker. Herein, we investigate the utility of a novel positron emission tomography (PET) tracer, [18F]ROStrace, as a biomarker of oxidative stress throughout the course of AD in the well-established APP/PS1 double-mutant mouse model. PET imaging studies were conducted in wild-type (WT) and APP/PS1 mice at 3 different time points, representing early (5 mo.), middle (10 mo.), and advanced (16 mo.) life (n = 6-12, per sex). Semi-quantitation SUVRs of the plateau phase (40-60 min post-injection; SUVR40-60) of ten brain subregions were designated by the Mirrione atlas and analyzed by Pmod. Statistical parametric mapping (SPM) was used to distinguish brain regions with elevated ROS in APP/PS1 relative to WT in both sexes. The PET studies were validated by ex vivo autoradiography and immunofluorescence with the parent compound, dihydroethidium. RESULTS: [18F]ROStrace retention was increased in the APP/PS1 brain compared to age-matched controls by 10 mo. of age (p < 0.0001) and preceded the accumulation of oxidative damage in APP/PS1 neurons at 16 mo. (p < 0.005). [18F]ROStrace retention and oxidative damages were higher and occurred earlier in female APP/PS1 mice as measured by PET (p < 0.001), autoradiography, and immunohistochemistry (p < 0.05). [18F]ROStrace differences emerged midlife, temporally and spatially correlating with increased Aß burden (r2 = 0.36; p = 0.0003), which was also greatest in the female brain (p < 0.001). CONCLUSIONS: [18F]ROStrace identifies increased oxidative stress and neuroinflammation in APP/PS1 female mice, concurrent with increased amyloid burden midlife. Differences in oxidative stress during this crucial time may partially explain the sexual dimorphism in AD. [18F]ROStrace may provide a long-awaited tool to stratify at-risk patients who may benefit from antioxidant therapy prior to irreparable neurodegeneration.

Preclinical Development of [211At]meta- astatobenzylguanidine ([211At]MABG) as an Alpha Particle Radiopharmaceutical Therapy for Neuroblastoma.

PURPOSE: [131I]meta-iodobenzylguanidine ([131I]MIBG) is a targeted radiotherapeutic administered systemically to deliver beta particle radiation in neuroblastoma. However, relapses in the bone marrow are common. [211At]meta-astatobenzylguanidine ([211At] MABG) is an alpha particle emitter with higher biological effectiveness and short path length which effectively sterilizes microscopic residual disease. Here we investigated the safety and antitumor activity [211At]MABG in preclinical models of neuroblastoma. EXPERIMENTAL DESIGN: We defined the maximum tolerated dose (MTD), biodistribution, and toxicity of [211At]MABG in immunodeficient mice in comparison with [131I]MIBG. We compared the antitumor efficacy of [211At]MABG with [131I]MIBG in three murine xenograft models. Finally, we explored the efficacy of [211At]MABG after tail vein xenografting designed to model disseminated neuroblastoma. RESULTS: The MTD of [211At]MABG was 66.7 MBq/kg (1.8 mCi/kg) in CB17SC scid-/- mice and 51.8 MBq/kg (1.4 mCi/kg) in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice. Biodistribution of [211At]MABG was similar to [131I]MIBG. Long-term toxicity studies on mice administered with doses up to 41.5 MBq/kg (1.12 mCi/kg) showed the radiotherapeutic to be well tolerated. Both 66.7 MBq/kg (1.8 mCi/kg) single dose and fractionated dosing 16.6 MBq/kg/fraction (0.45 mCi/kg) × 4 over 11 days induced marked tumor regression in two of the three models studied. Survival was significantly prolonged for mice treated with 12.9 MBq/kg/fraction (0.35 mCi/kg) × 4 doses over 11 days [211At]MABG in the disseminated disease (IMR-05NET/GFP/LUC) model (P = 0.003) suggesting eradication of microscopic disease. CONCLUSIONS: [211At]MABG has significant survival advantage in disseminated models of neuroblastoma. An alpha particle emitting radiopharmaceutical may be effective against microscopic disseminated disease, warranting clinical development.

Pre-clinical investigation of astatine-211-parthanatine for high-risk neuroblastoma.

Astatine-211-parthanatine ([211At]PTT) is an alpha-emitting radiopharmaceutical therapeutic that targets poly(adenosine-diphosphate-ribose) polymerase 1 (PARP1) in cancer cells. High-risk neuroblastomas exhibit among the highest PARP1 expression across solid tumors. In this study, we evaluated the efficacy of [211At]PTT using 11 patient-derived xenograft (PDX) mouse models of high-risk neuroblastoma, and assessed hematological and marrow toxicity in a CB57/BL6 healthy mouse model. We observed broad efficacy in PDX models treated with [211At]PTT at the maximum tolerated dose (MTD 36 MBq/kg/fraction x4) administered as a fractionated regimen. For the MTD, complete tumor response was observed in 81.8% (18 of 22) of tumors and the median event free survival was 72 days with 30% (6/20) of mice showing no measurable tumor >95 days. Reversible hematological and marrow toxicity was observed 72 hours post-treatment at the MTD, however full recovery was evident by 4 weeks post-therapy. These data support clinical development of [211At]PTT for high-risk neuroblastoma.

Cell-Proliferation Imaging for Monitoring Response to CDK4/6 Inhibition Combined with Endocrine-Therapy in Breast Cancer: Comparison of [18F]FLT and [18F]ISO-1 PET/CT.

PURPOSE: Cyclin-dependent kinase 4/6 (CDK4/6) inhibitors in combination with endocrine-therapy have emerged as an important regimen of care for estrogen receptor (ER)-positive metastatic breast cancer, although identifying predictive biomarkers remains a challenge. We assessed the ability of two PET-proliferation tracers, [18F]FLT and [18F]ISO-1, for evaluating response to CDK4/6-inhibitor (palbociclib) and ER-antagonist (fulvestrant). EXPERIMENTAL DESIGN: To determine the effect of CDK4/6 inhibition combined with estrogen-blockade, we assessed cell proliferation in six breast cancer cell lines after 1, 3, and 6 days of treatment with palbociclib and/or fulvestrant. These data were correlated to in vitro radiotracer assays and results were verified by longitudinal [18F]FLT and [18F]ISO-1 micro-PET imaging performed in MCF7 tumor-bearing mice. RESULTS: All palbociclib-sensitive cell lines showed decreased [18F]FLT accumulation and S-phase depletion after treatment, with both measures augmented by combination therapy. In contrast, these cells showed changes in [18F]ISO-1 analogue-binding and G0 arrest only after prolonged treatment. MicroPET imaging of MCF7 xenografts showed a significant decrease in [18F]FLT but no changes in [18F]ISO-1 uptake in all treated mice on day 3. On day 14, however, mice treated with combination therapy showed a significant decrease in [18F]ISO-1, corresponding to G0 arrest, while maintaining reduced [18F]FLT uptake, which corresponded to S-phase depletion. CONCLUSIONS: Our data suggest complementary roles of [18F]FLT and [18F]ISO-1 PET in evaluating tumor-proliferation after combined CDK4/6 inhibitor and endocrine therapy in breast cancer. [18F]FLT is more sensitive to immediate changes in S-phase, whereas [18F]ISO-1 can assess more delayed changes related to cell-cycle arrest and transition to G0 quiescence from combination therapy. These data suggest a potential role for early prediction of long-term response using these imaging biomarkers.

Targeting PARP-1 with Alpha-Particles Is Potently Cytotoxic to Human Neuroblastoma in Preclinical Models.

Alpha-emitters can be pharmacologically delivered for irradiation of single cancer cells, but cellular lethality could be further enhanced by targeting alpha-emitters directly to the nucleus. PARP-1 is a druggable protein in the nucleus that is overexpressed in neuroblastoma compared with normal tissues and is associated with decreased survival in high-risk patients. To exploit this, we have functionalized a PARP inhibitor (PARPi) with an alpha-emitter astatine-211. This approach offers enhanced cytotoxicity from conventional PARPis by not requiring enzymatic inhibition of PARP-1 to elicit DNA damage; instead, the alpha-particle directly induces multiple double-strand DNA breaks across the particle track. Here, we explored the efficacy of [211At]MM4 in multiple cancers and found neuroblastoma to be highly sensitive in vitro and in vivo Furthermore, alpha-particles delivered to neuroblastoma show antitumor effects and durable responses in a neuroblastoma xenograft model, especially when administered in a fractionated regimen. This work provides the preclinical proof of concept for an alpha-emitting drug conjugate that directly targets cancer chromatin as a therapeutic approach for neuroblastoma and perhaps other cancers.

In vivo imaging of beta-cell mass in rats using 18F-FP-(+)-DTBZ: a potential PET ligand for studying diabetes mellitus.

UNLABELLED: Recent studies on gene expression of beta-cell mass (BCM) in the pancreas showed that vesicular monoamine transporter 2 (VMAT2) is highly expressed in the BCM (mainly in the islets of Langerhans). Imaging pancreatic BCM may provide an important tool for understanding the relationship between loss of insulin-secreting beta-cells and onset of diabetes mellitus. In this article, 9-fluoropropyl-(+)-dihydrotetrabenazine (FP-(+)-DTBZ), which is a VMAT2 imaging agent, was evaluated as a PET agent for estimating BCM in vivo. METHODS: Organ biodistribution after an intravenous injection of (18)F-FP-(+)-DTBZ (active isomer) and (18)F-FP-(-)-DTBZ (inactive isomer) was evaluated in normal rats. The specificity of uptake of (18)F-FP-(+)-DTBZ was assessed by a pretreatment (3.8 mg of (+)-DTBZ per kilogram and 3.5 mg of FP-(+)-DTBZ per kilogram, intravenously, 5 min prior) or coadministration (2 mg of (+)-DTBZ per kilogram). PET studies were performed in normal rats. RESULTS: The in vivo biodistribution of (18)F-FP-(+)-DTBZ in rats showed the highest uptake in the pancreas (5% dose/g at 30 min after injection), whereas (18)F-FP-(-)-DTBZ showed a very low pancreas uptake. Rats pretreated with FP-(+)-DTBZ displayed a 78% blockade of pancreas uptake. PET studies in normal rats demonstrated an avid pancreas uptake of (18)F-FP-(+)-DTBZ. CONCLUSION: The preliminary data obtained with (18)F-FP-(+)-DTBZ suggest that this fluorinated derivative of DTBZ shows good pancreas specificity and has the potential to be useful for quantitative measurement of VMAT2 binding sites reflecting BCM in the pancreas.

In vivo imaging of vesicular monoamine transporter 2 in pancreas using an (18)F epoxide derivative of tetrabenazine.

OBJECTIVES: Development of imaging agents for pancreatic beta cell mass may provide tools for studying insulin-secreting beta cells and their relationship with diabetes mellitus. In this paper, a new imaging agent, [(18)F](+)-2-oxiranyl-3-isobutyl-9-(3-fluoropropoxy)-10-methoxy-2,3,4,6,7,11b-hexahydro-1H-pyrido[2,1-a]isoquinoline [(18)F](+)4, which displays properties targeting vesicular monoamine transporter 2 (VMAT2) binding sites of beta cells in the pancreas, was evaluated as a positron emission tomography (PET) agent for estimating beta cell mass in vivo. The hydrolyzable epoxide group of (+)4 may provide a mechanism for shifting biodistribution from liver to kidney, thus reducing the background signal. METHODS: Both (18)F- and (19)F-labeled (+) and (-) isomers of 4 were synthesized and evaluated. Organ distribution was carried out in normal rats. Uptake of [(18)F](+)4 in pancreas of normal rats was measured and correlated with blocking studies using competing drugs, (+)dihydrotetrabenazine [(+)-DTBZ] or 9-fluoropropyl-(+)dihydro tetrabenazine [FP-(+)-DTBZ, (+)2]. RESULTS: In vitro binding study of VMAT2 using rat brain striatum showed a K(i) value of 0.08 and 0.15 nM for the (+)4 and (+/-)4, respectively. The in vivo biodistribution of [(18)F](+)4 in rats showed the highest uptake in the pancreas (2.68 %ID/g at 60 min postinjection). In vivo competition experiments with cold FP-(+)-DTBZ, (+)2, (3.5 mg/kg, 5 min iv pretreatment) led to a significant reduction of pancreas uptake (85% blockade at 60 min). The inactive isomer [(18)F](-)4 showed significantly lower pancreas uptake (0.22 %ID/g at 30 min postinjection). Animal PET imaging studies of [(18)F](+)4 in normal rats demonstrated an avid pancreatic uptake in rats. CONCLUSION: The preliminary results suggest that the epoxide, [(18)F](+)4, is highly selective in binding to VMAT2 and it has an excellent uptake in the pancreas of rats. The liver uptake was significantly reduced through the use of the epoxide group. Therefore, it may be potentially useful for imaging beta cell mass in the pancreas.

Synthesis and evaluation of indolinyl- and indolylphenylacetylenes as PET imaging agents for beta-amyloid plaques.

Two new phenylacetylene derivatives, 5-((4-(2-(2-(2-fluoroethoxy)ethoxy)ethoxy)phenyl)ethynyl)indoline 8 and 5-((4-(2-(2-(2-fluoroethoxy)ethoxy)ethoxy)phenyl)ethynyl)-1H-indole 14, targeting beta-amyloid (Abeta) plaques have been prepared. In vitro binding carried out in tissue homogenates prepared from postmortem AD brains with [(125)I]IMPY (6-iodo-2-(4'-dimethylamino-)phenyl-imidazo[1,2-a]pyridine) as the radioligand indicated good binding affinities (K(i)=4.0 and 1.5nM for 8 and 14, respectively). Brain penetration of the corresponding radiofluorinated ligands, evaluated in the normal mice, showed good initial brain penetration (4.50 and 2.43% ID/g (injected dose/gram) for [(18)F]8 and [(18)F]14 at 2min after injection) with moderate to low washout rates from the brain (1.71% ID/g at 2h and 2.10% ID/g at 3h, respectively). Autoradiography and homogenate binding studies demonstrated the high specific binding of [(18)F]14 to the Abeta plaques; however, [(18)F]8 showed low specific binding. These preliminary results identified that indolylphenylacetylene, 14, may be a good lead for further structural modification to develop a useful Abeta plaque imaging agent.