Potential Positron Emission Tomography Tracer Research Articles

Synthesis and preclinical evaluation of [18F]AlF-NODA-MP-C6-CTHRSSVVC as a PET tracer for CD163-positive tumor-infiltrating macrophages

Positron emission tomography (PET) can provide information about tumor-associated macrophage (TAM) infiltration, as long as a suitable tracer is available. This study aimed to evaluate the radiolabeled peptide [18F]AlF-NODA-MP-C6-CTHRSSVVC as a potential PET tracer for imaging of the CD163 receptor, which is expressed on M2-type tumor-associated macrophages. The conjugated peptide NODA-MP-C6-CTHRSSVVC was labeled with aluminum [18F]fluoride. Tracer binding and its biodistribution were evaluated in an in vitro binding assay and in healthy BALB/c mice, respectively. In addition, different treatments with cyclophosphamide in tumor-bearing mice were used to assess whether the tracer could detect differences in CD163 expression caused by differential TAM infiltration. After 7 days of treatment, animals were injected with [18F]AlF-NODA-MP-C6-CTHRSSVVC, and a 60-min dynamic PET scan was performed, followed by an ex vivo biodistribution study. [18F]AlF-NODA-MP-C6-CTHRSSVVC was prepared in 23 ± 6 % radiochemical yield and showed approximately 50 % of specific receptor-mediated binding in an in vitro binding assay on human CD163-expressing tissue homogenates. No CD163-mediated binding of [18F]AlF-NODA-MP-C6-CTHRSSVVC was detected by PET under normal physiological conditions in healthy BALB/c mice. On the other hand, CD163-positive xenograft tumors were clearly visualized with PET and a positive correlation was found between CD163 levels and the [18F]AlF-NODA-MP-C6-CTHRSSVVC tumor-to-muscle ratio (TMR) obtained from the PET images (Pearson r = 0.76, p = 0.002). No significant differences in the CD163 protein level and in the tracer uptake between treatment groups were found in the tumors. Taken together, [18F]AlF-NODA-MP-C6-CTHRSSVVC appears a promising candidate PET tracer for M2-type TAM, as it binds specifically to CD163 in vitro and its tumor uptake correlates well with CD163 expression in vivo.

Discovery and Evaluation of Imidazo[2,1-b][1,3,4]thiadiazole Derivatives as New Candidates for α-Synuclein PET Imaging.

α-synuclein (α-syn) pathologies are central to the development of synucleinopathies including Parkinson's disease (PD). Positron emission tomography (PET) imaging of α-syn pathologies is one strategy to facilitate the diagnosis, understanding, and treatment of synucleinopathies, but has been restricted by the lack of specific α-syn PET probes. In this work, we identified 2,6-disubstituted imidazo[2,1-b][1,3,4]thiadiazole (ITA) as a new α-syn-binding scaffold. Through autoradiography studies, we discovered an iodinated lead compound [125I]ITA-3, with moderate binding affinity (IC50 = 55 nM) to α-syn pathologies in human PD brain sections. Modified from [125I]ITA-3, we developed a potential PET tracer, [18F]FITA-2 (radiochemical yield >25%, molar activity >110 GBq/μmol), which demonstrated clear signals in α-syn-rich regions in human PD brain tissues (IC50 = 245 nM), good brain uptake (SUVpeak = 2.80 ± 0.45), and fast clearance rate in rats. Overall, [18F]FITA-2 appears to be a promising candidate for α-syn PET imaging and merits further development.

Synthesis of 18F-labeled Aryl Trifluoromethyl Sulfones, -Sulfoxides, and -Sulfides for Positron Emission Tomography.

Positron emission tomography (PET) is becoming increasingly important in nuclear medicine and drug discovery. To date, the development of many potential PET tracers is hampered by the lack of suitable synthetic pathways for their preparation. This is particularly true for the highly desired radiolabeling of compounds bearing [18F]CF3-groups. For instance, S(O)nCF3-groups (n=0, 1, 2) serve as structural motif in a range of biologically active compounds, but their radiosynthesis remains largely unprecedented (for n=1, 2). Herein, we describe general methods for the radiosynthesis of 18F-labeled aryl trifluoromethyl sulfones, -sulfoxides, and -sulfides. All three methods are operationally straightforward, start from widely available precursors, i.e., sulfonyl fluorides and thiophenols, and make use of the recently established [18F]Ruppert-Prakash reagent. Further, the syntheses display good functional group tolerance as demonstrated by the 18F-labeling of more than 40 compounds. The applicability of the new method is demonstrated by the radiolabeling of three bioactive molecules, optionally to be used as PET tracers. In a broader context, this work presents a substantial expansion of the chemical space of radiofluorinated structural motifs to be used for the development of new PET tracers.

8-Bicycloalkyl-CPFPX derivatives as potent and selective tools for in vivo imaging of the A1 adenosine receptor

Imaging of the A1 adenosine receptor (A1R) by positron emission tomography (PET) with 8-cyclopentyl-3-(3-[18F]fluoropropyl)-1-propyl-xanthine ([18F]CPFPX) has been widely used in preclinical and clinical studies. However, this radioligand suffers from rapid peripheral metabolism and subsequent accumulation of radiometabolites in the vascular compartment. In the present work, we prepared four derivatives of CPFPX by replacement of the cyclopentyl group with norbornane moieties. These derivatives were evaluated by competition binding studies, microsomal stability assays and LC-MS analysis of microsomal metabolites. In addition, the 18F-labeled isotopologue of 8-(1-norbornyl)-3-(3-fluoropropyl)-1-propylxanthine (1-NBX) as the most promising candidate was prepared by radiofluorination of the corresponding tosylate precursor and the resulting radioligand ([18F]1-NBX) was evaluated by permeability assays with Caco-2 cells and in vitro autoradiography in rat brain slices. Our results demonstrate that 1-NBX exhibits significantly improved A1R affinity and selectivity when compared to CPFPX and that it does not give rise to lipophilic metabolites expected to cross the blood-brain-barrier in microsomal assays. Furthermore, [18F]1-NBX showed a high passive permeability (Pc = 6.9 ± 2.9 × 10−5 cm/s) and in vitro autoradiography with this radioligand resulted in a distribution pattern matching A1R expression in the brain. Moreover, a low degree of non-specific binding (5%) was observed. Taken together, these findings identify [18F]1-NBX as a promising candidate for further preclinical evaluation as potential PET tracer for A1R imaging.

Multi-parameter optimization: Development of a morpholin-3-one derivative with an improved kinetic profile for imaging monoacylglycerol lipase in the brain

Monoacylglycerol lipase (MAGL) is a gatekeeper in regulating endocannabinoid signaling and has gained substantial attention as a therapeutic target for neurological disorders. We recently discovered a morpholin-3-one derivative as a novel scaffold for imaging MAGL via positron emission tomography (PET). However, its slow kinetics in vivo hampered the application. In this study, structural optimization was conducted and eleven novel MAGL inhibitors were designed and synthesized. Based on the results from MAGL inhibitory potency, in vitro metabolic stability and surface plasmon resonance assays, we identified compound 7 as a potential MAGL PET tracer candidate. [11C]7 was synthesized via direct 11CO2 fixation method and successfully mapped MAGL distribution patterns on rodent brains in in vitro autoradiography. PET studies in mice using [11C]7 demonstrated its improved kinetic profile compared to the lead structure. Its high specificity in vivo was proved by using MAGL KO mice. Although further studies confirmed that [11C]7 is a P-glycoprotein (P-gp) substrate in mice, its low P-gp efflux ratio on cells transfected with human protein suggests that it should not be an issue for the clinical translation of [11C]7 as a novel reversible MAGL PET tracer in human subjects. Overall, [11C]7 ([11C]RO7284390) showed promising results warranting further clinical evaluation.

Discovery, synthesis and evaluation of novel reversible monoacylglycerol lipase radioligands bearing a morpholine-3-one scaffold

Monoacylglycerol lipase (MAGL) is a serine hydrolase that plays an important role in the endocannabinoid degradation in the brain. It has recently emerged as a promising therapeutic target in the treatment of neuroinflammatory and neurodegenerative diseases, such as multiple sclerosis, Alzheimer's disease and Parkinson's disease. Development of MAGL-specific radioligands for non-invasive imaging by positron-emission tomography (PET) would deepen our knowledge on the relevant pathological changes in diseased states and accelerate drug discovery. In this study, we report the selection and synthesis of two morpholine-3-one derivatives as potential reversible MAGL PET tracer candidates based on their multiparameter optimization scores. Both compounds ([11C]1, [11C]2) were radiolabeled by direct [11C]CO2 fixation and the in vitro autoradiographic studies demonstrated their specificity and selectivity towards MAGL. Dynamic PET imaging using MAGL knockout and wild-type mice confirmed the in vivo specificity of [11C]2. Our preliminary results indicate that morpholine-3-one derivative [11C]2 ([11C]RO7279991) binds to MAGL in vivo, and this molecular scaffold could serve as an alternative lead structure to image MAGL in the central nervous system.

Evaluation of [68Ga]Ga-NODAGA-RGD for PET Imaging of Rat Autoimmune Myocarditis.

The 68Gallium-labeled 1,4,7-triazacyclononane-1-glutaric acid-4,7-diacetic acid conjugated radiolabelled arginine-glycine-aspartic acid peptide ([68Ga]Ga-NODAGA-RGD) is a positron emission tomography (PET) tracer binding to cell surface receptor αvβ3 integrin that is upregulated during angiogenesis and inflammation. We studied whether αvβ3 targeting PET imaging can detect myocardial inflammation in a rat model of autoimmune myocarditis. To induce myocarditis, rats (n = 8) were immunized with porcine cardiac myosin in complete Freund's adjuvant on days 0 and 7. Control rats (n = 8) received Freund's adjuvant alone. On day 21, in vivo PET/CT imaging with [68Ga]Ga-NODAGA-RGD followed by ex vivo autoradiography and immunohistochemistry were carried out. Inflammatory lesions were detected histologically in the myocardium of 7 out of 8 immunized rats. In vivo PET images showed higher [68Ga]Ga-NODAGA-RGD accumulation in the myocardium of rats with inflammation than the non-inflamed myocardium of control rats (SUVmean 0.4 ± 0.1 vs. 0.1 ± 0.02; P = 0.00006). Ex vivo autoradiography and histology confirmed that [68Ga]Ga-NODAGA-RGD uptake co-localized with inflammatory lesions containing αvβ3 integrin-positive capillary-like structures. A non-specific [68Ga]Ga-DOTA-(RGE)2 tracer showed 76% lower uptake than [68Ga]Ga-NODAGA-RGD in the inflamed myocardium. Our results indicate that αvβ3 integrin-targeting [68Ga]Ga-NODAGA-RGD is a potential PET tracer for the specific detection of active inflammatory lesions in autoimmune myocarditis.

Novel Thienopyrimidine-Based PET Tracers for P2Y12 Receptor Imaging in the Brain.

The P2Y12 receptor (P2Y12R) is uniquely expressed on microglia in the brain, and its expression level directly depends on the microglial activation state. Therefore, P2Y12R provides a promising imaging marker for distinguishing the pro- and anti-inflammatory microglial phenotypes, both of which play crucial roles in neuroinflammatory diseases. In this study, three P2Y12R antagonists were selected from the literature, radiolabeled with carbon-11 or fluorine-18, and evaluated in healthy Wistar rats. Brain imaging was performed with and without blocking of efflux transporters P-glycoprotein and breast cancer resistance protein using tariquidar. Low brain uptake in healthy rats was observed for all tracers at baseline conditions, whereas blocking of efflux transporters resulted in a strong (6–7 fold) increase in brain uptake for both of them. Binding of the most promising tracer, [18F]3, was further evaluated by in vitro autoradiography on rat brain sections, ex vivo metabolite studies, and in vivo P2Y12R blocking studies. In vitro binding of [18F]3 on rat brain sections indicated high P2Y12R targeting with approximately 70% selective and specific binding. At 60 min post-injection, over 95% of radioactivity in the brain accounted for an intact tracer. In blood plasma, still 40% intact tracer was found, and formed metabolites did not enter the brain. A moderate P2Y12R blocking effect was observed in vivo by positron emission tomography (PET) imaging with [18F]3 (p = 0.04). To conclude, three potential P2Y12R PET tracers were obtained and analyzed for P2Y12R targeting in the brain. Unfortunately, the brain uptake appeared low. Future work will focus on the design of P2Y12R inhibitors with improved physicochemical characteristics to reduce efflux transport and increase brain penetration.

Design, Synthesis, and Evaluation of [18F]T-914 as a Novel Positron-Emission Tomography Tracer for Lysine-Specific Demethylase 1.

Histone methylation is associated with the pathophysiology of neurodevelopmental disorders. Lysine-specific demethylase 1 (LSD1) catalyzes histone demethylation in a flavin adenine dinucleotide (FAD)-dependent manner. Thus, inhibiting LSD1 enzyme activity could offer a novel way to treat neurodevelopmental disorders. Assessing LSD1 target engagement using positron-emission tomography (PET) imaging could aid in developing therapeutic LSD1 inhibitors. In this study, PET probes based on 4-(2-aminocyclopropyl)benzamide derivatives that bind irreversibly to FAD found in LSD1 were examined. By optimizing the profiles of brain penetrance and brain-penetrant metabolites, T-914 (1g) was identified as a suitable PET tracer candidate. PET studies in nonhuman primates demonstrated that [18F]1g had heterogeneous brain uptake, which corresponded to known LSD1 expression levels. Moreover, brain uptake of [18F]1g was reduced by coadministration of unlabeled 1g, demonstrating blockable binding. These data suggest that [18F]1g warrants further investigation as a potential PET tracer candidate for assessing target engagement of LSD1.

Astroglial tracer BU99008 detects multiple binding sites in Alzheimer\u2019s disease brain

With reactive astrogliosis being established as one of the hallmarks of Alzheimer’s disease (AD), there is high interest in developing novel positron emission tomography (PET) tracers to detect early astrocyte reactivity. BU99008, a novel astrocytic PET ligand targeting imidazoline-2 binding sites (I2BS) on astrocytes, might be a suitable candidate. Here we demonstrate for the first time that BU99008 could visualise reactive astrogliosis in postmortem AD brains and propose a multiple binding site [Super-high-affinity (SH), High-affinity (HA) and Low-affinity (LA)] model for BU99008, I2BS specific ligands (2-BFI and BU224) and deprenyl in AD and control (CN) brains. The proportion (%) and affinities of these sites varied significantly between the BU99008, 2-BFI, BU224 and deprenyl in AD and CN brains. Regional binding studies demonstrated significantly higher 3H-BU99008 binding in AD brain regions compared to CN. Comparative autoradiography studies reinforced these findings, showing higher specific binding for 3H-BU99008 than 3H-Deprenyl in sporadic AD brain compared to CN, implying that they might have different targets. The data clearly shows that BU99008 could detect I2BS expressing reactive astrocytes with good selectivity and specificity and hence be a potential attractive clinical astrocytic PET tracer for gaining further insight into the role of reactive astrogliosis in AD.

Nucleophilic 18F-fluorination of phosphorofluoridates and phosphonofluoridic acids via imidazole-activated precursors

18F-Labeled organofluorophosphates are important radiosynthons that have only been previously accessible via18/19F-isotope-exchange with limited molar activities. Herein, a novel 18F-fluorination methodology has been developed to prepare 18F-labeled phosphorofluoridates and phosphonofluoridic acids via the [18F]F− nucleophilic substitution of imidazole-activated precursors. The efficient one-step 18F-fluorination affords stable products in the presence of Zn(Ⅱ) with high radiochemical yields and high molar activities. This 18F-fluorination method could be used to prepare various phosphorofluoridate and phosphonofluoridic acid analogs for use as 18F-radiosynthons and potential positron emission tomography tracers.

Synthesis and evaluation of N-isopropyl-p-[11C]methylamphetamine as a novel cerebral blood flow tracer for positron emission tomography

IntroductionIncreases in fasting plasma glucose (PG) levels lead to a decrease in 2-deoxy-2-[18F]fluoro-d-glucose ([18F]FDG) uptake in the normal brain, especially in the precuneus, resulting in an Alzheimer’s disease (AD)-like uptake pattern. Therefore, patients with higher PG levels, such as those with diabetes, can be erroneously diagnosed with AD when positron emission tomography (PET) imaging is done using [18F]FDG, due to reduced uptake of [18F]FDG in the precuneus. To help avoid an erroneous diagnosis of AD due to differences in glucose metabolism, evaluating cerebral blood flow (CBF) in the brain is useful. However, current techniques such as single photon emission computed tomography (SPECT) and [15O]H2O PET have limitations regarding early diagnosis of AD because the images they produce are of low resolution. Here, we developed a novel CBF PET tracer that may be more useful than [18F]FDG for diagnosis of AD.MethodsWe synthesized and evaluated N-isopropyl-p-[11C]methylamphetamine ([11C]4) as a carbon-11-labeled analogue of the standard CBF SPECT tracer N-isopropyl-p-[123I]iodoamphetamine. Fundamental biological evaluations such as biodistribution, peripheral metabolism in mice, and brain kinetics of [11C]4 in non-human primates with PET with successive measurement of [15O]H2O were performed.Results[11C]4 was synthesized by methylation of the corresponding tributyltin precursor (2) with [11C]MeI in a palladium-promoted Stille cross-coupling reaction. The brain uptake of [11C]4 in mice peaked at 5–15 min after injection and then promptly decreased. Most radioactivity in the brain was detected in the unchanged form, although in the periphery, [11C]4 was rapidly metabolized to hydrophilic components. Acetazolamide (AZM) treatment significantly increased the brain uptake of [11C]4 without affecting the blood levels of radioactivity in mice. Preliminary kinetics analysis showed that the K1 of [11C]4 reflected regional CBF in a vehicle-treated monkey, but that the K1 did not reflect CBF in higher flow regions after AZM loading.Conclusion[11C]4 is a potential novel CBF PET tracer. Further validation studies are needed before [11C]4 can be used in humans.

Increased Tumoral Microenvironmental pH Improves Cytotoxic Effect of Pharmacologic Ascorbic Acid in Castration-Resistant Prostate Cancer Cells

BackgroundThe anticancer potential of pharmacologic ascorbic acid (AA) has been detected in a number of cancer cells. However, in vivo study suggested a strongly reduced cytotoxic activity of AA. It was known that pH could be a critical influencing factor for multiple anticancer treatments. In this study, we explored the influence of pH on the cytotoxicity of ascorbic acid. We employed castration-resistant prostate cancer (CRPC) cell lines PC3 and DU145 to observe the therapeutic effect of AA on PCa cells that were cultured with different pH in vitro. We also analyzed the influence of pH and extracellular oxidation on cytotoxicity of AA in cancer cells using reactive oxygen species (ROS) assay, cellular uptake of AA, and NADPH assay. Male BALB/c nude mice bearing prostate carcinoma xenografts (PC3 or DU145) were used to assess treatment response to AA with or without bicarbonate in vivo. The cellular uptake of AA in PCa xenografts was detected using positron emission tomography (PET). Small animal PET/CT scans were performed on mice after the administration of 6-deoxy-6-[18F] fluoro-L-ascorbic acid (18F-DFA).ResultsOur in vitro studies demonstrate that acidic pH attenuates the cytotoxic activity of pharmacologic ascorbic acid by inhibiting AA uptake in PCa cells. Additionally, we found that the cancer cell-selective toxicity of AA depends on ROS. In vivo, combination of AA and bicarbonate could provide a significant better therapeutic outcome in comparison with controls or AA single treated mice. 18F-DFA PET imaging illustrated that the treatment with NaHCO3 could significantly increase the AA uptake in tumor.ConclusionsThe alkalinity of tumor microenvironment plays an important role in anticancer efficiency of AA in CRPC. 18F-DFA PET/CT imaging could predict the therapeutic response of PCa animal model through illustration of tumoral uptake of AA. 18F-DFA might be a potential PET tracer in clinical diagnosis and treatment for CRPC.

18f]-(2S,4R)-4-Fluoroglutamine As a New Positron Emission Tomography Tracer in Multiple Myeloma

High glycolitic activity of multiple myeloma (MM) cells is the rationale for the use of Positron Emission Tomography (PET) with 18F-fluorodeoxyglucose ([18F]FDG) to detect both medullary and extramedullary disease. However, FDG-PET has some limitations, since there is a good portion of MM patients who are false-negative. Besides enhanced glycolysis, glutamine (Gln) addiction has been recently described as a metabolic feature of MM by our group. To sustain high Gln demand, MM cells increase the expression of several Gln transporters (ASCT2, SNAT1, LAT1) and are endowed with fast Gln uptake. Yet, at variance with other Gln-addicted cancers, the possible exploitation of Gln as a PET tracer in MM has never been assessed and was investigated in this study. To this purpose, we have firstly synthesized enantiopure (2S,4R)-4-Fluoroglutamine (4-FGln) and validated it as a Gln analogue in human MM cell lines (RPMI8226 and JJN3) comparing its uptake with that of 3H-labelled Gln. The intracellular levels of 4-FGln were determined by HPLC-MS/MS employing a HILIC gradient separation and multiple reaction monitoring (MRM) detection. Both Gln and 4-FGln were actively accumulated by MM cells and exhibited a strong reciprocal competition, pointing to shared transporters. Inhibition analysis revealed that ASCT2 was the major entry route of both compounds, with minor contributions from the other transporters. However, compared with Gln, 4-FGln exhibited higher affinity for both ASCT2 and LAT1 transporters. On the basis of these results, we then tested [18F]4-FGln uptake for MM detection by Positron Emission Tomography (PET) in two different in vivo murine models. Firstly, to investigate sensitivity of human MM to [18F]4-FGln in vivo, JJN3 cells were subcutaneously injected in immunodeficient NSG mice In this xenograft model, [18F]4-FGln- and[18F]FDG-PET scans were performed after plasmacytomas became palpable and repeated after one week. All the tumours were positive for [18F]FDG and displayed [18F]4-FGln uptake with Standard Uptake Values (SUV) of 1.21±1.9 and 0.99±0.07 after 2 weeks, respectively. Thereafter, the effect of bortezomib (BOR) was investigated to evaluate the potential use of [18F]4-FGln to monitor anti-MM treatment. Ten NGS mice were injected with JJN3 cells and, after 14 days, treated twice weekly with BOR, 1mg/kg, or vehicle for two weeks. PET scans were performed before and after 5 and 12 days of BOR treatment. As expected, BOR reduced tumour size as compared to vehicle. At the first post-BOR PET scan, [18F]4-FGln (SUV mean: pre 0.85±0.31; post 0.45±0.10, P<0.05), but not [18F]FDG (SUV mean: pre 0.97±0.38, post 0.75±0.14) was already significantly reduced: [18F]FDG and [18F]4-FGln uptake was reduced of 22 and 45% respectively. With both radiotracers, BOR treated animals displayed SUV mean values significantly lower than those of vehicle treated animals at post treatment PET (SUV means [18F ]FDG: BOR 0.75±0.14; vehicle 1.27±0.34, P<0.05; SUV mean [18F]4-FGln: BOR 0.45±0.10 ; vehicle: 0.73±0.18 ; P <0.05). Thereafter, to mimic BOR-resistant MM in a syngeneic mouse model, C57BL/6 mice were injected intravenously with Vk12598 cells obtained from transgenic Vk*MYC mice repeatedly treated with sub-optimal doses of BOR. Upon injection into C57BL/6 mice, Vk12598 cells colonize the BM without lytic lesions and extensively colonize the spleen generating an aggressive MM that brings animals to death within five weeks. PET scans were performed with [18F]4-FGln and [18F]FDG before Vk*MYC MM cells injection and after three, four and five weeks. Blood samples for M-spike evaluation were obtained in parallel. Four weeks after MM cells injection a significant increase of both [18F]4-FGln and [18F]FDG uptake was detected in spleens (SUV mean: 1.14±0.23, P=0.018; 0.94±0.24, P= 0.005). In both MM models, the volume of distribution of [18F]4-F-Gln did not overlap that of [18F]FDG. In conclusion, our data indicate that [18F]-(2S,4R)-4-Fluoroglutamine is a new potential PET tracer in pre-clinical MM models especially of extramedullary disease, either in a BOR-sensitive or in a BOR-resistant context, supporting the exploitation of Gln addiction for diagnostic purposes in MM patients. Disclosures Giuliani: Janssen: Research Funding.

Synthesis and evaluation of novel potent TSPO PET ligands with 2-phenylpyrazolo[1,5-a]pyrimidin-3-yl acetamide

Translocator protein (TSPO) expression is closely related with neuroinflammation and neuronal damage which might cause several central nervous system diseases. Herein, a series of TSPO ligands (11a–c and 13a–d) with a 2-phenylpyrazolo[1,5-a]pyrimidin-3-yl acetamide structure were prepared and evaluated via an in vitro binding assay. Most of the novel ligands exhibited a nano-molar affinity for TSPO, which was better than that of DPA-714. Particularly, 11a exhibited a subnano-molar TSPO binding affinity with suitable lipophilicity for in vivo brain studies. After radiolabeling with fluorine-18, [18F]11a was used for a dynamic positron emission tomography (PET) study in a rat LPS-induced neuroinflammation model; the inflammatory lesion was clearly visualized with a superior target-to-background ratio compared to [18F]DPA-714. An immunohistochemical examination of the dissected brains confirmed that the uptake location of [18F]11a in the PET study was consistent with a positively activated microglia region. This study proved that [18F]11a could be employed as a potential PET tracer for detecting neuroinflammation and could give possibility for diagnosis of other diseases, such as cancers related with TSPO expression.

Novel 18F-Labeled α-Methyl-Phenylalanine Derivative with High Tumor Accumulation and Ideal Pharmacokinetics for Tumor-Specific Imaging.

Positron emission tomography (PET) imaging with 18F-labeled α-methyl-substituted amino acids exerts significant influence on differential diagnosis of malignant tumors and tumor-like lesions. Exclusive uptake via L-type amino acid transporter 1 (LAT1), a tumor-specific transporter, accounts for their excellent tumor specificity and low background accumulation. However, further refinement and optimization in their tumor accumulation and pharmacokinetics are sorely needed. To address these issues, we newly designed 18F-labeled α-methyl-phenylalanine (18F-FAMP) regioisomers (2-, 3-, or 4-18F-FAMP) and stereoisomers (L- or D-form), and we comprehensively evaluated their potential as tumor-imaging agents. 18F-FAMPs were prepared from α-methyl phenylalanine by electrophilic radiofluorination and purified by reversed-phase HPLC. In biodistribution studies on normal mice, L-2-18F-FAMP and the three D-18F-FAMPs showed faster blood clearance and lower renal accumulation than L-3-18F-FAMP or L-4-18F-FAMP. In LS180 human colorectal cancer cell line xenograft mice, L-2-18F-FAMP exhibited significantly higher tumor accumulation than the D-18F-FAMPs or a clinically relevant tracer, L-3-18F-α-methyl-tyrosine (18F-FAMT) (p < 0.05). The renal accumulation levels of L-2-18F-FAMP were significantly lower than that of 18F-FAMT (p < 0.01). LAT-1 specificity of L-2-18F-FAMP was validated in the cellular uptake studies. The PET imaging with L-2-18F-FAMP clearly visualized the tumor as early as 1 h after injection, and the high tumor accumulation level was retained for 3 h. These findings suggest that L-2-18F-FAMP constitutes a potential PET tracer for tumor-specific imaging.

Biological evaluation of histidine and tryptophan labeled with gallium-68 as potential tumor imaging agents

Radiolabeled with gallium-68 amino acids can be interesting and promising probes for tumor imaging using positron emission tomography (PET). Radiolabeled amino acids target the increased amino acid transport in cancer cells compared with normal tissues. In this study we labeled two amino acids histidine and tryptophan with 68Ga, investigated its biodistribution in Wistar rats with subcutaneously transplanted cholangioma RS-1 by gamma counting and compared them with 68GaCl3. The accumulation of 68Ga-histidine in tumor tissue increased approximately 6 fold from 0.11±0.04 %ID/g at 5 min post injection (p.i.) to 0.67±0.07 %ID/g at 3 h p.i. The amount of 68Ga-tryptophan in tumor was also risen 2.4 fold from 0.34±0.18 %ID/g to 0.80±0.06 %ID/g at 5 min and 3 h p.i., respectively. In contrast, the uptake of 68GaCl3 decreased throughout the study from 0.34±0.07 %ID/g to 0.13±0.04 %ID/g. The uptake and retention of 68Ga-histidine and 68Ga-tryptophan in non-target organs, except kidney and femur, were higher than those of 68GaCl3. In conclusion, the obtained results suggest that 68Ga-histidine and 68Ga-tryptophan could serve as potential new PET tracers for tumor imaging.

New fluoroethyl phenylalanine analogues as potential LAT1-targeting PET tracers for glioblastoma

The use of O-(2-[18F]fluoroethyl)-l-tyrosine ([18F]FET) as a positron emission tomography (PET) tracer for brain tumor imaging might have some limitations because of the relatively low affinity for the L-type amino acid transporter 1 (LAT1). To assess the stereospecificity and evaluate the influence of aromatic ring modification of phenylalanine LAT1 targeting tracers, six different fluoroalkylated phenylalanine analogues were synthesized. After in vitro Ki determination, the most promising compound, 2-[18F]-2-fluoroethyl-l-phenylalanine (2-[18F]FELP), was selected for further evaluation and in vitro comparison with [18F]FET. Subsequently, 2-[18F]FELP was assessed in vivo and compared with [18F]FET and [18F]FDG in a F98 glioblastoma rat model. 2-[18F]FELP showed improved in vitro characteristics over [18F]FET, especially when the affinity and specificity for system L is concerned. Based on our results, 2-[18F]FELP is a promising new PET tracer for brain tumor imaging.

PET Imaging of the P2X7 Ion Channel with a Novel Tracer [18F]JNJ-64413739 in a Rat Model of Neuroinflammation

PurposeThe P2X7 receptor, an adenosine triphosphate (ATP)-gated purinoreceptor, has emerged as one of the key players in neuroinflammatory processes. Therefore, developing a positron emission tomography (PET) tracer for imaging of P2X7 receptors in vivo presents a promising approach to diagnose, monitor, and study neuroinflammation in a variety of brain disorders. To fulfill the goal of developing a P2X7 PET ligand as a biomarker of neuroinflammation, [18F]JNJ-64413739 has been recently disclosed.ProceduresWe evaluated [18F]JNJ-64413739 in a rat model of neuroinflammation induced by an intracerebral injection of lipopolysaccharide (LPS). In vivo brain uptake was determined by PET imaging. Upregulation of neuroinflammatory biomarkers was determined by quantitative polymerase chain reaction (qPCR). Distribution of the tracer in the brain was determined by ex vivo autoradiography (ARG). The specificity of [18F]JNJ-64413739 was confirmed by performing blocking experiments with the P2X7 antagonist JNJ-54175446.ResultsBrain regions of rats injected with LPS had a significantly increased uptake (34 % ± 3 % s.e.m., p = 0.036, t test, standardized uptake value measured over the entire scanning period) of [18F]JNJ-64413739 relative to the corresponding brain regions of control animals injected with phosphate-buffered saline (PBS). The uptake in the contralateral regions and cerebellum was not significantly different between the groups of animals. The increase in uptake of [18F]JNJ-64413739 at the LPS-injected site observed by PET imaging was concordant with ex vivo ARG, upregulation of neuroinflammatory biomarkers, and elevated P2X7 expression levels.ConclusionsWhile further work is needed to study [18F]JNJ-64413739 in other types of neuroinflammation, the current results favorably characterize [18F]JNJ-64413739 as a potential PET tracer of central neuroinflammation.

Exploring pitfalls of 64Cu-labeled EGFR-targeting peptide GE11 as a potential PET tracer.

The epidermal growth factor receptor (EGFR) represents an important molecular target for both radiotracer-based diagnostic imaging and radionuclide therapy of various cancer entities. For the delivery of radionuclides to the tumor, peptides hold great potential as a transport vehicle. With respect to EGFR, the peptide YHWYGYTPQNVI (GE11) has been reported to bind the receptor with high specificity and affinity. In the present study, GE11 with β-alanine (β-Ala-GE11) was conjugated to the chelating agent p-SCN-Bn-NOTA and radiolabeled with 64Cu for the first radio pharmacological evaluation as a potential probe for positron emission tomography (PET)-based cancer imaging. For better water solubility, an ethylene glycol-based linker was introduced between the peptide'sN terminus and the radionuclide chelator. The stability of the 64Cu-labeled peptide conjugate and its binding to EGFR-expressing tumor cells was investigated in vitro and in vivo, and then compared with the 64Cu-labeled EGFR-targeting antibody conjugate NOTA-cetuximab. The GE11 peptide conjugate [64Cu]Cu-NOTA-linker-β-Ala-GE11 ([64Cu]Cu-1) was stable in a buffer solution for at least 24h but only 50% of the original compound was detected after 24h of incubation in human serum. Stability could be improved by amidation of the peptide's C terminus (β-Ala-GE11-NH2 (2)). Binding assays with both conjugates, [64Cu]Cu-1 and [64Cu]Cu-2, using the EGFR-expressing tumor cell lines A431 and FaDu showed no specific binding. A pilot small animal PET investigation in FaDu tumor-bearing mice revealed only low tumor uptake (standard uptake value (SUV) < 0.2) for both conjugates. The best tumor-to-muscle ratio determined was 3.75 for [64Cu]Cu-1, at 1h post injection. In conclusion, the GE11 conjugates in its present form are not suitable for further biological investigations, since they presumably form aggregates.