Autoradiographic Analysis Research Articles

Design, Synthesis, and Biological Evaluation of Novel Nonsteroidal 18F-Labeled PET Probes Specifically Targeting Estrogen Receptor α.

The estrogen receptor α positive (ERα+) subtype represents nearly 70% of all breast cancers (BCs), which seriously threaten women's health. Positron emission computed tomography (PET) characterizes its superiority in detecting the recurrence and metastasis of BC. In this article, an array of novel PET probes ([18F]R-1, [18F]R-2, [18F]R-3, and [18F]R-4) targeting ERα based on the tetrahydropyridinyl indole scaffold were developed. Among them, [18F]R-3 and [18F]R-4 showed good target specificity toward ERα and could distinguish MCF-7 (ERα+) and MDA-MB-231 (ERα-) tumors efficiently. Especially, [18F]R-3 could differentiate the ERα positive/negative tumors successfully with a higher tumor-to-muscle uptake ratio (T/M) than that of [18F]R-4. The radioactivity of [18F]R-3 in the MCF-7 tumor was 5.24 ± 0.84%ID/mL and its T/M ratio was 2.49 ± 0.62 at 25 min postinjection, which might be the optimal imaging time point in PET scanning. On the contrary, [18F]R-3 did not accumulate in the MDA-MB-231 tumor at all. The autoradiography analysis of [18F]R-3 on the MCF-7 tumor-bearing mice model was consistent with the PET imaging results. [18F]R-3 exhibited the pharmacokinetic property of rapid distribution and slow clearance, making it suitable for use as a diagnostic PET probe. Overall, [18F]R-3 was capable of serving as a PET radiotracer to delineate the ERα+ tumor and was worthy of further exploitation.

Nicotine and fluoxetine alter adolescent dopamine-mediated behaviors via 5-HT1A receptor activation.

Abuse or misuse of tobacco, e-cigarettes, or antidepressants may have serious clinical consequences during adolescence, a sensitive period during brain development when the distinct neurobiology of adolescent serotonin (5-HT) and dopamine (DA) systems create unique behavioral vulnerabilities to drugs of abuse. Using a pharmacological approach, we modeled the behavioral and neurochemical effects of subchronic (4-day) nicotine (60µg/kg, i.v.) or fluoxetine (1mg/kg, i.v.) exposure in adolescent and adult male rats. Nicotine and fluoxetine significantly enhance quinpirole-induced locomotor activity and initial cocaine self-administration in adolescents, but not adults. These effects were blocked by serotonin 5-HT1A receptor antagonists, WAY-100,635 (100 µg/kg, i.v.) or S-15535 (300 µg/kg, i.v.). Neurochemical and anatomical autoradiographic analysis of 8-OH-DPAT-stimulated [35S]GTPγS reveal that prior exposure to nicotine and fluoxetine results in both overlapping and distinct effects on regional 5-HT1A receptor activity. Both fluoxetine and nicotine enhance adolescent 5-HT1A receptor activity in the primary motor cortex (M1), whereas fluoxetine alone targets prefrontal cortical neurocircuitry and nicotine alone targets the amygdala. Given their different pharmacological profiles, comparison between WAY-100,635 and S-15535 indicates that postsynaptic 5-HT1A receptors mediate the behavioral effects of prior nicotine and fluoxetine exposure. In addition, within the adolescent M1, maladaptive changes in 5-HT signaling and 5-HT1A activity after nicotine or fluoxetine exposure may potentiate hyper-responsiveness to dopaminergic drugs and prime adolescent vulnerability for future substance abuse.

Synthesis and evaluation of TSPO-targeting radioligand [18F]F-TFQC for PET neuroimaging in epileptic rats

The translocator protein (TSPO) positron emission tomography (PET) can noninvasively detect neuroinflammation associated with epileptogenesis and epilepsy. This study explored the role of the TSPO-targeting radioligand [18F]F-TFQC, an m-trifluoromethyl ER176 analog, in the PET neuroimaging of epileptic rats. Initially, [18F]F-TFQC was synthesized with a radiochemical yield of 8%–10% (EOS), a radiochemical purity of over 99%, and a specific activity of 38.21 ± 1.73 MBq/nmol (EOS). After determining that [18F]F-TFQC exhibited good biochemical properties, [18F]F-TFQC PET neuroimaging was performed in epileptic rats at multiple time points in various stages of disease progression. PET imaging showed specific [18F]F-TFQC uptake in the right hippocampus (KA-injected site, i.e., epileptogenic zone), which was most pronounced at 1 week (T/NT 1.63 ± 0.21) and 1 month (T/NT 1.66 ± 0.20). The PET results were further validated using autoradiography and pathological analysis. Thus, [18F]F-TFQC can reflect the TSPO levels and localize the epileptogenic zone, thereby offering the potential for monitoring neuroinflammation and guiding anti-inflammatory treatment in patients with epilepsy.

Acute Myelomonoblastic Leukemia (My1/De): A Preclinical Rat Model.

Since acute myeloid leukemias still represent the most aggressive type of adult acute leukemias, the profound understanding of disease pathology is of paramount importance for diagnostic and therapeutic purposes. Hence, this study aimed to explore the real-time disease fate with the establishment of an experimental myelomonoblastic leukemia (My1/De) rat model using preclinical positron emission tomography (PET) and whole-body autoradiography. In vitro [18F]F-FDG uptake studies were performed to compare the tracer accumulation in the newly cultured My1/De tumor cell line (blasts) with that in healthy control and My1/De bone marrow suspensions. Post transplantation of My1/De cells under the left renal capsule of Long-Evans rats, primary My1/De tumorigenesis, and metastatic propagation were investigated using [18F]F-FDG PET imaging, whole-body autoradiography and phosphorimage analyses. To assess the organ uptake profile of the tumor-carrying animals we accomplished ex vivo biodistribution studies. The tracer accumulation in the My1/De culture cells exceeded that of both the tumorous and the healthy bone marrow suspensions (p<0.01). Based on in vivo imaging, the subrenally transplanted My1/De cells resulted in the development of leukemia in the abdominal organs, and metastasized to the mesenterial and thoracic parathymic lymph nodes (PTLNs). The lymphatic spread of metastasis was further confirmed by the significantly higher %ID/g values of the metastatic PTLNs (4.25±0.28) compared to the control (0.94±0.34). Cytochemical staining of the peripheral blood, autopsy findings, and wright-Giemsa-stained post-mortem histological sections proved the leukemic involvement of the assessed tissues/organs. The currently established My1/De model appears to be well-suited for further leukemia-related therapeutic and diagnostic investigations.

Detection of Intestinal Inflammation by Vascular Adhesion Protein-1-Targeted [68Ga]Ga-DOTA-Siglec-9 Positron Emission Tomography in Murine Models of Inflammatory Bowel Disease

PurposeInflammatory bowel disease (IBD) can be imaged with positron emission tomography (PET), but existing PET radiopharmaceuticals have limited diagnostic accuracy. Vascular adhesion protein-1 (VAP-1) is an endothelial cell surface molecule that controls leukocyte extravasation into sites of inflammation. However, the role of inflammation-induced VAP-1 expression in IBD is still unclear. Therefore, this study investigated the utility of VAP-1-targeted [68Ga]Ga-DOTA-Siglec-9 positron emission tomography/computed tomography (PET/CT) for assessing inflammation in two mouse models of IBD.ProceduresStudies were performed using K8−/− mice that develop a chronic colitis-phenotype and C57Bl/6NCrl mice with acute intestinal inflammation chemically-induced using 2.5% dextran sodium sulfate (DSS) in drinking water. In both diseased and control mice, uptake of the VAP-1-targeting peptide [68Ga]Ga-DOTA-Siglec-9 was assessed in intestinal regions of interest using in vivo PET/CT, after which ex vivo gamma counting, digital autoradiography, and histopathological analyses were performed. Immunofluorescence staining was performed to determine VAP-1-expression in the intestine, including in samples from patients with ulcerative colitis.ResultsIntestinal inflammation could be visualized by [68Ga]Ga-DOTA-Siglec-9 PET/CT in two murine models of IBD. In both models, the in vivo PET/CT and ex vivo studies of [68Ga]Ga-DOTA-Siglec-9 uptake were significantly higher than in control mice. The in vivo uptake was increased on average 1.4-fold in the DSS model and 2.0-fold in the K8−/− model. Immunofluorescence staining revealed strong expression of VAP-1 in the inflamed intestines of both mice and patients.ConclusionsThis study suggests that the VAP-1-targeting [68Ga]Ga-DOTA-Siglec-9 PET tracer is a promising tool for non-invasive imaging of intestinal inflammation. Future studies in patients with IBD and evaluation of the potential value of [68Ga]Ga-DOTA-Siglec-9 in diagnosis and monitoring of the disease are warranted.

Detection of metabolic adaptation in a triple-negative breast cancer animal model with [18F]choline-PET imaging as a surrogate for drug resistance.

Test the feasibility of an image-based method to identify taxane resistance in mouse bearing triple-negative breast cancer (TNBC) tumor xenografts. Xenograft tumor-bearing mice from paclitaxel-sensitive and paclitaxel-resistant TNBC cells (MDA-MD-346) were generated by orthotopic injection into female NOD-SCID mice. When tumors reached 100-150 mm3, mice were scanned using [18F]choline PET/CT. Tumors were collected and sliced for autoradiography and immunofluorescence analysis. Quantitative data was analyzed accordingly. From fifteen mice scanned, five had taxane-sensitive cell line tumors of which two underwent taxol-based treatment. From the remaining 10 mice with taxane-resistant cell line tumors, four underwent taxol-based treatment. Only 13 mice had the tumor sample analyzed histologically. When normalized to the blood pool, both cell lines showed differences in metabolic uptake before and after treatment. Treated and untreated taxane-sensitive and taxane-resistant cell lines have different metabolic properties that could be leveraged before the start of chemotherapy.

Multifunctional Probe Based on "Chemical Antibody-Aptamer" for Noninvasive Detection of PD-L1 Expression in Cancer.

Immune checkpoint inhibitors (ICIs) therapy based on programmed cell death ligand 1 (PD-L1) has shown significant development in treating several carcinomas, but not all patients respond to this therapy due to the heterogeneity of PD-L1 expression. The sensitive and accurate quantitative analysis of in vivo PD-L1 expression is critical for treatment decisions and monitoring therapy. In the present study, an aptamer-based dual-modality positron emission tomography/near-infrared fluorescence (PET/NIRF) imaging probe was developed, and its specificity and sensitivity to PD-L1 were assessed in vitro and in vivo. The probe precursor NOTA-Cy5-R1 was prepared by using automated solid-phase oligonucleotide synthesis. PET/NIRF dual-modality probe [68Ga]Ga-NOTA-Cy5-R1 was successfully synthesized and radiolabeled. The binding specificity of [68Ga]Ga-NOTA-Cy5-R1 to PD-L1 was evaluated by flow cytometry, fluorescence imaging, and cellular uptake in A375-hPD-L1 and A375 cells, and it showed good fluorescence properties and stability in vitro. In vivo PET/NIRF imaging studies illustrated that [68Ga]Ga-NOTA-Cy5-R1 can sensitively and specifically bind to PD-L1 positive tumors. Meanwhile, the rapid clearance of probes from nontarget tissues achieved a high signal-to-noise ratio. In addition, changes of PD-L1 expression in NCI-H1299 xenografts treated with cisplatin (CDDP) were sensitivity monitored by [68Ga]Ga-NOTA-Cy5-R1 PET imaging, and ex vivo autoradiography and western blot analyses correlated well with the change of PD-L1 expression in vivo. Overall, [68Ga]Ga-NOTA-Cy5-R1 showed notable potency as a dual-modality PET/NIRF imaging probe for visualizing tumors and monitoring the dynamic changes of PD-L1 expression, which can help to direct and promote the clinical practice of ICIs therapy.

Quantitative Analysis of S1PR1 Expression in the Postmortem Multiple Sclerosis Central Nervous System.

Multiple sclerosis (MS) is an immune-mediated disease that is characterized by demyelination and inflammation in the central nervous system (CNS). Previous studies demonstrated that sphingosine-1-phosphate receptor (S1PR) modulators effectively inhibit S1PR1 in immune cell trafficking and reduce entry of pathogenic cells into the CNS. Studies have also implicated a nonimmune, inflammatory role of S1PR1 within the CNS in MS. In this study, we explored the expression of S1PR1 in the development and progression of demyelinating pathology of MS by quantitative assessment of S1PR1 expression using our S1PR1-specific radioligand, [3H]CS1P1, in the postmortem human CNS tissues including cortex, cerebellum, and spinal cord of MS cases and age- and sex-matched healthy cases. Immunohistochemistry with whole slide scanning for S1PR1 and various myelin proteins was also performed. Autoradiographic analysis using [3H]CS1P1 showed that the expression of S1PR1 was statistically significantly elevated in lesions compared to nonlesion regions in the MS cases, as well as normal healthy controls. The uptake of [3H]CS1P1 in the gray matter and nonlesion white matter did not significantly differ between healthy and MS CNS tissues. Saturation autoradiography analysis showed an increased binding affinity (Kd) of [3H]CS1P1 to S1PR1 in both gray matter and white matter of MS brains compared to healthy brains. Our blocking study using NIBR-0213, a S1PR1 antagonist, indicated [3H]CS1P1 is highly specific to S1PR1. Our findings demonstrated the activation of S1PR1 and an increased uptake of [3H]CS1P1 in the lesions of MS CNS. In summary, our quantitative autoradiography analysis using [3H]CS1P1 on human postmortem tissues shows the feasibility of novel imaging strategies for MS by targeting S1PR1.

99mTc-HFAPi imaging identifies early myocardial fibrosis in the hypertensive heart.

This study aimed to explore whether 99mTc-radiolabeled fibroblast activation protein inhibitor (99mTc-HFAPi) imaging can detect early myocardial fibrosis in the hypertensive heart. In the experimental model, spontaneously hypertensive rats (SHRs) and age-matched Wistar Kyoto rats (WKYs) were randomly divided into three groups (8, 16, and 28 weeks). The animals underwent 99mTc-HFAPi imaging and echocardiography. Autoradiography and histological analyses were performed in the left ventricle. The mRNA and protein expression level of the fibroblast activation protein (FAP) and collagen I were measured using quantitative PCR and western blot. In the clinical investigation, a total of 106 patients with essential hypertension and 20 gender-matched healthy controls underwent 99mTc-HFAPi imaging and echocardiography. In-vivo and in-vitro autographic images demonstrated diffusely enhanced 99mTc-HFAPi uptake in the SHR heart starting at week 8, before irreversible collagen deposition. The mRNA and protein levels of FAP in SHRs began to increase from week 8, whereas changes in collagen I levels were not detected until week 28. In the clinical investigation, even in hypertensive patients with normal diastolic indicators, normal left ventricular geometry, and normal global longitudinal strain (GLS), the prevalence of increased 99mTc-HFAPi uptake reached 34, 41, and 20%, respectively, indicating that early fibrogenesis precedes structural and functional myocardial abnormalities. In hypertension, 99mTc-HFAPi imaging can detect early fibrotic process before myocardial functional and structural changes.

Characterization of a S1PR2 specific 11C-labeled radiotracer in streptozotocin-induced diabetic murine model

BackgroundDiabetes mellitus is a chronic progressive metabolic disorder that affects millions of people worldwide. Emerging evidence suggests the important roles of sphingolipid metabolism in diabetes. In particular, sphingosine-1-phosphate (S1P) and S1P receptor 2 (S1PR2) have important metabolic functions and are involved in several metabolic diseases. In diabetes, S1PR2 can effectively preserve β cells and improve glucose/insulin tolerance in high-fat diet induced and streptozotocin (STZ)-induced diabetic mouse models. We previously developed a group of potent and selective S1PR2 ligands and radioligands. MethodsIn this study, we continued our efforts and characterized our leading S1PR2 radioligand, [11C]TZ34125, in a STZ-induced diabetic mouse model. [11C]TZ34125 was radiosynthesized in an automated synthesis module and in vitro saturation binding assay was performed using recombinant human S1PR2 membrane. In vitro saturation autoradiography analysis was also performed to determine the binding affinity of [11C]TZ34125 against mouse tissues. Type-1 diabetic mouse model was developed following a single high dose of STZ in C57BL/6 mice. Ex vivo biodistribution was performed to evaluate the distribution and amount of [11C]TZ34125 in tissues. In vitro autoradiography analysis was performed to compare the uptake of [11C]TZ34125 between diabetic and control animals in mouse spleen and pancreas. ResultsOur in vitro saturation binding assay using [11C]TZ34125 confirmed [11C]TZ34125 is a potent radioligand to recombinant human S1PR2 membrane with a Kd value of 0.9 nM. Saturation autoradiographic analysis showed [11C]TZ34125 has a Kd of 67.5, 45.9, and 25.0 nM to mouse kidney, spleen, and liver tissues respectively. Biodistribution study in STZ-induced diabetic mice showed the uptake of [11C]TZ34125 was significantly elevated in the spleen (~2 fold higher) and pancreas (~1.4 fold higher) compared to normal controls. The increased uptake of [11C]TZ34125 was further confirmed using autoradiographic analysis in the spleen and pancreases of STZ-induced diabetic mice, indicating S1PR2 can potentially act as a biomarker of diabetes in pancreases and inflammation in spleen. Future mechanistic analysis and in vivo quantitative assessment using non-invasive PET imaging in large animal model of diabetes is worthwhile. ConclusionsOverall, our data showed an increased uptake of our lead S1PR2-specific radioligand, [11C]TZ34125, in the spleen and pancreases of STZ-induced diabetic mice, and demonstrated [11C]TZ34125 has a great potential for preclinical and clinical usage for assessment of S1PR2 in diabetes and inflammation.

Preclinical Characterization of the Tau PET Tracer [18F]SNFT-1: Comparison of Tau PET Tracers.

Tau PET tracers are expected to be sufficiently sensitive to track the progression of age-related tau pathology in the medial temporal cortex. The tau PET tracer N-(4-[18F]fluoro-5-methylpyridin-2-yl)-7-aminoimidazo[1,2-a]pyridine ([18F]SNFT-1) has been successfully developed by optimizing imidazo[1,2-a]pyridine derivatives. We characterized the binding properties of [18F]SNFT-1 using a head-to-head comparison with other reported 18F-labeled tau tracers. Methods: The binding affinity of SNFT-1 to tau, amyloid, and monoamine oxidase A and B was compared with that of the second-generation tau tracers MK-6240, PM-PBB3, PI-2620, RO6958948, JNJ-64326067, and flortaucipir. In vitro binding properties of 18F-labeled tau tracers were evaluated through the autoradiography of frozen human brain tissues from patients with diverse neurodegenerative disease spectra. Pharmacokinetics, metabolism, and radiation dosimetry were assessed in normal mice after intravenous administration of [18F]SNFT-1. Results: In vitro binding assays demonstrated that [18F]SNFT-1 possesses high selectivity and high affinity for tau aggregates in Alzheimer disease (AD) brains. Autoradiographic analysis of tau deposits in medial temporal brain sections from patients with AD showed a higher signal-to-background ratio for [18F]SNFT-1 than for the other tau PET tracers and no significant binding with non-AD tau, α-synuclein, transactiviation response DNA-binding protein-43, and transmembrane protein 106B aggregates in human brain sections. Furthermore, [18F]SNFT-1 did not bind significantly to various receptors, ion channels, or transporters. [18F]SNFT-1 showed a high initial brain uptake and rapid washout from the brains of normal mice without radiolabeled metabolites. Conclusion: These preclinical data suggest that [18F]SNFT-1 is a promising and selective tau radiotracer candidate that allows the quantitative monitoring of age-related accumulation of tau aggregates in the human brain.

Chronic Δ9-tetrahydrocannabinol treatment has dose-dependent effects on open field exploratory behavior and [3H] SR141716A receptor binding in the rat brain

AimsAcute and chronic Δ9-THC exposure paradigms affect the body differently. More must be known about the impact of chronic Δ9-THC on cannabinoid-1 (CB1R) and mu-opioid (MOR) receptor levels in the brain. The present study examined chronic Δ9-THC's effects on CB1R and MOR levels and locomotor activity. Main methodsAdolescent Sprague-Dawley rats were given daily intraperitoneal injections of Δ9-THC [0.75mg/kg (low dose or LD) or 2.0 mg/kg (high dose or HD)] or vehicle for 24 days, and locomotion in the open field was tested after the first and fourth weeks of chronic Δ9-THC exposure. Brains were harvested at the end of treatment. [3H] SR141716A and [3H] DAMGO autoradiography assessed CB1R and MOR levels, respectively. Key findingsRelative to each other, chronic HD rats showed reduced vertical plane (VP) entries and time, while LD rats had increased VP entries and time for locomotion, as assessed by open-field testing; no effects were found relative to the control. Autoradiography analyses showed that HD Δ9-THC significantly decreased CB1R binding relative to LD Δ9-THC in the cingulate (33%), primary motor (42%), secondary motor (33%) somatosensory (38%), rhinal (38%), and auditory (50%) cortices; LD Δ9-THC rats displayed elevated binding in the primary motor (33% increase) and hypothalamic (33% increase) regions compared with controls. No significant differences were observed in MOR binding for the LD or HD compared to the control. SignificanceThese results demonstrate that chronic Δ9-THC dose-dependently altered CB1R levels throughout the brain and locomotor activity in the open field.

Preclinical evaluation of 68Ga-radiolabeled trimeric affibody for PDGFRβ-targeting PET imaging of hepatocellular carcinoma

PurposeHepatocellular carcinoma (HCC) is a highly vascularized solid carcinoma and tumor vessel–targeted molecular imaging might be effective for early diagnosis of HCC. Herein, we developed a novel trimeric affibody (ZTRI) with highly specific binding to the platelet-derived growth factor receptor beta (PDGFRβ). The aim of this study is to evaluate the feasibility of 68Ga-radiolabeled ZTRI ([68Ga]Ga-DOTA-ZTRI) as PET tracer for diagnosis of HCC.MethodsThe bioinformatics analysis of clinical database and immunoblotting of clinical specimens were performed to validate the potential of PDGFRβ as HCC biomarker. The trimeric affibody ZTRI was conjugated with DOTA-NHS-ester and radiolabeled with 68Ga to produce [68Ga]Ga-DOTA-ZTRI conjugate. Immunoreactivity and specific uptake of [68Ga]Ga-DOTA-ZTRI were assessed by dose-dependent cell binding, autoradiography, and biodistribution analysis. [68Ga]Ga-DOTA-ZTRI PET/CT scanning of diethylnitrosamine (DEN)-induced primary HCC rats and a rare case of idiopathical HCC rhesus monkey was performed to evaluate the imaging capability and radiation dosimetry of [68Ga]Ga-DOTA-ZTRI in vivo.ResultsExcessive PDGFRβ was validated as a representative biomarker of HCC neovascularization. The radiolabeling of [68Ga]Ga-DOTA-ZTRI was achieved at more than 95% radiochemical yield. In vitro assays showed specific uptake of [68Ga]Ga-DOTA-ZTRI in HCC tumor vessels by autoradiography. Animal PET/CT imaging with [68Ga]Ga-DOTA-ZTRI successfully visualized the tumor lesions in primary HCC rats and rhesus monkey, and indicated radiation absorbed dose of 2.03E-02 mSv/MBq for each scanning.ConclusionsOur results demonstrated that [68Ga]Ga-DOTA-ZTRI conjugate could be applied as a promising PET tracer for early diagnosis of hepatocellular carcinoma.

Synthesis and Preliminary Evaluations of [18F]fluorinated Pyridine-2- carboxamide Derivatives for Targeting PD-L1 in Cancer.

Treatment with immune checkpoint inhibitors has improved both progressionfree survival and overall survival in a subset of patients with tumors. However, the selection of patients who benefit from immune checkpoint inhibitor treatment remains challenging. Positron Emission Tomography (PET) is a non-invasive molecular imaging tool that offers a promising alternative to the current IHC for detecting the PD-L1 expression in malignant cells in vivo, enabling patient selection and predicting the response to individual patient immunotherapy treatment. Herein, we report the development of novel [18F]labeled pyridine-2-carboxamide derivatives [18F]2 and [18F]3 as small-molecule probes for imaging immune checkpoint (PD-1/PD-L1) in cancer using PET. [18F]2 and [18F]3 were prepared by a one-step radiofluorination in 44 ± 5% and 30 ± 4% radiochemical yield and > 98% radiochemical purity for a potential clinical translation. The total synthesis time, including HPLC purification, was less than 45 min. [18F]2 and [18F]3 showed excellent stability in injection solution and a significant accumulation and retention in PD-1/PD-L1 expressing MDA-MB-231 breast cancer and in HeLa cervix carcinoma cells (2- 5 cpm/1000 cells). In addition, autoradiographic analysis and inhibition experiments on tumor slices confirm the potential of both compounds as specific imaging probes for the PD-1/PD-L1 axis in tumors. The in vitro evaluation in PD-L1 expressing cells together with results from autoradiographic analysis in PD-L1 positive tumor sections, suggest that [18F]2 and [18F]3 could be potential imaging probes for assessing PD-L1 expression in tumors and warrant further biological evaluations in vivo.

Assessment of the Pharmacokinetics, Disposition, and Duration of Action of the Tumour-Targeting Peptide CEND-1

CEND-1 (iRGD) is a bifunctional cyclic peptide that can modulate the solid tumour microenvironment, enhancing the delivery and therapeutic index of co-administered anti-cancer agents. This study explored CEND-1's pharmacokinetic (PK) properties pre-clinically and clinically, and assessed CEND-1 distribution, tumour selectivity and duration of action in pre-clinical tumour models. Its PK properties were assessed after intravenous infusion of CEND-1 at various doses in animals (mice, rats, dogs and monkeys) and patients with metastatic pancreatic cancer. To assess tissue disposition, [3H]-CEND-1 radioligand was administered intravenously to mice bearing orthotopic 4T1 mammary carcinoma, followed by tissue measurement using quantitative whole-body autoradiography or quantitative radioactivity analysis. The duration of the tumour-penetrating effect of CEND-1 was evaluated by assessing tumour accumulation of Evans blue and gadolinium-based contrast agents in hepatocellular carcinoma (HCC) mouse models. The plasma half-life was approximately 25 min in mice and 2 h in patients following intravenous administration of CEND-1. [3H]-CEND-1 localised to the tumour and several healthy tissues shortly after administration but was cleared from most healthy tissues by 3 h. Despite the rapid systemic clearance, tumours retained significant [3H]-CEND-1 several hours post-administration. In mice with HCC, the tumour penetration activity remained elevated for at least 24 h after the injection of a single dose of CEND-1. These results indicate a favourable in vivo PK profile of CEND-1 and a specific and sustained tumour homing and tumour penetrability. Taken together, these data suggest that even single injections of CEND-1 may elicit long-lasting tumour PK improvements for co-administered anti-cancer agents.

Therapeutic Efficacy and Radiobiological Effects of Boric-Acid-Mediated BNCT in an Osteosarcoma-Bearing SD Rat Model.

Osteosarcoma (OS) is the most common primary malignancy of the bone and is notoriously resistant to radiation therapy. High-dose cytotoxic chemotherapy and surgical resection have improved the survival rate and prognosis of patients with OS. Nonetheless, treatment challenges remain when the tumor cannot be removed by surgery. Boron neutron capture therapy (BNCT) provides high linear energy transfer (LET) radiation, and its internal targeted characteristics make BNCT a novel therapy for removing OS and reducing radiation damage to adjacent healthy tissues. In this study, a UMR-106-grafted OS rat model was developed, and boric acid (BA) was used as the boron drug for BNCT. The pharmacokinetics of BA, following intravenous injection, were evaluated to determine the optimal time window for neutron irradiation. OS-bearing rats were irradiated by an epithermal neutron beam at Tsing Hua Open-Pool Reactor (THOR). The therapeutic efficacy of and tissue response after BNCT were evaluated by radiographic and histopathological observations. OS-bearing rats were irradiated by neutrons in the first hour following the intravenous injection of BA. The prescription-absorbed doses in the tumor regions were 5.8 and 11.0 Gy. BNCT reduced the body weight of the tumor-bearing rats, but they recovered after a few days. The BA-mediated BNCT effectively controlled the orthotopic OS tumor, reduced osteolysis, and induced bone healing. Autoradiography and histological analysis confirmed that the BA retention region is consistent with the calcification region in OS tissue. BA is specifically retained in OS, and the BA-mediated BNCT can significantly reduce the tumor burden and osteolysis in OS-bearing rats.

Unlocking the mystery of lysine toxicity on Microcystis aeruginosa

Lysine toxicity on certain groups of bacterial cells has been recognized for many years, but the detailed molecular mechanisms that drive this phenomenon have not been elucidated. Many cyanobacteria including Microcystis aeruginosa cannot efficiently export and degrade lysine, although they have evolved to maintain a single copy of the lysine uptake system through which arginine or ornithine can also be transported into the cytoplasm. Autoradiographic analysis using 14C-l-lysine confirmed that lysine was competitively uptaken into cells with arginine or ornithine, which explained the arginine or ornithine-mediated alleviation of lysine toxicity in M. aeruginosa. A relatively non-specific MurE amino acid ligase could incorporate l-lysine into the 3rd position of UDP-N-acetylmuramyl-tripeptide by replacing meso-diaminopimelic acid during the stepwise addition of amino acids on peptidoglycan (PG) biosynthesis. However, further transpeptidation was blocked because lysine substitution at the pentapeptide of the cell wall inhibited the activity of transpeptidases. The leaky PG structure caused irreversible damage to the photosynthetic system and membrane integrity. Collectively, our results suggest that a lysine-mediated coarse-grained PG network and the absence of concrete septal PG lead to the death of slow-growing cyanobacteria.

18F]RO948 tau positron emission tomography in genetic and sporadic frontotemporal dementia syndromes

PurposeTo examine [18F]RO948 retention in FTD, sampling the underlying protein pathology heterogeneity.MethodsA total of 61 individuals with FTD (n = 35), matched cases of AD (n = 13) and Aβ-negative cognitively unimpaired individuals (n = 13) underwent [18F]RO948PET and MRI. FTD included 21 behavioral variant FTD (bvFTD) cases, 11 symptomatic C9orf72 mutation carriers, one patient with non-genetic bvFTD-ALS, one individual with bvFTD due to a GRN mutation, and one due to a MAPT mutation (R406W). Tracer retention was examined using a region-of-interest and voxel-wise approaches. Two individuals (bvFTD due to C9orf72) underwent postmortem neuropathological examination. Tracer binding was additionally assessed in vitro using [3H]RO948 autoradiography in six separate cases.Results[18F]RO948 retention across ROIs was clearly lower than in AD and comparable to that in Aβ-negative cognitively unimpaired individuals. Only minor loci of tracer retention were seen in bvFTD; these did not overlap with the observed cortical atrophy in the cases, the expected pattern of atrophy, nor the expected or verified protein pathology distribution. Autoradiography analyses showed no specific [3H]RO948 binding. The R406W MAPT mutation carriers were clear exceptions with AD-like retention levels and specific in-vitro binding.Conclusion[18F]RO948 uptake is not significantly increased in the majority of FTD patients, with a clear exception being specific MAPT mutations.

Testing mosses exposed in bags as biointerceptors of airborne radiocaesium after the Fukushima Dai-ichi Nuclear Power Station accident

Eight years after the Fukushima nuclear accident, mosses exposed in bags were used to investigate their ability to accumulate radiocaesium and therefore to act as biointerceptors of 134Cs and 137Cs in the evacuated area of the Fukushima territory. Bags were filled with 3 widely studied moss species (Sphagnum palustre, Hypnum cupressiforme, and Hypnum plumaeforme) and exposed for 3, 6 or 9 weeks at 5 former residential sites within the Fukushima area and, for comparison, at three background sites located 700 km away. The radiocaesium activity concentrations found in moss bags were evaluated as function of exposure time, site conditions and moss species. In the Fukushima area, the moss bags accumulated 137Cs at all exposure sites and in all exposure periods, with S. palustre having the highest 137Cs accumulation ability. The 137Cs activity concentrations (from 28 to 4700 Bq kg−1) measured in moss bags increased with the exposure time and were consistent with the decontamination status of each exposure site, highlighting the big potential of moss bags to discriminate among exposure sites. Time dependency of 137Cs activity concentrations measured in mosses allowed the calculation of location-specific and species-specific factors, which can be used to predict radiocaesium accumulation trends in future biomonitoring surveys performed in the same area with the same experimental design. Autoradiography and electron microscopy analyses of the moss surfaces revealed a prevalence of soil-derived particulate form of radiocaesium, suggesting the use of moss bags as warning sensors of resuspended particles potentially harmful for local residents.

Using CD69 PET Imaging to Monitor Immunotherapy-Induced Immune Activation.

Immune checkpoint inhibitors (ICI) have been effective in treating a subset of refractory solid tumors, but only a small percentage of treated patients benefit from these therapies. Thus, there is a clinical need for reliable tools that allow for the early assessment of response to ICIs, as well as a preclinical need for imaging tools that aid in the future development and understanding of immunotherapies. Here we demonstrate that CD69, a canonical early-activation marker expressed on a variety of activated immune cells, including cytotoxic T cells and natural killer (NK) cells, is a promising biomarker for the early assessment of response to immunotherapies. We have developed a PET probe by radiolabeling a highly specific CD69 mAb, H1.2F3, with Zirconium-89 (89Zr), [89Zr]-deferoxamine (DFO)-H1.2F3. [89Zr]-DFO-H1.2F3 detected changes in CD69 expression on primary mouse T cells in vitro and detected activated immune cells in a syngeneic tumor immunotherapy model. In vitro uptake studies with [89Zr]-DFO-H1.2F3 showed a 15-fold increase in CD69 expression for activated primary mouse T cells, relative to untreated resting T cells. In vivo PET imaging showed that tumors of ICI-responsive mice had greater uptake than the tumors of nonresponsive and untreated mice. Ex vivo biodistribution, autoradiography, and IHC analyses supported the PET imaging findings. These data suggest that the CD69 PET imaging approach detects CD69 expression with sufficient sensitivity to quantify immune cell activation in a syngeneic mouse immunotherapy model and could allow for the prediction of therapeutic immune responses to novel immunotherapies.