A ROI-based quantitative pipeline for 18F-FDG PET metabolism and pCASL perfusion joint analysis: Validation of the 18F-FDG PET line.

In Mild Cognitive Impairment (MCI), the study of brain metabolism, provided by 18F-FluoroDeoxyGlucose Positron Emission Tomography (18F-FDG PET) can be integrated with brain perfusion through pseudo-Continuous Arterial Spin Labeling Magnetic Resonance sequences (MR pCASL). Cortical hypometabolism identification generally relies on wide control group datasets; pCASL control groups are instead not publicly available yet, due to lack of standardization in the acquisition parameters. This study presents a quantitative pipeline to be applied to PET and pCASL data to coherently analyze metabolism and perfusion inside 16 matching cortical regions of interest (ROIs) derived from the AAL3 atlas. The PET line is tuned on 36 MCI patients and 107 healthy control subjects, to agree in identifying hypometabolic regions with clinical reference methods (visual analysis supported by a vendor tool and Statistical Parametric Mapping, SPM, with two parametrizations here identified as SPM-A and SPM-B). The analysis was conducted for each ROI separately. The proposed PET analysis pipeline obtained accuracy 78 % and Cohen's к 60 % vs visual analysis, accuracy 79 % and Cohen's к 58 % vs SPM-A, accuracy 77 % and Cohen's к 54 % vs SPM-B. Cohen's к resulted not significantly different from SPM-A and SPM-B Cohen's к when assuming visual analysis as reference method (p-value 0.61 and 0.31 respectively). Considering SPM-A as reference method, Cohen's к is not significantly different from SPM-B Cohen's к as well (p-value = 1.00). The complete PET-pCASL pipeline was then preliminarily applied on 5 MCI patients and metabolism-perfusion regional correlations were assessed. The proposed approach can be considered as a promising tool for PET-pCASL joint analyses in MCI, even in the absence of a pCASL control group, to perform metabolism-perfusion regional correlation studies, and to assess and compare perfusion in hypometabolic or normo-metabolic areas.

Effects of Metformin as Add-On Therapy against Glioblastoma: An Old Medicine for Novel Oncology Therapeutics.

BACKGROUND: Glioblastoma is the most aggressive primary brain malignancy in adults, with a poor prognosis of about 14 months. Recent evidence ascribed to metformin (MET), an antihyperglycemic drug, the potential to reduce cancer incidence and progression, but the molecular mechanisms underlying these effects need to be better investigated. METHODS: Here, we tested the efficacy of MET on n = 10 primary glioblastoma endothelial cells (GECs), by viability and proliferation tests, as MTT and Live/Dead assays, apoptosis tests, as annexin V assay and caspase 3/7 activity, functional tests as tube-like structure formation and migration assay and by mRNA and protein expression performed by quantitative real-time PCR analysis (qRT-PCR) and Western Blot, respectively. RESULTS: Data resulting revealed a time- and µ-dependent ability of MET to decrease cell viability and proliferation, increasing pro-apoptotic mechanisms mediated by caspases 3/7. Also, MET impacted GEC functionality with a significant decrease of angiogenesis and invasiveness potential. Mechanistically, MET was able to interfere with sphingolipid metabolism, weakening the oncopromoter signaling promoted by sphingosine-1-phosphate (S1P) and shifting the balance toward the production of the pro-apoptotic ceramide. CONCLUSIONS: These observations ascribed to MET the potential to serve as add-on therapy against glioblastoma, suggesting a repurposing of an old, totally safe and tolerable drug for novel oncology therapeutics.

Endovascular administration of magnetized nanocarriers targeting brain delivery after stroke.

The increasing use of mechanical thrombectomy in stroke management has opened the window to local intraarterial brain delivery of therapeutic agents. In this context, the use of nanomedicine could further improve the delivery of new treatments for specific brain targeting, tracking and guidance. In this study we take advantage of this new endovascular approach to deliver biocompatible poly(D-L-lactic-co-glycolic acid) (PLGA) nanocapsules functionalized with superparamagnetic iron oxide nanoparticles and Cy7.5 for magnetic targeting, magnetic resonance and fluorescent molecular imaging. A complete biodistribution study in naïve (n = 59) and ischemic (n = 51) mice receiving intravenous or intraarterial nanocapsules, with two different magnet devices and imaged from 30 min to 48 h, showed an extraordinary advantage of the intraarterial route for brain delivery with a specific improvement in cortical targeting when using a magnetic device in both control and ischemic conditions. Safety was evaluated in ischemic mice (n = 69) showing no signs of systemic toxicity nor increasing mortality, infarct lesions or hemorrhages. In conclusion, the challenging brain delivery of therapeutic nanomaterials could be efficiently and safely overcome with a controlled endovascular administration and magnetic targeting, which could be considered in the context of endovascular interventions for the delivery of multiple treatments for stroke.

Acute Inescapable Stress Rapidly Increases Synaptic Energy Metabolism in Prefrontal Cortex and Alters Working Memory Performance.

Brain energy metabolism actively regulates synaptic transmission and activity. We have previously shown that acute footshock (FS)-stress induces fast and long-lasting functional and morphological changes at excitatory synapses in prefrontal cortex (PFC). Here, we asked whether FS-stress increased energy metabolism in PFC, and modified related cognitive functions. Using positron emission tomography (PET), we found that FS-stress induced a redistribution of glucose metabolism in the brain, with relative decrease of [18F]FDG uptake in ventro-caudal regions and increase in dorso-rostral ones. Absolute [18F]FDG uptake was inversely correlated with serum corticosterone. Increased specific hexokinase activity was also measured in purified PFC synaptosomes (but not in total extract) of FS-stressed rats, which positively correlated with 2-Deoxy [3H] glucose uptake by synaptosomes. In line with increased synaptic energy demand, using an electron microscopy-based stereological approach, we found that acute stress induced a redistribution of mitochondria at excitatory synapses, together with an increase in their volume. The fast functional and metabolic activation of PFC induced by acute stress, was accompanied by rapid and sustained alterations of working memory performance in delayed response to T-maze test. Taken together, the present data suggest that acute stress increases energy consumption at PFC synaptic terminals and alters working memory.

Molecular imaging of neuroinflammation in preclinical rodent models using positron emission tomography.

Neuroinflammation (NI) is an adaptive response to different noxious stimuli, involving microglia, astrocytes and peripheral immune cells. NI is a hallmark of several acute and chronic diseases of central nervous system (CNS) and contributes to both damage and repair of CNS tissue. Interventional or genetically modified rodent models mimicking human neuropathologies may provide valuable insights on basic mechanisms of NI, but also for improving the development of new diagnostic and therapeutic strategies. Preclinical positron emission tomography (PET) allows to investigate noninvasively the inflammatory response in CNS of rodent models at a molecular level, validating innovative probes for early diagnosis, and characterizing the time course of neuroinflammatory changes and their relationship with disease progression, as well as the effects of experimental treatments with high translational potential. In particular, recent efforts of preclinical PET field are intended to develop specific and selective radiotracers that target the activation of innate immune system in CNS. Here, we have reviewed the state of art for PET in relevant rodent models of acute and chronic neuropathologies associated with NI, with particular regard on imaging of activated microglia and astrocytes.

Cancer cell growth and survival as a system-level property sustained by enhanced glycolysis and mitochondrial metabolic remodeling.

Systems Biology holds that complex cellular functions are generated as system-level properties endowed with robustness, each involving large networks of molecular determinants, generally identified by "omics" analyses. In this paper we describe four basic cancer cell properties that can easily be investigated in vitro: enhanced proliferation, evasion from apoptosis, genomic instability, and inability to undergo oncogene-induced senescence. Focusing our analysis on a K-ras dependent transformation system, we show that enhanced proliferation and evasion from apoptosis are closely linked, and present findings that indicate how a large metabolic remodeling sustains the enhanced growth ability. Network analysis of transcriptional profiling gives the first indication on this remodeling, further supported by biochemical investigations and metabolic flux analysis (MFA). Enhanced glycolysis, down-regulation of TCA cycle, decoupling of glucose and glutamine utilization, with increased reductive carboxylation of glutamine, so to yield a sustained production of growth building blocks and glutathione, are the hallmarks of enhanced proliferation. Low glucose availability specifically induces cell death in K-ras transformed cells, while PKA activation reverts this effect, possibly through at least two mitochondrial targets. The central role of mitochondria in determining the two investigated cancer cell properties is finally discussed. Taken together the findings reported herein indicate that a system-level property is sustained by a cascade of interconnected biochemical pathways that behave differently in normal and in transformed cells.

Characterization of preclinical models of prostate cancer using PET-based molecular imaging.

PURPOSE: Transgenic adenocarcinoma of the mouse prostate (TRAMP) mice spontaneously develop hormone-dependent and hormone-independent prostate cancer (PC) that potentially resembles the human pathological condition. The aim of the study was to validate PET imaging as a reliable tool for in vivo assessment of disease biology and progression in TRAMP mice using radioligands routinely applied in clinical practice: [(18)F]FDG and [(11)C]choline. METHODS: Six TRAMP mice were longitudinally evaluated starting at week 11 of age to visualize PC development and progression. The time frame and imaging pattern of PC lesions were subsequently confirmed on an additional group of five mice. RESULTS: PET and [(18)F]FDG allowed detection of PC lesions starting from 23 weeks of age. [(11)C]Choline was clearly taken up only by TRAMP mice carrying neuroendocrine lesions, as revealed by post-mortem histological evaluation. CONCLUSION: PET-based molecular imaging represents a state-of-the-art tool for the in vivo monitoring and metabolic characterization of PC development, progression and differentiation in the TRAMP model.

11C-labeling of n-[4-[4-(2,3-dichlorophenyl)piperazin-1-yl]butyl]arylcarboxamide derivatives and evaluation as potential radioligands for PET imaging of dopamine D3 receptors.

The selective dopamine D(3) receptor ligands N-4-[4-[(2,3-dichlorophenyl)piperazin-1-yl]butyl]1-methoxy-2-naphthalencarboxamide (1) and N-4-[4-[(2,3-dichlorophenyl)piperazin-1-yl]butyl]-7-methoxy-2-benzofurancarboxamide (2) were labeled with (11)C (t(1/2) = 20.4 min) as potential radioligands for the noninvasive assessment of the dopamine D(3) neurotransmission system in vivo with positron emission tomography (PET). The radiosynthesis consisted in an O-methylation of the des-methyl precursors N-[4-[4-(2,3-dichlorophenyl)piperazin-1-yl]butyl]-1-hydroxy-2-naphthalenecarboxamide (3) and N-[4-[4-(2,3-dichlorophenyl)piperazin-1-yl]butyl]-7-hydroxy-2-benzofurancarboxamide (4) with [(11)C]methyl iodide using tBuOK/HMPA and KOH/DMSO, respectively. The radiotracers [(11)C]1 and [(11)C]2 were obtained in 35 min with over 99% radiochemical purity, 74 +/- 37 GBq/mumol of specific radioactivity, 13% and 26% radiochemical yield (EOB, decay-corrected). Distribution studies in rats demonstrated that the new tracers [(11)C]1 and [(11)C]2 cross the blood-brain barrier and localize in the brain. However, the kinetics of cerebral uptake did not reflect the regional expression of the D(3) receptors. Despite their in vitro pharmacological profile, [(11)C]1 and [(11)C]2 do not display an in vivo behavior suitable to image D(3) receptor expression using PET.

Basal ganglia and language: phonology modulates dopaminergic release.

Basal ganglia have been implicated in syntactic and phonological processes, but direct evidence has been scarce. Here, we used [11C]raclopride and positron emission tomography to measure modulations of the dopaminergic system induced by phonological or syntactic processing. Two significant effects were found. First, the level of accuracy in phonological processing significantly correlated with tracer binding potential in the left caudate nucleus. Second, the speed in phonological processing significantly correlated with tracer binding potential in the left putamen. Thus, a more accurate and fast phonological processing was associated with a reduced dopamine requirement in the left striatum. These findings show that the striatal dopaminergic system plays an essential role in grammatical processes that form the core of human language.

[11C]RN5: a new agent for the in vivo imaging of myocardial alpha1-adrenoceptors.

The radiolabelling with the positron-emitter Carbon-11 and the biological evaluation in rats of 3-[2-[4-(2-[11C]methoxyphenyl)piperazin-1-yl]ethyl]pyrimido[5,4-b]indole-2,4-dione ([11C]RN5), alpha1-adrenoceptor antagonist (K(i)=0.21 nM), as a putative radioligand for the non-invasive assessment of alpha1-adrenoceptors with positron emission tomography (PET) is reported. The radiosynthesis procedure consisted of O-methylation of des-methyl precursor with [11C]methyl iodide in the presence of potassium hydroxide in dimethylformamide (DMF) at 80 degrees C. [11C]RN5 was obtained in >99% radiochemical purity in 25 min with a radiochemical yield in the 20-30% range, end of synthesis (EOS) (non-decay corrected) and a specific radioactivity of 92.5+/-18.5 GBq/micromol. Pre-clinical studies in rats showed a high uptake of [11C]RN5 in heart, spleen, adrenal gland, lung and kidney but not in the brain. Inhibition studies with high doses of different adrenergic antagonists indicate that more than 70% of myocardial uptake of [11C]RN5 is due to specific binding to alpha1-adrenoceptors. Our results indicate that [11C]RN5 is suitable to be further developed as a potential radioligand for the in vivo PET imaging of myocardial alpha1-adrenoceptors in humans.