Early Detection of Cardiotoxicity Using [64Cu]Cu-NODAGA-E[(cRGDyK)]2 PET Imaging in a Rat Model of Doxorubicin-Induced Heart Failure.

Doxorubicin (DOX) is a common and highly effective chemotherapeutic. However, its use is limited by cardiotoxic effects and the lack of methods to detect these at early time points. In the present study, we evaluated if [64Cu]Cu-NODAGA-E[(cRGDyK)]2 positron emission tomography-computed tomography ([64Cu]Cu-RGD PET/CT) could detect cardiotoxicity in a rat model of DOX-induced heart failure. Male Lewis rats were divided into two groups and treated with either a cumulative dose of 15 mg/kg of DOX or left untreated. Cardiac anatomy and function were assessed using magnetic resonance imaging at baseline and in week 8. [64Cu]Cu-RGD PET/CT scans were performed in week 4. DOX treatment led to a decline in pump function as well as an increase in cardiac and thymic uptake of [64Cu]Cu-RGD. In addition, DOX altered cardiac gene expression, led to infiltration of immune cells, reduced endothelial content, and increased interstitial fibrosis. Furthermore, concentrations of inflammatory plasma proteins were increased in the DOX group. In conclusion, DOX treatment resulted in the development of cardiotoxicity and heart failure, which could be detected using [64Cu]Cu-RGD PET/CT at early time points. [64Cu]Cu-RGD uptake in the myocardial septum and thymus predicted a low left ventricular ejection fraction in week 8.

The cellular composition of chronic subdural hematoma.

INTRODUCTION: The pathophysiology of chronic subdural hematoma (CSDH) remains to be fully understood. Basic knowledge of the composition and features of cells in the CSDH fluid may contribute to the understanding of the seemingly complex processes involved in CSDH formation and recurrence. This study is the first to examine the composition of cells and of cellular features in both systemic blood and subdural fluid from CSDH patients. We hypothesized that the cellular composition and features in the hematoma fluid may be; 1) different from that in the systemic blood; 2) different between patients with and without recurrence; 3) and different between the first and second operation in patients with recurrent CSDH. METHODS: Systemic blood and subdural hematoma fluid were collected from CSDH patients with and without recurrent CSDH at the time of primary and secondary surgery. Analyses of cells and cellular features included total number of white blood cells, erythroblasts, reticulocytes, platelets, neutrophilocytes, lymphocytes, monocytes, eosinophils, basophils, reticulocytes, immature granulocytes, mean corpuscular cell volume (MCV), mean corpuscular hemoglobin, mean corpuscular hemoglobin concentration, hemoglobin and hematocrit. RESULTS: Of the 85 included patients, 20 patients were operated for a recurrent CSDH within 90 days follow-up. All cells found in the systemic blood were present in the CSDH fluid, but the composition was different (p < 0.0001). MCV was higher in the hematoma fluid from the primary operation of patients later developing a recurrent CSDH compared to patients not developing recurrence (p = 0.009). Also, the percentage distribution of inflammatory cells in hematoma fluid from patients with recurrent CSDH was different between the first and second operation (p = 0.0017). CONCLUSION: This study is the first to investigate the cellular composition of CSDH fluid. Compared to systemic blood and to a reference distribution, an increased number of immune cells were present in the hematoma fluid, supporting an inflammatory component of the CSDH pathophysiology. MCV was higher in the subdural fluid at time of the first operation of CSDH patients later developing recurrence. CLINICAL TRIAL REGISTRATION: The study was approved by the Scientific Ethical Committee of the Capital Region of Denmark (Journal no. H-20051073.

[68Ga]Ga-NODAGA-E[(cRGDyK)]2 angiogenesis PET following myocardial infarction in an experimental rat model predicts cardiac functional parameters and development of heart failure.

BACKGROUND: Angiogenesis has increasingly been a target for imaging and treatment over the last decade. The integrin αvß3 is highly expressed in cells during angiogenesis and are therefore a promising target for imaging. In this study, we aimed to investigate the PET tracer [68Ga]Ga-RGD as a marker of angiogenesis following MI and its ability to predict cardiac functional parameters. METHODS: First, the real-time interaction between [68Ga]Ga-RGD and integrin αvß3 was investigated using surface plasmon resonance (SPR). Second, an animal study was performed to investigate the [68Ga]Ga-RGD uptake in the infarcted area after one and four weeks following MI in a rat model (MI = 68, sham surgery = 36). Finally, the specificity of the [68Ga]Ga-RGD tracer was evaluated ex vivo using histology, autoradiography, gamma counting and flow cytometry. RESULTS: SPR showed that [68Ga]Ga-RGD has a high affinity for integrin αvß3, forming a strong and stable binding. PET/CT showed a significantly higher uptake of [68Ga]Ga-RGD in the infarcted area compared to sham one week (p < 0.001) and four weeks (p < 0.001) after MI. The uptake of [68Ga]Ga-RGD after one week correlated to end diastolic volume (r = 0.74, p < 0.001) and ejection fraction (r = - 0.71, p < 0.001) after four weeks. CONCLUSION: This study demonstrates that [68Ga]Ga-RGD has a high affinity for integrin αvß3, which enables the evaluation of angiogenesis and remodeling. The [68Ga]Ga-RGD uptake after one week indicates that [68Ga]Ga-RGD may be used as an early predictor of cardiac functional parameters and possible development of heart failure after MI. These encouraging data supports the clinical translation and future use in MI patients.

Multimodal Positron Emission Tomography Imaging to Quantify Uptake of 89Zr-Labeled Liposomes in the Atherosclerotic Vessel Wall.

Nanotherapy has recently emerged as an experimental treatment option for atherosclerosis. To fulfill its promise, robust noninvasive imaging approaches for subject selection and treatment evaluation are warranted. To that end, we present here a positron emission tomography (PET)-based method for quantification of liposomal nanoparticle uptake in the atherosclerotic vessel wall. We evaluated a modular procedure to label liposomal nanoparticles with the radioisotope zirconium-89 (89Zr). Their biodistribution and vessel wall targeting in a rabbit atherosclerosis model was evaluated up to 15 days after intravenous injection by PET/computed tomography (CT) and PET/magnetic resonance imaging (PET/MRI). Vascular permeability was assessed in vivo using three-dimensional dynamic contrast-enhanced MRI (3D DCE-MRI) and ex vivo using near-infrared fluorescence (NIRF) imaging. The 89Zr-radiolabeled liposomes displayed a biodistribution pattern typical of long-circulating nanoparticles. Importantly, they markedly accumulated in atherosclerotic lesions in the abdominal aorta, as evident on PET/MRI and confirmed by autoradiography, and this uptake moderately correlated with vascular permeability. The method presented herein facilitates the development of nanotherapy for atherosclerotic disease as it provides a tool to screen for nanoparticle targeting in individual subjects' plaques.

In vivo imaging of cell proliferation in meningioma using 3'-deoxy-3'-[18F]fluorothymidine PET/MRI.

PURPOSE: Positron emission tomography (PET) with 3'-deoxy-3'-[18F]fluorothymidine ([18F]FLT) provides a noninvasive assessment of tumour proliferation in vivo and could be a valuable imaging modality for assessing malignancy in meningiomas. We investigated a range of static and dynamic [18F]FLT metrics by correlating the findings with cellular biomarkers of proliferation and angiogenesis. METHODS: Seventeen prospectively recruited adult patients with intracranial meningiomas underwent a 60-min dynamic [18F]FLT PET following surgery. Maximum and mean standardized uptake values (SUVmax, SUVmean) with and without normalization to healthy brain tissue and blood radioactivity obtained from 40 to 60 min summed dynamic images (PET40-60) and ~ 60-min blood samples were calculated. Kinetic modelling using a two-tissue reversible compartmental model with a fractioned blood volume (VB) was performed to determine the total distribution volume (VT). Expressions of proliferation and angiogenesis with key parameters including Ki-67 index, phosphohistone-H3 (phh3), MKI67, thymidine kinase 1 (TK1), proliferating cell nuclear antigen (PCNA), Kirsten RAt Sarcoma viral oncogene homolog (KRAS), TIMP metallopeptidase inhibitor 3 (TIMP3), and vascular endothelial growth factor A (VEGFA) were determined by immunohistochemistry and/or quantitative polymerase chain reaction. RESULTS: Immunohistochemistry revealed 13 World Health Organization (WHO) grade I and four WHO grade II meningiomas. SUVmax and SUVmean normalized to blood radioactivity from PET40-60 and blood sampling, and VT were able to significantly differentiate between WHO grades with the best results for maximum and mean tumour-to-whole-blood ratios (sensitivity 100%, specificity 94-95%, accuracy 99%; P = 0.003). Static [18F]FLT metrics were significantly correlated with proliferative biomarkers, especially Ki-67 index, phh3, and TK1, while no correlations were found with VEGFA or VB. Using Ki-67 index with a threshold > 4%, the majority of [18F]FLT metrics showed a high ability to identify aggressive meningiomas with SUVmean demonstrating the best performance (sensitivity 80%, specificity 81%, accuracy 80%; P = 0.024). CONCLUSION: [18F]FLT PET could be a useful imaging modality for assessing cellular proliferation in meningiomas.

Pharmacokinetic analysis of [68Ga]Ga-DOTA-TOC PET in meningiomas for assessment of in vivo somatostatin receptor subtype 2.

PURPOSE: DOTA-D-Phe1-Tyr3-octreotide with gallium-68 ([68Ga]Ga-DOTA-TOC) is one of the PET tracers that forms the basis for peptide receptor radionuclide therapy based on somatostatin receptor subtype 2 (SSTR2) expression in meningiomas. Yet, the quantitative relationship between [68Ga]Ga-DOTA-TOC accumulation and SSTR2 is unknown. We conducted a correlative analysis of a range of [68Ga]Ga-DOTA-TOC PET metric(s) as imaging surrogate(s) of the receptor binding in meningiomas by correlating the PET results with SSTR2 expression from surgical specimens. We additionally investigated possible influences of secondary biological factors such as vascularization, inflammation and proliferation. METHODS: Fifteen patients with MRI-presumed or recurrent meningiomas underwent a 60-min dynamic [68Ga]Ga-DOTA-TOC PET/CT before surgery. The PET data comprised maximum and mean standardized uptake values (SUVmax, SUVmean) with and without normalization to reference regions, and quantitative measurements derived from kinetic modelling using a reversible two-tissue compartment model with the fractional blood volume (VB). Expressions of SSTR2 and proliferation (Ki-67, phosphohistone-H3, proliferating cell nuclear antigen) were determined by immunohistochemistry and/or quantitative polymerase chain reaction (qPCR), while biomarkers of vascularization (vascular endothelial growth factor A (VEGFA), endothelial marker CD34) and inflammation (cytokine interleukin-18, microglia/macrophage-specific marker CD68) by qPCR. RESULTS: Histopathology revealed 12 World Health Organization (WHO) grade I and three WHO grade II meningiomas showing no link to SSTR2. The majority of [68Ga]Ga-DOTA-TOC PET metrics showed significant associations with SSTR2 protein, while all PET metrics were positively correlated with SSTR2 mRNA with the best results for mean tumour-to-blood ratio (TBRmean) (r = 0.757, P = 0.001) and SUVmean (r = 0.714, P = 0.003). Significant positive correlations were also found between [68Ga]Ga-DOTA-TOC PET metrics, and VEGFA and VB. SSTR2 mRNA was moderately correlated with VEGFA (r = 0.539, P = 0.038). Neither [68Ga]Ga-DOTA-TOC PET metrics nor SSTR2 were correlated with proliferation or inflammation. CONCLUSION: [68Ga]Ga-DOTA-TOC accumulation in meningiomas is associated with SSTR2 binding and vascularization with TBRmean being the best PET metric for assessing SSTR2.

P53, Somatostatin receptor 2a and Chromogranin A immunostaining as prognostic markers in high grade gastroenteropancreatic neuroendocrine neoplasms.

BACKGROUND: High grade gastroenteropancreatic (GEP) neuroendocrine neoplasms (NEN) with a Ki67 proliferation index > 20%, include well-differentiated tumours grade 3 (NET G3) and poorly differentiated (PD) neuroendocrine carcinomas (NEC). Abnormal p53-expression is a feature of PD tumours, while expression of chromogranin A (CgA) and somatostatin-receptor 2a (SSTR-2a) may be a feature of well-differentiated tumours. The aim of this study was to elucidate the expression and prognostic value of these three markers in 163 GEP-NEN patients with a Ki67-index > 20%. METHOD: Clinical data, histopathology and overall survival were analysed according to Kaplan-Meier's method and Cox regression. The expression of SSTR-2a, CgA and synaptophysin was analysed in tumour specimens by immunohistochemistry, and semi-quantitatively scored as negative (< 5%), heterogeneously positive (5-30%) or strongly positive (> 30%). P53 was defined as normal when scored as heterogeneously positive (1-30%), and abnormal when negative (0%) or strongly positive (> 30%). RESULTS: In multivariate analysis, better survival was observed among patients with heterogeneously positive p53 compared to strongly positive (p < 0.001). When dichotomised, tumours with a heterogeneously positive p53 vs. negative and strongly positive p53 also showed a significantly better survival (p = 0.002). Survival was significantly worse for negative CgA compared to heterogeneously positive CgA (p = 0.02). Strongly positive SSTR-2a expression was found in 26% of the 163 included patients. Well-differentiated morphology correlated with strong expression of SSTR-2a and CgA, and heterogeneously positive p53-staining, and was more frequent in pancreatic primaries. In pancreatic primaries, strongly positive SSTR-2a was associated with longer survival (univariate analysis, p = 0.02). A significantly lower Ki67 proliferation index was found in patients with a heterogeneously positive p53, a positive SSTR-2a and CgA expression. CONCLUSION: Our results suggest that abnormal p53-expression is an independent negative prognostic marker in GEP-NEN with a Ki67-index > 20%. Patients with heterogeneously positive p53 had the best prognosis. SSTR-2a was a positive prognostic marker in pancreatic NEN. Negative CgA was associated with a significantly worse OS compared to heterogeneously positive CgA-expression in a multivariate sub-analysis. Lower Ki67 index correlated significantly with heterogeneously positive p53, positive SSTR-2a and CgA expression.

Imaging Macrophage and Hematopoietic Progenitor Proliferation in Atherosclerosis.

RATIONALE: Local plaque macrophage proliferation and monocyte production in hematopoietic organs promote progression of atherosclerosis. Therefore, noninvasive imaging of proliferation could serve as a biomarker and monitor therapeutic intervention. OBJECTIVE: To explore (18)F-FLT positron emission tomography-computed tomography imaging of cell proliferation in atherosclerosis. METHODS AND RESULTS: (18)F-FLT positron emission tomography-computed tomography was performed in mice, rabbits, and humans with atherosclerosis. In apolipoprotein E knock out mice, increased (18)F-FLT signal was observed in atherosclerotic lesions, spleen, and bone marrow (standardized uptake values wild-type versus apolipoprotein E knock out mice, 0.05 ± 0.01 versus 0.17 ± 0.01, P<0.05 in aorta; 0.13 ± 0.01 versus 0.28 ± 0.02, P<0.05 in bone marrow; 0.06 ± 0.01 versus 0.22 ± 0.01, P<0.05 in spleen), corroborated by ex vivo scintillation counting and autoradiography. Flow cytometry confirmed significantly higher proliferation of macrophages in aortic lesions and hematopoietic stem and progenitor cells in the spleen and bone marrow in these mice. In addition, (18)F-FLT plaque signal correlated with the duration of high cholesterol diet (r(2)=0.33, P<0.05). Aortic (18)F-FLT uptake was reduced when cell proliferation was suppressed with fluorouracil in apolipoprotein E knock out mice (P<0.05). In rabbits, inflamed atherosclerotic vasculature with the highest (18)F-fluorodeoxyglucose uptake enriched (18)F-FLT. In patients with atherosclerosis, (18)F-FLT signal significantly increased in the inflamed carotid artery and in the aorta. CONCLUSIONS: (18)F-FLT positron emission tomography imaging may serve as an imaging biomarker for cell proliferation in plaque and hematopoietic activity in individuals with atherosclerosis.

PET imaging with copper-64 as a tool for real-time in vivo investigations of the necessity for cross-linking of polymeric micelles in nanomedicine.

Polymeric micelles in nanomedicine are often cross-linked to prevent disintegration in vivo. This typically requires clinically problematic chemicals or laborious procedures. In addition, cross-linking may interfere with advanced release strategies. Despite this, it is often not investigated whether cross-linking is necessary for efficient drug delivery. We used positron emission tomography (PET) imaging with 64 Cu to demonstrate general methodology for real-time in vivo investigations of micelle stability. Triblock copolymers with 4-methylcoumarin cores of ABC-type (PEG-PHEMA-PCMA) were functionalized in the handle region (PHEMA) with CB-TE2A chelators. Polymeric micelles were formed by dialysis and one half was core cross-linked (CL) by UV light and the other half was not (nonCL). Both CL and nonCL were radiolabeled with 64 Cu and compared in vivo in tumor-bearing mice, with free 64 Cu as control. Accumulation in relevant organs was quantified by region of interest analysis on PET images and ex vivo counting. It was observed that CL and nonCL showed limited differences in biodistribution from each other, whereas both differed markedly from control (free 64 Cu). This demonstrated that 4-methylcoumarin core micelles may form micelles that are stable in circulation even without cross-linking. The methodology presented here where individual unimers are radiolabeled is applicable to a wide range of polymeric micelle types.

In vivo evaluation of PEGylated 64Cu-liposomes with theranostic and radiotherapeutic potential using micro PET/CT.

PURPOSE: The objective of this study was to evaluate the potential of PEGylated (64)Cu-liposomes in clinical diagnostic positron emission tomography (PET) imaging and PEGylated (177)Lu-liposomes in internal tumor radiotherapy through in vivo characterization and dosimetric analysis in a human xenograft mouse model. METHODS: Liposomes with 5 and 10 mol% PEG were characterized with respect to size, charge, and (64)Cu- and (177)Lu-loading efficiency. The tumor imaging potential of (64)Cu-loaded liposomes was evaluated in terms of in vivo biodistribution, tumor accumulation and tumor-to-muscle (T/M) ratios, using PET imaging. The potential of PEGylated liposomes for diagnostic and therapeutic applications was further evaluated through dosimetry analysis using OLINDA/EXM software. The (64)Cu-liposomes were used as biological surrogates to estimate the organ and tumor kinetics of (177)Lu-liposomes. RESULTS: High remote loading efficiency (>95 %) was obtained for both (64)Cu and (177)Lu radionuclides with PEGylated liposomes, and essentially no leakage of the encapsulated radionuclide was observed upon storage and after serum incubation for 24 h at 37 °C. The 10 mol% PEG liposomes showed higher tumor accumulation (6.2 ± 0.2 %ID/g) than the 5 mol% PEG liposomes, as evaluated by PET imaging. The dosimetry analysis of the (64)Cu-liposomes estimated an acceptable total effective dose of 3.3·10(-2) mSv/MBq for diagnostic imaging in patients. A high absorbed tumor dose (114 mGy/MBq) was estimated for the potential radiotherapeutic (177)Lu-liposomes. CONCLUSION: The overall preclinical profile of PEGylated (64)Cu-liposomes showed high potential as a new PET theranostic tracer for imaging in humans. Dosimetry results predicted that initial administered activity of 200 MBq of (64)Cu-liposomes should be acceptable in patients. Work is in progress to validate the utility of PEGylated (64)Cu-liposomes in a clinical research programme. The high absorbed tumor dose (114 mGy/MBq) estimated for (177)Lu-liposomes and the preliminary dosimetric studies justify further therapeutic and dosimetry investigation of (177)Lu-liposomes in animals before potential testing in man.

Somatostatin-Immunoreactive Pancreaticoduodenal Neuroendocrine Neoplasms: Twenty-Three Cases Evaluated according to the WHO 2010 Classification.

BACKGROUND/OBJECTIVE: Neuroendocrine neoplasms of the pancreas and duodenum with predominant or exclusive immunoreactivity for somatostatin (pdSOMs) are rare, and knowledge about tumour biology, treatment, survival and prognostic factors is limited. This study aims to describe clinical, pathological and biochemical features as well as treatment and prognosis of pdSOMs. DESIGN: Twenty-three patients with pdSOM (9 duodenal, 12 pancreatic and 2 unknown primary tumours) were identified from our prospective neuroendocrine tumour database, and data according to the study aims were recorded. RESULTS: Among the 9 patients with duodenal SOM, the male/female ratio was 4/5. All males and 1 female had neurofibromatosis type 1. Seven patients had stage 1A/B and 2 had stage 2B disease. The Ki-67 index was 1-5% (median 2%). Plasma somatostatin was elevated in the patients with 2B disease. Of the 14 patients with pancreatic SOM or an unknown primary tumour, the male/female ratio was 2/12. One male had multiple endocrine neoplasia type 1. Five had stage 1A/2B and 9 had stage 4. The Ki-67 index was 1-40% (median 7%). Plasma somatostatin was elevated in 7 patients. Patients reported symptoms related to the somatostatinoma syndrome, but none fulfilled the criteria for a full syndrome. Primary tumour in the pancreas, metastatic disease at diagnosis and higher tumour grade were all associated with significantly poorer survival. CONCLUSION: None of the patients with pdSOM presented with the full somatostatinoma syndrome. Prognostic factors are localisation of the primary tumour, dissemination and tumour grade. A Ki-67 index of 5% may discriminate the course of the disease.

FoxO3A promotes metabolic adaptation to hypoxia by antagonizing Myc function.

Exposure of metazoan organisms to hypoxia engages a metabolic switch orchestrated by the hypoxia-inducible factor 1 (HIF-1). HIF-1 mediates induction of glycolysis and active repression of mitochondrial respiration that reduces oxygen consumption and inhibits the production of potentially harmful reactive oxygen species (ROS). Here, we show that FoxO3A is activated in hypoxia downstream of HIF-1 and mediates the hypoxic repression of a set of nuclear-encoded mitochondrial genes. FoxO3A is required for hypoxic suppression of mitochondrial mass, oxygen consumption, and ROS production and promotes cell survival in hypoxia. FoxO3A is recruited to the promoters of nuclear-encoded mitochondrial genes where it directly antagonizes c-Myc function via a mechanism that does not require binding to the consensus FoxO recognition element. Furthermore, we show that FoxO3A is activated in human hypoxic tumour tissue in vivo and that FoxO3A short-hairpin RNA (shRNA)-expressing xenograft tumours are decreased in size and metabolically changed. Our findings define a novel mechanism by which FoxO3A promotes metabolic adaptation and stress resistance in hypoxia.

Positron emission tomography based analysis of long-circulating cross-linked triblock polymeric micelles in a U87MG mouse xenograft model and comparison of DOTA and CB-TE2A as chelators of copper-64.

Copolymers of ABC-type (PEG-PHEMA-PCMA) architecture were prepared by atom transfer radical polymerization and formulated as micelles with functionalizable primary alcohols in the shell-region (PHEMA-block) to which the metal-ion chelators DOTA or CB-TE2A were conjugated. Using this micelle system we compared the in vivo stabilities of DOTA and CB-TE2A as chelators of (64)Cu in micelle nanoparticles. The coumarin polymer (PCMA-block) micelle core was cross-linked by UV irradiation at 2 W/cm(2) for 30 min. The cross-linked micelles were labeled with (64)Cu at room temperature for 2 h (DOTA) or 80 °C for 3 h (CB-TE2A), giving labeling efficiencies of 60-76% (DOTA) and 40-47% (CB-TE2A). (64)Cu-micelles were injected into tumor-bearing mice (8 mg/kg) and PET/CT scans were carried out at 1, 22, and 46 h postinjection. The micelles showed good blood stability (T1/2: 20-26 h) and tumor uptake that was comparable with other nanoparticle systems. The DOTA micelles showed a biodistribution similar to the CB-TE2A micelles and the tumor uptake was comparable for both micelle types at 1 h (1.9% ID/g) and 22 h (3.9% ID/g) but diverged at 46 h with 3.6% ID/g (DOTA) and 4.9% ID/g (CB-TE2A). On the basis of our data, we conclude that cross-linked PEG-PHEMA-PCMA micelles have long circulating properties resulting in tumor accumulation and that DOTA and CB-TE2A (64)Cu-chelates show similar in vivo stability for the studied micelle system.

Positron emission tomography/computed tomography for optimized colon cancer staging and follow up.

OBJECTIVES: Optimal management of colon cancer (CC) requires detailed assessment of extent of disease. This study prospectively investigates the diagnostic accuracy of 2-deoxy-2-[18F]fluoro-D-glucose positron emission tomography/computed tomography (PET/CT) for staging and detection of recurrence in primary CC. MATERIAL AND METHODS: PET/CT for preoperative staging was performed in 66 prospectively included patients with primary CC. Diagnostic accuracy for PET/CT and CT was analyzed. In addition to routine follow up, 42 stages I-III CC patients had postoperative PET/CT examinations every 6 months for 2 years. Serological levels of tissue inhibitor of metalloproteinase-1 (TIMP-1), carcinoembryonic antigen, and liberated domain I of urokinase plasminogen activator receptor were analyzed. RESULTS: Accuracy for tumor, nodal, and metastases staging by PET/CT were 82% (95% confidence interval [CI]: 70; 91), 66% (CI: 51; 78), and 89% (CI: 79; 96); for CT the accuracy was 77% (CI: 64; 87), 60% (CI: 46; 73), and 69% (CI: 57; 80). Cumulative relapse incidences for stages I-III CC at 6, 12, 18, and 24 months were 7.1% (CI: 0; 15); 14.3% (CI: 4; 25); 19% (CI: 7; 31), and 21.4% (CI: 9; 34). PET/CT diagnosed all relapses detected during the first 2 years. High preoperative TIMP-1 levels were associated with significant hazards toward risk of recurrence and shorter overall survival. CONCLUSIONS: This study indicates PET/CT as a valuable tool for staging and follow up in CC. TIMP-1 provided prognostic information potentially useful in selection of patients for intensive follow up.

Prognostic thresholds of fluorine-18 fluorodeoxyglucose-positron emission tomography mean and maximum standardized uptake values for survival and nodal involvement in lung neuroendocrine neoplasms.

OBJECTIVES: The mean standardized uptake value (SUVmean) and maximum standardized uptake value (SUVmax) on fluorine-18 fluorodeoxyglucose-positron emission tomography are prognostic biomarkers for survival and nodal involvement in non-small-cell lung cancer but their prognostic value in lung neuroendocrine neoplasms (NENs) remains unexplored. In this study, we aimed to examine whether they are also prognostic biomarkers for survival and nodal involvement in lung NENs. METHODS: We retrospectively studied patients with typical carcinoid, atypical carcinoid or large cell neuroendocrine carcinoma who had been radically resected at our institution between 2008 and 2020. We measured SUVmean and SUVmax on all primary tumours and lymph nodes that were clinically and/or pathologically involved. We dichotomized the patients into groups of high or low SUVmean and SUVmax of the primary tumour using time-dependent receiver operating characteristic curves and compared their overall survival using Kaplan-Meier curves and Cox models. Lastly, we predicted the patients' pathological nodal status with SUVmean and SUVmax of the lymph nodes using binomial logistic models. RESULTS: The study included 245 patients. Patients died earlier if their SUVmean of the primary tumour exceeded 3.9 [hazard ratio 1.97, 95% confidence interval (CI) 1.27-3.04, P = 0.002] or SUVmax exceeded 5.3 (hazard ratio 1.85, 95% CI 1.20-2.87, P = 0.006). Likewise, patients had a higher risk of pathological nodal involvement if their SUVmean of the lymph nodes exceeded 3.3 (odds ratio 10.00, 95% CI 2.59-51.01, P = 0.002) or SUVmax exceeded 4.2 (odds ratio 4.00, 95% CI 1.20-14.65, P = 0.028). CONCLUSIONS: The fluorine-18 fluorodeoxyglucose-positron emission tomography SUVmean and SUVmax are strong prognostic biomarkers for survival and nodal involvement in lung NENs and could be important guides for making treatment decisions.

Urokinase Plasminogen Activator Receptor: An Important Focal Player in Chronic Subdural Hematoma?

Chronic subdural hematoma (CSDH) development involves inflammatory, angiogenetic, and fibrinolytic mechanisms, several components of which are now unraveled through intensive research. The urokinase plasminogen activator receptor (uPAR) is part of the plasminogen activator system and possesses inflammatory, angiogenetic, and fibrinolytic capabilities. As a first, this study aims to identify uPAR in the hematoma fluid, hematoma membrane, dura mater, and systemic blood from patients with CSDH and, if present, to investigate if the uPAR level at the time of surgery may be a predictor for later developing recurrent CSDH. uPAR expression in the hematoma membrane and dura mater was analyzed using immunohistochemistry and presented as the H-score of the positive immunostaining. The uPAR levels in the hematoma fluid and systemic blood were determined using a multiplex antibody bead kit (Luminex). Samples were collected at the time of the first CSDH surgery, and in the case of recurrent CSDH within 90 days, the samples were again collected at reoperation. A comparison of uPAR expression between the hematoma membrane and dura mater, as well as uPAR levels in systemic blood and hematoma fluid, was performed using the Wilcoxon rank sum test. We included 112 patients, 26 of whom had recurrent CSDH. The median hematoma uPAR level was 22,125 (14,845-33,237) and significantly higher than the median systemic blood level of 789 pg/L (465-2,088) (p < 0.001). Similarly, the uPAR level of the hematoma membrane was 14.3 (7.54-44.8) and significantly higher than the dural uPAR level of 0.81 (0.3-1.98) (p < 0.001). For the first time, we identified uPAR in the subdural fluid, hematoma membrane, dura mater, and systemic blood from patients with CSDH. The high expression of uPAR in the subdural fluid and hematoma membrane indicates that the mechanisms of CSDH are predominantly in the subdural fluid collection and surrounding hematoma membrane.

Longitudinal imaging of murine atherosclerosis with 2-deoxy-2-[18F]fluoro-D-glucose and [18F]-sodium fluoride in genetically modified Apolipoprotein E knock-out and wild type mice.

In a longitudinal design, four arterial segments in mice were followed by positron emission tomography/computed tomography (PET/CT) imaging. We aimed to determine how the tracers reflected the development of atherosclerosis via the uptake of 2-deoxy-2-[18F]fluoro-D-glucose ([18F]FDG) for imaging inflammation and [18F]-sodium fluoride (Na[18F]F) for imaging active microcalcification in a murine model of atherosclerosis. Apolipoprotein E knock-out (ApoE) mice and C57 BL/6NtaC (B6) mice were divided into four groups. They received either normal chow (N = 7, ApoE mice and N = 6, B6 mice) for 32 weeks or a high-fat diet (N = 6, ApoEHFD mice and N = 9, B6HFD mice) for 32 weeks. The mice were scanned with [18F]FDG and Na[18F]F using a dedicated small animal PET/CT scanner at three timepoints. The tracer uptakes in four aortic segments (abdominal aorta, aortic arch, ascending aorta, and thoracic aorta) were measured and reported as SUVmax values. The uptake of [18F]FDG (SUVmax: 5.7 ± 0.5 vs 1.9 ± 0.2, 230.3%, p = < 0.0001) and Na[18F]F (SUVmax: 9.6 ± 1.8 vs 4.0 ± 0.3, 175%, p = 0.007) was significantly increased in the abdominal aorta of ApoEHFD mice at Week 32 compared to baseline abdominal aorta values of ApoEHFD mice. [18F]FDG uptake in the aortic arch, ascending aorta and the thoracic aorta of B6HFD mice at Week 32 showed a robust resemblance to the abdominal aorta uptake whereas the Na[18F]F uptake only resembled in the thoracic aorta of B6HFD mice at Week 32 compared to the abdominal aorta. The uptake of both [18F]FDG and Na[18F]F increased as the disease progressed over time, and the abdominal aorta provided a robust measure across mouse strain and diet. Therefore, it seems to be the preferred region for image readout. For [18F]FDG-PET, both B6 and ApoE mice provide valuable information and either mouse strain may be used in preclinical cardiovascular studies, whereas for Na[18F]F -PET, ApoE mice should be preferred.

Imaging of atherosclerosis with [64Cu]Cu-DOTA-TATE in a translational head-to-head comparison study with [18F]FDG, and Na[18F]F in rabbits.

Atherosclerosis is a chronic inflammatory disease of the larger arteries that may lead to cardiovascular events. Identification of patients at highest risk of cardiovascular events is challenging, but molecular imaging using positron emission tomography (PET) may prove useful. The aim of this study was to evaluate and compare head-to-head three different PET tracers. Furthermore, tracer uptake is compared to gene expression alterations of the arterial vessel wall. Male New Zealand White rabbits (control group; n = 10, atherosclerotic group; n = 11) were used for the study. Vessel wall uptake was assessed with the three different PET tracers: [18F]FDG (inflammation), Na[18F]F (microcalcification), and [64Cu]Cu-DOTA-TATE (macrophages), using PET/computed tomography (CT). Tracer uptake was measured as standardized uptake value (SUV), and arteries from both groups were analyzed ex vivo by autoradiography, qPCR, histology, and immunohistochemistry. In rabbits, the atherosclerotic group showed significantly higher uptake of all three tracers compared to the control group [18F]FDG: SUVmean 1.50 ± 0.11 versus 1.23 ± 0.09, p = 0.025; Na[18F]F: SUVmean 1.54 ± 0.06 versus 1.18 ± 0.10, p = 0.006; and [64Cu]Cu-DOTA-TATE: SUVmean 2.30 ± 0.27 versus 1.65 ± 0.16; p = 0.047. Of the 102 genes analyzed, 52 were differentially expressed in the atherosclerotic group compared to the control group and several genes correlated with tracer uptake. In conclusion, we demonstrated the diagnostic value of [64Cu]Cu-DOTA-TATE and Na[18F]F for identifying atherosclerosis in rabbits. The two PET tracers provided information distinct from that obtained with [18F]FDG. None of the three tracers correlated significantly to each other, but [64Cu]Cu-DOTA-TATE and Na[18F]F uptake both correlated with markers of inflammation. [64Cu]Cu-DOTA-TATE was higher in atherosclerotic rabbits compared to [18F]FDG and Na[18F]F.

Subdural Levels of Interleukin 1-receptor Antagonist are Elevated in Patients with Recurrent Chronic Subdural Hematomas.

Anti-inflammatory treatment reduces the risk of recurrent chronic subdural hematoma (CSDH), but clinical implementation is improper due to side effects. Exact knowledge of subdural molecules involved in recurrent CSDH may lead to targeted medical treatment and possibly improve the prospect of a personalized approach by eliminating the broad use of anti-inflammatory drugs on the entire CSDH population. With this study, we aim to (1) describe the associations between cytokine levels at the primary surgery and the risk of subsequent recurrence and (2) describe the association between cytokines in patients with recurrent CSDH between the first and second operations. Systemic and subdural levels of pro- and anti-inflammatory cytokines were measured and compared between patients with the first-time CSDH and recurrent CSDH. Cytokine levels were analyzed using a multiplex antibody bead kit. In case of recurrent CSDH within 90 days of follow-up, the samples were re-collected and analyzed. We included 101 adult CSDH patients of which 20 had a recurrence. The levels of cytokines in the CSDH fluid from patients who were operated on for the first-time CSDH were not associated with the risk of later developing a recurrence. We found interleukin-1 receptor antagonist (IL-1ra) to be elevated in subdural fluid in patients with recurrent CSDH at the time of their second operation (p = 0.0005). This study provides knowledge on cytokine composition in the subdural fluid in patients with CSDH with and without recurrence. IL-1ra is elevated in subdural fluid in patients with recurrent CSDH at the time of the second operation, identifying a possible medical target.

Urokinase-Type Plasminogen Activator Receptor (uPAR) Expression and [64Cu]Cu-DOTA-AE105 uPAR-PET/CT in Patient-Derived Xenograft Models of Oral Squamous Cell Carcinoma.

PURPOSE: [64Cu]Cu-DOTA-AE105 urokinase-type plasminogen activator receptor (uPAR)-PET/CT is a novel and promising imaging modality for cancer visualization, although it has not been tested in head and neck cancer patients nor in preclinical models that closely resemble these heterogenous tumors, i.e., patient-derived xenograft (PDX) models. The aim of the present study was to establish and validate oral squamous cell carcinoma (OSCC) PDX models and to evaluate [64Cu]Cu-uPAR-PET/CT for tumor imaging in these models. PROCEDURES: PDX flank tumor models were established by engrafting tumor tissue from three patients with locally advanced OSCC into immunodeficient mice. [64Cu]Cu-DOTA-AE105 was injected in passage 2 (P2) mice, and [64Cu]Cu-uPAR-PET/CT was performed 1 h and 24 h after injection. After the last PET scan, all animals were euthanized, and tumors dissected for autoradiography and immunohistochemical (IHC) staining. RESULTS: Three PDX models were established, and all of them showed histological stability and unchanged heterogenicity, uPAR expression, and Ki67 expression through passages. A significant correlation between uPAR expression and tumor growth was found. All tumors of all models (n=29) showed tumor uptake of [64Cu]Cu-DOTA-AE105. There was a clear visual concordance between the distribution of uPAR expression (IHC) and [64Cu]Cu-DOTA-AE105 uptake pattern in tumor tissue (autoradiography). No significant correlation was found between IHC (H-score) and PET-signal (SUVmax) (r=0.34; p=0.07). CONCLUSIONS: OSCC PDX models in early passages histologically mimic donor tumors and could serve as a valuable platform for the development of uPAR-targeted imaging and therapeutic modalities. Furthermore, [64Cu]Cu-uPAR-PET/CT showed target- and tumor-specific uptake in OSCC PDX models demonstrating the diagnostic potential of this modality for OSCC patients.