Multimodal imaging of tumor response to sorafenib combined with radiation therapy: comparison between diffusion‐weighted MRI,choline spectroscopy and 18F‐FLT PET imaging

The purpose of this study was to determine the value of different imaging modalities, that is, magnetic resonance imaging/spectroscopy (MRI/MRS) and positron emission tomography (PET), to assess early tumor response to sorafenib with or without radiotherapy. Diffusion‐weighted (DW)‐MRI, choline ¹H MRS at 11.7 T, and ¹⁸F‐FLT PET imaging were used to image fibrosarcoma (FSaII) tumor‐bearing mice over time. The imaging markers were compared with apoptosis cell death and cell proliferation measurements assessed by histology. Anti‐proliferative effects of sorafenib were evidenced by ¹H MRS and ¹⁸F‐FLT PET after 2 days of treatment with sorafenib, with no additional effect of the combination with radiation therapy, results that are in agreement with Ki67 staining. Apparent diffusion coefficient calculated using DW‐MRI was not modified after 2 days of treatment with sorafenib, but showed significant increase 24 h after 2 days of sorafenib treatment combined with consecutive irradiation. The three imaging markers were able to show early tumor response as soon as 24 h after treatment initiation, with choline MRS and ¹⁸F‐FLT being sensitive to sorafenib in monotherapy as well as in combined therapy with irradiation, whereas DW‐MRI was only sensitive to the combination of sorafenib with radiotherapy. Copyright © 2013 John Wiley & Sons, Ltd.     

18F‐FDG PET/CT and MRI in the follow‐up of head and neck squamous cell carcinoma

We evaluated the diagnostic performance of ¹⁸F‐FDG PET/CT and MRI for the assessment of head and neck squamous cell carcinoma (HNSCC) relapse. Since early treatment might prevent inoperable relapse, we also evaluated THE performance of early unenhanced ¹⁸F‐FDG PET/CT in residual tumor detection. The study was prospectively performed on 32 patients who underwent ¹⁸F‐FDG PET/CT and MRI before treatment and at 4 and 12 months after treatment. ¹⁸F‐FDG PET/CT was also performed 2 weeks after the end of radiotherapy. Histopathology or a minimum of 18 months follow‐up were used as gold standard. Before treatment ¹⁸F‐FDG PET/CT and MRI detected all primary tumors except for two limited vocal fold lesions (sensitivity 94%). MRI was more sensitive than ¹⁸F‐FDG PET/CT for the detection of local extension sites (sensitivity 75 vs 58%), but at the cost of a higher rate of false positive results (positive predictive value 74 vs 86%). For relapse detection at 4 months, sensitivity was significantly higher for ¹⁸F‐FDG PET/CT (92%) than for MRI (70%), but the diagnostic performances were not significantly different at 12 months. For the detection of residual malignant tissue 2 weeks post‐radiotherapy, sensitivity and specificity of ¹⁸F‐FDG PET/CT were respectively 86 and 85% (SUV cut‐off value 5.8). ¹⁸F‐FDG PET/CT is effective in the differentiation between residual tumor and radiation‐induced changes, as early as 2 weeks after treatment of a primary HNSCC. For follow‐up, performance of ¹⁸F‐FDG PET/CT and MRI are similar except for a higher sensitivity of ¹⁸F‐FDG PET/CT at 4 months. Copyright © 2011 John Wiley & Sons, Ltd.     

Monitoring Response to Radiotherapy in Human Squamous Cell Cancer Bearing Nude Mice: Comparison of 2′-deoxy-2′-[<Superscript>18</Superscript>F]fluoro-<Emphasis Type="SmallCaps">d</Emphasis>-glucose (FDG) and 3′-[<Superscript>18</Superscript>F]fluoro-3′-deoxythymidine (FLT)

Objective The uptake of 3′-[¹⁸F]fluoro-3′-deoxythymidine (FLT), a proliferation marker, was measured before and during fractionated radiotherapy to evaluate the potential of FLT-positron emission tomography (PET) imaging as an indicator of tumor response compared to 2′-deoxy-2′-[¹⁸F]fluoro-d-glucose (FDG). Materials and Methods Nude mice bearing established human head and neck xenografts (HNX-OE; nu/nu mice) were locally irradiated (three fractions/week; 22 Gy) using a 150-kV_p unit. Multiple FDG- and FLT-PET scans were acquired during treatment. Tumor volume was determined regularly, and tissue was analyzed for biomarkers involved in tracer uptake. Results Both groups revealed a significant decline in tumor volume (P < 0.01) compared to untreated tumors. For FDG as well as for FLT, a significant decline in retention was observed at day 4. For FLT, most significant decline in retention was observed at day 12; whereas, for FDG, this was already noted at day 4. Maximum decline in tumor-to-nontumor ratios (T/NT) for FDG and FLT was 42 ± 18% and 49 ± 16% (mean ± SD), respectively. FLT uptake was higher then that of FDG. For FLT, statistical significant correlations were found for both tumor volume at baseline and at day 29 with T/NT and ΔT/NT. All tumors demonstrated expression of glucose transporter-1, thymidine kinase-1, and hexokinase II. No differences were found for amount of tumor cells and necrosis at the end of treatment. Conclusion This new experimental in vivo model supports the promise of using FLT-PET, as with FDG-PET, to monitor response to external radiotherapy. This warrants further clinical studies to compare these two tracers especially in cancers treated with radiotherapy.     

Assessment of the biodistribution of an [18F]FDG‐loaded perfluorocarbon double emulsion using dynamic micro‐PET in rats

Perfluorocarbon (PFC) double emulsions loaded with a water‐soluble, therapeutic agent can be triggered by ultrasound in a process known as acoustic droplet vaporization. Elucidating the stability and biodistribution of these sonosensitive vehicles and encapsulated agents is critical in developing targeted drug delivery strategies using ultrasound. [¹⁸F]fluorodeoxyglucose (FDG) was encapsulated in a PFC double emulsion and the in vitro diffusion of FDG was assessed using a Franz diffusion cell. Using dynamic micro‐positron emission tomography and direct tissue sampling, the biodistribution of FDG administered as a solution (i.e. non‐emulsified) or as an emulsion was studied in Fisher 344 rats (n = 6) bearing subcutaneous 9L gliosarcoma. Standardized uptake values (SUVs) and area under the curve of the SUV (AUC_SUV) of FDG were calculated for various tissues. The FDG flux from the emulsion decreased by up to a factor of 6.9 compared with the FDG solution. FDG uptake, calculated from the AUC_SUV, decreased by 36% and 44% for brain and tumor, respectively, when comparing FDG solution vs FDG emulsion (p < 0.01). Decreases in AUC_SUV in highly metabolic tissues such as brain and tumor demonstrated retention of FDG within the double emulsion. No statistically significant differences in lung AUC_SUV were observed, suggesting minimal accumulation of the emulsion in the pulmonary capillary bed. The liver AUC_SUV increased by 356% for the FDG emulsion, thus indicating significant hepatic retention of the emulsion. Copyright © 2013 John Wiley & Sons, Ltd.     

DW‐MRI and 18F‐FLT PET for early assessment of response to radiation therapy associated with hypoxia‐driven interventions. Preclinical studies using manipulation of oxygenation and/or dose escalation

Early markers of treatment response may help in the management of patients by predicting the outcome of a specific therapeutic intervention. Here, we studied the potential value of diffusion‐weighted MRI (DW‐MRI) and ¹⁸F‐fluorothymidine (¹⁸F‐FLT), markers of cell death and cell proliferation respectively, to predict the response to irradiation. In addition, dose escalation and/or carbogen breathing were used to modulate the response to irradiation. The studies were performed on two hypoxic rat tumor models: rhabdomyosarcoma and 9L‐glioma. The rats were imaged using MRI and PET before and two days after the treatment. In both tumor models, changes in ADC (apparent diffusion coefficient) and ¹⁸F‐FLT SUV (standardized uptake value) were significantly correlated with the tumor growth delay. For both tumor models, the ADC values increased in all irradiated groups two days after the treatment while they decreased in the untreated groups. At the same time, the uptake of ¹⁸F‐FLT increased in the untreated groups and decreased in all treated groups. Yet, ADC values were not sensitive enough to predict the added value of dose escalation or carbogen breathing in either model. Change in ¹⁸F‐FLT uptake was able to predict the higher tumor response when using increased dose of irradiation, but not when using a carbogen breathing challenge. Our results also emphasize that the magnitude of change in ¹⁸F‐FLT uptake was strongly dependent on the tumor model. Copyright © 2015 John Wiley & Sons, Ltd.     

L‐Amino acid load to enhance PET differentiation between tumor and inflammation: an in vitro study on 18F‐FET uptake

Labeled amino acids (AA) are tumor tracers for use in nuclear medecine. O‐(2‐[¹⁸F]fluoroethyl)‐L ‐tyrosine (FET) is transported by the L ‐system, known to function as an exchanger. In vitro utilization of FET, after a preload or prior to an afterload of non radioactive L ‐amino acids, was evaluated in order to measure the potential effects of AA content on the distinction between tumor and inflammatory lesions. Cellular uptake of FET was studied on rat osteosarcoma cells (ROS 17/2.8) and human leukocytes, initially loaded with nonradioactive L ‐tyrosine or L ‐methionine. FET efflux was evaluated from cells loaded with nonradioactive L ‐phenylalanine after tracer uptake. ROS 17/2.8 showed a higher sensitivity to preload and afterload effects on cellular FET content as compared with the leukocytes. We conclude that preload with L ‐tyrosine, prior to the administration of FET, may be a potential procedure to improve PET differentiation between tumor and inflammatory lesions. Copyright © 2006 John Wiley & Sons, Ltd.     

In vitro effects of the cyclooxygenase inhibitor indomethacin and of the phospholipase‐C inhibitor U‐73122 on carbachol‐induced contractions of porcine detrusor muscle

Prostaglandin synthetase inhibitors belong to one substance class additionally used in the treatment of bladder dysfunctions associated with involuntary bladder contractions. However, the mechanism of action of non‐steroidal anti‐inflammatory drugs (NSAIDs) on the detrusor muscle is not clear. In this study, it was examined in vitro whether the NSAID indomethacin exhibited an inhibitory effect on carbachol‐induced contractions of the porcine detrusor muscle. Additionally, the inhibitory effect of the phospholipase‐C inhibitor U‐73122 on carbachol‐induced contractions of the porcine detrusor muscle was investigated. Experiments were performed on the muscle strips of the porcine detrusor muscle suspended in a tissue bath. Effects of indomethacin at 10^?6 and 10^?5 m on the maximum carbachol‐induced contraction and on the carbachol–response curve were investigated. Additionally, the inhibitory influence of U‐73122 at a concentration of 10^?5.5 m on the carbachol–response curve was investigated. Pretreatment with indomethacin at both concentrations did not result in a significant reduction in the maximum contraction compared with the control. In the experiments in which carbachol concentration‐response curves were generated, indomethacin exhibited at both concentrations a very small but significant change at carbachol concentrations of 10^?8 and 10^?7.5 m . In the experiments with U‐73122, a significant change was found in the concentration–response curve of carbachol at all concentrations of carbachol from 10^?6.5 to 10^?4 m . The mean maximum carbachol‐induced contraction was 141.8 ± 6.8% after incubation with U‐73122 and 166.0 ± 6.4% in the control group (P < 0.05). Indomethacin did not inhibit the carbachol‐induced contractions of the porcine detrusor muscle. The cyclooxygenase does not play a significant role in the carbachol‐induced bladder contraction of the porcine detrusor muscle. The inhibitory action of the phospholipase‐C inhibitor U‐73122 on the carbachol‐induced contraction was significant, but small. The results point to an inferior role of this pathway.     

Prediction of Short-term Survival in Patients with Advanced Nonsmall Cell Lung Cancer Following Chemotherapy Based on 2-Deoxy-2-[F-18]fluoro-<Emphasis Type="SmallCaps">d</Emphasis>-glucose-Positron Emission Tomography: A Feasibility Study

INTRODUCTION: Dynamic positron emission tomography (PET) studies with 2-deoxy-2-[F-18]fluoro-D-glucose (FDG) were performed in patients with advanced nonsmall cell lung cancer (NSCLC) who received palliative chemotherapy to evaluate the impact of full kinetic analysis and assess its value with regard to short or long survival. MATERIALS AND METHODS: The evaluation includes 42 metastatic lesions in 14 patients with NSCLC. All patients received a combined chemotherapeutic protocol consisting of vinorelbin and oxaliplatin. The survival data served as reference for the PET data. All patients were examined before onset of chemotherapy and on day 15-21 after onset of the first cycle. The following parameters were retrieved from the dynamic PET studies: standardized uptake value (SUV), fractal dimension, two-compartment model with computation of k1, k2, k3, k4 (unit: 1/min), the fractional blood volume, and the FDG-influx according to Patlak was calculated using the formula (k1 x k3) / (k2 + k3). We used a two-group classification, namely, a short- and long-term survival group based on the median survival time (193 days) as a cutoff. A support vector machines (SVM) analysis was used for classification of the two a prior defined groups. RESULTS: The observed survival times varied from 40 to 392 days with a median survival time of 193 days. Most kinetic parameters demonstrated only small changes mostly declining after one cycle. The change in all kinetic parameters did not correlate to the survival-based classification. The change in SUV was significant between the first and second study (p = 0.006) but without an impact on the prediction of short or long survival. SVM-based analysis revealed the highest correct classification rate (CCR) between short and long survival for the combination of SUV and influx of the first study and SUV, influx, k2, and k4 of the second study with a CCR of 95.2%. CONCLUSION: The results demonstrate that a full kinetic analysis of the FDG kinetics in NSCLC is helpful for the classification into short or long survival and may be used to identify those patients who may benefit from this palliative chemotherapeutic protocol.     

Performance of a new fluorescence‐labeled MMP inhibitor to image tumor MMP activity in vivo in comparison to an MMP‐activatable probe

Matrix metalloproteinases (MMPs) are proteolytic enzymes that degrade structural components of the extracellular matrix and participate in pathologies such as cancer and inflammatory disorders. The development of novel contrast agents for optical imaging of MMP activity in vivo is of great interest. The commonly used near‐infrared fluorescence‐compatible agents are dye‐quenched probes that do not emit fluorescence until they interact with MMPs. In contrast, fluorescent synthetic low‐molecular‐weight MMP inhibitors have not been systematically employed. The aim of this study was to evaluate the performance of our recently developed Cy5.5‐labeled MMP inhibitor to image MMP activity in tumors in vivo compared with activatable fluorescent MMP‐sensing probes. The dynamic uptake of Cy5.5‐AF489 into four different tumor entities was analyzed in xenografted mice by intravenous injection and subsequent fluorescence reflectance imaging. Tumors were characterized in regard to their MMP‐2 and ?9 mRNA expressions (qRT‐PCR analysis), proteins (immunohistochemistry) and gelatinase/collagenase activities (in situ zymography). Cy5.5‐AF489 was compared with MMPSense^TM 680 and MMPSense^TM 750 FAST, two commercially available MMP‐activatable probes. Cy5.5‐AF489 showed a specific tumor uptake, which was blocked by pre‐injection of the unlabeled MMPI, and discriminated between tumors with high or low MMP‐2/‐9 expressions. Our optical probe facilitated faster visualization of MMP‐active tumors accompanied by excellent tumor‐to‐background ratios when compared with activatable probes. The MMP inhibitor Cy5.5‐AF489 permits fast in vivo imaging of MMP expression/activity in tumors. Given its small molecular weight and non‐peptidic structure, translational imaging from a preclinical application to a diagnostic tool for MMP‐related diseases seems feasible. Copyright © 2012 John Wiley & Sons, Ltd.     

The hyperfibrinolytic state of mice with combined thrombin‐activatable fibrinolysis inhibitor (TAFI) and plasminogen activator inhibitor‐1 gene deficiency is critically dependent on TAFI deficiency

Summary. Background: Mice with single gene deficiency of thrombin‐activatable fibrinolysis inhibitor (TAFI) or plasminogen activator inhibitor‐1 (PAI‐1) have an enhanced fibrinolytic capacity. Objectives: To unravel the function and relevance of both antifibrinolytic proteins through the generation and characterization of mice with combined TAFI and PAI‐1 gene deficiency. Results: Mating of TAFI knockout (KO) mice with PAI‐1 KO mice resulted in the production of TAFI/PAI‐1 double‐KO mice that were viable, were fertile, and developed normally. In a tail vein bleeding model, the bleeding time and hemoglobin content of the TAFI/PAI‐1 double‐KO mice did not deviate significantly from those of the single‐KO mice or of the wild‐type (WT) counterparts. Interestingly, in ex vivo rotational thromboelastometry measurements with whole blood samples, TAFI KO mice and TAFI/PAI‐1 double‐KO mice were more sensitive to fibrinolytic activation with tissue‐type plasminogen activator than WT or PAI‐1 KO mice. This enhanced fibrinolytic capacity was confirmed in vivo in a mouse thromboembolism model, as shown by decreased fibrin deposition in the lungs of TAFI KO mice and TAFI/PAI‐1 double‐KO mice as compared with WT or PAI‐1 KO mice.Conclusions: TAFI gene inactivation predominantly contributes to the increased fibrinolytic capacity of TAFI and PAI‐1 double‐gene‐deficient mice, as observed in some basic thrombosis models.     

Clinical pharmacology of single‐ and multiple‐ascending doses of ACT‐178882, a new direct renin inhibitor,and its pharmacokinetic interaction with food and midazolam

This study investigated the tolerability, safety, pharmacokinetics, and pharmacodynamics of ACT‐178882, a new direct renin inhibitor, as well as its interaction with food and midazolam. Healthy male subjects received either single (10–1000 mg) or multiple doses (30–600 mg) administered once daily for 14 days of ACT‐178882, placebo, or 20 mg enalapril in the fasted state. Following a 2‐week washout, the single dose of 30 mg ACT‐178882 was also administered in the fed state. In the multiple‐ascending‐dose part, subjects were dosed with midazolam on days ?2, 2, and 12 to investigate interactions with CYP3A4. Dizziness and headache were the most frequently reported adverse events. No clinically relevant changes occurred for body weight, vital signs, clinical laboratory variables, and ECG although both enalapril and ACT‐178882 tended to decrease systolic blood pressure. Following single doses of ACT‐178882, t_1/2 and t_max varied from 18.7 to 24.7 h and from 3 to 5 h, respectively, and food had no significant effect. Steady‐state conditions were achieved after 4–6 days of dosing and accumulation was minimal. ACT‐178882 pharmacokinetics were dose proportional. ACT‐178882 but not enalapril dose‐dependently increased C_max and area under the concentration‐time curve of midazolam. Single and multiple doses of ACT‐178882 dose‐dependently increased active renin and decreased plasma renin activity, whereas enalapril increased both variables. No effects on urinary excretion of creatinine, potassium, and the 6β‐hydroxycortisol/cortisol ratio were observed, whereas sodium and aldosterone excretion was decreased by both ACT‐178882 and enalapril. The current results with ACT‐178882 warrant further clinical investigation of this renin inhibitor in hypertensive patients.     

Antibody‐functionalized nanoparticles for imaging cancer: influence of conjugation to gold nanoparticles on the biodistribution of 89Zr‐labeled cetuximab in mice

Antibody‐labeled gold nanoparticles represent a promising novel tool regarding cancer imaging and therapy. Nevertheless, the characterization of biodistribution of such immunonanocarriers has been poorly documented. In this study, the biodistribution of ⁸⁹Zr‐labeled cetuximab before and after the coupling reaction to gold nanoparticles (AuNPs) was compared and the quantitative imaging performance of ⁸⁹Zr immuno‐PET was evaluated. Cetuximab was functionalized with the desferal moiety and labeled with ⁸⁹Zr (⁸⁹Zr–Df–Bz–NCS–cetuximab). AuNPs with a mean diameter of 5 nm were synthesized according a new method developed in the laboratory, and conjugated to ⁸⁹Zr–Df–Bz–NCS–cetuximab using carbodiimide chemistry (AuNPs–PPAA–cetuximab–⁸⁹Zr). The two tracers were injected in A431 xenograft‐bearing mice. Tumor and liver uptakes were assessed at different times after injection using quantitative PET imaging. The in vivo specificity of the binding was investigated using a saturating dose of unlabeled cetuximab. Radiolabeled cetuximab was conjugated to AuNPs with a coupling reaction yield >75%. All conjugates were stable in vitro and to a lesser extent in plasma. In vivo distribution studies revealed no significant difference in tumor uptake for cetuximab conjugated to nanoparticles up to 72 h after injection, compared with unconjugated cetuximab. Immuno‐PET studies showed that AuNPs–PPAA–cetuximab–⁸⁹Zr provided high tumor‐to‐background ratio. The liver uptake of AuNPs–PPAA–cetuximab–⁸⁹Zr was higher, compared with ⁸⁹Zr–Df–Bz–NCS–cetuximab. In vivo blocking experiments demonstrated selective tumor targeting after coupling reaction. This study showed that the conjugation of AuNPs to cetuximab did not affect its tumor accumulation and that the efficacy of EGFR‐targeted nanoparticles was unaltered. The ⁸⁹Zr‐labeled cetuximab‐targeted gold nanoparticles could be a valuable tool for theranostic purposes. Copyright © 2013 John Wiley & Sons, Ltd.