In vivo mouse imaging and spectroscopy in drug discovery

Imaging modalities such as micro-computed tomography (micro-CT), micro-positron emission tomography (micro-PET), high-resolution MRI, optical imaging, and high-resolution ultrasound have become invaluable tools in preclinical pharmaceutical research. They can be used to non-invasively investigate, in vivo, rodent biology and metabolism, disease models, and pharmacokinetics and pharmacodynamics of drugs. The advantages and limitations of each approach usually determine its application, and therefore a small-rodent imaging laboratory in a pharmaceutical environment should ideally provide access to several techniques. In this paper we aim to illustrate how these techniques may be used to obtain meaningful information for the phenotyping of transgenic mice and for the analysis of compounds in murine models of disease.     

Time-resolved contrast-enhanced three-dimensional pulmonary MR-angiography: 1.0 M gadobutrol vs. 0.5 M gadopentetate dimeglumine

PURPOSE: To compare contrast characteristics and image quality of 1.0 M gadobutrol with 0.5 M Gd-DTPA for time-resolved three-dimensional pulmonary magnetic resonance angiography (MRA). MATERIALS AND METHODS: Thirty-one patients and five healthy volunteers were examined with a contrast-enhanced time-resolved pulmonary MRA protocol (fast low-angle shot [FLASH] three-dimensional, TR/TE = 2.2/1.0 msec, flip angle: 25 degrees, scan time per three-dimensional data set = 5.6 seconds). Patients were randomized to receive either 0.1 mmol/kg body weight (bw) or 0.2 mmol/kg bw gadobutrol, or 0.2 mmol/kg bw Gd-DTPA. Volunteers were examined three times, twice with 0.2 mmol/kg bw gadobutrol using two different flip angles and once with 0.2 mmol/kg bw Gd-DTPA. All contrast injections were performed at a rate of 5 mL/second. Image analysis included signal-to-noise ratio (SNR) and contrast-to-noise ratio (CNR) measurements in lung arteries and veins, as well as a subjective analysis of image quality. RESULTS: In patients, significantly higher SNR and CNR were observed with Gd-DTPA compared to both doses of gadobutrol (SNR: 35-42 vs.17-25; CNR 33-39 vs. 16-23; P < or = 0.05). No relevant differences were observed between 0.1 mmol/kg bw and 0.2 mmol/kg bw gadobutrol. In volunteers, gadobutrol and Gd-DTPA achieved similar SNR and CNR. A significantly higher SNR and CNR was observed for gadobutrol-enhanced MRA with an increased flip angle of 40 degrees. Image quality was rated equal for both contrast agents. CONCLUSION: No relevant advantages of 1.0 M gadobutrol over 0.5 M Gd-DTPA were observed for time-resolved pulmonary MRA in this study. Potential explanations are T2/T2*-effects caused by the high intravascular concentration when using high injection rates.     

Contrast-enhanced three-dimensional pulmonary perfusion magnetic resonance imaging: intraindividual comparison of 1.0 M gadobutrol and 0.5 M Gd-DTPA at three dose levels

RATIONALE AND OBJECTIVES: To compare 1.0 M gadobutrol and 0.5 M Gd-DTPA for contrast-enhanced three-dimensional pulmonary perfusion magnetic resonance imaging (3D MRI). MATERIALS AND METHODS: Ten healthy volunteers (3 females; 7 males; median age, 27 years; age range, 18-31 years) were examined with contrast-enhanced dynamic 3D MRI with parallel acquisition technique (FLASH 3D; reconstruction algorithm: generalized autocalibrating partially parallel acquisitions; acceleration factor: 2; TE/TR/alpha: 0.8/1.9 milliseconds/40 degrees; FOV: 500 x 375 mm; matrix: 256 x 86; slab thickness: 180 mm; 36 partitions; voxel size: 4.4 x 2 x 5 mm; TA: 1.48 seconds). Twenty-five consecutive data sets were acquired after intravenous injection of 0.025, 0.05, and 0.1 mmol/kg body weight of gadobutrol and Gd-DTPA. Quantitative measurements of peak signal-to-noise ratios (SNR) of both lungs were performed independently by 3 readers. Bolus transit times through the lungs were assessed from signal intensity time curves. RESULTS: The peak SNR in the lungs was comparable between gadobutrol and Gd-DTPA at all dose levels (15.7 vs. 15.5 at 0.1 mmol/kg bw; 12.9 vs. 12.5 at 0.05 mmol/kg bw; 7.6 vs. 8.9 at 0.025 mmol/kg bw). A dose of 0.1 mmol/kg achieved the highest peak SNR compared with all other dose levels (P < 0.05). A higher peak SNR was observed in gravity dependent lung (P < 0.05). Despite different injection volumes, transit times of the contrast bolus did not differ between both agents. CONCLUSION: Higher concentrated gadolinium chelates offer no advantage over standard 0.5 M Gd-DTPA for contrast-enhanced 3D MRI of lung perfusion.     

Evaluation of chest motion and volumetry during the breathing cycle by dynamic MRI in healthy subjects: comparison with pulmonary function tests

RATIONALE AND OBJECTIVES: To investigate diaphragm and chest wall motion during the whole breathing cycle using magnetic resonance imaging (MRI) and a volumetric model in correlation with spirometry. MATERIALS AND METHODS: Breathing cycles of 15 healthy volunteers were examined using a trueFISP sequence (5 slices in 3 planes, 3 images per second). Time-distance curves were calculated and correlated to spirometry. A model for vital capacity (VC), continuous time-dependent vital capacity (tVC), and investigating the influence of horizontal and vertical parameters on tVC was introduced. RESULTS: Time-distance curves of the breathing cycle using MRI correlated highly significant with spirometry (P < 0.0001). VC calculated by the model was similar to VC measured in spirometry (5.00 L vs. 5.15 L). tVC correlated highly significantly with spirometry (P < 0.0001). Vertical parameters had a more profound influence on tVC change than horizontal parameters. CONCLUSIONS: Dynamic MRI is a simple noninvasive method to evaluate local chest wall motion and respiratory mechanics. It widens the repertoire of tools for lung examination with a high temporal resolution.     

Low mechanical index contrast-enhanced ultrasound better reflects high arterial perfusion of liver metastases than arterial phase computed tomography

RATIONALE AND OBJECTIVES: We investigated whether observing the arterial vascularization of liver metastases by contrast-enhanced ultrasound with low mechanical index (low-MI) imaging offers additional diagnostic information for the characterization of the liver lesions. METHODS: Twenty nine patients with untreated liver metastases of different primaries were examined. Measurements were performed using a low frame rate, low-MI pulse inversion technique after injection of 2.4 mL SonoVue. The relative maximum signal intensity of the liver lesions related to the normal liver tissue was quantified. Ultrasound findings were compared with contrast-enhanced, dual-phase computed tomography (CT) using a pattern-based classification scheme. RESULTS: Compared with contrast-enhanced CT, this modality better detects arterial perfusion. Metastases, even those usually considered hypovascularized, often showed homogeneous enhancement (66%) and higher arterial vascularization than normal liver tissue. CT did not show a comparable vascularization pattern (P < 0.001) or any similarly early signal intensity (P < 0.001). CONCLUSIONS: Contrast-enhanced CT may not be able to visualize short-lasting but large differences of the arterial perfusion of liver metastases, as does contrast-enhanced low-MI ultrasound. This offers new methods for their characterization and for monitoring of therapeutic effects.     

Quantitative 3D pulmonary MR-perfusion in patients with pulmonary arterial hypertension: correlation with invasive pressure measurements

PURPOSE: Pathological changes of the peripheral pulmonary arteries induce pulmonary arterial hypertension (PAH). Aim of this study was to quantitatively assess the effect of PAH on pulmonary perfusion by 3D-MR-perfusion techniques and to compare findings to healthy controls. Furthermore, quantitative perfusion data were correlated with invasive pressure measurements. MATERIAL AND METHODS: Five volunteers and 20 PAH patients (WHO class II or III) were examined using a 1.5T MR scanner. Measurement of pulmonary perfusion was done in an inspiratory breathhold (FLASH3D; 3.5 mm x 1.9 mm x 4mm; TA per 3D dataset 1.5s). Injection of contrast media (0.1 mmol Gd-DTPA/kg BW) and image acquisition were started simultaneously. Evaluation of 3D perfusion was done using singular value decomposition. Lung borders were outlined manually. Each lung volume was divided into three regions (anterior, middle, posterior), and the following parameters were assessed: Time-to-Peak (TTP), blood flow (PBF), blood volume (PBV), and mean transit time (MTT). In 10 patients invasive pulmonary artery pressure measurements were available and correlated to the perfusion measurements. RESULTS: In both, controls and patients, an anterior-to-posterior gradient with higher PBF and PBV posterior was observed. In the posterior lung region, a significant difference (p<0.05) was found for TTP (12s versus 16s) and MTT (4s versus 6s) between volunteers and patients. PBF and PBV were lower in patients than in volunteers (i.e. dorsal regions: 124 versus 180 ml/100 ml/min and 10 versus 12 ml/100 ml), but the difference failed to be significant. The ratio of PBF and PBV between the posterior and the middle or ventral regions showed no difference between both groups. A moderate linear correlation between mean pulmonary arterial pressure (mPAP) and PBV (r=0.51) and MTT (r=0.56) was found. CONCLUSION: The only measurable effect of PAH on pulmonary perfusion is a prolonging of the MTT. There is only a moderate linear correlation of invasive mPAP with PBV and MTT.     

Prediction of glomerular filtration rate in renal transplant recipients: cystatin C or Modification of Diet in Renal Disease equation?

BACKGROUND: To overcome disadvantages of serum creatinine two strategies have been suggested to identify patients with reduced glomerular filtration rate (GFR). On the one hand, the Modification of Diet in Renal Disease (MDRD) equation is now recommended to classify the stage of chronic kidney disease. On the other hand, cystatin C (Cys C) has been investigated in numerous studies, finding a higher sensitivity than creatinine in detecting diminished GFR. To date, no comparison of both strategies in patients after renal transplantation has been performed. METHODS: One hundred and five consecutive renal transplant recipients underwent (99m)Tc-DTPA-- clearance measurement. Simultaneously, MDRD estimates were calculated and Cys C serum levels were determined. ROC analyses were performed at different decision points from 20 to 70 mL/min/1.73 m(2). RESULTS: Although the area under the curve did not differ significantly between MDRD and Cys C within the tested GFR range, the AUC for Cys C tended to be higher when GFR exceeded 55 mL/min/1.73 m(2). A significantly higher diagnostic accuracy for Cys C compared with MDRD (p = 0.045 at 65 mL/min/1.73 m(2)) was found when investigating the subgroup of patients with well-functioning grafts (GFR>40 mL/min/1.73 m(2)). CONCLUSION: MDRD equation is equivalent to Cys C measurement in renal transplant recipients. As availability of MDRD is superior to Cys C, we recommend GFR estimation using the MDRD equation. Nevertheless, Cys C may serve as a confirmation test of high MDRD estimates in patients with well-functioning grafts because of superior accuracy in these patients.     

Quantitative CT detects progression in COPD patients with severe emphysema in a 3-month interval

Chronic obstructive pulmonary disease (COPD) is characterized by variable contributions of emphysema and airway disease on computed tomography (CT), and still little is known on their temporal evolution. We hypothesized that quantitative CT (QCT) is able to detect short-time changes in a cohort of patients with very severe COPD. Two paired in- and expiratory CT each from 70 patients with avg. GOLD stage of 3.6 (mean age = 66 ± 7.5, mean FEV1/FVC = 35.28 ± 7.75) were taken 3 months apart and analyzed by fully automatic software computing emphysema (emphysema index (EI), mean lung density (MLD)), air-trapping (ratio expiration to inspiration of mean lung attenuation (E/I MLA), relative volume change between − 856 HU and − 950 HU (RVC856–950)), and parametric response mapping (PRM) parameters for each lobe separately and the whole lung. Airway metrics measured were wall thickness (WT) and lumen area (LA) for each airway generation and the whole lung. The average of the emphysema parameters (EI, MLD) increased significantly by 1.5% (p < 0.001) for the whole lung, whereas air-trapping parameters (E/I MLA, RVC856–950) were stable. PRMEmph increased from 34.3 to 35.7% (p < 0.001), whereas PRMNormal decrased from 23.6% to 22.8% (p = 0.012). WT decreased significantly from 1.17 ± 0.18 to 1.14 ± 0.19 mm (p = 0.036) and LA increased significantly from 25.08 ± 4.49 to 25.84 ± 4.87 mm2 (p = 0.041) for the whole lung. The generation-based analysis showed heterogeneous results. QCT detects short-time progression of emphysema in severe COPD. The changes were partly different among lung lobes and airway generations, indicating that QCT is useful to address the heterogeneity of COPD progression. • QCT detects short-time progression of emphysema in severe COPD in a 3-month period. • QCT is able to quantify even slight parenchymal changes, which were not detected by spirometry. • QCT is able to address the heterogeneity of COPD, revealing inconsistent changes individual lung lobes and airway generations.     

Therapeutic strategies in male breast cancer: Clinical implications of chromosome 17 gene alterations and molecular subtypes

Male breast cancer (MBC) is a rare disease. To date, therapy is mainly based on studies and clinical experiences with breast cancer in women. Only little is known about molecular typing of MBC, particularly with regard to potential biological predictors for adjuvant therapy. In female breast cancer tumors with chromosome 17 centromere (CEP17) duplication, HER2 and/or Topoisomerase II alpha (Topo II-α) gene alterations have been suggested to be associated with poor prognosis and increased sensitivity to anthracycline-containing regimens.In a well characterized cohort of 96 primary invasive MBC, we studied CEP17, HER2 and Topo II-α alterations by fluorescence in-situ hybridization (FISH), and expression of hormone receptors (HR), HER2 and Ki67 by immunohistochemistry to define molecular subtypes. Tumor characteristics and follow-up data were available and correlated with molecular findings.HER2 amplification and Topo II-α amplification/deletion were exceptionally rare in MBC (6.3% and 3.1%, respectively). CEP17 polysomy were found in 9.4% of tumors. HER2, Topo II-α and CEP17 gene alterations were not correlated to patients outcome. 96.9% of our cases were HR positive. Triple negative tumors were found in only 3.1% of the cases. In nodal negative tumors luminal A subtypes were significantly associated with better overall survival.Our results provide evidence for a predominant male breast cancer phenotype, characterized by HR expression and a lack of HER2/Topo II-α alterations and CEP17 duplicates. Therefore, the impact of anthracycline sensitivity linked to HER2/Topo II-α alterations as found in female breast cancer has low clinical significance for this specific male breast cancer phenotype.