In Vivo PET Detection of Lung Micrometastasis in Mice by Targeting Endothelial VCAM-1 Using a Dual-Contrast PET/MRI Probe

Current clinical diagnostic imaging methods for lung metastases are sensitive only to large tumours (1–2 mm cross-sectional diameter), and early detection can dramatically improve treatment. We have previously demonstrated that an antibody-targeted MRI contrast agent based on microparticles of iron oxide (MPIO; 1 μm diameter) enables the imaging of endothelial vascular cell adhesion molecule-1 (VCAM-1). Using a mouse model of lung metastasis, upregulation of endothelial VCAM-1 expression was demonstrated in micrometastasis-associated vessels but not in normal lung tissue, and binding of VCAM-MPIO to these vessels was evident histologically. Owing to the lack of proton MRI signals in the lungs, we modified the VCAM-MPIO to include zirconium-89 (89Zr, t1/2 = 78.4 h) in order to allow the in vivo detection of lung metastases by positron emission tomography (PET). Using this new agent (89Zr-DFO-VCAM-MPIO), it was possible to detect the presence of micrometastases within the lung in vivo from ca. 140 μm in diameter. Histological analysis combined with autoradiography confirmed the specific binding of the agent to the VCAM-1 expressing vasculature at the sites of pulmonary micrometastases. By retaining the original VCAM-MPIO as the basis for this new molecular contrast agent, we have created a dual-modality (PET/MRI) agent for the concurrent detection of lung and brain micrometastases

Improving delineation of true tumour volume with multimodal MRI in a rat model of brain metastasis

Purpose: Brain metastases are almost universally lethal with short median survival times. Despite this they are often potentially curable, with therapy failing only because of local relapse. One key reason relapse occurs is because treatment planning did not delineate metastasis margins sufficiently accurately, allowing residual tumour to regrow. The aim of this study was to determine the extent to which multimodal MRI, with a simple and automated analysis pipeline, could improve upon current clinical practice of single modality independent observer tumour delineation. Methods and Materials: We used a single rat model of brain metastasis (ENU1564 breast carcinoma cells in BD-IX rats), with and without radiotherapy. Multimodal MRI data were acquired using sequences either in current clinical use or in clinical trial, and included post gadolinium T1-weighted images and maps of blood flow, blood volume, T1 and T2 relaxation times, and apparent diffusion coefficient. Results: In all cases, independent observers underestimated the true size of metastases from single modality gadolinium-enhanced MRI (85±36µL vs. 131±40µL histological measurement), whilst multimodal MRI more accurately delineated tumour volume (132±41µL). Multimodal MRI offered increased sensitivity compared to independent observer for detecting metastasis (0.82 vs. 0.61, respectively), with only a slight decrease in specificity (0.86 vs. 0.98). Blood flow maps conferred the greatest improvements in margin detection for late-stage metastases after radiotherapy. Gadolinium-enhanced T1 weighted images conferred the greatest increase in accuracy of detection for smaller metastases. Conclusions: These findings suggest that multimodal MRI of brain metastases could significantly improve the visualisation of brain metastasis margins, beyond current clinical practice, with the potential to decrease relapse rates and increase patient survival. This finding now needs validation in additional tumour models or clinical cohorts.</p

Optimization of molecularly targeted MRI in the brain: empirical comparison of sequences and particles

Background: Molecular MRI is an evolving field of research with strong translational potential. Selection of the appropriate MRI sequence, field strength and contrast agent depend largely on the application. The primary aims of the current study were to: 1) assess the sensitivity of different MRI sequences for detection of iron oxide particles in mouse brain; 2) determine the effect of magnetic field strength on detection of iron oxide particles in vivo; and 3) compare the sensitivity of targeted microparticles of iron oxide (MPIO) or ultra-small superparamagnetic iron oxide (USPIO) for detection of vascular cell adhesion molecule-1 (VCAM-1) in vivo. Methods: Mice were injected intrastriatally with interleukin 1β to induce VCAM-1 expression on the cerebral vasculature. Subsequently, animals were injected intravenously with either VCAM-MPIO or VCAM-USPIO and imaged 1 or 13 hours post-injection, respectively. MRI was performed at 4.7, 7.0, or 9.4 T, using three different T2*-weighted sequences: single gradient echo 3D (GE3D), multi-gradient echo 3D (MGE3D) and balanced steady-state free precession 3D (bSSFP3D). Results: MGE3D yielded the highest signal-to-noise ratio (SNR) and contrast-to-noise ratio (CNR) for the detection of iron oxide particles. All sequences showed a significant increase in SNR and CNR from 4.7 to 7.0 T, but no further improvement at 9.4 T. However, whilst targeted MPIO enabled sensitive detection of VCAM-1 expression on the cerebral vasculature, the long half-life (16.5 h vs 1.2 min) and lower relaxivity per particle (1.29×10–14 vs 1.18×10–9 Hz L/particle) of USPIO vs. MPIO rendered them impractical for molecular MRI. Conclusion: These findings demonstrate clear advantages of MPIO compared to USPIO for molecularly-targeted MRI, and indicate that the MGE3D sequence is optimal for MPIO detection. Moreover, higher field strengths (7.0/9.4 T) showed enhanced sensitivity over lower field strengths (4.7 T). With the development of biodegradable MPIO, these agents hold promise for clinical translation

Optimization of molecularly targeted MRI in the brain: empirical comparison of sequences and particles

Background: Molecular MRI is an evolving field of research with strong translational potential. Selection of the appropriate MRI sequence, field strength and contrast agent depend largely on the application. The primary aims of the current study were to: 1) assess the sensitivity of different MRI sequences for detection of iron oxide particles in mouse brain; 2) determine the effect of magnetic field strength on detection of iron oxide particles in vivo; and 3) compare the sensitivity of targeted microparticles of iron oxide (MPIO) or ultra-small superparamagnetic iron oxide (USPIO) for detection of vascular cell adhesion molecule-1 (VCAM-1) in vivo. Methods: Mice were injected intrastriatally with interleukin 1β to induce VCAM-1 expression on the cerebral vasculature. Subsequently, animals were injected intravenously with either VCAM-MPIO or VCAM-USPIO and imaged 1 or 13 hours post-injection, respectively. MRI was performed at 4.7, 7.0, or 9.4 T, using three different T2*-weighted sequences: single gradient echo 3D (GE3D), multi-gradient echo 3D (MGE3D) and balanced steady-state free precession 3D (bSSFP3D). Results: MGE3D yielded the highest signal-to-noise ratio (SNR) and contrast-to-noise ratio (CNR) for the detection of iron oxide particles. All sequences showed a significant increase in SNR and CNR from 4.7 to 7.0 T, but no further improvement at 9.4 T. However, whilst targeted MPIO enabled sensitive detection of VCAM-1 expression on the cerebral vasculature, the long half-life (16.5 h vs 1.2 min) and lower relaxivity per particle (1.29×10–14 vs 1.18×10–9 Hz L/particle) of USPIO vs. MPIO rendered them impractical for molecular MRI. Conclusion: These findings demonstrate clear advantages of MPIO compared to USPIO for molecularly-targeted MRI, and indicate that the MGE3D sequence is optimal for MPIO detection. Moreover, higher field strengths (7.0/9.4 T) showed enhanced sensitivity over lower field strengths (4.7 T). With the development of biodegradable MPIO, these agents hold promise for clinical translation

Biomimetic screening of class-B G protein-coupled receptors.

The 41-amino acid peptide corticotropin releasing factor (CRF) is a major modulator of the mammalian stress response. Upon stressful stimuli, it binds to the corticotropin releasing factor receptor 1 (CRF(1)R), a typical member of the class-B G-protein-coupled receptors (GPCRs) and a prime target in the treatment of mood disorders. To chemically probe the molecular interaction of CRF with the transmembrane domain of its cognate receptor, we developed a high-throughput conjugation approach that mimics the natural activation mechanism of class-B GPCRs. An acetylene-tagged peptide library was synthesized and conjugated to an azide-modified high-affinity carrier peptide derived from the CRF C-terminus using copper-catalyzed dipolar cycloaddition. The resulting conjugates reconstituted potent agonists and were tested in situ for activation of the CRF(1) receptor in a cell-based assay. By use of this approach we (i) defined the minimal sequence motif that is required for full receptor activation, (ii) identified the critical functional groups and structure-activity relationships, (iii) developed an optimized, highly modified peptide probe with high potency (EC(50) = 4 nM) that is specific for the activation domain of the receptor, and (iv) probed the behavioral role of CRF receptors in living mice. The membrane recruitment by a high-affinity carrier enhanced the potency of the tethered peptides by &gt;4 orders of magnitude and thus allowed the testing of very weak initial fragments that otherwise would have been inactive on their own. As no chromatography purification of the test peptides was necessary, a substantial increase in screening throughput was achieved. Importantly, the peptide conjugates can be used to probe the endogenous receptor in its native environment in vivo