Imaging with radiolabelled anti-membrane type 1 matrix metalloproteinase (MT1-MMP) antibody: potentials for characterizing atherosclerotic plaques.

PURPOSE: Membrane type 1 matrix metalloproteinase (MT1-MMP) activates pro-MMP-2 and pro-MMP-13 to their active forms and plays important roles in the destabilization of atherosclerotic plaques. This study sought to determine the usefulness of (99m)Tc-labelled monoclonal antibody (mAb), recognizing MT1-MMP, for imaging atherosclerosis in a rabbit model (WHHLMI rabbits). METHODS: Anti-MT1-MMP monoclonal IgG(3) and negative control IgG(3) were radiolabelled with (99m)Tc after derivatization with 6-hydrazinonicotinic acid (HYNIC) to yield (99m)Tc-MT1-MMP mAb and (99m)Tc-IgG(3), respectively. WHHLMI and control rabbits were injected with these radio-probes. The aorta was removed and radioactivity was measured at 24 h after the injection. Autoradiography and histological studies were performed. RESULTS: (99m)Tc-MT1-MMP mAb accumulation in WHHLMI rabbit aortas was 5.4-fold higher than that of control rabbits. Regional (99m)Tc-MT1-MMP mAb accumulation was positively correlated with MT1-MMP expression (r = 0.59, p < 0.0001), while (99m)Tc-IgG(3) accumulation was independent of MT1-MMP expression (r = 0.03, p = NS). The highest (99m)Tc-MT1-MMP mAb accumulation was found in atheromatous lesions (4.8 ± 1.9, %ID×BW/mm(2) × 10(2)), followed in decreasing order by fibroatheromatous (1.8 ± 1.3), collagen-rich (1.6 ± 1.0) and neointimal lesions (1.5 ± 1.5). In contrast, (99m)Tc-IgG(3) accumulation was almost independent of the histological grade of lesions. CONCLUSION: Higher (99m)Tc-MT1-MMP mAb accumulation in grade IV atheroma was shown in comparison with neointimal lesions or other more stable lesions. Nuclear imaging with (99m)Tc-MT1-MMP mAb, in combination with CT and MRI, could provide new diagnostic imaging capabilities for detecting vulnerable plaques, although further investigations to improve target to blood ratios are strongly required.

Radioimmunodetection of membrane type-1 matrix metalloproteinase relevant to tumor malignancy with a pre-targeting method.

Since membrane type-1 matrix metalloproteinase (MT1-MMP) is exclusively expressed in tumors and is closely associated with metastasis and invasion, MT1-MMP is a potential target of radiotracers for the evaluation of tumor malignancy. In this study, we planned to visualize MT1-MMP in vivo by a two-step pre-targeting strategy using a streptavidin (SAv)-biotin system combined with anti-MT1-MMP monoclonal immunoglobulin (IgG) (anti-MT1-MMP monoclonal antibody (mAb)). Streptavidinylated anti-MT1-MMP mAb was synthesized by reacting biotinylated anti-MT1-MMP mAb with SAv. In the pre-targeting study, FM3A mouse breast carcinoma-implanted mice were injected with anti-MT1-MMP mAb-SAv, followed 72 h later with radioiodinated biotin, (3-[123/125I]iodobenzoyl)norbiotinamide (123/125I-IBB). Biodistribution and imaging (single photon emission computed tomography (SPECT)/CT) data were collected at several time points in the 24 h period following introduction of the tracer. The comparison groups were injected with 125I-IBB alone or with 125I-IBB pre-targeted with negative control IgG-SAv. In the pre-targeting study for MT1-MMP, within 1 h of tracer injection, rapid tumor uptake and abrupt clearance from the blood of radioactivity (2.22, 0.87% injected dose/g at 1 h) were observed. The tumor to blood (T/B) radioactivity ratios were significantly higher than those from mice dosed with the pre-targeting negative control (p<0.0001). 125I-IBB alone did not accumulate in tumors. SPECT/CT image analysis of FM3A bearing mice showed high-contrast tumor images after 3 h with minimal blood-pool activity. The present study that uses a pre-targeting method showed high T/B radioactivity ratios and clear tumor images of MT1-MMP. This imaging method may be useful for the clinical diagnosis of malignant tumors.

Targeting of lectinlike oxidized low-density lipoprotein receptor 1 (LOX-1) with 99mTc-labeled anti-LOX-1 antibody: potential agent for imaging of vulnerable plaque.

UNLABELLED: Lectinlike oxidized low-density lipoprotein (LDL) receptor 1 (LOX-1), a cell surface receptor for oxidized LDL, has been implicated in vascular cell dysfunction related to plaque instability, which could be a potential target for an atherosclerosis imaging tracer. In this study, we designed and prepared (99m)Tc-labeled anti-LOX-1 monoclonal IgG and investigated its usefulness as an atherosclerosis imaging agent. METHODS: Anti-LOX-1 monoclonal IgG and control mouse IgG2a were labeled with (99m)Tc after derivatization with 6-hydrazinonicotinic acid to yield (99m)Tc-LOX-1-mAb and (99m)Tc-IgG2a, respectively. Myocardial infarction-prone Watanabe heritable hyperlipidemic (WHHLMI) rabbits (atherosclerosis model) and control rabbits were injected intravenously with these probes, and in vivo planar imaging was performed. At 24 h after the injection, the aortas were removed, and radioactivity was measured. Autoradiography and histologic studies were performed with serial aortic sections. RESULTS: The level of (99m)Tc-LOX-1-mAb accumulation was 2.0-fold higher than the level of (99m)Tc-IgG2a accumulation in WHHLMI rabbit aortas, and the level of (99m)Tc-LOX-1-mAb accumulation in WHHLMI rabbit aortas was 10.0-fold higher than the level of (99m)Tc-LOX-1-mAb accumulation in control rabbit aortas. In vivo imaging clearly visualized the atherosclerotic aortas of WHHLMI rabbits. Autoradiography and histologic studies revealed that regional (99m)Tc-IgG2a accumulation was independent of the histologic grade of the lesions; however, regional (99m)Tc-LOX-1-mAb accumulation was significantly correlated with LOX-1 expression density and the vulnerability index. The highest level of (99m)Tc-LOX-1-mAb accumulation, expressed as {radioactivity in region of interest (Bq/mm(2))/[injected radioactivity (Bq)/animal body weight (g)]} x 10(2), was found in atheromatous lesions (3.8 +/- 1.1 [mean +/- SD]), followed in decreasing order by fibroatheromatous lesions (2.0 +/- 1.0), collagen-rich lesions (1.6 +/- 0.8), and neointimal lesions (1.4 +/- 0.7). CONCLUSION: The level of (99m)Tc-LOX-1-mAb accumulation in grade IV atheroma was higher than that in neointimal lesions or other, more stable lesions. Nuclear imaging of LOX-1 expression with (99m)Tc-LOX-1-mAb may be a useful means for predicting atheroma at high risk for rupture.

PET O-15 cerebral blood flow and metabolism after acute stroke in spontaneously hypertensive rats.

Hypertension is a major stroke risk factor and is correlated with worse outcome after stroke. Thus, the effects of hypertension on cerebral hemodynamics and metabolism within an hour after stroke must be evaluated in detail. Cerebral blood flow (CBF), oxygen extraction fraction (OEF), cerebral metabolic rate for oxygen (CMRO2) and cerebral metabolic rate for glucose (CMRglc) were measured 1 h after the occlusion of the right middle cerebral artery (MCA) in male spontaneously hypertensive rats (SHR) and male normotensive Wistar Kyoto rats (WKY). Physiological responses were determined by positron emission tomography (PET) using 15O-H2O and radiolabeled 15O-O2 blood (methodology previously developed in this laboratory) and by autoradiography (ARG) using 18F-FDG. The right hemisphere of SHR showed lower CBF values than the left hemisphere after stroke (right: 0.17+/-0.07 mL/min/g; left: 0.29+/-0.08 mL/min/g), CMRO2 (right: 2.55+/-0.80 mL/min/100 g; left: 4.11+/- 0.84 mL/min/100 g) and CMRglc (right: 52.4+/-16.2 mg/min/100 g; left: 65.6+/-10.2 mg/min/100 g). WKY rats exhibited significant decreases only in CBF and CMRO2. These results suggest greater underlying physiologic disturbances in SHR. Also, the occlusion significantly reduced CBF in both hemispheres of SHR compared with WKY, suggesting a disturbance of the autoregulatory mechanism in SHR. In summary, our results indicate that hypertension intensifies metabolic disturbances after the onset of stroke, at least in the first hour. Therefore, we suggest that hypertension not only increases the incidence of stroke but also exacerbates stroke-mediated damage.

Development of a radiolabeled probe for detecting membrane type-1 matrix metalloproteinase on malignant tumors.

Membrane type-1 matrix metalloproteinase (MT1-MMP) expressed on the tumor cell surface activates pro-MMP-2 and pro-MMP-13 to exacerbate the malignancy, suggesting its suitability as a target molecule for diagnosis by in vivo molecular imaging. Thus, we prepared radiolabeled anti-MT1-MMP monoclonal antibody (mAb) as a novel radiolabeled probe for detecting MT1-MMP in vivo and evaluated its usefulness in breast tumor-bearing rodents. (99m)Tc-anti-MT1-MMP mAb was prepared using HYNIC as a bifunctional chelating agent and immunoreactivity was evaluated by flow cytometry. MT1-MMP expression in breast carcinoma cells (rat: Walker-256 and MRMT-1, mouse: FM3A) was measured by Western blotting. In vivo biodistribution was examined for 48 h using tumor-implanted rodents followed by estimation of radiation absorbed by a standard quantitation platform Organ Level Internal Dose Assessment (OLINDA). (99m)Tc-anti-MT1-MMP mAb was obtained with 84% immunoreactivity to MT1-MMP and more than 92% radiochemical purity. MT1-MMP was highly expressed in all malignant cells. Tumor radioactivity increased with time after administration and reached 3 to 5 times higher values at 24 h post-injection than those at 1 h. Other organs, including the stomach, showed decreasing values over time. Tumor to blood ratios increased with time and reached more than 1.3 at 48 h. The effective dose was <5.0 muSv/MBq. The results suggest that (99m)Tc-anti-MT1-MMP mAb is a promising probe for future diagnosis of breast tumors by in vivo nuclear medical imaging.