In vivo Magnetic Resonance Imaging of Tumor Protease Activity (original) (raw)
Related papers
2011
The papain family of cysteine cathepsins are actively involved in multiple stages of tumorigenesis. Because elevated cathepsin activity can be found in many types of human cancers, they are promising biomarkers that can be used to target radiological contrast agents for tumor detection. However, currently there are no radiological imaging agents available for these important molecular targets. We report here the development of positron emission tomography (PET) radionuclidelabeled probes that target the cysteine cathepsins by formation of an enzyme activity-dependent bond with the active site cysteine. These probes contain an acyloxymethyl ketone (AOMK) functional group that irreversibly labels the active site cysteine of papain family proteases attached to a 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) tag for labeling with 64 Cu for PET imaging studies. We performed biodistribution and microPET imaging studies in nude mice bearing subcutaneous tumors expressing various levels of cysteine cathepsin activity and found that the extent of probe uptake by tumors correlated with overall protease activity as measured by biochemical methods. Furthermore, probe signals could be reduced by pre-treatment with a general cathepsin inhibitor. We also found that inclusion of a Cy5 tag on the probe increased tumor uptake relative to probes lacking this fluorogenic dye. Overall, these results demonstrate that small molecule activity-based probes carrying radio-tracers can be used to image protease activity in living subjects.
Proteases in brain tumour progression
Acta Neurochirurgica, 2003
Local invasion of tumour cells is characteristic of brain tumour progression. It is associated with increased motility and a potential to hydrolyse macromolecular components of the extracellular matrix. The peptidases that have been most investigated, and are induced during this process, are reviewed: the plasminogen activators (PAs), matrix metalloproteinases (MMPs) and lysosomal cysteine peptidases called cathepsins (Cats). Increased levels of urokinase-type PA (uPA) are observed mainly at the invasive margins of a tumour, whereas the data on the expression of tissue-type PA (tPA) are still controversial. It has been shown that the endogenous inhibitor of PAs, PAI-1, is localised in both tumour and tumour-associated endothelial cells. Among MMPs, the expression of the gelatinases, MMP2 and MMP9, strongly correlates with glioma progression. Membrane bound MT-MMPs, in particular MT1-and MT2-MMP, seem to play a major role in activating MMP-2. Several members of the ADAMTS family have also been detected in brain tumours, the most relevant being ADAMTS4, due to its cleavage of CNS specific proteins. Lysosomal cathepsin B is highly expressed in malignant glial cells and in endothelial cells of vascularised glioblastomas and is a predictor of a shorter survival. In addition to invasion, cathepsin L may play a role in decreased susceptibility of anaplastic glioma cells to apoptosis. Finally, cathepsin B was proposed as a marker for malignancy in the more aggressive type of meningiomas. Each of these peptidases may act alone, or in concert with the others, to support malignant behaviour of brain tumour cells; the development of new inhibitors of invasion, therefore, should contribute to the control of local spread of a tumour.
Detection of cathepsin S cysteine protease in human brain tumour microdialysates in vivo
British Journal of Neurosurgery, 2007
Microdialysis enables the chemistry of extracellular fluid in body tissues to be measured. Extracellular proteases such as the cysteine protease, cathepsin S (CatS), are thought to facilitate astrocytoma invasion. Microdialysates obtained from human brain tumours in vivo were subjected to cathepsin S activity and ELISA assays. Cathepsin S ELISA expression was detected in five out of 10 tumour microdialysates, while activity was detected in five out of 11 tumour microdialysates. Cathepsin S expression was also detected in microdialysate from the normal brain control although no activity was found in the same sample. While some refinements to the technique are necessary, the authors demonstrate the feasibility and safety of microdialysis in human astrocytomas in vivo. Characterisation of the extracellular environment of brain tumours in vivo using microdialysis may be a useful tool to identify the protease profile of brain tumours.
Molecular Imaging of Proteases in Cancer
Cancer Growth and Metastasis, 2009
Proteases play important roles during tumor angiogenesis, invasion, and metastasis. Various molecular imaging techniques have been employed for protease imaging: optical (both fluorescence and bioluminescence), magnetic resonance imaging (MRI), single-photon emission computed tomography (SPECT), and positron emission tomography (PET). In this review, we will summarize the current status of imaging proteases in cancer with these techniques. Optical imaging of proteases, in particular with fluorescence, is the most intensively validated and many of the imaging probes are already commercially available. It is generally agreed that the use of activatable probes is the most accurate and appropriate means for measuring protease activity. Molecular imaging of proteases with other techniques (i.e. MRI, SPECT, and PET) has not been well-documented in the literature which certainly deserves much future effort. Optical imaging and molecular MRI of protease activity has very limited potential f...
Clinical and Experimental Metastasis, 1996
Recent studies suggest that cysteine proteinase cathepsin L is involved in the process of tumor invasion and metastasis. We examined cathepsin L activity in brain tumor tissue samples by an enzymatic assay, and cathepsin L protein content by enzyme-linked immunoadsorbent assays and Western blotting to determine whether increased levels of cathepsin L correlate with the progression of human gliomas. Native and acid-activatable cathepsin L activities were highest in glioblastomas followed by anaplastic astrocytomas and were lowest in low-grade gliomas and normal brain tissues. Significantly higher amounts of an Mr 29 000 cathepsin L were present in glioblastomas and anaplastic astrocytomas than in normal brain tissues and low-grade glioma tissue extracts. Using specific antibodies to cathepsin L, we also studied its cellular distribution by immunohistochemical procedures. Higher diffuse cathepsin L immunoreactivity was found in glioblastomas than in low-grade gliomas and normal brain tissue samples. Finally, the addition of cathepsin L antibody inhibits the invasion of glioblastoma cell lines through Matrigel invasion assay. These results suggest the expression of cathepsin L is dramatically upregulated in malignant gliomas and correlates with the malignant progression of human gliomas in vivo.
Expression Analysis of All Protease Genes Reveals Cathepsin K to Be Overexpressed in Glioblastoma
Background: Cancer genome and transcriptome analyses advanced our understanding of cancer biology. We performed transcriptome analysis of all known genes of peptidases also called proteases and their endogenous inhibitors in glioblastoma multiforme (GBM), which is one of the most aggressive and deadly types of brain cancers, where unbalanced proteolysis is associated with tumor progression.
Improved Cathepsin Probes for Sensitive Molecular Imaging
Molecules, 2022
Cysteine cathepsin proteases are found under normal conditions in the lysosomal compartments of cells, where they play pivotal roles in a variety of cellular processes such as protein and lipid metabolism, autophagy, antigen presentation, and cell growth and proliferation. As a consequence, aberrant localization and activity contribute to several pathologic conditions such as a variety of malignancies, cardiovascular diseases, osteoporosis, and other diseases. Hence, there is a resurgence of interest to expand the toolkit to monitor intracellular cathepsin activity and better ascertain their functions under these circumstances. Previous fluorescent activity-based probes (ABPs) that target cathepsins B, L, and S enabled detection of their activity in intact cells as well as non-invasive detection in animal disease models. However, their binding potency is suboptimal compared to the cathepsin inhibitor on which they were based, as the P1 positive charge was capped by a reporter tag. H...
Journal of Molecular Biomarkers & Diagnosis, 2017
Background: Substantial miss-rate of gastric cancer by conventianl endoscope was reported, indicating the importance of novel biomarkers for early detection of precancerous and cancerous lesions. Objective: To identify potential biomarker molecules for early gastric cancer detection and evaluation of their application for molecular in vivo imaging. Method: Besides the CEA-TAG+/-mouse model, 3 human and 2 murine gastric cancer cell lines were applied in this study. Cells lines were cultured at standard conditions. To determine the expression pattern of the targets, total RNA was extracted from cultured cells; cDNA was synthesised and qPCR was performed using spcefic primers. The murine stomach was removed and washed with PBS. The stomach was macroscopically categorized into tumor region and normal tissues. Histopathology and imunohistochemitry was performed in one half of the tumor tissues using specific antibodies. In the other half of the tumour tissue, RNA was extracted, reverse transcribed into cDNA and qPCR was performed. Histopathology, immhunohistochemistry and qPCR was performed for tumor-adjescent normal tissues to compare the difference in expression pattern of the target. Finally, a cathepsin activatable probe was injected into mice intravenously. Ex vivo fluorescent imaging was performed after 24 hours of intravenous injection of a cathepsin activatable probe. Gene expression was quantified by standard method with normalizing CT values to the housekeeping gene (Cyclophilin A). Results: Significantly elevated expression of cathepsins B, H and MMP2 in human and murine gastric cancer cell lines was detected. In addition, increased expression of cathepsins and MMPs was observed in murine primary gastric tumour specimens compared to the corresponding adjacent normal gastric mucosa (p<0.05, tumor versus normal mucosa). Accordingly, the NIRF probe was specifically activated in stomach tumor of CEA-TAG+/-mice and allowed ex vivo detection of the stomach tumor by Odyssey planar near-infrared scanner. Furthermore, immunohistochemical stainings with antibodies specific for cathepsins and MMPs indicated a stronger signal towards the tumor tissues compared to the adjacent normal mucosa. Conclusion: Cathepsins and MMPs are potential biomarkers for detection of high grade dysplasia and early gastric cancer in mouse models, revealing the potential applicability of the biomarkers in tumor imaging and therapeutic monitoring.
Molecular Cancer Therapeutics, 2008
Human cathepsin L along with cathepsin S, K, and V are collectively known as cathepsin L-like proteases due to their high homology. The overexpression and aberrant activity of each of these proteases has been implicated in tumorigenesis. These proteases contain propeptide domains that can potently inhibit both their cognate protease and other proteases within the cathepsin L-like subfamily. In this investigation, we have produced the cathepsin S propeptide recombinantly and have shown that it is a potent inhibitor of the peptidolytic, elastinolytic, and gelatinolytic activities of the cathepsin L-like proteases. In addition, we show that this peptide is capable of significantly attenuating tumor cell invasion in a panel of human cancer cell lines. Furthermore, fusion of an IgG Fc-domain to the COOH terminus of the propeptide resulted in a chimeric protein with significantly enhanced ability to block tumor cell invasion. This Fc fusion protein exhibited enhanced stability in cell-based assays in comparison with the unmodified propeptide species. This approach for the combined inhibition of the cathepsin L-like proteases may prove useful for the further study in cancer and other conditions where their aberrant activity has been implicated. Furthermore, this strategy for simultaneous inhibition of multiple cysteine cathepsins may represent the basis for novel therapeutics to attenuate tumorigenesis.