Haiming Luo1, Hao Hong1, Michael R Slater2, Stephen A Graves3, Sixiang Shi4, Yunan Yang1, Robert J Nickles4, Frank Fan2, Weibo Cai5. 1. Department of Radiology, University of Wisconsin-Madison, Madison, Wisconsin. 2. Promega Corp., Madison, Wisconsin. 3. Department of Medical Physics, University of Wisconsin-Madison, Madison, Wisconsin. 4. Materials Science Program, University of Wisconsin-Madison, Madison, Wisconsin; and. 5. Department of Radiology, University of Wisconsin-Madison, Madison, Wisconsin Department of Medical Physics, University of Wisconsin-Madison, Madison, Wisconsin Materials Science Program, University of Wisconsin-Madison, Madison, Wisconsin; and University of Wisconsin Carbone Cancer Center, Madison, Wisconsin wcai@uwhealth.org.
Abstract
UNLABELLED: The hepatocyte growth factor (HGF) and its receptor, c-Met, are actively involved in tumor progression and metastasis and are closely associated with a poor prognostic outcome for cancer patients. Thus, the development of PET agents that can assess c-Met expression would be extremely useful for diagnosing cancer and subsequently monitoring response to c-Met-targeted therapies. Here, we report the characterization of recombinant human HGF (rh-HGF) as a PET tracer for detection of c-Met expression in vivo. METHODS: rh-HGF was expressed in human embryonic kidney 293 cells and purified by nickel-nitrilotriacetic acid affinity chromatography. The concentrated rh-HGF was conjugated to 2-S-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid and labeled with (64)Cu. c-Met binding evaluation by flow cytometry was performed on both U87MG and MDA-MB-231 cell lines, which have a high level and a low level, respectively, of c-Met. PET imaging and biodistribution studies were performed on nude mice bearing U87MG and MDA-MB-231 xenografted tumors. RESULTS: The rh-HGF expression yield was 150-200 μg of protein per 5 × 10(6) cells after a 48-h transfection, with purity of approximately 85%-90%. Flow cytometry examination confirmed that rh-HGF had a strong and specific capacity to bind to c-Met. After (64)Cu labeling, PET imaging revealed specific and prominent uptake of (64)Cu-NOTA-rh-HGF in c-Met-positive U87MG tumors (percentage injected dose per gram, 6.8 ± 1.8 at 9 h after injection) and significantly lower uptake in c-Met-negative MDA-MB-231 tumors (percentage injected dose per gram, 1.8 ± 0.6 at 9 h after injection). The fact that sonication-denatured rh-HGF had significantly lower uptake in U87MG tumors, along with histology analysis, confirmed the c-Met specificity of (64)Cu-NOTA-rh-HGF. CONCLUSION: This study provided initial evidence that (64)Cu-NOTA-rh-HGF visualizes c-Met expression in vivo, an application that may prove useful for c-Met-targeted cancer therapy.
UNLABELLED: The hepatocyte growth factor (HGF) and its receptor, c-Met, are actively involved in tumor progression and metastasis and are closely associated with a poor prognostic outcome for cancerpatients. Thus, the development of PET agents that can assess c-Met expression would be extremely useful for diagnosing cancer and subsequently monitoring response to c-Met-targeted therapies. Here, we report the characterization of recombinant humanHGF (rh-HGF) as a PET tracer for detection of c-Met expression in vivo. METHODS:rh-HGF was expressed in humanembryonic kidney 293 cells and purified by nickel-nitrilotriacetic acid affinity chromatography. The concentrated rh-HGF was conjugated to 2-S-(4-isothiocyanatobenzyl)-1,4,7-triazacyclononane-1,4,7-triacetic acid and labeled with (64)Cu. c-Met binding evaluation by flow cytometry was performed on both U87MG and MDA-MB-231 cell lines, which have a high level and a low level, respectively, of c-Met. PET imaging and biodistribution studies were performed on nude mice bearing U87MG and MDA-MB-231 xenografted tumors. RESULTS: The rh-HGF expression yield was 150-200 μg of protein per 5 × 10(6) cells after a 48-h transfection, with purity of approximately 85%-90%. Flow cytometry examination confirmed that rh-HGF had a strong and specific capacity to bind to c-Met. After (64)Cu labeling, PET imaging revealed specific and prominent uptake of (64)Cu-NOTA-rh-HGF in c-Met-positive U87MGtumors (percentage injected dose per gram, 6.8 ± 1.8 at 9 h after injection) and significantly lower uptake in c-Met-negative MDA-MB-231tumors (percentage injected dose per gram, 1.8 ± 0.6 at 9 h after injection). The fact that sonication-denatured rh-HGF had significantly lower uptake in U87MGtumors, along with histology analysis, confirmed the c-Met specificity of (64)Cu-NOTA-rh-HGF. CONCLUSION: This study provided initial evidence that (64)Cu-NOTA-rh-HGF visualizes c-Met expression in vivo, an application that may prove useful for c-Met-targeted cancer therapy.
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