In Vivo Molecular Targeted Imaging of Cancer

Hepatocellular carcinoma (HCC) presents a global healthcare problem. It is the second most lethal cancer worldwide, causing 745,000 deaths annually. HCC accounts for 70% to 90% of primary liver cancer cases with rising incidence in developed countries. Newly diagnosed cases in the U.S. are expected to increase by 10% in three years. Symptoms of HCC typically do not appear until advanced stage, leaving surgical resection the primary therapy. However, HCC patients suffer from dire prognosis of less than 5% five-year survival rate and >50% incidence of tumor recurrence, due to poor contrast of HCC against surrounding liver tissue limiting resection accuracy. Using a molecular imaging system that targets differentially expressed tumor specific surface biomarkers may help detect HCC neoplasm missed by surgeons and preserve viable liver tissue to reduce recurrence and improve patient recovery. This dissertation presents the HCC targeting and imaging methods developed to specifically identify HCC neoplasm with high contrast, fast kinetics and deep penetration. Two cancer cell surface biomarkers, epidermal growth factor receptor (EGFR) and glypican-3 (GPC3), are important in the development of HCC. To create a molecular imaging strategy for HCC detection, short peptide sequences specifically binding to these biomarkers have been selected and validated. They demonstrated high target affinities (kd < 75 nM) and fast cellular binding kinetics (<10 min). After conjugating with near-infrared organic dye, these molecular targeting probes were able to home to the HCC tumor xenograft in vivo after intravenous administration. Ex vivo and in vivo optical imaging was conducted with fluorescent laparoscopy, whole body fluorescent imaging, and hand held dual-axis confocal microscopy. In vivo cell surface binding of peptide probe to HCC xenograft in mice was observed at subcellular resolution in both horizontal (1000×1000 µm2) and vertical (1000×430 µm2) planes. Tumor margins were automatically detected with computerized segmentation algorithm. High target-to-background ratios (2.99 and 6.2 respectively) were achieved at tumor sites after 6 and 2 hours respectively, and targeting probes were cleared from the animal system within 24 hours. In addition, targeted in vivo photoacoustic tomography (PAT) imaging visualized probe penetration inside the tumor 1.8 cm beneath intact skin. Plasmonic nanoparticles absorb light more efficiently than organic dyes. By coating GPC3 targeting peptide onto gold nanoshell (GNS) surface, in vivo photoacoustic imaging contrast was improved from 2.25 to 4.45 and imaging depth reached 2.1 cm. Peak probe uptake in vivo occurred at 2 hours and clearance took place within 12 hours, which are desirable pharmacokinetics for clinical settings of intraoperative imaging guidance. Specific binding, biodistribution and toxicity were investigated in cultured cells, ex vivo tissues (human and mouse) as well as in mouse models. The GPC3 targeting probe was able to distinguish HCC from non-HCC human patient biopsies (n=41) at 93% sensitivity and 88% specificity, with area under the receiver operator characteristic curve (AUC) value reaching 0.98. These studies showed that affinity peptide based molecular imaging is an enabling technology which will allow clinicians to perform functional imaging during surgery to identify resection margin with high contrast, sensitivity and speed.PHDBiomedical EngineeringUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttps://deepblue.lib.umich.edu/bitstream/2027.42/138522/1/zhouquan_1.pd