Browsing by Subject "Molecular Imaging"
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Item Development of a Lung Cancer Targeting Peptide for Imaging and Drug Delivery(2018-04-12) Allred, Curtis Arlin; Corbin, Ian R.; Brown, Kathlynn C.; Kohler, Jennifer J.; Minna, John D.Lung cancer kills more people in the United Stated than the next three biggest cancer killers combined. Non-small cell lung cancer (NSCLC) accounts for 85% of lung cancer patients and has an overall estimated 5-year survival of 18% in 2017. The emergence of targeted therapies (molecularly guided treatment) has improved the overall survival and quality of life of ~20% of NSCLC patients. A different molecularly guided treatment modality, targeting therapy, has shown success in other cancer types, but has not yet been successfully applied in NSCLC. Peptides are a class of molecules that have demonstrated exquisite targeting of cancer cells. HCC15.2 peptide was identified from a phage display library screen on NSCLC cells. It binds to an unknown receptor on a specific subset of NSCLC patient samples (24%) and cell lines (50%), which is not present in immortalized but non-transformed human bronchial epithelial cells (HBEC). Not only does HCC15.2 have a high affinity of ~5 nM and high specificity, it also triggers internalization and delivers its cargo into the cancer cell. HCC15.2 was optimized by altering the multimerization and amino acid content and protecting it from serum degradation. After the optimizations, HCC15.2 was shown to deliver many cargo types into cancer cells which traffic to and accumulate in lysosomes. HCC15.2 also demonstrated its ability to home to a subcutaneous xenograft in mice ~30-fold better than non-targeted dye after systemic delivery. This peptide used in molecular imaging could aid in earlier tumor detection, which correlates with better patient survival. HCC15.2 conjugated to saporin, a ribosome inactivating protein, showed an IC50 of 5.4 nM in in vitro viability assays. Delivery of saporin by HCC15.2 significantly slowed the growth of tumor xenografts. HCC15.2 is a perfect candidate for molecularly guided imaging and therapy.Item An Imaging Approach to Examine Telomere Dynamics and Regulation of Gene Expression with Aging(2020-08-01T05:00:00.000Z) Zhang, Ning; Xie, Yang; Danuser, Gaudenz; Shay, Jerry W.; Jaqaman, Khuloud; Siegwart, Daniel J.Telomeres are repetitive non-coding nucleotide sequences (TTAGGG)n capping the ends of chromosomes. Improved methods to measure the shortest (not just average) telomere lengths (TLs) are needed. Progressive telomere shortening with increasing age has been associated with shifts in gene expression through models such as the telomere position effect (TPE), which suggests reduced interference of the telomere with transcriptional activity of increasingly more distant genes. A modification of the TPE model, referred to as Telomere Position Effects over Long Distance (TPE-OLD), explains why some genes 1-10 MB from a telomere are still affected by TPE, but genes closer to the telomere are not. Therefore, demonstrating the regulatory roles of telomere length shortening on genes with accurate TL measurement will improve our understanding to the 3D genomic DNA landscape including telomeres. In this doctoral dissertation, I developed a user-friendly software for automatic electrophoresis gel quantification and contributed to developing the Telomere Shortest Length Assay (TeSLA), a technique that detects telomeres from all chromosome ends from <1 kb to 18 kb using small amounts of input DNA. Using cells with more TL information provided by TeSLA, I conducted an imaging approach to systematically examine the occurrence of TPE-OLD at the single cell level. Compared to existing methods, the pipeline allows rapid analysis of hundreds to thousands of cells, which is necessary to establish TPE-OLD as an acceptable mechanism of gene expression regulation. I examined two human genes, for which TPE-OLD has been described before, ISG15 (Interferon Stimulated Gene 15) and TERT (TElomerase Reverse Transcriptase). For both genes I found less interaction with the telomere on the same chromosome in old cells compared to young cells. Experimentally elongated telomeres in old cells rescued the level of telomere interaction for both genes. However, the dependency of the interactions on the age progression from young to old cells varied. One model for the differences between ISG15 and TERT may relate to the markedly distinct interstitial telomeric sequence arrangement in the two genes. Overall, this provides a strong rationale for the role of telomere length shortening in the regulation of gene expression.Item Molecular Imaging in Noninvasive Assessment of Diabetes Progression(2013-07-16) Lo, Su-Tang; Lenkinski, Robert; Sun, Xiankai; Öz, Orhan K.; Lingvay, Ildiko; Ahn, Jung-MoMolecular imaging is a recently emerged multidisciplinary scientific field comprised of diverse technologies. The goal of this branch of science is to understand molecular mechanism of diseases and facilitate drug development, namely the interplay of non-invasive imaging techniques with molecular biology and medicine. The goal of this dissertation is to reflect the roles of molecular imaging in biomedical applications by longitudinal and non-invasive detection of the initiation and progression of diabetes. Diabetes is a chronic disease caused by a gradual loss of pancreatic β-cell mass (BCM). The current clinical diagnosis measures the parameters of the β-cell function (BCF) post the onset of the disease, which cannot accurately reflect the BCM loss during the initiation and progression of the disease. Therefore, a non-invasive imaging technique that enables the direct assessment of BCM change would be highly desirable. In this dissertation, Positron Emission Tomography (PET) imaging was conducted to non-invasively and longitudinally monitor the change of glucagon-like peptide 1 receptor (GLP-1R), a specific biomarker of pancreatic β-cell, during disease progression. Through a two-fold screening method, a modified bicyclic GLP-1 analog, which enhances biological stability while maintaining the receptor binding affinity, was selected. An imaging probe was therefore developed based on this selected stable GLP-1 analog for noninvasive imaging assessment of BCM. In vivo evaluation was carried out to determine the BCM targeting properties of the probe followed by ex vivo PET imaging and histology. Further, a reliable and reproducible multimodality imaging technique was developed by combining PET imaging with the GLP-1R targeted probe with anatomical imaging techniques including BaSO4-enhanced CT and MRI for quantitative analysis of BCM imaging in mouse models. To test the potential of the imaging technique for longitudinal monitoring of BCM change during the diabetes initiation and progression, a serial PET/CT imaging was performed in a streptozotocin (STZ)-induced diabetic mouse model. With the successful development of the BCM imaging probe, a valid imaging technique has been established for noninvasive assessment of the progression of diabetes, which may find applications in early diagnosis of diabetes and monitoring therapeutic interventions of the disease.Item Molecular Imaging of αvβ6–Positive Tumors and Pancreatic β-Cell Mass by Radiolabeled Peptides(2011-02-01) Lin, Mai; Sun, XiankaiConventional diagnostic methodologies of lung cancer and diabetes are limited by sensitivity and specificity. In consequence, patients usually get diagnosed when the symptoms appear and the diseases are at the advanced stages. As the expressions of the avß6 integrin and glucagon-like peptide-1 receptor (GLP-1R) are highly related to aggressive tumor phenotypes and functional pancreatic ß-cells, this work has been set to develop peptide-based radiotracers that can specifically bind to avß6 or GLP-1R for noninvasively monitoring the progression of lung cancer and diabetes. By phage display, a peptide sequence that specific binds to avß6 was identified. In the evaluation of its truncated forms with similar binding affinity, a polyethylene glycol chain (PEG11) was inserted to the C-terminus and a bifunctional chelator (DOTA) was conjugated to either end of the peptides. The conjugates were labeled with 111In (t1/2: 2.8 d) under mild conditions with the highest achievable specific activity of 1.15 × 104 MBq/µmol. The in vivo evaluation was performed in a lung adenocarcinoma xenograft mouse model. Of the six conjugates, 10PD showed the best tissue contrast of the H2009 (avß6+) tumor. However, it also yielded to have the highest renal accumulation. The high kidney uptake of 10PD was found to be alleviated by conjugating DOTA at the N-terminus or reducing the peptide net charges. To evaluate GLP-1-based radiotracers for imaging pancreatic ß-cell mass (BCM), GLP-1, [D-Ala8]GLP-1, two bicyclic GLP-1 analogs (EM2196 and EM2198), and exendin-4 were synthesized and compared for their biological properties. All peptide constructs were tagged with an 6-aminohexanoic linker (Ahx) followed by DOTA conjugation at C-terminus and labeled with 64Cu (t1/2: 12.7 h). The specific activity of the labeled peptide conjugates was up to 1.0 × 106 MBq/µmol with radiochemical purity over 97%. Compared to GLP-1, [D-Ala8]GLP-1 revealed strong resistance against DPP-IV. In addition, EM2198 demonstrated high stability against NEP 24.11 presumably by the shielding effects from the two lactam bridges. All peptide conjugates were highly selective to the GLP-1R with the IC50 values in 0.1-0.4 nM. However, only 64Cu-EM2198 showed clear pancreas area on the microPET/CT studies. The signal of 64Cu-EM2198 from the pancreas was confirmed by the ex vivo imaging scans. The potential of 64Cu-EM2198 for imaging BCM was further supported by co-injecting a blocking dose of unlabeled exendin-4 and performing imaging studies in the STZ-treated diabetic mice.