Browsing by Subject "Lung Neoplasms"
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Item Alpha(V)Beta(3)-Targeted Nanoprobes for In Vivo Imaging of Tumor Angiogenesis(2010-11-02T18:20:26Z) Kessinger, Chase William; Gao, JinmingLung cancer is the leading cause of cancer-related deaths in the US and abroad (WHO, 2010). Early detection of the disease has increased patients’ five-year survival rates from 4% to over 50% (NCI SEER, 2010). Angiogenesis plays a critical role in the carcinogenesis and cancer metastasis of solid tumors. Integrin αvβ3 is a well-established overexpressed biomarker of angiogenesis and has recently been exploited in the clinical stratification of different types of cancer. Although magnetic resonance imaging (MRI) is not a first-line clinical imaging modality for the detection and diagnosis of lung cancer, recent advances in theranostic polymeric nanoplatforms and the development of ultrasensitive contrast agents, such as superparamagnetic iron oxide (SPIO) nanoparticles, has greatly broadened the application of MRI in cancer detection and molecular imaging. The objective of this work is to develop superparamagnetic polymeric micelle (SPPM) nanoprobes that can noninvasively image tumor angiogenesis in vivo using conventional T2/T2*-weighted and off-resonance saturation (ORS) MRI methods. Therefore, we hypothesized that the inclusion of an αvβ3-specific ligand, cRGD (cyclic Arg-Gly-Asp) peptide and encapsulation of a cluster of SPIO in the SPPM nanoprobe formulation would allow the specific imaging of tumor angiogenesis. SPPM nanoprobes are small (50-70 nm), and contain a cluster of SPIO in the core while maintaining the micelle core-shell architecture. In vitro examination of αvβ3-targeted cRGD-SPPM demonstrate an increase cellular uptake in αvβ3 overexpressing cells over control SPPM formulations. Upon translation to in vivo subcutaneous lung tumor models in mice, cRGD-SPPM is able to noninvasively image and quantitate tumor angiogenesis and demonstrate in vivo colocalization with αvβ3 integrin. In parallel with SPPM characterization, the ORS imaging method was validated in vitro and was successfully applied in vivo for imaging tumor angiogenesis and demonstrated an increased sensitivity and specificity for SPPM over conventional T2*-weighted MR imaging. Application of high temporal resolution (HTR) - MRI, combined with the ultrasensitivity of the SPPM nanoprobe, allows for the kinetic analysis of cRGD-SPPM targeting to angiogenic regions in vivo. Finally, SPPM shows the ability to detect small lung cancer nodules (<700 μm - 3 mm) in tail-vein induced orthotopic lung cancer models using convention T2-weighted and ORS MRI. The results presented herein, provide the characterization and proof-of-principle experiments that point towards the diagnostic potential of SPPM nanoprobes for the early detection of lung cancer.Item Identification and Characterization of Non Small Cell Lung Cancer Stem Cells(2011-02-01T19:34:43Z) Sullivan, James Patrick; Minna, John D.The discovery of rare tumor cells with stem cell features first in myeloproliferative disease and later in solid tumors has emerged as an important area in cancer research. Through these studies the cancer stem cell model has emerged, which postulates that many tumors are initiated and progressed by a population of self-renewing malignant stem cells, referred to as cancer stem cells. This new tumor growth paradigm suggests that tumor metastasis and recurrence may be driven by a residual population of highly aggressive cancer stem cells. Furthermore this model argues that complete cancer remission may only be achieved by eradicating the malignant stem cell population charged as the source of tumor cell renewal. Lung cancer is the most commonly lethal form of cancer in the world with about 90% of the nearly one million new cases succumbing to the disease. While progress is being made in understanding lung cancer pathogenesis and improving therapy, prognosis remains poor. One approach to improving outcome in lung cancer has been to therapeutically target a unique, phenotypically defined lung cancer stem cell population. However despite the relatively rapid pace of cancer stem cell research in solid tumors such as breast, brain and colon cancers, similar progress in lung cancer remains hampered in part due to an incomplete understanding of lung stem cell hierarchy and the complex heterogeneity of the disease. To address this challenge, putative lung cancer stem cells were prospectively isolated from patient lung tumors and lung tumor cell lines using methods that have been reported to enrich for other stem cell populations in other cancers. As a result, a subpopulation of cells with elevated aldehyde dehydrogenase (ALDH) activity within many NSCLCs was identified with properties indicative of a cancer stem cell population including enhanced tumorigenicity in xenograft models, clonogenicity in culture and the capacity for self-renewal. In support of this, analysis of 282 clinically annotated non small cell lung cancer samples found elevated ALDH1A1 expression, the protein that drives ALDH in lung cancer, was associated with poor patient prognosis. Finally, molecular characterization of isolated ALDH+ lung cancer cells revealed elevated expression of stem cell transcripts including Notch signaling transcripts, suggesting enhanced pathway activity. Suppression of Notch signaling through chemical inhibition or knockdown of the proto-oncogene NOTCH3 resulted in a significant reduction in clonogenic ALDH+ cells indicating the importance of Notch signaling in lung cancer stem cell homeostasis and as a potential target for lung cancer stem cell directed therapy.Item Molecular Imaging of αvβ6–Positive Tumors and Pancreatic β-Cell Mass by Radiolabeled Peptides(2011-02-01T19:35:51Z) 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.Item Moving Phantom Study of Stereotactic Body Radiation Therapy for Lung Cancer(2011-02-01T19:34:50Z) Huang, Long; Papiez, LechTo assess the accuracy of current stereotactic body radiation therapy (SBRT) lung treatment methodologies, we performed a systematic evaluation using phantoms that simulated motions from real patients (irregular motions) as well as sinusoidal motions (regular motions). The irregular patterns investigated in this study were of two types: small range irregular breathing motions (=10mm) and large range irregular breathing motions (=20mm). Four-dimensional computed tomography (4DCT) and cone beam computed tomography (CBCT) are important methodologies for SBRT, but previously have only been used to evaluate regular patterns. For targets moving regularly or irregularly within a small range (7.0 ± 1.8 mm, n = 6), we observed good agreement between the measured and computed dose distributions. However, for targets moving irregularly with a larger range (20.8 ± 2.6 mm, n = 4), the measured isodose lines were found to be shifted relative to the planned distribution, resulting in an underdosing (over 20%) in a portion of the PTV. In this underdosed volume, 1-2% of the PTV is underdosed by over 18 Gy, causing a 35-40% drop in the local control rate. We further observed that the discrepancy between the planned and measured dose distribution was due to the inaccurate representation of the irregular target motion in the maximum intensity projection (MIP) images generated from 4DCT, which could not be corrected by CBCT. A method of Extended Distance Virtual Isocenter (EDVI) was developed to lower the toxicity of healthy tissues. In all, caution should be used when planning from 4DCT images in the presence of large and irregular target motion. The inaccuracy inherent in 4DCT MIP and CBCT images can be mitigated through the application of methodologies to reduce respiratory motion, such as abdominal compression, and through the use of volumetric image guidance to assure precise targeting with minimal shifts.Item Part 1: Steroid Receptor CO-Activator 3 (SRC-3) Expression in Lung Cancer and its Role in Regulating Cancer Cell Survival and Proliferation; Part II: Development of Phosphospecific Peptoid Ligand(2009-09-04) Cai, Di; Kodadek, ThomasPART I: STEROID RECEPTOR CO-ACTIVATOR 3 (SRC-3) EXPRESSION IN LUNG CANCER AND ITS ROLE IN REGULATING CANCER CELL SURVIVAL AND PROLIFERATION Steroid receptor coactivator-3 (SRC-3) is a histone acetyltransferase and nuclear hormone receptor (NHR) coactivator, located on 20q12, which is amplified in several epithelial cancers and well studied in breast cancer, however, its role in lung tumorigenesis is unknown. We found that SRC-3 is over-expressed in 27% of the NSCLC patients, and SRC-3 high expression correlates with poor disease-free survival and overall survival. We also studied DNA copy number, mRNA and protein expression of SRC-3 in a large panel of lung (55 non-small cell lung cancers and 23 small cell lung cancers) and breast cancers (N=31) and also evaluated the functional consequences of altering its expression in lung cancer cell lines. There are significant alterations in lung cancers in SRC-3 gene copy number, including examples of both gene amplification and deletion. SRC-3 mRNA and protein expression varied dramatically among lung cancer cell lines. On average, lung cancer cell lines express higher levels of SRC-3 than immortalized human bronchial epithelial cells, which themselves express higher level of SRC-3 than cultured primary human bronchial epithelial cells. We found that ~27% of NSCLCs exhibited SRC-3 gene amplification and expressed SRC-3 mRNA at very high levels, suggesting that the expression of SRC-3 played a role in the malignant phenotype of these cancers. siRNA-mediated down-regulation of SRC-3 in high-expressing tumor cells significantly inhibited tumor cell growth and induced apoptosis. The effect of SRC-3 down-regulation on cell phenotypes correlated with a cell line's endogenous expression level of the gene. Finally, we show that SRC-3 knockdown is "synthetically lethal" to EGFR-TKI-resistant cells. Together these data indicate that SRC-3 is an important new oncogene and therapeutic target for lung cancer. PART II: DEVELOPMENT OF PHOSPHOSPECIFIC PEPTOID LIGAND Most proteins can exist in a variety of post-translationally modified forms. Chemical methods that would allow one to specifically purify or pharmacologically target a particular form of the protein would be of great interest. Here, we report the first peptidomimetic compounds that bind specifically to a serine-phosphorylated PDID domain of Brd4 protein, identified by screening a library of 40,000 peptoids for PDID binders. The isolated hit peptoids are only specific to phos-PDID, but not the non-phosphorylated form of the protein, or other phosphoserine- or phosphothreonine-containing proteins. Phos-PDID-binding peptoids can specifically capture a recombinant phos-PDID from a crude insect cell extract, without binding to the unmodified PDID in bacteria lysate. Moreover, the phosphospecific peptoid ligand engineered with a Biotin tag and a DOPA crosslinker can specifically detect phos-PDID from whole cell lysate, demonstrating its potential as a "Western blotting"-like reagent. Furthermore, GST pull-down assay and reporter gene assay reveal that the peptoid ligand can specifically disrupt the interaction between phos-PDID and high-risk HPV 18E2, and hence inhibits the Brd4-dependent transcription activation in human cervical cancer cells. Taken together, these data showed that our phosphospecific peptoid ligand is able to substitute phosphospecific antibodies for the detection and isolation of phosphoproteins; it can also perhaps be developed as a drug-like compound targeting the active form of protein in cells.