Part I: 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

dc.contributor.advisorKodadek, Thomas J.en
dc.contributor.advisorMinna, John D.en
dc.contributor.committeeMemberWang, Xiaodongen
dc.contributor.committeeMemberKliewer, Steven A.en
dc.creatorCai, Dien
dc.description.abstractPART 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.en
dc.format.digitalOriginborn digitalen
dc.subjectCarcinoma, Non-Small-Cell Lungen
dc.subjectNuclear Receptor Coactivator 3en
dc.subjectLung Neoplasmsen
dc.titlePart I: 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 Liganden
dc.type.materialTexten School of Biomedical Sciencesen Chemistryen Southwestern Medical Centeren of Philosophyen


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