Overexpression of the epidermal growth factor receptor (EGFR), a member of the ErbB family, and its closest homologue HER2, have been associated with aggressive tumour progression and reduced sensitivity to DNA-damaging agents. In order to block the proliferation of refractory tumors overexpressing EGFR, a novel strategy has been developed that sought to design molecules capable of not only blocking EGFR-TK, but also damaging DNA. These molecules, termed combi-molecules (CMs), have been shown to degrade under physical conditions to release another inhibitor of EGFR, and to be potent against tumor cells of various origins including breast, prostate and carcinoma of the vulva. However, despite their potency, their growth inhibitory IC50 values were still in the high micromolar range. In order to augment the potency of the CMs, here they were re-designed to contain two quinazoline moieties and a central N,N-bis(2-aminoethyl)methylamine spacer which, following degradation, could yield higher concentrations of free inhibitors and a more cytotoxic bifunctional DNA damaging species. Here, we describe the mechanism of action of the first prototype of this approach, JDE52, which we now classify as a double-arm CM, in comparison with ZRBA1, its closest single-arm counterpart. The results indicated that JDE52 was capable of inducing significant blockade of EGFR, DNA single-strand breaks and inter-strand cross-links. ZRBA1, its single-arm counterpart, was capable of only forming DNA single-strand breaks. The fluorescent property of FD105, the secondary inhibitor that both JDE52 and ZRBA1 are capable of releasing, has permitted the analysis of its levels in tumor cells by UV flowcytometry. It was found that JDE52 was indeed capable of significantly releasing higher levels of fluorescence (p<0.05) in human tumor cells, compared with levels of fluorescence released by ZRBA1. More importantly, JDE52 induced higher levels of apoptosis and cell killing than ZRBA1. Apoptosis was triggered by JDE52 at a faster rate than ZRBA1. The results in toto suggest that the superior potency of JDE52, when compared with ZRBA1, may be imputed to mechanisms associated with the generation of higher levels of FD105 intracellularly, and the induction of DNA cross-links, which are known to be more cytotoxic. These combined mechanisms (blockade of EGFR-TK and formation of cross-links) contributed to an accelerated rate of apoptosis in cells treated with JDE52. This study conclusively demonstrated that designing molecules as prodrugs of high levels of quinazoline inhibitors of EGFR and bifunctional DNA cross-linking species is a valid strategy to enhance the potency of CMs against refractory tumors.

Lung cancer is the leading cause of cancer-related deaths among men and women. Non-Small Cell Lung Cancer (NSCLC) constitutes the most common type of lung cancer and is frequently diagnosed at advanced stages. In the past decade, discovery of Epidermal Growth Factor Receptor (EGFR) mutations have heralded a new paradigm of personalized treatment for NSCLC. Clinical studies have shown that molecular targeted therapies increase survival and improve quality of life in patients. Despite these advances, the realization of personalized therapies for NSCLC faces a number of challenges including the integration of clinical and genetic data and a lack of clinical decision support tools to assist physicians with patient selection. This thesis demonstrates the development of a predictive computational model for personalized therapeutic interventions in advanced NSCLC. The findings suggest that the combination of clinical and genetic data significantly improves the model’s predictive performance for tumor response than clinical data alone.

Membrane rafts are small plasma membrane domains that contain high levels of cholesterol and sphingolipids. They have been implicated in processes as diverse as signal transduction, endocytosis and cholesterol trafficking. Traditional methods for the biochemical preparation of lipid rafts involve the extraction of membranes with nonionic detergents followed by the separation of a low-density, detergent-resistant membrane fraction on density gradients. Because of concerns regarding the possible introduction of artifacts through the use of detergents, several methods were developed for the isolation of lipid rafts that do not involve detergent extraction. / In this study, we compared three different biochemical methodologies of membrane rafts preparation from purified rat liver PM. Only detergent-resistant membranes (DRMs) fulfill the requirements of membrane rafts. We subsequently found using the low dose of EGF (1 ug/100 g BW); the content of EGFR in PM-DRMs did not changed significantly following EGF administration. When a higher dose of EGF (5 ug/100 g BW) is administrated we observed a rapid and almost complete disappearance of EGFR (around 80%) from both PM and DRMs fractions. Interestingly, following the administration of a low or high dose of EGF, the pool of EGFR in the PM-DRMs fraction becomes highly Tyr-phosphorylated. In accordance with the higher level of EGFR Tyr-Phosphorylation, EGF induced an augmented recruitment of Grb2 and Shc proteins to PM-DRMs compared with whole PM. / Furthermore neither high nor low dose of EGF affects the caveolin content in DRMs and PM. These observations suggest that EGFR located in DRM are competent for signaling and non-caveolae PM rafts are involved in the compartmentalization and presumably internalization of the EGFR.

Diss. (sammanfattning) Stockholm : Karolinska institutet, 2007. / Härtill 5 uppsatser.

Thesis (Ph. D.)--University of Alabama at Birmingham, 2007. / Title from first page of PDF file (viewed Oct. 31, 2007). Includes bibliographical references.

Diss. (sammanfattning) Stockholm : Karol. inst., 2005. / Härtill 4 uppsatser.

Diss. (sammanfattning) Uppsala : Uppsala universitet, 2006. / Härtill 6 uppsatser.

Diss. (sammanfattning) Umeå : Umeå universitet, 2006. / Härtill 5 uppsatser.

Thesis (Ph. D.)--Case Western Reserve University, 2005. / [School of Medicine] Department of Physiology and Biophysics. Includes bibliographical references. Available online via OhioLINK's ETD Center.

Thesis (Ph. D.)--Medical College of Ohio, 2004. / "In partial fulfillment of the requirements for the degree of Doctor of Philosophy in Medical Sciences." Major advisor: Sonia Najjar. Includes abstract. Document formatted into pages: iv, 181 p. Title from title page of PDF document Includes bibliographical references (p. 123-180).