Enhancing Direct Tumor Cell Cytotoxicity By Manipulating Growth Factor Signaling

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The broad, long-term goal of this application is to optimize serosal photodynamic therapy (PDT) through the inhibition of relevant components of the signal transduction pathways thereby improving the outcome of patients with pleural and peritoneal spread of cancers. Activation of the epidermal growth factor receptor (EGFR) signaling pathway is thought to limit the effectiveness of PDT as a therapeutic approach. Preliminary data suggest that inhibition of EGFR will increase the therapeutic index of PDT, increasing tumor cell kill without a concomitant increase in normal tissue toxicity. Moreover, while PDT is thought to kill cells primarily through apoptotic mechanisms, augmentation of cancer cell death through inhibition of EGFR signaling may occur through stimulation of necrotic/non-apoptotic cell death pathways. Therefore, this project will focus on defining and characterizing the impact of inhibiting growth factor signaling on the signaling, cytotoxicity and mechanisms of cell death following BPD-mediated PDT of ovarian and lung cancer cells. To maximize the external validity, generalizability and clinical applicability of these findings, experiments will be performed using a panel of ovarian cancer (QvCa) and non-small cell lung cancer (NSCLC) cell lines that includes OVCAR-5 and H460 and also using OVCAR-5 and H460 tumor xenografts. We will use chemical inhibitors and siRNA to inhibit EGFR and its post-receptor signaling partners to investigate the molecular mechanism by which the inhibition of EGFR signaling enhances BPD-mediated PDT cytotoxicity. We will also examine the sequencing and timing PDT and EGFR pathway inhibition in order to maximize the enhancement of BPD-mediated PDT cytotoxicity. The specific endpoints that will be analyzed in these studies are: 1) Levels and activation of EGFR and related signaling proteins 2} The nucleo-cytoplasmic distribution of EGFR and its association with nuclear STAT3: 3) The clonogenic survival of cells and tumors. Lastly, we will determine the effect of modulating growth factor signaling on the mechanism of cell death following BPD-mediated PDT in order to further characterize this novel treatment approach and to further identify areas for enhancement of cytotoxicity. The specific endpoints that will be analyzed in these studies are: 1) The percent of cells with apoptotic vs necrotic cell death pheotypes; 2) The expression and activation of cell death-related proteins: 3) The effect on p53. Bax and Bak protein expression on clonogenic cell survival and the percent of cells with apoptotic vs necrotic ceil death phenotypes These studies should help us to understand the potential clinical application of combination PDT + growth factor inhibition as a therapeutic approach for patients with serosal malignancies. Given the extremely poor prognosis for patients with spread of either ovarian cancer within the abdominal cavity or NSCLC within the chest cavity, successful clinical application of this treatment approach could lead to a new standard of clinical care in these populations.