Congratulations to our 2013 Pilot Project Awardees (Women’s Cancers)!
Deri Morgan, PhD
Qiping Zheng, PhD
Title: The Role of Voltage-Gated Proton Channels in pH Regulation and Reactive Oxygen Species in Metastatic Breast Cancer Cells
PI: Deri Morgan PhD, Associate Professor, Department of Biophysics
Abstract: Cancer cells are highly metabolic cells that often grow in hypoxic environments of low pH and generate a large amount of reactive oxygen species (ROS). The ability of these cells to extrude acid to maintain a high cytosolic pH enables their survival. Prevention of acid extrusion in malignant cells thus would serve as a method to target cancerous tissue, which produces excessive acid due to the Warburg effect. It was shown recently that highly metastatic breast cancer cells have substantially higher levels of voltage gated proton channel (Hv1) expression than healthy breast tissue. Furthermore, high Hv1 levels were correlated with increased tumor size, more severe classification, more advanced clinical stage, lower recurrence-free survival, and shorter overall survival. Inhibition of gene expression using siRNA inhibited cancer growth and suppressed the cancer cell proliferation. Down-regulation of Hv1 prevented extracellular acidification and increased intracellular acidification in some cancer cell lines. These results indicate that Hv1 is a highly promising target to ameliorate breast cancer. However, because this is the first report of Hv1 involvement in cancer, very little is known about the contribution of the voltage gated proton channel to pH homeostasis in cancer cells. In fact, there is no direct electrophysiological confirmation that functioning Hv1 are present in plasma membranes of breast cancer cells. We will apply our expertise in Hv1 electrophysiology to measure the membrane activity of Hv1 in breast cancer cells directly via the patch clamp technique. We further hypothesize that the “enhanced gating mode” - profound enhancement of Hv1 activity that occurs in phagocytosing leukocytes – may contribute to the ability of Hv1 to exacerbate breast cancer by facilitating acid extrusion by cancer cells. Cancer cells produce a large amount of ROS and NADPH oxidase has been implicated in cancer cell survival and metastasis. We have shown that Hv1 regulates NADPH oxidase in cells such as neutrophils and B cells. We have evidence that highly metastatic cancer cells release ROS via the NADPH oxidase and we hypothesize that Hv1 enables the oxidase to function in an increased capacity in metastatic cancer cells enabling their survival, migration and invasion. We intend to utilize cellular assays and molecular knockdown techniques to determine the presence of NADPH oxidase in breast cancer cell lines and the nature of the involvement Hv1 with the activity of the enzyme. We hypothesize that the mechanism for at least part of the effects that Hv1 has on breast cancer cells involves modulation of NADPH oxidase activity.
Title: URI1 variants in female cancer
PI: Qiping Zheng, PhD, Associate Professor, Department of Anatomy
Co-Is: Drs. Sumner Dewdney, MD, Junxia Gu, MD, PhD and Carl Maki, PhD
Abstract: The overall objective of this project is to define the correlation of URI1 with cervical cancer carcinogenesis. Cervical cancer is the second most common female cancer worldwide and one of the leading causes of death in relatively young women. Previously, cervical cancer has been attributed to human papillomavirus (HPV) infection, which may take years to turn precancerous cervical lesions into invasive cervical cancer. This makes it possible to identify cervical lesions or cancerous changes at early stages, and thus, greatly decreased the incidence and mortality of cervical cancer after proper intervention and treatment. However, among the more than 120 types of HPV, only 15 are considered high risk HPVs that are linked to cervical cancer carcinogenesis. The fact that not everyone infected with high-risk HPVs develops cervical cancer suggests that additional oncogenic biomarkers or genetic changes may also contribute to this cancer progression. Based on ours and others’ published work and our preliminary studies, here, our hypothesis is that URI1 is such a biomarker that is involved in cervical cancer oncogenesis. URI1, or URI/RMP, a member of the prefoldin family of molecular chaperones, has recently been implicated a role in the progression of hepatocellular carcinoma and ovarian cancer. Interestingly, by tissue microarray (TMA) and immunohistochemistry (IHC) analysis, we have observed upregulated URI1 expression in tissues from precancerous cervical intraepithelial neoplasia (CIN) and invasive cervical cancer compared to normal cervical tissue. This result suggests an early diagnostic and possibly, a prognostic value of URI1 in cervical cancer clinics. URI1 is known to consist of several alternatively spliced transcript variants that encode multiple URI1 isoforms. How these variants contribute to precancerous changes of the cervix and invasive cervical cancer is largely unknown. Surprisingly, our preliminary results support a role of URI1 over- or mis-expression in cancer formation due to mutation or gene copy number variation (CNV). We therefore propose to genetically determine the correlation of different URI1 variants with cervical cancer initiation and progression. By examining the URI1 transcript variants in precancerous CINs, we expect to identify URI1 isoforms, in addition to HPV, as an alternative biomarker that will facilitate early detection of precancerous changes of the cervix. Moreover, we have previously demonstrated a correlation of URI1 in P53 related cell apoptosis signaling pathway with hepatocellular carcinoma tumor growth. We therefore propose to delineate the putative involvement of URI1 in the canonical HPV-P53-RB pathway in cervical cancer oncogenesis. Our long-term goal is to define the mechanism and correlation of URI1 with cervical cancer initiation and progression, and ultimately promote the diagnosis, as well as the therapeutics and treatment of cervical cancer.