Date of Graduation
Eberly College of Arts and Sciences
Scott A. Weed
Head and neck squamous cell carcinoma (HNSCC) is a highly invasive cancer with a five-year overall survival of approximately 50%. HNSCC incidence is increasing in WV and other Appalachian areas due to high tobacco consumption and human papilloma virus exposure. Genomic instability is a hallmark of tobacco carcinogen exposure, resulting in amplification of chromosomal region 11q13 in 25% of all HNSCC cases. 11q13 amplification correlates with poorer prognosis and increased tumor aggressiveness. The CTTN locus resides in the 11q13 amplicon and encodes the actin-regulatory protein cortactin. Increased cortactin expression resultant from CTTN amplification drives HNSCC invasiveness by increasing tumor cell migration and invadopodia formation. Invadopodia are tumor cell membrane protrusions responsible for degrading the basement membrane and stromal extracellular matrix (ECM) proteins, facilitating regional tumor dissemination. Prior studies have demonstrated that loss of cortactin expression by RNA interference blocks invadopodia formation, suggesting that cortactin is an essential invadopodia protein. However, recent work from our group utilizing 3D in vitro and transgenic knockout mouse models indicates that cortactin removal fails to block HNSCC invasion. The purpose of this study is to unequivocally determine the role of cortactin in HNSCC invadopodia function and tumor invasion using clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9 knockout technology in HNSCC cell lines. The overall hypothesis is that HNSCC may have compensatory cortactin and/or invadopodia-independent mechanisms capable of driving tumor invasiveness independent from cortactin. We have produced multiple cortactin knockout HNSCC clonal cell lines using CRISPR-Cas9 that lack detectable cortactin expression. HNSCC lines lacking cortactin form mature invadopodia and degrade ECM to the same extent as cortactin-expressing control cells. In addition, cortactin knockout cells have the same degree of invasiveness in 3D collagen-based tumor spheroid and organotypic models. These results suggest that HNSCC utilizes one or more cortactin-independent alternative/compensatory mechanisms to generate invadopodia in order to maintain the capacity for tumor cells to invade. Future work will identify these compensatory mechanisms as a first step towards crafting effective anti-invasive therapies.
Hames, River Austin, "CRISPR-Cas9 mediated genetic ablation on the actin regulatory protien cortactin does not impain HNSCC invasive processes" (2018). Capstones and Honors Theses. 12.