The structural integrity of the human epidermis crucially depends on attachment of basal keratinocytes to the basement membrane through integrin receptors. Integrin ?? heterodimers respond to a variety of extracellular matrix cues to promote adhesion and epidermal stratification. Several integrin subunits are aberrantly over-expressed over the course of squamous cell carcinoma (SCC) progression, leading to altered subunit heterodimerization and intracellular signaling that can promote tumor growth and invasion. While some integrin subunits are known to contribute to both skin homeostasis and SCC, the functional roles for most subunits in skin are unknown. The Ridky lab has developed a novel, three-dimensional human skin model that is ideal for the study of integrin signaling. We reconstitute skin in organotypic culture (OTC), using primary human dermal and epidermal cells and native human dermis with a functional and intact basement membrane zone. The primary keratinocytes used in this skin model can be transduced with oncogenes to generate invasive neoplastic tissue. Additionally, this OTC skin can be grafted onto an immunocompromised mouse to generate viable human skin or SCC tumors in vivo. Using this model, I have shown for the first time that integrin ?v is necessary for basal keratinocyte proliferation and epidermal stratification. I have also shown that integrin ?v is essential for squamous cell carcinoma invasion through the basement membrane. The experiments described in this proposal aim to elucidate the specific signaling mechanisms that mediate this loss of skin proliferation (Aim 1) and SCC invasion (Aim 2) seen upon integrin ?v knock down in OTC and in vivo. In cultured cells, integrin ?v knock down correlates with a decrease in EGFR, TGF? and FAK activation.
Aims 1 A and 2A seek to uncover whether these pathways functionally mediate the loss of epidermal proliferation and stratification and SCC invasion upon integrin ?v knock down. This will be done by functionally rescuing activation of these pathways using lentivirus transduction, and determining whether the phenotype is reversed. Experiments proposed in Aims1B and 2B will determine the specific ? binding partners that complex with integrin ?v to mediate its role in both normal skin and neoplasia. This will be done by performing co-immunoprecipitation in cultured cells and tissues, and knocking down expression of each ? binding partner to determine if this can phenocopy loss of integrin ?v. Two of integrin ?v's ? binding partners-?1 and ?6-are up-regulated in a similar model of SCC, suggesting a heterodimeric switch during the course of tumor progression. My proposed work will determine if this heterodimeric switch exists, and if specific ?v heterodimers are viable therapeutic targets.
Tumor cell invasion through the basement membrane and dissemination to other tissues and organs is the major cause of death for patients suffering from epithelial cancers. I have discovered that a specific integrin subunit-integrin ?v-is necessary for skin squamous cell carcinoma (SCC) cells to invade through the basement membrane. My proposal aims to study how this integrin signals in both normal skin tissue and in invasive SCC tissue in order to determine how to effectively target integrin ?v heterodimers for anti-metastatic therapy.
