The model system we study is a transgenic mouse model of squamous cell carcinoma (SCC) development where human papillomavirus type 16 (HPV16) oncogenes are expressed in mitotically active basal keratinocytes in skin epidermis, e.g., K14-HPV16 transgenic mice (Coussens et al., 1996). While HPV16 oncogenes act as ‘initiators’ of neoplastic progression in this model, we have identified ‘host’ inflammatory cells, e.g., mast cells and neutrophils (Coussens et al 1999; deVisser et al 2004, 2005), and specific stromal-derived extracellular proteinases, e.g., matrix metalloproteinases (MMPs) (Coussens et al., 2000; vanKempen et al., 2002), chymase/mMCP-4 (Coussens et al., 1999), tryptase/mMCP-6 (Coussens et al., 1999) and their endogenous tissue inhibitors (Rhee et al., 2004) as critical extrinsic regulators of epithelial neoplastic evolution. By genetically modulating the host immune response and/or presence of various extracellular proteinases with specific gene knockouts and double transgenic mice, we have been able to manipulate the temporal dynamics, overall incidence and malignant potential of carcinomas that develop in HPV16 mice. Our findings that inflammatory leukocytes contribute in a dominant manner to the activation of neoplasia-associated angiogenesis functionally link inflammation as a potentiator of the neoplastic process (Coussens et al., 2000; van Kempen et al., 2002; de Visser et al, 2005). More recently, we demonstrated that B lymphocytes are required for establishing chronic inflammatory states and promotion of de novo epithelial carcinogenesis (de Visser et al., 2005). Adaptive immune-deficiency in HPV16 mice results in failure to recruit innate immune cells into premalignant tissue, and as a consequence, responding pathways downstream of inflammatory cell activation, e.g. tissue remodeling, angiogenesis, keratinocyte hyperproliferation and cancer development, are significantly attenuated. Importantly, these necessary characteristics of premalignant progression are restored by transfer of B lymphocytes or serum from HPV16 mice into T and B lymphocyte-deficient/HPV16 mice (deVisser et al, 2005), indicating that B lymphocytes play a crucial role as initiators of chronic inflammation associated with premalignant progression, and thus potentiate the neoplastic cascade downstream of oncogene expression. While there have been many studies on the role of B lymphocytes and/or immunoglobulins as mediators of acute inflammation, this is the first to provide clear and convincing functional proof, via the combined use of gene knock out mice, adoptive B cell transfer and serum transfer, to demonstrate that B cells play such important roles in epithelial carcinogenesis, and particularly at the earliest stages of carcinogenesis, during premalignant progression. Altogether, these studies indicate that molecules downstream of inflammation and/or protease action play fundamental roles in discrete stages of tumor development.

Based on our data implicating inflammatory leukocytes and their products, e.g., proteases, as important regulators of epithelial carcinogenesis, we hypothesized that altering the susceptibility of critical MMP substrates to proteolytic cleavage might similarly alter neoplastic development. Thus, we tested whether the equilibrium between synthesis, accumulation and degradation of a key MMP target, e.g., type I collagen, was central for the cascade of cellular and genetic changes necessary to manifest carcinoma development and/or metastasis in HPV16 mice (vanKempen et al., Manuscript in preparation). Our studies reveal that both neoplastic progression to SCC and metastatic capability of SCCs is significantly reduced in an environment where matrix remodeling is restricted. In combination, HPV16/immune altered, HPV16/protease altered and HPV16/protease-substrate altered mice have allowed us to test the hypothesis that the ‘host response’ to neoplastic progression represents a fundamental aspect of carcinogenesis and reveals important new targets for therapeutic intervention.