In colitis, TH17 cell-mediated inflammation can damage mucosal barrier surfaces and cause chronic inflammation, creating a tumor-permissive microenvironment. Individuals with colitis are therefore at increased risk for developing colorectal cancers. Accordingly, negative regulation of TH17-type immune responses is an important component in prevention and attenuation of colitis-associated colorectal cancer (CAC). Nedd4-family E3 ubiquitin ligases are known regulators of T cell activation and cytokine production. These ligases can be activated by interaction with adaptor proteins like Nedd4-family interacting protein 2 (Ndfip2). In vitro data indicate that Ndfip2 can bindto and activate the catalytic function of several Nedd4-family E3 ligases. While it has been shown that Ndfip2 expression is increased upon T cell activation, a physiological role for Ndfip2 has not been described. Newly generated Ndfip2-/- mice do not exhibit overt immunopathology, but show an accumulation of lymphocytes in the small bowel and colon. In a T cell transfer model of autoimmune colitis, transfer of Ndfip2-/- CD4+ T cells caused more severe colon pathology in Rag1-/- recipients than WT control cells. Helping to explain this increase pathology, more Ndfip2-/- CD4+ T cells produced IL-17 compared to WT CD4+ T cells. These data suggest that Ndfip2, through interaction with Nedd4-family E3 ligases, is a negative regulator of mucosal immune responses and TH17 cell pathogenicity. This proposal will test the hypothesis that Ndfip2 activates specific Nedd4-family E3 ligases to limit the effector function and persistence of pathogenic TH17 cells, thereby promoting mucosal barrier integrity and attenuating CAC progression. Ndfip2 could regulate the pathogenicity of mucosal TH17 cells through limiting cytokine production or the expansion/persistence of activated T cells.
In aim 1, the role of Ndfip2 in regulation of TH17 cell is examined using in vitro cultures and by in vivo induction of TH17-mediated inflammation.
This aim will determine effects of Ndfip2 on cytokine type and quantity, and examine cell viability and division to determine if Ndfip2 affects the persistence and/or function of activated TH17 cells. These experiments will be repeated with CD4+ T cells deficient in two relevant Nedd4-family E3 ligases to determine if these ligases affect the pathogenic potential of TH17 cells.
Aim 2 will determine the physiological relevance of Ndfip2 in TH17-mediated mucosal inflammation using a mouse model of CAC. The impact of Ndfip2 deficiency on mucosal inflammation will be determined by quantifying the magnitude of inflammation and tumor burden during progression of disease relative to WT animals; the contribution of T cels and innate cells to disease will be analyzed. Together, these aims will characterize a novel ubiquitin-mediated pathway that limits gastrointestinal inflammation and colon cancer, and is likely to reveal previously unappreciated pathogenic pathways in inflammatory cancers that might be targeted therapeutically. Additionally, studies detailed in this proposal will lay the foundation for future work aimed at identifying substrates ubiquitylated in a Ndfip2-dependent manner.
Individuals with colitis are at increased risk for developing colorectal cancer. Chronic T cell-mediated inflammation and TH17 cytokine production creates an environment permissive for tumor growth. Limiting TH17 cytokine production and inflammation is therefore an important component in prevention and attenuation of colitis-associated colorectal cancer. My preliminary data indicate that Nedd4-family interacting protein 2 (Ndfip2) is a negative regulator of mucosal immune responses and pathogenic TH17 cells. In this proposal we will determine how Ndfip2 limits TH17 pathogenicity and test whether loss of Ndfip2 leads to increased gastrointestinal inflammation and colon cancer progression. These studies will enhance the current understanding of TH17-cell mediated pathology in inflammatory cancers, and highlight new molecular pathways for therapeutic intervention.