Project Summary/Abstract Outcomes among patients with Nodal Lymphomas of Follicular-Helper T cell (TFH) origin are poor, with overall survival rates of <35% at 5 years. These lymphomas are characterized by the recurrent hotspot RHOA G17V and TET2 loss-of-function (LOF) mutations. The causative molecular contributions of these mutations to TFH lymphomagenesis remain undefined. Rho family proteins are molecular switches that are active when bound to GTP and inactive when bound to GDP. The G17V mutation locks RhoA in an inactive state that has dominant negative and likely neomorphic function through sequestration of multi-substrate binding partners. TET2 LOF mutations are predicted to cause increased 5' methylation (5-mC) and decreased 5' hydroxymethylation (5-hmC) of DNA cytosines, both of which are predicted to result in diminished expression from affected genetic loci. These inactivating properties make these mutations difficult to target. Recently we and others have determined that RHOA G17V expression causes increased signaling through the Akt/mTOR pathway during T-cell receptor (TCR) and co-stimulatory pathway stimulation. We hypothesize that RhoAG17V-mediated disruption of signaling through multi-substrate small GTPase regulatory proteins combined with gene expression and epigenetic aberrancies caused by TET2 loss drive lymphomagenesis in T cells. We will test this with two specific aims. 1. Determine the molecular intermediaries of RhoA G17V-mediated TCR and co-stimulatory receptor dysfunction. We will determine direct binding partners of RhoA G17V and define how they modulate TCR and co-stimulatory pathways to promote lymphomagenesis. 2. Determine the epigenetic and transcriptional aberrancies mediated by TET2 loss in T cells. We will use a unique Nodal TFH lymphoma cell line to define the epigenetic modifications associated with TET2 loss and to determine whether genes that are silenced by these mutations can act as tumor suppressors. There are no current therapies that directly target these mutations, and thus, these studies serve an unmet need for these diseases. Dr. Ng has outlined a five-year development plan to achieve his goal of becoming an independent investigator in translational cancer research. He has assembled an advisory committee of internationally recognized experts in antigen-receptor signaling, malignant T-cell, and lymphoma biology. He has enlisted collaborators who are experts in epigenetics and computational biology to provide experimental advice and for specific training in the field. Dana- Farber Cancer Institute is the ideal environment for completion of his scientific and career goals given its outstanding research community and substantial record for training independent physician- scientists.