New PROTACs Targeting Tissue Transglutaminase in Ovarian Cancer

Background/Readiness: Ovarian cancer (OC) peritoneal metastasis: The main cause of fatality in OC is related to peritoneal metastasis which leads to the common complications associated with the disease, bowel obstruction and ascites accumulation. There are unique traits involved in OC peritoneal dissemination which differ from the process of metastasis in other solid tumors where loss of cell-cell contact with breakdown of the basement membrane followed by tumor invasion into lymphatic or vascular channels are required1. By contrast, hematogenous dissemination is uncommon in OC, tumor spread occurring predominantly intraperitoneally (ip) 2. OC cells are in direct contact with the overlying peritoneal surface and fluid and simple dislodgement from the primary tumor allows cells to float in the peritoneal space, where they adhere and form metastatic implants. In the peritoneal fluid, cells aggregate as spheroids, which provide protection from the stress imposed by the extracellular environment. Interactions between cancer cells and the peritoneal mesothelium activate “outside-in” signaling3 which stimulates cell proliferation, survival and tumor angiogenesis. Identifying key mechanisms that regulate this process could lead to new treatments to attack and block metastasis leading to decreased burden of the disease and thus responding to the call of the 2021 Department of Defense-OC Research Program. Over the past decade we reported upregulation of tissue transglutaminase (TG2) and established its critical role in OC peritoneal metastasis 4 5. Our group demonstrated sequentially that TG2 regulates epithelial to mesenchymal transition (EMT) 6, migration of cells from the primary tumor site5, formation of spheroids in the peritoneal fluid 7, survival of cancer stem cells (CSCs)7,8, and invasion into the peritoneum 5,9. These steps, regulated by TG2 partly through its interaction with fibronectin (FN), contribute to establishment of peritoneal implants 5,6 and render TG2 an attractive new cancer target. We showed that TG2 knock down blocked peritoneal dissemination in orthotopic OC models 6. Based on our discoveries over the past decade 5-7,9-12 mechanistically linking the TG2 and the TG2/FN/integrin β1 interaction to peritoneal metastasis and cancer stemness, we propose TG2 as a new target in OC. TG2—structure/function relationship: TG2 is a 76-kD protein belonging to the transglutaminase family, which includes TG1-7, Factor XIII and erythrocyte protein 4.2. They have similar catalytic activities, but restricted substrate specificity and distinct mechanisms of transcriptional regulation, leading to a tissue specific pattern of expression. TG2 has four domains: an N-terminus β-sandwich domain, which binds to FN, a catalytic triad C277H335D358, which carries out the acyl-transfer function, and two β-barrel domains 13,14 (Fig. 1). A GTP/GDP-binding site is located between the catalytic and the first barrel domain, suggesting that TG2 functions as a GTP-ase. However, TG2 does not have a classical switch region, characteristic of G-proteins, and it remains unclear whether and how GTP/GDP binding affects signaling. TG2 also interacts with PLCγ through a region mapped at its C-terminus, supporting signaling from adrenergic receptors 15. The functions of the protein are modulated through large allosteric changes in the protein structure, regulated by external factors (GTP and Ca2+ concentrations). The crystal structure of GTP bound TG2 (PDB ID 1KV3) showed a compact conformation where the 2 β-barrel domains folded over the catal