SUMMARY The goal of our project it to provide a structural and function framework for the development of specific therapeutics targeting an understudied kinase HIPK1. HIPK1 is a member of the homeodomain-interacting protein kinase (HIPK) family of nuclear dual specificity serine/threonine and tyrosine kinases that belong to the CMGC group of human protein kinases. HIPK kinases are involved in regulation of transcription and play essential roles in human development and adult organism physiology. Their involvement in several types of cancer, including lung, breast, colorectal and pancreatic cancer, as well as in lung injury and fibrosis has recently elevated them to exciting new potential therapeutic targets. In particular, HIPK1 has been recently emerging as a potent oncogene, capable of transforming normal cells in the in vitro assays. HIPK1 levels and kinase activity are found to be significantly elevated in a number of human cancers, including lung adenocarcinoma. In the lung, HIPK1 also plays a role in control of inflammatory response in acute lung injury model. Hence in lung pathologies HIPK1 is a promising new potential therapeutic target. However, with the exception of HIPK2, the remaining members of the HIPK family, including HIPK1, are categorized as understudied by the Illuminating the Druggable Genome (IDG) initiative. Our team has recently solved the first crystal structure of a HIPK kinase: HIPK2. This structure elucidated unique structural features of HIPK2, including kinase active site bound to a small molecule inhibitor and a conformation of a domain inserted within the HIPK2 kinase module, called the CMGC insert. In other CMGC kinases, the CMGC insert region is critical for signaling via protein-protein interactions, and our analysis showed that all HIPK kinases likely have uniquely structured CMGC insert regions. In this proposal, we aim to leverage our experience gained via the structural studies of HIPK2 and through combined biochemical, structural, computational and cell-based approaches characterize the structure and inhibitor selectivity of the understudied HIPK1 kinase. We have recently identified candidate small molecule HIPK1 binders that we will optimize to identify potent and selective HIPK1 inhibitors to interrogate HIPK1 function and the benefit of its inhibition in lung cancer cells. The results of our work have potential to advance known strategies for therapies of several types of cancer and diseases associate with lung injury.