Project Summary/Abstract We aim to improve the cure rates for metastatic cancers. To achieve this we propose a combined modality approach to stimulate and diversify an endogenous anti-tumor immune response at all tumor sites to recognize and destroy tumor cells in a manner that will prevent recurrence and enable long-term cancer free survival. Immune checkpoint inhibitors (ICI; e.g. anti-PD-L1), are a class of immunotherapies that modulate immune tolerance of a tumor by blocking specific inhibitory receptor-ligand interactions on the surface of T cells and thereby overcoming T cell inhibition or exhaustion. In patients with immunologically “hot” tumors, characterized by a pre-existing but exhausted anti-tumor immune response, ICIs can restore efficacy to the anti-tumor immune response, sometimes resulting in complete and durable tumor regression. However, ICIs have not shown clinical benefit in the treatment of immunologically “cold” cancers that are characterized by low levels of T cell infiltrate and low mutation burden resulting in few mutation-created neo-antigens. To overcome immunotherapy treatment barriers posed by immunologically cold tumors, we propose to combine systemic delivery of ICIs with systemic delivery of radiation by targeted radionuclide therapy (TRT). To date, nearly all approaches to combining radiation and immunotherapy have used external beam radiotherapy (EBRT), which promotes tumor immune cell infiltration through activation of type I interferon (IFN) responses. Administration of EBRT to multiple tumor sites or to the whole body (to target radiographically occult or microscopic disease) would result in prohibitive toxicity, including lymphopenia. TRT is a systemic method of delivering a therapeutic radionuclide to a tumor, which poses an alternative option for delivery of immunomodulatory radiation to metastatic tumor sites without causing immunosuppression. The Weichert lab at the University of Wisconsin-Madison has developed a novel class of TRT, known as NM600, an alkylphosphocholine analog that is selectively taken up and retained in nearly any tumor type in any location. Our broad hypothesis is that unique physical properties of radionuclides (e.g. emission type, linear energy transfer, half-life, tissue range) differentially impact immunomodulation by TRT. In this study, the immunomodulatory capacity of alpha- (225Ac) and beta- (90Y) particle emitting radionuclides will be compared directly. In a project that builds upon the ongoing collaborative progress of the Morris and Weichert labs, we will now determine the radionuclide-specific potency of combining TRT with immunotherapy to enhance the immune response against immunologically cold tumors. In murine models, we will: 1) expand on preliminary data showing potent synergy with the combination of TRT and ICI, 2) evaluate therapeutic mechanisms of TRT and ICI using the intrinsic properties of 225Ac- and 90Y-NM600, focusing on type I IFN response activation and 3) in...