Glioblastoma (GBM), the most common primary malignant brain tumor of adults, is a significant cause of patient morbidity and mortality for which effective treatments are lacking. The cyclin D1-cyclin dependent kinase 4/6-retinoblastoma (cyclinD1-CDK4/6-Rb) signaling axis is genetically activated in majority of GBM (~80%) via genomic loss of CDKN2A/B, amplification of CDK4/6 or deletion/mutation of RB1. CDK4/6 has been targeted based on the notion that suppressing the phosphorylation of pRB by CDK4/6 will lead to cell cycle arrest. Beyond suppressing cell cycle progression, we recently found that CDK4/6 antagonists promote anti-tumor immunity. The molecular mechanisms underlying this are exerted at two levels: (i) a tumor cellautonomous enhancement of the antigen processing and presentation machinery and (ii) a non-tumor cellautonomous, systemic decrease of the Treg/CD8+ ratio. Collectively, these effects promote cytotoxic T cellmediated clearance of tumor cells, which is further enhanced by the addition of immune checkpoint blockade therapeutics. Notably the actions of the combination of CDK4/6 inhibition and checkpoint blockade was much greater than additive in our preclinical models. CDK4/6 inhibitors are FDA-approved for the treatment of estrogen receptor (ER)-positive metastatic breast cancer, where they now present a well-tolerated, first-line therapy that improves progression-free survival. Their efficacy against GBM is unknown. However, early unpublished clinical data suggest that, like most targeted therapies, CDK4/6 inhibitors as single agents may have only modest benefit. Similarly, early data on immune checkpoint blockade have not been promising in recurrent GBM in which recently this class of drug failed to improve survival as single agent therapy. Building upon our recent findings, we hypothesize that brain penetrant CDK4/6 inhibitors could augment immunotherapy approaches for GBM including PD-1 checkpoint inhibitors for recurrent GBM. This proposal has three specific aims designed to investigate the therapeutic approach of combined CDK4/6 inhibition and immune checkpoint blockade (ICB) in GBM in both preclinical and clinical settings: (Aim 1) To assess the effects of CDK4/6 inhibition on GBM cell-intrinsic immune response; (Aim 2) To assess the effects of CDK4/6 inhibition on enhancing immunotherapy in syngeneic models of GBM; and (Aim 3) To evaluate the impact of CDK4/6 inhibitors on immune function and clinical outcome for GBM patients. By using patient-derived GBM tumors and syngeneic mouse models of GBM, we will determine the preclinical efficacy of CDK4/6 inhibitors in combination with immunotherapy against GBM, further solidifying the preclinical rationale to design clinical trials for patients with GBM.