Enterovirus-D68 (EV-D68) is a non-enveloped, positive-sense single-stranded RNA virus. Although EV-D68 infections usually cause mild cold-like symptoms in rare cases the virus can cause Acute Flaccid Myelitis (AFM) in children, a paralysis disease that has no known cure or therapeutics. Like many RNA viruses, EV-D68 utilizes the cellular autophagy pathway to promote its own replication. HMGB1 (High-mobility group B1) is a multifunctional DNA binding nuclear protein with established roles in autophagy regulation. Because of this, we posited a role for HMGB1 in EV-D68 production. We observed that when HMGB1 is knocked down in cells, those cells infected with EV-D68 show a significant decrease in both intraand extracellular viral titers, suggesting that HMGB1 is beneficial to viral replication. Its overexpression however, is associated with cancer. HMGB1 can bind to at least two known cell surface receptors: TLR4 (Toll-like receptor 4); and RAGE (Receptor for Advanced Glycation End products) both promising targets for cancer therapeutics. While we have been unable to show a specific role in virus replication for TLR4 binding by HMGB1, the RAGE inhibitor FPS-ZM1 caused a large decrease in intracellular titers and viral RNA replication. We have evidence that multiple steps in EV-D68 production are inhibited in the absence of functional RAGE signaling. Our data suggest multiple roles for HMGB1 and its surface receptor RAGE in EV-D68 replication.