Hepatitis C virus (HCV) is a hepatotropic positive-sense RNA virus of the Flaviviridae virus family and is a major cause of chronic liver disease worldwide. Like all obligate parasites, HCV relies on host pathways to enable its pathogenesis. HCV, in particular, has a clear link with hepatic lipid metabolism, promoting a lipid-rich environment for its proliferation. This manifests as liver steatosis in many patients harboring chronic HCV infection. Based on our recent findings regarding an immunometabolic and HCV antiviral microRNA (miRNA), miRNA-185 targeting and down regulating serine hydrolases (SH) involved in lipid and endocannabinoid metabolism, here we investigate HCV and its dependency on certain metabolic serine hydrolases involved in lipid and endocannabinoid metabolism. Serine hydrolases are one of the largest and most diverse enzyme families. This enzyme family has emerged as a center of therapeutic potential due to its implications in many metabolic roles. Here, we demonstrate that pharmacological inhibition of metabolic serine hydrolases alpha-beta hydrolyzing domain 6 (ABHD6), carboxylesterase 1 (CES1), and monoacylglycerol lipase (MGLL), enzymes involved in the hydrolysis of the endogenous cannabinoid receptor 1 (CB1) agonist 2-arachidonoyl glycerol (2-AG) are potently antiviral against HCV. Serine hydrolase inhibition with the MGLL inhibitor MJN110 paired with endocannabinoid signaling antagonization led to additive antiviral effects against HCV and has revealed modulation of the viral pathogenic phenotype to be its key course of action. MGLL inhibitor MJN110 transcriptomic characterization revealed modulations in humoral immunity and phagocytosis and acts antiviraly against HCV independent of CB1 antagonization. This provides an avenue for future investigation, assessing the viability of CB1 antagonization, and MGLL as a key host targeted antiviral factor in affecting HCV viral life cycle.