Effective therapies for the treatment of obesity, a key element of metabolic syndrome, are urgently needed but currently lacking. Stearoyl-CoA desaturase-1 (SCD1) is the rate-limiting enzyme catalyzing the conversion of saturated long-chain fatty acids into monounsaturated fatty acids, which are major components of triglycerides. In the current study, we tested the efficacy of pharmacological inhibition of SCD1 in controlling lipogenesis and body weight in mice. SCD1-specific antisense oligonucleotide inhibitors (ASOs) reduced SCD1 expression, reduced fatty acid synthesis and secretion, and increased fatty acid oxidization in primary mouse hepatocytes. Treatment of mice with SCD1 ASOs resulted in prevention of diet-induced obesity with concomitant reductions in SCD1 expression and the ratio of oleate to stearoyl-CoA in tissues and plasma. These changes correlated with reduced body adiposity, hepatomegaly and steatosis, and postprandial plasma insulin and glucose levels. Furthermore, SCD1 ASOs reduced de novo fatty acid synthesis, decreased expression of lipogenic genes, and increased expression of genes promoting energy expenditure in liver and adipose tissues. Thus, SCD1 inhibition represents a new target for the treatment of obesity and related metabolic disorders.