Hyperinsulinism is commonly caused by genetic mutations and as a consequence of hypobaric surgery. Hyperinsulinism can lead to hypoglycemia, psychiatric disorders and insulin resistance, a cause of type two diabetes. An effective avenue for therapeutic treatment of hyperinsulinism is antagonism of the Glucagon-Like Peptide 1 Receptor (GLP1R). Previously, we identified a highly potent antagonist antibody, called TB-001-003, which was from our synthetic antibody libraries designed to target G protein-coupled receptors (GPCRs). Here, we designed a library to optimize the activity of TB-001-003 against GLP1R and performed phage-display on cells overexpressing GLP1R. We found many optimized antagonists, including partial agonists. One antagonist, called TB-222-023, is a β-arrestin biased inverse agonist. In vivo, TB-222-023 effectively decreased insulin secretion in mouse pancreatic islets and increased plasma glucose in a mouse model of hyperinsulinism. We show that targeting GLP1R with an antibody antagonist is an effective strategy for treatment of hyperinsulinism.