BackgroundHNF1Ais an essential component of the transcription factor network that controls pancreatic β-cell differentiation, maintenance, and glucose stimulated insulin secretion (GSIS). A continuum of protein malfunction is caused by variations in theHNF1Agene, from severe loss-of-function (LOF) variants that cause the highly penetrant Maturity Onset Diabetes of the Young (MODY) to milder LOF variants that are far less penetrant but impart a population-wide risk of type 2 diabetes that is up to five times higher. Before classifying and reporting the discovered variations as relevant in clinical diagnosis, a critical review is required. Functional investigations offer substantial support for classifying a variant as pathogenic, or otherwise as advised by the American College of Medical Genetics and Genomics (ACMG) and the Association for Molecular Pathology (AMP) ACMG/AMP criteria for variant interpretation.ObjectiveTo determine the molecular basis for the variations in theHNF1Agene found in patients with monogenic diabetes in India.MethodsWe performed functional protein analyses such as transactivation, protein expression, DNA binding, nuclear localization, and glucose stimulated insulin secretion (GSIS) assay, along with structural prediction analysis for 14HNF1Avariants found in 20 patients with monogenic diabetes.ResultsOf the 14 variants, 4 (28.6%) were interpreted as pathogenic, 6 (42.8%) as likely pathogenic, 3 (21.4%) as variants of uncertain significance, and 1 (7.14%) as benign. Patients harboring the pathogenic/likely pathogenic variants were able to successfully switch from insulin to sulfonylureas (SU) making these variants clinically actionable.ConclusionOur findings are the first to show the need of using additive scores during molecular characterization for accurate pathogenicity evaluations ofHNF1Avariants in precision medicine.