ABSTRACT A new, simple, rapid, precise, accurate and reproducible RP-HPLC method for estimation of Letermovir in bulk form and marketed formulation. Separation of Letermovir was successfully achieved on a Phenomenex Gemini ODS C18 (4.6mm x 250mm, 5mm) column in an isocratic mode of separation utilizing Methanol: Phosphate Buffer in the ratio of 25:75% v/v at a flow rate of 1.0 mL/min and the detection was carried out at 272nm. The method was validated according to ICH guidelines for linearity, sensitivity, accuracy, precision, specificity and robustness. The response was found to be linear in the drug concentration range of 10-50mcg/mL for Letermovir.The correlation coefficient was found to be 0.999 for Letermovir. The LOD and LOQ for Letermovir were found to be 1.1µg/mL and 3.2µg/mL respectively. The proposed method was found to be good percentage recovery for Letermovir, which indicates that the proposed method is highly accurate. The specificity of the method shows good correlation between retention times of standard solution with the sample solution. Therefore, the proposed method specifically determines the analyte in the sample without interference from excipients of pharmaceutical dosage forms. Keywords:Letermovir, RP-HPLC, Method Development, Accuracy, ICH Guidelines, Validation.