<p>This study presents the development and validation of a stability-indicating UPLC-MS/MS method for the simultaneous determination of trifluridine, tipiracil hydrochloride and their impurities. The chromatographic separation was achieved on a Waters UPLC using the Acquity BEH Phenyl column (100&#xa0;mm ×2.1&#xa0;mm, 1.7&#xa0;µm) with a mobile phase comprising methanol and ammonium formate buffer (50:50, v/v) at a flow rate of 0.5&#xa0;mL/min. The detection was carried out at the wavelength of 257&#xa0;nm. The validation of the developed method adhered to ICH guidelines, evaluating parameters such as specificity, linearity, precision, accuracy, robustness and forced degradation. The method exhibited excellent linearity (<i>R</i>² &gt; 0.999), high precision (%RSD &lt; 1%) and recoveries ranging from 99.54% to 100.90%. The method was demonstrated to be robust against variations in flow rate and mobile phase composition, showing reproducibility across analysts. Forced degradation studies under acidic, alkaline, oxidative, thermal, photolytic and hydrolytic conditions confirmed the method’s stability-indicating capability. Degradation products were identified and characterized using LC–MS/MS, providing enhancing its application for stability testing, which is crucial for ensuring the quality, safety and efficacy of pharmaceutical &#xa0;formulations. The developed method has been reported to be reliable and suitable for routine analysis of trifluridine, tipiracil hydrochloride and their impurities/related substances in both bulk drug and dosage forms.</p>