Background and Purpose:This study aimed to differentially assess the frequency and severity of late radiation-induced toxicity following adjuvant whole-breast irradiation for early breast cancer with conventional fractionation (CF) and moderate hypofractionation (mHF).Materials and Methods:Patients recruited in a previous randomised controlled trial comparing acute toxicity between CF and mHF without disease recurrence were included in a post hoc analysis. Spectrophotometric and ultrasonographic examinations were performed for an objective evaluation and subsequent comparison of long-term skin toxicity. Furthermore, patient- and clinician-reported outcomes were recorded.Results:Sixty-four patients with a median age of 58 (37–81) years were included. The median follow-up was 57 (37–73) months. A total of 55% underwent CF and 45% mHF. A total of 52% received a sequential boost to the tumour bed. A significant decrease in mean L* (p= 0.011) and an increase in a* (p= 0.040) and b* values (p< 0.001) were observed, indicating hyperpigmentation. In comparison with the non-irradiated breast, there was a significant increase in both cutis (+14%;p< 0.001) and subcutis (+17%;p= 0.011) thickness, significantly more pronounced in CF patients (p= 0.049). In CF patients only, a sequential boost significantly increased the local cutis thickness and oedema compared to non-boost regions in the same breast (p= 0.001 andp< 0.001, respectively).Conclusions:mHF objectively resulted in reduced long-term skin toxicity compared to CF. A sequential boost increased the local fibrosis rate in CF, but not in mHF. This might explain the subjectively reported better cosmetic outcomes in patients receiving mHF and reinforces the rationale for favouring mHF as the standard of care.