Objective The aim of this study was to determine the sex differences in lower extremity biomechanics of amateur runners with patellofemoral pain (PFP) and without PFP. Lower extremity kinematic, kinetic, and electromyographic data in running were compared between male and female amateur runners with and without PFP. Methods 15 male and 10 female amateur runners with PFP were identified and enrolled in the PFP group, 25 healthy amateur runners matched with the PFP group were recruited as the control group. Three-dimensional kinematics and kinetics, as well as electromyographic data in the running task, were collected. A 2 × 2 analysis of variance was performed to compare for each lower extremity biomechanics between groups and between sexes. Results Male participants with PFP showed a greater peak knee flexion angle during the landing phase of running when compared to the control group. Participants with PFP showed a greater peak hip adduction angle during the landing phase of running when compared to the control group for both males and females, while the peak hip adduction angle of female participants was significantly greater than that of male participants. Conclusion Amateur runners with PFP showed sex-specific lower extremity biomechanics in the running task, the clinical intervention for PFP, therefore, should be sex-specific.