The overall crash statistics in the EU still show a very significant number of car–cyclist crashes. Within the Horizon 2020 project Safe-Up, countermeasures have been developed to reduce this number. One of these countermeasures involves a V2X-enhanced on-board unit for cycles, which can provide on-time warning triggers. The research assumption was based on studying the benefits of connectivity in enhancing cyclists’ safety. This study assessed the performance of this potential technology both qualitatively by analyzing volunteer feedback during physical testing and quantitatively by virtual simulations. The volunteers’ study showed positive findings on system’s safety relevance, user experience, and user acceptance. The method applied for the virtual simulation is a prospective safety performance assessment with reconstructed accident scenarios based on the GIDAS database and cyclist behavior models, obtained from physical testing. The results using a warning trigger 4 s prior to the collision showed a potential safety benefit of approximately 98%. It should be noted that this trigger time was found to be quite early in both physical testing and virtual simulation. Further research is required to evaluate the system’s performance in more complex urban scenarios, as well as to design the human–machine interaction strategies for optimal accident avoidance.