CD8+ T cell killing of tumor cells is suppressed by the tumor microenvironment. Inhibitory receptors, prominently PD-1, are key mediators of this suppression. To discover cellular defects triggered by tumor exposure and associated PD-1 signaling, we have established an ex vivo imaging approach to investigate CD8+ tumor infiltrating lymphocytes (TILs) interacting with tumor targets. Whilst TIL:tumor cell couples still formed, couple stability deteriorated within 1-2 minutes. This was associated with impaired F-actin clearing from the center of the cellular interface, reduced calcium signaling, increased TIL locomotion and impaired tumor cell killing. Interaction of CD8+ lymphocytes with tumor cell spheroids in vitro induced a similar phenotype, supporting a critical role of direct T cell tumor cell contact. Diminished engagement of PD-1 within the tumor, but not acute ex vivo blockade, partially restored cell couple maintenance and killing. PD-1 thus suppresses TIL function by inducing a polarization-impaired state.