Phorbol 12-myristate 13-acetate (PMA) stimulates the differentiation of promyelocytic leukemia HL-60 cells by inducing adhesion followed by cell aggregation and, importantly, apoptosis. p130Cas (Crk-associated substrate) is an adapter molecule that controls cell growth, attachment and apoptotic programs. Notably, elevated p130Cas activity is associated with leukemias and lymphomas. Since p130Cas regulates cell adhesion, we tested the hypothesis that it participates in the differentiation of hematopoietic cells. Here we show that PMA mediates the late induction of p130Cas expression in HL-60 cells, which coincided with cell aggregation and the onset of apoptosis. Ectopic p130Cas expression led to increased cell adhesion and earlier cell aggregation potentially contributing to the observed increased cell viability in these transductants. p130Cas expression concurred with the induction of its own regulator the transcription factor EGR1, its coregulator NAB2, and apoptosis. NF-κB inhibition in PMA-treated HL-60 cells promoted the loss of cell aggregation and cell death. We further showed a reduction of p130Cas, EGR1, and NAB2 levels in response to NF-κB inhibition during PMA treatment. Hence, p130Cas acts as survival factor by limiting PMA-mediated cell cluster disruption and resulting cell death in HL-60 cells.