We have previously shown preferential tumor-homing and therapeutic efficacy of adoptively transferred type 1 CTL (Tc1) when compared with type 2 CTL (Tc2) in mice bearing intracranial ovalbumin-transfected melanoma (M05). Further characterizing the expression of a panel of homing receptors on Tc1 and Tc2 cells, we found that very late antigen (VLA)-4 (a heterodimer of CD49d and CD29), but none of other receptors evaluated, was expressed at significantly higher levels on Tc1 cells than on Tc2 cells. Although CD49d (alpha(4) integrin) can form heterodimers with both beta(1) (CD29) and beta(7) integrins, alpha(4)beta(7) complexes were not expressed by either Tc1 or Tc2 cells, suggesting that CD49d is solely expressed in VLA-4 complexes. VLA-4 expression on Tc2 cells was down-regulated in an interleukin (IL)-4 dose-dependent manner but not by other type 2 cytokines, such as IL-10 and IL-13, suggesting that IL-4 uniquely down-regulates VLA-4 expression on these cells. In accordance with the differential expression of VLA-4 on Tc1 versus Tc2 cells, Tc1 cells alone were competent to adhere to plate-bound VCAM-1-Ig fusion protein. Finally, the efficient trafficking of Tc1 cells into intracranial M05 lesions in vivo was efficiently blocked by administration of monoclonal antibodies against CD49d or VCAM-1 or small interfering RNA-mediated silencing of CD49d on Tc1 cells. Collectively, these data support the critical role of VLA-4 in the effective intracranial tumor homing of adoptive-transferred, antigen-specific Tc1 cells and suggest that more effective vaccine and/or ex vivo T-cell activation regimens may be developed by promoting the generation of VLA-4(+) antitumor Tc1 cells.