Multiple sclerosis (MS) is a common and devastating chronic inflammatory disease of the CNS. CD4 T cells are assumed to be the first to cross the blood-central nervous system (CNS) barrier and trigger local inflammation. Here, we explored how pathogenicity-associated effector programs define CD4 T cell subsets with brain-homing ability in MS. Runx3- and Eomes-, but not T-bet-expressing CD4 memory cells were diminished in the blood of MS patients. This decline reversed following natalizumab treatment and was supported by a Runx3 Eomes T-bet enrichment in cerebrospinal fluid samples of treatment-naïve MS patients. This transcription factor profile was associated with high granzyme K (GZMK) and CCR5 levels and was most prominent in Th17.1 cells (CCR6 CXCR3 CCR4 ). Previously published CD28 CD4 T cells were characterized by a Runx3 Eomes T-bet phenotype that coincided with intermediate CCR5 and a higher granzyme B (GZMB) and perforin expression, indicating the presence of two separate subsets. Under steady-state conditions, granzyme K Th17.1 cells spontaneously passed the blood-brain barrier in vitro. This was only found for other subsets including CD28 cells when using inflamed barriers. Altogether, CD4 T cells contain small fractions with separate pathogenic features, of which Th17.1 seems to breach the blood-brain barrier as a possible early event in MS.