AbstractMultiple 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^−/dim). 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^high 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.