Background: Abnormal insulin signaling in the brain has been linked to Alzheimer's disease (AD). Objective: To evaluate whether cerebrospinal fluid (CSF) insulin levels are associated with cognitive performance and CSF amyloid- and Tau. Additionally, we explore whether any such association differs by sex or APOE 4 genotype. Methods: From 258 individuals participating in the Parelsnoer Institute Neurodegenerative Diseases, a nationwide multicenter memory clinic population, we selected 138 individuals (mean age 669 years, 65.2% male) diagnosed with subjective cognitive impairment (n = 45), amnestic mild cognitive impairment (n = 44), or AD (n = 49), who completed a neuropsychological assessment, including tests of global cognition and memory performance, and who underwent lumbar puncture. We measured CSF levels of insulin, amyloid-1-42, total (t-)Tau, and phosphorylated (p-)Tau. CSF insulin levels did not differ between the diagnostic groups (p = 0.136). Across the whole study population, CSF insulin was unrelated to cognitive performance and CSF biomarkers of AD, after adjustment for age, sex, body mass index, diabetes status, and clinic site (all p=0.131). Importantly, however, we observed effect modification by sex and APOE 4 genotype. Specifically, among women, higher insulin levels in the CSF were associated with worse global cognition (standardized regression coefficient -0.483; p = 0.008) and higher p-Tau levels (0.353; p = 0.040). Among non-carriers of the APOE 4 allele, higher CSF insulin was associated with higher t-Tau (0.287; p = 0.008) and p-Tau (0.246; p = 0.029). Conclusion: Our findings provide further evidence for a relationship between brain insulin signaling and AD pathology. It also highlights the need to consider sex and APOE 4 genotype when assessing the role of insulin.