Background: ICIs are a common therapeutic option for many solid tumors. While prior studies have shown that ATB exposure may negatively impact ICI outcomes through gut microbiome changes, many were small studies with heterogeneity in ATB classes and exposure windows. Here, we performed a population level retrospective cohort study to evaluate the impact of ATB exposure prior to ICI on OS. Methods: We used administrative data to identify a cohort of cancer patients > 65 years of age receiving ICIs from June 2012 to October 2018 in Ontario, Canada and deterministically linked with databases to obtain socio-demographic and clinical co-variates and ATB prescription claims. Multivariable cox-proportional hazard models evaluated the impact of ATB exposure both within 1 year and 60 days prior to starting ICI on OS, adjusted for age, gender, body mass index, comorbidities, autoimmune history, hospitalization in the past year and treatment facility level at start of ICI therapy. Results: Among 2737 patients, median age 73; 43% received Nivolumab, 41% Pembrolizumab and 13% Ipilimumab; 53% were lung cancer, 34% melanoma. Median ATB treatment duration for patients receiving ATB within 1 year (59%) and 60 days (19%) prior to ICI were 14 days (SD = 32) and 9 days (SD = 13) respectively. Median OS estimate was 306 days. Any ATB exposure within 1 year prior to ICI was associated with worse OS (aHR = 1.12 95% CI [1.12-1.23] p = 0.03). A non significant dose effect was seen based on weeks of ATB exposure 1 year prior to ICI (aHR = 1.01 per week [1.00-1.02] p = 0.10). ATB class analysis identified fluoroquinolone exposure within 1 year (aHR = 1.26 [1.13-1.40] p < 0.001) and 60 days before ICI (aHR = 1.20 [0.99-1.45] p = 0.06) were associated with worse OS; with a dose effect based on total weeks of exposure over 1 year (aHR = 1.07 per week [1.03-1.11] p < 0.001) and 60 days (aHR = 1.12 per week [1.03-1.23] p = 0.01). Subgroup analysis showed similar results for patients receiving anti-PD1 ICIs, where patients exposed to fluoroquinolone both 1 year (aHR = 1.28 [1.15-1.44] p < 0.001) and 60 days (aHR = 1.19 [0.98-1.44] p = 0.08) before ICIs had poorer OS with dose effects observed based on weeks of fluoroquinolone exposure. Similarly, subgroup analyses based on disease site identified that lung cancer patients exposed to fluoroquinolones 1 year before starting ICIs (aHR = 1.22 [1.06-1.39] p = 0.005) and melanoma patients exposed to fluoroquinolones 60 days before starting ICIs (aHR = 1.66 [1.12-2.47] p = 0.01) had poorer OS. Conclusions: Exposure to ATBs and specifically fluoroquinolones prior to ICI therapy is associated with worse OS. Interventions aimed at altering the gut microbiome may be required to help improve outcomes for patients on ICIs with prior ATB exposure.