Background We investigated the putative associations of alcohol, cigarette, e-cigarette, and marijuana use with kidney function and proteinuria among adolescents and young adults (AYA) with pediatric-onset chronic kidney disease (CKD) enrolled in the Chronic Kidney Disease in Children (CKiD) study. Methods Participants responded to questions about past year and 30-day substance use. Associations between each substance and kidney function, proteinuria, nephrotic range proteinuria, and high blood pressure were separately estimated using repeated measures regression models, adjusting for sociodemographic characteristics. Models controlled for covariates at the present visit (contemporaneous) and additionally controlled for disease severity at the year prior to reporting substance use (lagged). Results A total of 441 participants ≥16 years contributed 1,245 person-visits with 39% reporting alcohol and 16%, 17%, and 15% reporting cigarette, e-cigarette, and marijuana use, respectively, over the previous year. In adjusted lagged models, past year and 30-day cigarette use were significantly associated with higher levels of proteinuria (+18.6%, 95%CI: +2.8%, +36.9%; and +20.0%, 95%CI: +0.7%, +43.1%, respectively). Inferences were similar when controlling for secondhand smoke exposure. Conclusions In a cohort of AYA with pediatric kidney diseases, substance use was non-trivial, and cigarette use was associated with higher proteinuria, although the prevalence of use was low. Occasional alcohol, e-cigarette, and marijuana use were not associated with proteinuria, disease progression, or elevated blood pressure. Pediatric nephrologists as specialty care providers are well-positioned to discuss substance use and should encourage tobacco prevention/treatment efforts among AYA at high risk for use in order to preserve kidney function and promote well-being.