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Significant gaps exist in current scholarship on the relationship between substance abuse and sexual/gender identity. Transgender populations, especially transgender women (TW), seem to be at greater risk of substance abuse as a result of stigmatization and discrimination, the pathologizing of their gender identity and families, and social and work exclusion. This cross-sectional study aimed to describe the factors associated with polysubstance use in TW in three cities in Colombia, South America. This cross-sectional study used Respondent-Driven Sampling (RDS) yielding 688 TW participants (M=32.9; 46.7% sex workers/webcam; 79% single, and mostly low-SES) from Bogotá, Medellín and Santiago de Cali. Descriptive analyses, bivariate associations, and a binomial regression were performed. Alcohol was the most used substance with 75% TW reporting at least one drinking day in the past six months, followed by marihuana 58.6%, cocaine 34.6%, and ecstasy or Popper 25.4%; while 60.5% TW consumed 2 or more substances combined. Factors associated with polysubstance included transgender discrimination (1,45; IC- 1,061-2,120), low-education (2,09; IC- 1,306-3,366), being a sex worker/webcam (4,051; IC- 2,543-6,455) and having sex before age 15 (1,585; IC- 1,109-2,265). Results will be discussed as they relate to prevention/intervention efforts and social policies aiming to reduce polysubstance use in TW in Colombia.