Background: Road-traffic injuries are a key cause of death and disability in low-income and middle-income countries, but the effect of city characteristics on road-traffic mortality is unknown in these countries. The aim of this study was to determine associations between city-level built environment factors and road-traffic mortality in large Latin American cities. Methods: We selected cities from Argentina, Brazil, Chile, Colombia, Costa Rica, El Salvador, Guatemala, Mexico, Panama, and Peru; cities included in the analysis had a population of at least 100 000 people. We extracted data for road-traffic deaths that occurred between 2010 and 2016 from country vital registries. Deaths were grouped by 5-year age groups and sex. Road-traffic deaths were identified using ICD-10 codes, with adjustments for ill-defined codes and incomplete registration. City-level measures included population, urban development, street design, public transportation, and social environment. Associations were estimated using multilevel negative binomial models with robust variances. Findings: 366 cities were included in the analysis. There were 328 408 road-traffic deaths in nearly 3·5 billion person-years across all countries, with an average crude rate of 17·1 deaths per 100 000 person-years. Nearly half of the people who died were younger than 35 years. In multivariable models, road-traffic mortality was higher in cities where urban development was more isolated (rate ratio [RR] 1·05 per 1 SD increase, 95% CI 1·02–1·09), but lower in cities with higher population density (0·94, 0·90–0·98), higher gross domestic product per capita (0·96, 0·94–0·98), and higher intersection density (0·92, 0·89–0·95). Cities with mass transit had lower road mortality rates than did those without (0·92, 0·86–0·99). Interpretation: Urban development policies that reduce isolated and disconnected urban development and that promote walkable street networks and public transport could be important strategies to reduce road-traffic deaths in Latin America and elsewhere. Funding: Wellcome Trust.