Conversion of active lesions to black holes has been associated with disability progression in subjects with relapsing-remitting multiple sclerosis (RRMS) and represents a complementary approach to evaluating clinical efficacy. The objective of this study was to assess the conversion of new active magnetic resonance imaging (MRI) lesions, identified 6 months after initiating treatment with glatiramer acetate 40 mg/mL three-times weekly (GA40) or placebo, to T1-hypointense black holes in subjects with RRMS. Subjects received GA40 (n = 943) or placebo (n = 461) for 12 months. MRI was obtained at baseline and Months 6 and 12. New lesions were defined as either gadolinium-enhancing T1 or new T2 lesions at Month 6 that were not present at baseline. The adjusted mean numbers of new active lesions at Month 6 converting to black holes at Month 12 were analyzed using a negative binomial model; adjusted proportions of new active lesions at Month 6 converting to black holes at Month 12 were analyzed using a logistic regression model. Of 1,292 subjects with complete MRI data, 433 (50.3 %) GA-treated and 247 (57.2 %) placebo-treated subjects developed new lesions at Month 6. Compared with placebo, GA40 significantly reduced the mean number (0.31 versus 0.45; P = .0258) and proportion (15.8 versus 19.6 %; P = .006) of new lesions converting to black holes. GA significantly reduced conversion of new active lesions to black holes, highlighting the ability of GA40 to prevent tissue damage in RRMS.