A heat treatment process was proposed to enhance the anti-scratch performance of nanostructured anti-reflection (AR) PMMA films fabricated by thermal nanoimprint lithography (t-NIL). Nanostructured AR films were fabricated using the high-temperature/high-pressure flat pressing (FP) to control free volume and rapid cooling (RC) to induce a compressive residual stress, which are compatible with t-NIL. The effects of the heat treatment process on the optical and the mechanical properties of the nanostructured films were examined by evaluating the optical properties, such as reflectance and transmittance, and measuring the mechanical properties with a pencil hardness test and a nanoindentation. It was found that the anti-scratch performance of the film was significantly improved while the optical properties are degraded slightly by a combination of FP and RC processes. It was further confirmed that the enhanced anti-scratch performance is due to the improved elastic modulus of the film by the optimized sequence of the process consisting of FP, RC and t-NIL. The enhanced antiscratch performance is attributed to the compressive residual stress on the surface and the minimization of the unevenly distributed free volume in the polymer film.