Foot-and-mouth disease (FMD) is one of the most economically important viral diseases that can affect livestock. In the last 70 years, use of an inactivated whole antigen vaccine has contributed to the eradication of disease from many developed nations. However, recent outbreaks in Europe and Eastern Asia demonstrated that infection can spread as wildfire causing economic and social devastation. Therefore, it is essential to develop new control strategies that could confer early protection and rapidly stop disease spread. Live attenuated vaccines (LAV) are one of the best choices to obtain a strong early and long-lasting protection against viral diseases. In proof of concept studies, we previously demonstrated that "synonymous codon deoptimization" could be applied to the P1 capsid coding region of the viral genome to derive attenuated FMDV serotype A12 strains. Here, we demonstrate that a similar approach can be extended to the highly conserved non-structural P2 and P3 coding regions, providing a backbone for multiple serotype FMDV LAV development. Engineered codon deoptimized P2, P3 or P2, and P3 combined regions were included into the ACruzeiro infectious clone optimized for vaccine production, resulting in viable progeny that exhibited different degrees of attenuation in cell culture, in mice, and in the natural host (swine). Derived strains were thoroughly characterized and . Our work demonstrates that overall, the entire FMDV genome tolerates codon deoptimization, highlighting the potential of using this technology to derive novel improved LAV candidates.