Carbapenems are a powerful class of antibiotics, often used as a last-resort treatment to eradicate multidrug-resistant (MDR) infections. However, as carbapenem-resistant pathogens have become more common, the need for new strategies to combat resistance has emerged as a critical public health priority. Here, we show that the Facile Accelerated Specific Therapeutic (FAST) platform can use transcriptomic data for the rapid (<1 week) design, synthesis, and testing of antisense inhibitors that are able to re-sensitize a strain of MDR E. coli to carbapenem treatment. We used total and small RNA-seq data to determine genes that may be important to the meropenem (to which the strain is sensitive) and ertapenem (resistant) responses. These observations, combined with genome sequencing of the MDR E. coli, allowed us to use FAST to create peptide nucleic acids (PNA) to knock down the translation of specific genes. Using PNA in combination with both antibiotics, we demonstrated the ability to inhibit bacterial growth at antibiotic concentrations to which MDR E. coli strain was initially resistant. This work confirmed FAST’s ability to use transcriptomics to design novel antibiotic strategies, and identified gene expression-based resistance mechanisms that warrant further exploration.