KRAS is the most frequently mutated oncogene in human cancer, but its therapeutic targeting remains challenging. Here, we report a synthetic lethal screen with a library of deubiquitinases and identify USP39, which encodes an essential splicing factor, as a critical gene for the viability of KRAS-dependent cells. We show that splicing fidelity inhibitors decrease preferentially the proliferation rate of KRAS-active cells. Moreover, depletion of DHX38, encoding an USP39-interacting splicing factor, also reduces the viability of these cells. In agreement with these results, USP39 depletion caused a significant reduction in pre-mRNA splicing efficiency, as demonstrated through RNA-seq experiments. Furthermore, we show that USP39 is up-regulated in lung and colon carcinomas and its expression correlates with KRAS levels and poor clinical outcome. Accordingly, our work provides critical information for the development of splicing-directed antitumor treatments and supports the potential of USP39-targeting strategies as the basis of new anticancer therapies.