ARTICLE ABSTRACTThe TGF-β signaling mediator SMAD4 is frequently mutated or deleted in colorectal and pancreatic cancers. SMAD4 acts as a tumor suppressor and its loss is associated with poorer patient outcomes. The purpose of this study was to find synthetic lethal interactions with SMAD4 deficiency in order to find novel therapeutic strategies for the treatment of patients with SMAD4-deficient colorectal or pancreatic cancers. Using pooled lentiviral sgRNA libraries, we conducted genome-wide loss-of-function screens in Cas9-expressing colorectal and pancreatic cancer cells harboring altered or wild-type SMAD4. The small GTPase protein RAB10 was identified and validated as a susceptibility gene in SMAD4-altered colorectal and pancreatic cancer cells. Rescue assays showed that RAB10 re-introduction reversed the anti-proliferative effects of RAB10 KO in SMAD4 negative cell lines. Further investigation is necessary to shed light on the mechanism by which RAB10 inhibition decreases cell proliferation of SMAD4-negative cells.