Background: Resumption of imatinib is one of the available therapeutic options for patients with gastrointestinal stromal tumors (GIST) refractory to tyrosine kinase inhibitors (TKIs). Intermittent dosing of imatinib was suggested to delay outgrow of the imatinib-resistant clones in a preclinical study. Methods: A randomized phase 2 study was performed to evaluate the efficacy and safety of continuous or intermittent schedule of imatinib in GIST patients whose disease had progressed to at least imatinib and sunitinib. Patients were randomly assigned (1:1) to the intermittent dosing group that received 400mg daily imatinib based on a 1 week-on and 1 week-off schedule or the continuous dosing group that received imatinib 400mg daily without drug holiday. Disease control rate (DCR) rate at 12 weeks was the primary endpoint. Plasma samples were collected for circulating tumor DNA analysis at baseline, at 3 months of continuous or intermittent imatinib treatment and at the time of progressive disease. Results: Fifty patients received at least one dose of imatinib and were included in the full analysis set. Most patients (n = 46) had received prior regorafenib. DCR rate at 12 weeks was 34.8% and 43.5% in the continuous and intermittent groups, respectively. Median progression-free survival was 1.68 months and 1.57 months in the continuous and intermittent groups, respectively. The frequency of any grade anorexia, dysphagia, diarrhea, decreased neutrophil was lower in the intermittent group, whereas that of abdominal pain was higher in the intermittent group. Two patients in the continuous imatinib group underwent dose modification, and there was no patient who discontinued the study treatment because of adverse events related to study treatment. Conclusions: We confirmed that imatinib re-challenge was effective in TKI-refractory GIST patients. Intermittent dosing schedule may also be a clinically feasible treatment option for imatinib resumption in TKI-refractory GIST patients. Clinical trial information: NCT02712112.