Background: Tyrosine kinase inhibitors (TKIs) are the cornerstone treatment for advanced GIST via pharmacologic targeting of driver oncogenes such as KIT. Detection of KIT alterations through tissue-based next-generation sequencing (NGS) is common, but circulating tumor DNA (ctDNA)-based NGS is a less invasive alternative to identify driver and resistance mutations in advanced GIST. Patients (pts) with KIT-mutant GIST benefit from first-line (1L) imatinib; however, KIT resistance mutations may confer imatinib-resistance and differential sensitivity to subsequent TKIs. We sought to analyze ctDNA from GIST pts to determine whether certain resistance mutations were associated with superior outcomes with particular TKIs in the second-line and beyond (2L+). Methods: Under an approved institutional review board protocol, a retrospective analysis was performed with available ctDNA NGS results (Guardant360; Redwood City, CA) from pts (N = 104) who progressed on 1L imatinib between 2017-21. Using R statistical programming, we identified pts with primary KIT alterations (N = 64) and known resistance mutations in KIT exons 13 (N = 25) and 17 (N = 35). We studied the median time to treatment failure (mTTF), defined as the time from treatment start to treatment end (months) due to progressive disease or toxicity, for each 2L+ drug. Using Kaplan-Meier methods, we calculated Cox proportional-hazard ratios (HR) with confidence intervals (CI) and p-values for statistical significance. Results: 49% were male (median age 66; range, 31-94). Driver oncogenes were detected in 80% (N = 83), including KIT, NF1, PDGFRA and BRAF. Of those with a KIT alteration, 12 (19%) had KIT exon 9 mutations and 52 (81%) had KIT exon 11 mutations. KIT resistance mutations were observed in KIT exons 13 (N = 25; V654), 14 (N = 2; T670), and 17 (N = 45; D816, D820, N822, Y823). Pts with KIT resistance mutations received 2L+ therapy with avapritinib, dose-escalated imatinib, nilotinib, pazopanib, ponatinib, regorafenib, ripretinib, or sunitinib. mTTF for KIT exon 13 V654 pts treated with 2L+ sunitinib, imatinib 800mg, or other was 10.8, 7.5, and 3.7 months, respectively. TTF for sunitinib vs other 2L+ drugs showed a HR of 0.51 (CI 0.33-0.8), p = 0.003. mTTF for KIT exon 17 (non-V654) pts treated with 2L+ regorafenib, imatinib 800mg, or other was 4.6, 1.2, and 6.3 months, respectively. Comparison of mTTF for regorafenib vs other 2L+ drugs was not statistically significant. Conclusions: ctDNA is a noninvasive tool for detecting driver and resistance mutations in pts with advanced GIST. GIST pts with KIT exon 13 V654 resistance mutations had superior outcomes in the 2L+ setting with sunitinib. Regorafenib was not superior to other 2L+ TKIs in pts with KIT exon 17 resistance mutations, possibly due to their own activity against exon 17 resistance alterations. ctDNA-guided therapy warrants evaluation in a prospective clinical trial.