Emerging evidence has shown that long non-coding RNAs (lncRNAs) play crucial roles in human cancers. However, systematic characterization of lncRNAs and their roles in gastrointestinal stromal tumor (GIST) therapy have been lacking. We performed high-throughput RNA sequencing (RNA-seq) of 20 GIST and paired adjacent normal samples. We characterized the transcriptional landscape and dysregulation of lncRNAs in GIST. We identified 866 upregulated and 1,268 downregulated lncRNAs in GIST samples, the majority of which were GIST-specific over other cancer types. Most hallmarks were found to be dysregulated in GIST samples, and lncRNAs were highly associated with cancer-related hallmarks. RP11-616M22.7 was identified to increase in imatinib-resistant samples compared to those in non-resistant samples. Further analysis revealed that RP11-616M22.7 was closely associated with the Hippo signaling pathway. By treating GIST cells with different doses of imatinib, we verified that RP11-616M22.7 knockdown promotes the sensitivity of tumor cells, whereas RP11-616M22.7 overexpression induces resistance to imatinib. We further confirmed reducing of resistance to imatinib by knocking down RP11-616M22.7 in vivo. Additionally, RP11-616M22.7 was observed to interact with RASSF1 protein. Our study revealed that deficiency of RP11-616M22.7 was able to reduce resistance of the GIST cell response to imatinib treatment both in vitro and in vivo.