{"paper_id":"2cf1f772-fa8f-47da-ba88-0c0e222d909b","body_text":"Clinicopathologic and molecular spectrum of gastrointestinal stromal tumor (GIST) with NTRK fusion and marked response to Larotrectinib in GIST with NTRK fusion: a case report | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Clinicopathologic and molecular spectrum of gastrointestinal stromal tumor (GIST) with NTRK fusion and marked response to Larotrectinib in GIST with NTRK fusion: a case report Yanying Shen, Qi Peng, Jinxuan Wen, Linxi Yang, Lingyan Zhu, Yiming Chen, and 6 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7615382/v1 This work is licensed under a CC BY 4.0 License Status: Published Journal Publication published 19 Jan, 2026 Read the published version in Cellular Oncology → Version 1 posted 13 You are reading this latest preprint version Abstract Purpose This study aimed to characterize the clinicopathological, immunophenotypic, and molecular features of gastrointestinal stromal tumors (GISTs) harboring NTRK fusions and to evaluate their diagnostic, prognostic, and therapeutic implications. Methods Twenty-six cases of KIT/PDGFRA/SDH/BRAF wild-type GISTs were evaluated using pan-TRK immunohistochemistry (IHC), fluorescence in situ hybridization (FISH) for *NTRK1/2/3*, and next-generation sequencing (NGS). Transcriptome analysis was performed on all NTRK fusion-positive cases. Seven KIT -mutant GISTs served as controls. Clinicopathological parameters, IHC profiles, genetic alterations, and treatment responses were analyzed, supplemented by a literature review. Results Five of the 26 wild-type GISTs harbored NTRK fusions, all confirmed by NGS as ETV6::NTRK3 . Pan-TRK IHC showed 100% sensitivity and 66.7% specificity. All five patients were male; four tumors were intestinal and one gastric. Four cases were high-risk and one very low-risk. Two cases recurred post-resection, showing additional mutations and copy number variations (CNVs). Transcriptome analysis revealed molecular heterogeneity among NTRK fusion-positive GISTs, with profiles overlapping those of KIT -mutant GISTs. Both recurrent patients received multi-line TKI therapy (imatinib, sunitinib, regorafenib, ripretinib) with disease progression; one subsequently achieved remission with larotrectinib. Conclusion NTRK fusion-positive GISTs are rare and exhibit distinct clinicopathological characteristics. FISH and NGS are reliable detection methods, while pan-TRK IHC has limited specificity. Co-occurring genetic alterations may confer aggressive behavior. These tumors respond to TRK inhibition but are resistant to conventional TKIs, underscoring the need for molecularly guided therapy. gastrointestinal stromal tumor Pan-TRK NTRK3 fusion NTRK inhibitors Figures Figure 1 Figure 2 Figure 3 Figure 4 Introduction Gastrointestinal stromal tumor (GIST) is the most common mesenchymal neoplasm of the digestive system, accounting for 0.1% to 3% of all gastrointestinal malignancies. Its annual incidence is estimated at 1 to 2 cases per 100,000 people [ 1 ]. Genetically, GISTs are predominantly driven by mutations in the KIT (70%–85%) or PDGFRA (5%–15%) genes [ 1 ]. The advent of small-molecule targeted inhibitors, such as Imatinib, which selectively inhibit receptor tyrosine kinases, has markedly improved survival outcomes for patients with KIT or PDGFRA -mutant GISTs [ 2 , 3 ]. However, a subset of GISTs lacks mutations in both KIT and PDGFRA , classifying them as wild-type GISTs [ 4 – 6 ]. These patients present a clinical challenge as effective targeted therapies are currently lacking. In recent years, NTRK gene fusions have been identified in wild-type GISTs [ 7 ]. The NTRK family includes three genes— NTRK1 , NTRK2 , and NTRK3 —located on chromosomes 1q22, 9q21, and 15q25, respectively. These genes encode the highly homologous TrkA, TrkB, and TrkC receptor tyrosine kinases. Under physiological conditions, neurotrophic factors bind to these receptors, activating downstream signaling pathways such as RAS , which play critical roles in nervous system development and function [ 8 , 9 ]. However, NTRK gene fusions often lead to the expression of chimeric TRK proteins with constitutively active or overexpressed kinase activity, promoting oncogenesis. Such fusions have been implicated in a variety of cancers, including lung cancer, thyroid cancer, secretory breast cancer, and infantile fibrosarcoma [ 10 – 12 ]. Notably, TRK inhibitors such as Larotrectinib and Entrectinib have shown significant and durable antitumor efficacy in patients with NTRK fusion-positive tumors, including GISTs [ 13 , 14 ]. Therefore, detecting NTRK fusions in GISTs is essential for enabling targeted therapeutic interventions and improving patient prognosis. Despite the clinical significance of NTRK fusion-positive GISTs, reported cases remain scarce, with only 22 documented in the literature to date [ 7 , 14 – 22 ]. The clinicopathological characteristics, prognosis and treatment of this rare GIST subtype are not yet well defined. In this study, we reviewed all wild-type GIST cases from our institution over the past five years. Using immunohistochemistry, fluorescence in situ hybridization (FISH), and next-generation sequencing (NGS), we identified five cases harboring NTRK gene fusions. This study aims to further elucidate the clinicopathological and molecular features, diagnostic strategies, treatment options, and prognostic implications of this rare GIST subtype. Materials and Methods Patient Selection We reviewed pathological records from 1,273 patients diagnosed with GISTs between January 2019 and December 2023 at Renji Hospital and Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China. All cases were initially evaluated using immunohistochemical (IHC) staining for CD117 and DOG1, along with Sanger sequencing of KIT (exons 9, 11, 13, and 17) and PDGFRA (exons 12 and 18). Among these, 47 cases were initially classified as KIT/PDGFRA wild-type (WT) GISTs. Subsequent IHC analysis for SDHB and BRAF V600E led to the exclusion of 12 SDH -deficient and 9 BRAF -mutant GISTs. Thus, 26 cases of KIT/PDGFRA/SDH/BRAF wild-type GISTs were included for further evaluation. These cases underwent additional testing including IHC for TRK expression, fluorescence in situ hybridization (FISH) for NTRK1 , NTRK2 , and NTRK3 rearrangements, and next-generation sequencing (NGS). The screening workflow for GIST cases was summarized in Fig. 1 . Medical records and pathological slides were re-examined, and histologic features were independently evaluated by two pathologists. Clinical data were retrieved from an electronic medical database. Follow-up information was obtained via postoperative records and telephone interviews. Overall survival (OS) was defined as the interval from initial diagnosis to the last follow-up or death, and disease-free survival (DFS) as the period from surgery to tumor recurrence, metastasis, or last follow-up. Both OS and DFS were measured in months, with follow-up concluding in January 2025. This study was approved by the Ethics Committees of Renji Hospital and Ruijin Hospital, Shanghai Jiaotong University School of Medicine. IHC for TRK, SDHB, and BRAF V600E Expression All tissue specimens were fixed in 4% buffered formalin and embedded in paraffin. IHC was performed on 4-µm-thick formalin-fixed, paraffin-embedded (FFPE) sections using a BenchMark XT automated stainer (Roche, Basel, Switzerland). The primary antibody against pan-TRK (clone EPR17341; rabbit monoclonal; Abcam, Cambridge, MA, USA) was applied at a dilution of 1:100. Antigen retrieval was conducted at 95°C for 30 minutes, followed by incubation with the primary antibody at 37°C for 30 minutes. A known positive control and a negative control (with phosphate-buffered saline replacing the primary antibody) were included in each run. Pan-TRK positivity was defined as immunoreactivity (nuclear, cytoplasmic, or membranous) in ≥ 5% of tumor cells. Fluorescence In Situ Hybridization (FISH) FISH was performed on 3-µm FFPE sections using dual-color break-apart probes for NTRK1 , NTRK2 , and NTRK3 (LBP, Guangzhou, China), following the manufacturer’s instructions. After denaturation at 95°C for 5 minutes, hybridization was carried out at 37°C for 16 hours in a ThermoBrite automated hybridizer (Abbott, Chicago, IL, USA). Signal interpretation was performed under a fluorescence microscope (Leica DM2500, Germany) at 1000× magnification using appropriate filters. A positive result was defined by either of the following patterns in ≥ 30% of tumor cells: (1) a classic break-apart pattern, showing one fused signal plus separated 3′ (orange) and 5′ (green) signals (separation ≥ 2 signal diameters); or (2) an atypical pattern, exhibiting one fused signal and an isolated orange signal without a corresponding green signal. Next-Generation Target Sequencing Genomic DNA and total RNA were extracted from formalin-fixed paraffin-embedded (FFPE) tumor samples using the QIAamp DNA FFPE Tissue Kit (Qiagen) and the RNeasy DSP FFPE Kit (Qiagen), respectively, following the manufacturers’ protocols. A comprehensive genomic profiling was performed using a hybrid capture-based targeted sequencing panel covering 506 cancer-related genes at the DNA level and 201 genes at the RNA level (Geneseeq, Nanjing, China). Libraries were prepared according to the manufacturer's instructions and sequenced on an Illumina NextSeq 550 platform with 2×150 bp paired-end reads (Illumina, San Diego, CA, USA). For DNA sequencing, raw reads were aligned to the human reference genome (hg19) using BWA (v0.7.12). PCR duplicates were removed with Picard (v2.5.0), and base quality recalibration was performed using BaseRecalibrator from GATK (v3.1.1). Somatic variants and indels were detected using an in-house developed pipeline. Gene fusions were identified from RNA-seq data via the STAR aligner (v2.5.3) and STAR-Fusion (v0.8). A fusion event was considered valid if supported by ≥ 5 spanning reads. Transcriptome sequencing and data analysis RNA quantity was assessed using Qubit, with a minimum input requirement of 1 µg. Quality was evaluated with the Bioanalyzer 2100, and samples with an RNA Integrity Number (RIN) > 7 were retained. Library preparation was conducted using the KAPA Library Preparation Kit, which included RNA fragmentation, reverse transcription, end repair, A-tailing, adapter ligation, and PCR enrichment. Libraries were sequenced on the Illumina HiSeq platform with PE150 chemistry, yielding at least 30 million reads per sample. Raw sequencing data in BCL format were converted to FASTQ using bcl2fastq (v2.17.1.14). Quality control and adapter trimming were performed with Trimmomatic. Clean reads were aligned to the transcriptome using STAR (v2.5.3a), and gene expression quantification was carried out with RSEM (v1.3.0). Differential expression analysis was conducted using DESeq2 (v1.16.1) and edgeR (v3.18.1), with significance thresholds set at |fold change| >2 and adjusted p-value < 0.05. Functional enrichment analyses, including Gene Set Enrichment Analysis (GSEA), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses, were performed using the ClusterProfiler R package (v4.8.3). Results Screening of GISTs with NTRK Gene Fusions Pan-TRK immunohistochemistry (IHC) was performed on 26 cases of KIT/PDGFRA/SDH/BRAF wild-type GISTs. TRK expression was negative in 14 cases and positive in 12 cases. Among the positive cases, three exhibited perinuclear punctate staining (Fig. 2 , A2, C2- 4 , D2), two showed a mixed nuclear and perinuclear punctate pattern (Fig. 2 , B2- 4 , E2), and seven displayed weak to moderate cytoplasmic staining (Fig. 2 , F1). Two case presented weak and focal staining that was only detectable under high magnification (Fig. 2 , C2 and E2). Table 1 Characteristics of GIST patients harboring ETV6::NTRK3 fusions. Case 1 Case 2 Case 3 Case 4 Case 5 Age (years) 64 64 60 26 77 Sex Male Male Male Male Male Location Rectum Small intestine Small intestine Stomach Small intestine Size (cm) 1.5 6.5 8 12 10 Mitotic index (/50HPF) 0 2 2 > 10 > 10 Morphology Spindled Epithelioid Spindled Epithelioid Spindled Risk assessment VLR HR HR HR HR IHC (Pan-TRK) moderate perinuclear punctate staining moderate nuclei positive perinuclear punctate staining focal and weak perinuclear punctate staining moderate perinuclear punctate staining weak perinuclear punctate staining FISH (NTRK3) typical pattern atypical pattern typical pattern typical pattern typical pattern NTRK fusion ETV6::NTRK3 (E5::N14) ETV6::NTRK3 (E4::N14) ETV6::NTRK3 (E4::N15) ETV6::NTRK3 (E5::N14) ETV6::NTRK3 (E5::N15) concomitant mutation CDKN2C p.L65* ARID1A del TNFRSF14 del CDKN2A del CDKN2B del SETD2 p.V2223Tfs*22 ARID1A del TNFRSF14 del MDM2 amp DDIT3 amp CDK4 amp HMGA2 amp CNV analysis 1p del 1q and 7q amp 1p del 1q, 10q, 14q, 22q and 7p del 1p, 22q,4p, 4q, 9p and 9q del 11p amp MSI/TMB MSS/TMB:3.9 MSS/TMB:4.0 MSS/TMB:1.94 MSS/TMB:1.94 MSS/TMB:3.87 Surgery Yes Yes Yes Yes Yes Drugs No No IM IM, SU, Reg IM, SU, Rip TRK inhibitor No No No Lo No Relapse/Time(month) NED, 40 NED, 12 NED, 36 42 6 Death/Time(month) NED, 40 NED, 12 NED, 36 AWD, 63 AWD, 34 Abbreviation: VLR: Very low risk, HR: High risk, IHC: Immunohistochemistry, FISH: Fluorescence in Situ Hybridization, CNV: copy number variation, MSI: microsatellite instability, MSS: microsatellite stable, TMB: Tumor mutation burden, IM: Imatinib, SU: Sunitinib, Reg: Regorafenib, Rip: Ripretinib; Lo: Larotrectinib, NED: No evidence of disease, AWD: Alive with disease, del: deletion, amp: amplification; fs: frameshift Fluorescence in situ hybridization (FISH) was used to assess NTRK1, NTRK2, and NTRK3 fusions in all 26 cases. NTRK3 fusions were identified in five tumors, while no fusions were detected in NTRK1 or NTRK2. Among the NTRK3 fusion-positive cases, four displayed a classic break-apart signal pattern—one fused signal plus separated orange and green signals (Fig. 2 , A3, C5, D3)—and one exhibited an atypical pattern with one fused signal and a single orange signal (Fig. 2 , B5). All five cases with NTRK3 fusion were positive for pan-TRK IHC, showing either perinuclear punctate staining or a mixed nuclear and perinuclear pattern. The remaining seven pan-TRK-positive cases, which showed only weak to moderate cytoplasmic staining (Fig. 2 , F1–F4), were negative for NTRK fusions by FISH. Next-generation sequencing (NGS) analysis confirmed the presence of ETV6::NTRK3 fusions in all five FISH-positive cases, with ETV6 identified as the fusion partner. Additionally, NGS detected NF1 mutations in 11 of the 26 wild-type GISTs. Among the seven pan-TRK IHC-positive/FISH-negative cases, five harbored NF1 mutations, while the remaining two showed no definitive or clinically significant mutations. In summary, five cases of ETV6::NTRK3 fusion-positive GISTs were identified among the 26 wild-type tumors. Pan-TRK IHC demonstrated 100% sensitivity and 66.7% specificity for detecting NTRK fusion in this cohort. Clinicopathological Features of GISTs with ETV6::NTRK3 Fusion The clinicopathological characteristics of the five identified GIST patients harboring the ETV6::NTRK3 fusion were summarized in Table 1 . Histologic features, TRK expression patterns, and FISH results indicating NTRK fusion in these fusion-positive cases were presented in Fig. 2 , A–E. All five patients were male. Four were elderly, with ages of 60, 64, 64, and 77 years, respectively, while one patient was younger, aged 26 years. Each underwent curative surgical resection. Tumor sites included the small intestine (three cases), rectum (one case), and stomach (one case). Tumor sizes ranged from 1.5 to 12 cm in diameter. Mitotic counts were ≤ 5 per 50 high-power fields (HPF) in three cases, and exceeded 10/50 HPF in the remaining two. Morphologically, three tumors were of the spindle cell type (Fig. 2 , A1, C1, E1), and two exhibited epithelioid morphology (Fig. 2 , B1, D1). Based on the modified NIH risk classification, one tumor was categorized as very low risk and the other four as high risk for recurrence. No distinctive histological characteristics were specifically associated with NTRK fusion-positive GISTs. Genetic Features and Prognosis of GISTs with ETV6::NTRK3 Fusion The next-generation sequencing (NGS) results for the five ETV6::NTRK3 fusion-positive GISTs were summarized in Table 1 . Two cases harbored fusions between exons 1–5 of ETV6 and exons 14–19 of NTRK3. The remaining three cases exhibited the following fusion patterns: exons 1–4 of ETV6 with exons 15–19 of NTRK3; exons 1–4 of ETV6 with exons 14–19 of NTRK3; and exons 1–5 of ETV6 with exons 15–19 of NTRK3. In all cases, the kinase domain of NTRK3 remained intact. NGS analysis also revealed that all five tumors were microsatellite stable, with a tumor mutational burden (TMB) ranging from 1.25 to 4.0. Case 3 showed copy number deletions in ARID1A and TNFRSF14, along with a frameshift mutation in CDKN2C. Case 4 exhibited copy number deletions of CDKN2A and CDKN2B. Case 5 displayed more extensive genetic alterations, including a frameshift mutation in SETD2, copy number deletions of ARID1A and TNFRSF14, and amplifications of MDM2, CDK4, DDIT3, and HMGA2. No other clinically significant variants were detected in Cases 1 and 2. Arm-level chromosomal copy number variation (CNV) analysis across the five cases revealed recurrent deletions at 1p (60%), 22q (40%), 14q (20%), 4p (20%), 4q (20%), 9p (20%), 9q (20%), 1q (20%), 10q (20%), and 7p (20%), as well as amplifications at 11p (20%), 1q (20%), and 7q (20%). Overall, chromosomal deletions were more frequent than amplifications, particularly in Cases 4 and 5. The complete CNV profiles are summarized in Table 1 and Supplementary Table 1. Notably, Cases 1 and 2, which lacked additional mutations, did not receive adjuvant targeted therapy after complete resection. Both patients showed no recurrence during follow-up periods of 40 and 12 months, respectively. Case 3 received adjuvant imatinib (400 mg daily) and remained recurrence-free over 12 months of follow-up. CNV analysis indicated that these three fusion-positive GISTs had either no or minimal CNV alterations. In contrast, Cases 4 and 5, which carried more complex genomic profiles including additional mutations and CNVs, received adjuvant imatinib but experienced recurrence at 42 months and 6 months, respectively. Comparison of Transcriptome Profiles Between NTRK Fusion-Positive and KIT-Mutated GISTs We performed transcriptome analysis on five NTRK fusion-positive GISTs—to our knowledge for the first time—and compared their expression profiles with those of seven KIT-mutant GISTs (including three high-risk and four low-risk cases) that served as controls. The objective was to identify gene expression differences between NTRK fusion-driven GISTs and the more common KIT-mutant subtype. Contrary to expectations, principal component analysis (PCA) did not clearly segregate NTRK fusion-positive and KIT-mutant GISTs into two separate clusters. The five NTRK fusion positive GIST cases and seven KIT-mutant cases collectively formed three distinct groups in the PCA space (Fig. 3 ). Although NTRK-related genes showed marked upregulation in the fusion-positive group, our observations indicated that there could be considerable transcriptomic heterogeneity within these tumors. We further examined the clinicopathological characteristics of these three groups and conducted differential gene expression analyses. Group 1, located in the lower-right quadrant of the PCA plot, comprised three low-risk gastric GISTs with KIT mutations. Group 2, situated in the upper-left quadrant, included six intestinal GISTs (four high-risk and two low-risk). Group 3, clustered in the upper-right quadrant, consisted of three high-risk gastric GISTs. Among these, two carried KIT mutations with higher variant allele frequencies (VAF), suggestive of loss of heterozygosity (LOH), and one case harbored an NTRK fusion along with CDKN2A/CDKN2B deletion. Detailed clinicopathological information for all 12 GIST cases across these groups is provided in Supplementary Table 2. When compared to Group 1, the differentially expressed genes in Group 2 were significantly enriched in pathways related to circulatory system processes. In contrast, Group 3 exhibited prominent enrichment in pathways involved in the mitotic cell cycle process. Additionally, genes associated with interferon signaling were significantly upregulated in Group 1. Treatment with TRK Inhibitor in a GIST Patient Harboring NTRK Fusion In Case 4 (summarized in Table 1 ), tumor recurrence occurred 42 months after initial surgery while the patient was on Imatinib therapy, presenting as multiple masses near the gastric anastomosis. The patient underwent a second surgical procedure, followed by initiation of Sunitinib. However, a new subcapsular nodule (2.9 cm) appeared three months later, prompting a switch to Regorafenib. After 15 months, disease progression was noted with an increase in tumor size to 6.0 cm. Treatment was subsequently changed to Larotrectinib. Within three months of starting this agent, the subcapsular splenic lesion showed significant regression. Figure 4 illustrates the radiographic changes of the lesion throughout the course of these TKI treatments. Discussion Although NTRK gene fusions are exceptionally rare across all gastrointestinal stromal tumors (GISTs), their identification carries considerable clinical significance. A major diagnostic challenge lies in selecting an optimal screening strategy for NTRK fusions in wild-type (WT) GISTs—those lacking mutations in KIT, PDGFRA, RAS, and SDH. In this study, we identified the ETV6::NTRK3 fusion in five out of 26 WT GIST patients using next-generation sequencing (NGS), fluorescence in situ hybridization (FISH), and immunohistochemistry (IHC). Both NGS and FISH yielded consistent results in detecting NTRK fusions, confirming that either method was effective for identifying NTRK fusions in GIST. IHC, being more accessible, cost-effective, and rapid, serves as a practical initial screening tool in most clinical settings. The pan-TRK antibody used in IHC targeted the conserved C-terminal region common to all TRK proteins to detect TRK overexpression. Although all five NTRK3 fusion-positive cases in our study showed positive pan-TRK staining—exhibiting perinuclear punctate or nuclear patterns—with 100% sensitivity, reported sensitivity in the literature varies widely (0%–100%) [ 18 – 20 , 23 ]. It is noteworthy that in our study, two cases exhibited very weak and focal pan-TRK immunostaining, which posed a risk of misinterpretation. Accurate identification required examination under high magnification, underscoring the need for experienced pathologists in the evaluation. Additionally, the specificity of pan-TRK IHC remained limited. Our data indicated that pan-TRK immunoreactivity occured in a notable subset of KIT/PDGFRA wild-type GISTs, particularly those associated with neurofibromatosis type 1 (NF-1). This was consistent with reports by Hung et al. and Solomon et al., who observed reduced specificity of pan-TRK IHC in sarcomas—especially tumors with neural or smooth muscle differentiation—due to physiological TRK expression in these tissues [ 24 , 25 ]. Therefore, while pan-TRK IHC may be useful for initial screening, its reliability for definitively identifying NTRK fusions in WT GISTs is constrained by variable sensitivity and suboptimal specificity. All five patients with NTRK fusion-positive GIST in our cohort were male, with tumors primarily located in the small intestine, and most classified as high-risk. These findings imply that NTRK fusion-positive GISTs may possess distinct clinicopathological characteristics. Through a comprehensive PubMed review [ 7 , 14 – 22 ], we identified 22 previously reported cases of NTRK fusion-positive GISTs from 10 publications and two clinical studies. We summarized the clinicopathological, immunohistochemical, and genetic features of these 22 cases along with our five cases in Supplementary Table 3. Among the 27 total cases, clinicopathological data were available for 23. Of these, seven patients were ≤ 45 years old and 16 were older; 16 were male and seven female. The most common tumor site was the intestine (69.57%, 16/23), followed by the stomach (26.09%, 6/23), with one case (4.35%, 1/23) of unknown origin. Morphological data were available for 16 cases: seven exhibited spindle cell morphology, six had epithelioid morphology (including one with small round cell features), and three showed mixed morphology. Among 21 patients with documented disease extent, only one had metastatic disease at diagnosis; the rest had localized tumors. Based on modified NIH risk criteria, 16 patients were high-risk, one was at least intermediate-risk, one was low-risk, and two were very low-risk. These results suggested that NTRK fusion-positive GISTs tend to occur at a younger age, show male predominance, often exhibit epithelioid morphology, and arise primarily in the intestine. Importantly, most were high-risk, indicating that NTRK fusion-positive GISTs may follow a more aggressive clinical course compared to non-NTRK fusion-positive GISTs. We performed the first comprehensive analysis of co-occurring mutations and copy number variations (CNVs) in these five NTRK fusion-positive GISTs. Ident alterations included frameshift mutations in CDKN2C and SETD2, deletion of CDKN2A/2B, and amplification of MDM2, DDIT3, CDK4, and HMGA2. CNV analysis revealed recurrent deletions at 1p, 22q, 14q, 4p, 4q, 9p, 9q, 1q, 10q, and 7p, as well as amplifications at 11p, 1q, and 7q. Overall, chromosomal deletions were more frequent than amplifications. Similar CNV patterns have been reported in common KIT- or PDGFRA-mutated GISTs. Our findings also suggest that aggressive NTRK fusion-positive GISTs harbor more genomic aberrations, a trait commonly seen in advanced KIT/PDGFRA-mutant GISTs [ 26 – 30 ]. These shared molecular features imply that NTRK fusion GISTs may progress through mechanisms similar to those of conventional GIST subtypes. Therefore, established prognostic biomarkers—such as tumor site, size, mitotic rate, loss of chromosomes 1p and 22q, and cell cycle gene abnormalities—may also be relevant in NTRK fusion-positive GISTs. Initial analyses from transcriptome sequencing and gene expression profiling lead us to tentatively hypothesize that NTRK fusion-positive GISTs might form a molecularly heterogeneous group with a profile that bears resemblance to KIT-mutated GISTs. Using PCA, we classified our cohort (five NTRK-fusion positive and seven KIT-mutated GISTs) into three groups, each with distinct clinicopathological features. In a related study, Xie et al. classified common KIT- and PDGFRA-mutated GISTs into four molecular subtypes (C1–C4): C1 (genome-stable): mainly low/intermediate-risk gastric GISTs with KIT exon 11 mutations and good prognosis after surgery; C2 (CD8 + inflamed): primarily high-risk or metastatic intestinal GISTs, potentially responsive to TKIs plus immunotherapy; C3 (immune desert): mainly high-risk or metastatic gastric GISTs with KIT exon 11 mutations, unlikely to benefit from immunotherapy but potential candidates for CDK4/6 inhibitors and TKIs; C4 (PDGFRA-driven): all PDGFRA-mutated GISTs, suitable for Avapritinib [ 30 ]. Subtypes C1–C3 closely correspond to our groups 1–3. Our cohort did not include PDGFRA-mutated GISTs, so the C4 subtype was not represented. The five NTRK fusion-positive GISTs in our study fell into groups 2 and 3, aligning with Xie et al.’s C2 and C3 subtypes. The comparison of the molecular typing in our study with that in Xie et al. 's study is summarized in Supplementary Table 2. Whether the treatment strategies recommended for these subtypes apply to NTRK fusion GISTs merits further investigation. It is important to note that NTRK fusion-positive GISTs are exceedingly rare. Consequently, the small cohort size in this study, while valuable, limits the generalizability of our findings. Future studies with larger patient numbers are essential to validate the observations and hypotheses proposed here. NTRK fusion-positive GISTs are exceedingly rare, and reports of treatment with TRK inhibitors are even scarcer. Here, we presented an additional case of NTRK fusion-positive GIST that responded markedly to Larotrectinib. To date, eight patients with NTRK fusion GISTs have received TRK inhibitors, with detailed clinical data available for seven, all of whom responded positively [ 7 , 14 , 15 , 21 ]. However, due to the high cost and limited availability of TRK inhibitors, as well as insufficient awareness of NTRK fusions during treatment planning, some patients still receive Imatinib as first-line therapy—even in neoadjuvant or adjuvant settings. Previous case reports indicated that Imatinib failed to control tumor growth in NTRK fusion-positive GISTs when used preoperatively [ 7 ]. In this study, we reported two patients with NTRK fusion-positive GIST who experienced recurrence during adjuvant imatinib therapy after complete resection. These cumulative findings suggest that patients with NTRK fusion-positive GISTs are unlikely to benefit from Imatinib or other non-TRK TKIs but may achieve significant clinical responses with TRK inhibitor therapy. Declarations Human ethics and consent to participate This research involved the collection of FFPE sections from resection tissues. All related protocols were approved by the ethics committee of Renji Hospital (Ethics approval number: LY2024-264-B) and were carried out according to the Ethical Principles of the Declaration of Helsinki. All patients signed written informed consent forms. Competing interests The authors declare no competing interests. Funding This work was supported by the National Natural Science Foundation of China (No. 82070207 and 82300212). Author Contribution Y.S., A.L., Z.L., and Q.P., and J.W. participated in the literature search and in the writing. Y.S., L.Y., L.Z., and Y.C. participated in data analysis and visualization. Q.L., L.W., and J.Y. participated in data curation and methodology. A.L., M.W., and Z.L. supervised, coordinated the study, participated in the writing, and reviewed the final manuscript. All authors reviewed the manuscript. 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Supplementary Files Supplementarytable12025.09.16.xlsx Supplementarytable320250916.xlsx Supplementarytable220250916.xlsx Cite Share Download PDF Status: Published Journal Publication published 19 Jan, 2026 Read the published version in Cellular Oncology → Version 1 posted Editorial decision: Revision requested 23 Oct, 2025 Reviews received at journal 23 Oct, 2025 Reviewers agreed at journal 23 Oct, 2025 Reviews received at journal 14 Oct, 2025 Reviewers agreed at journal 14 Oct, 2025 Reviewers agreed at journal 09 Oct, 2025 Reviews received at journal 07 Oct, 2025 Reviewers agreed at journal 26 Sep, 2025 Reviewers agreed at journal 24 Sep, 2025 Reviewers invited by journal 17 Sep, 2025 Editor assigned by journal 17 Sep, 2025 Submission checks completed at journal 17 Sep, 2025 First submitted to journal 14 Sep, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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fusions.\\u003c/strong\\u003e\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage1.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7615382/v1/d51dcc462c0d2039fd205a2f.jpeg\"},{\"id\":92475972,\"identity\":\"8a904046-8222-4d26-a3d5-0509078ced4c\",\"added_by\":\"auto\",\"created_at\":\"2025-09-30 07:19:09\",\"extension\":\"jpeg\",\"order_by\":2,\"title\":\"Figure 2\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":902300,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eillustrated characteristic histological, immunohistochemical, and genetic features of six GIST cases, including five with \\u003cem\\u003eNTRK\\u003c/em\\u003e gene fusions (corresponding to Cases 1–5 in Table 1) and one with \\u003cem\\u003eNF1\\u003c/em\\u003e mutation. Histologically, spindle tumor cells were shown in Case 1 (A1), Case 3 (C1), and Case 5 (E1), while epithelioid morphology was observed in Case 2 (B1) and Case 4 (D1). Cases 4 and 5 (D1 and E1) displayed significant nuclear atypia and prominent mitotic figures (indicated by black arrows). Immunohistochemically, pan-TRK expression showed a perinuclear punctate pattern in Cases 1 (A2), 3 (C2–C4), and 4 (D2). Case 2 (B2) exhibited heterogeneous staining, with some tumor cells showing nuclear positivity and others showing perinuclear punctate expression. Regions B3 and B4 represented magnified views of B2. Case 5 (E2) demonstrated a mixed nuclear and perinuclear punctate pattern, similar to Case 2 (red arrow: nuclear staining; green arrow: perinuclear punctate). In Cases 3 (C2) and 5 (E2), pan-TRK staining was weak and focal in a subset of tumor cells, which could easily be misinterpreted as negative. Regions C3 and C4 represent magnified views of C2. Genetic analysis by FISH revealed \\u003cem\\u003eNTRK3\\u003c/em\\u003e fusion with a classic break-apart pattern (one fused and two separated orange/green signals) in Cases 1 (A3), 3 (C5), and 4 (D3). Case 2 (B5) showed an atypical pattern with one fused signal and a single orange signal. Notably, cytoplasmic pan-TRK immunoreactivity was also observed in the \\u003cem\\u003eNF1\\u003c/em\\u003e-mutant GIST (E1). No \\u003cem\\u003eNTRK1/2/3\\u003c/em\\u003efusions were detected in this case by FISH (E2–E4).\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage2.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7615382/v1/e52b76cd4462804be636c2f0.jpeg\"},{\"id\":92474335,\"identity\":\"fdf74149-80ed-452f-bb93-896aea772d10\",\"added_by\":\"auto\",\"created_at\":\"2025-09-30 07:11:09\",\"extension\":\"jpeg\",\"order_by\":3,\"title\":\"Figure 3\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":217693,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003eThe five \\u003cem\\u003eNTRK\\u003c/em\\u003efusion-positive and seven \\u003cem\\u003eKIT\\u003c/em\\u003e-mutant GIST cases were clustered into three distinct groups in the PCA plot, hinting at potential transcriptomic differences that might exist among these subtypes. Differentially expressed genes (DEGs) were identified between Group 1 and Group 2, as well as between Group 1 and Group 3. Subsequent functional enrichment analysis revealed upregulation of the IFN signaling pathway in Group 1, pathways associated with circulatory system processes in Group 2, and genes related to mitotic cell cycle process in Group 3. (A) Principal Component Analysis (PCA) of transcriptome data (B) DEGs between Group 1 and Group 2 (C) DEGs between Group 1 and Group 3.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage3.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7615382/v1/e94eab21aab94078d68d02ab.jpeg\"},{\"id\":92474340,\"identity\":\"337bfc2e-e479-4480-bb60-71e375a4f385\",\"added_by\":\"auto\",\"created_at\":\"2025-09-30 07:11:10\",\"extension\":\"jpeg\",\"order_by\":4,\"title\":\"Figure 4\",\"display\":\"\",\"copyAsset\":false,\"role\":\"figure\",\"size\":320617,\"visible\":true,\"origin\":\"\",\"legend\":\"\\u003cp\\u003e\\u003cstrong\\u003eCT Imaging of a Patient with \\u003c/strong\\u003e\\u003cem\\u003e\\u003cstrong\\u003eNTRK\\u003c/strong\\u003e\\u003c/em\\u003e\\u003cstrong\\u003eFusion-Positive GIST During Sequential TKI Therapy.\\u003c/strong\\u003e Serial CT images demonstrated the development of a new subcapsular splenic nodule (arrows in B, C, and D) during postoperative adjuvant treatment with Sunitinib, Regorafenib, and Larotrectinib. \\u003cstrong\\u003e(A) \\u003c/strong\\u003eBaseline scan after initial surgery. \\u003cstrong\\u003e(B)\\u003c/strong\\u003e After 3 months of Sunitinib treatment. \\u003cstrong\\u003e(C)\\u003c/strong\\u003e After 15 months of Regorafenib treatment. \\u003cstrong\\u003e(D)\\u003c/strong\\u003e After 3 months of Larotrectinib treatment, showing significant reduction of the lesion.\\u003c/p\\u003e\",\"description\":\"\",\"filename\":\"floatimage4.jpeg\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7615382/v1/0cc28c82a4df3cd1709cbb39.jpeg\"},{\"id\":101151852,\"identity\":\"987e3f3d-05d5-494a-9d0c-21e3c150f131\",\"added_by\":\"auto\",\"created_at\":\"2026-01-26 16:06:54\",\"extension\":\"pdf\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"manuscript-pdf\",\"size\":2703092,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"manuscript.pdf\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7615382/v1/7711ed74-6d6a-4e42-80f7-73e5237d7900.pdf\"},{\"id\":92474349,\"identity\":\"38eb1747-513f-424e-9d0d-1ce33310e540\",\"added_by\":\"auto\",\"created_at\":\"2025-09-30 07:11:10\",\"extension\":\"xlsx\",\"order_by\":0,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"supplement\",\"size\":11021,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Supplementarytable12025.09.16.xlsx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7615382/v1/628dbe5140723d2f25f1280e.xlsx\"},{\"id\":92474334,\"identity\":\"63460052-fe86-458c-bb62-abc48350fb2e\",\"added_by\":\"auto\",\"created_at\":\"2025-09-30 07:11:09\",\"extension\":\"xlsx\",\"order_by\":1,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"supplement\",\"size\":20428,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Supplementarytable320250916.xlsx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7615382/v1/0d236add39a97631abbe9413.xlsx\"},{\"id\":92474343,\"identity\":\"5046a5e1-de31-46dd-96ed-e24027d1509b\",\"added_by\":\"auto\",\"created_at\":\"2025-09-30 07:11:10\",\"extension\":\"xlsx\",\"order_by\":2,\"title\":\"\",\"display\":\"\",\"copyAsset\":false,\"role\":\"supplement\",\"size\":12313,\"visible\":true,\"origin\":\"\",\"legend\":\"\",\"description\":\"\",\"filename\":\"Supplementarytable220250916.xlsx\",\"url\":\"https://assets-eu.researchsquare.com/files/rs-7615382/v1/e7fb99fb7c8951ae041de636.xlsx\"}],\"financialInterests\":\"No competing interests reported.\",\"formattedTitle\":\"Clinicopathologic and molecular spectrum of gastrointestinal stromal tumor (GIST) with NTRK fusion and marked response to Larotrectinib in GIST with NTRK fusion: a case report\",\"fulltext\":[{\"header\":\"Introduction\",\"content\":\"\\u003cp\\u003eGastrointestinal stromal tumor (GIST) is the most common mesenchymal neoplasm of the digestive system, accounting for 0.1% to 3% of all gastrointestinal malignancies. Its annual incidence is estimated at 1 to 2 cases per 100,000 people [\\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e1\\u003c/span\\u003e]. Genetically, GISTs are predominantly driven by mutations in the \\u003cem\\u003eKIT\\u003c/em\\u003e (70%\\u0026ndash;85%) or \\u003cem\\u003ePDGFRA\\u003c/em\\u003e (5%\\u0026ndash;15%) genes [\\u003cspan citationid=\\\"CR1\\\" class=\\\"CitationRef\\\"\\u003e1\\u003c/span\\u003e]. The advent of small-molecule targeted inhibitors, such as Imatinib, which selectively inhibit receptor tyrosine kinases, has markedly improved survival outcomes for patients with \\u003cem\\u003eKIT\\u003c/em\\u003e or \\u003cem\\u003ePDGFRA\\u003c/em\\u003e-mutant GISTs [\\u003cspan citationid=\\\"CR2\\\" class=\\\"CitationRef\\\"\\u003e2\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR3\\\" class=\\\"CitationRef\\\"\\u003e3\\u003c/span\\u003e]. However, a subset of GISTs lacks mutations in both \\u003cem\\u003eKIT\\u003c/em\\u003e and \\u003cem\\u003ePDGFRA\\u003c/em\\u003e, classifying them as wild-type GISTs [\\u003cspan additionalcitationids=\\\"CR5\\\" citationid=\\\"CR4\\\" class=\\\"CitationRef\\\"\\u003e4\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR6\\\" class=\\\"CitationRef\\\"\\u003e6\\u003c/span\\u003e]. These patients present a clinical challenge as effective targeted therapies are currently lacking.\\u003c/p\\u003e\\u003cp\\u003eIn recent years, \\u003cem\\u003eNTRK\\u003c/em\\u003e gene fusions have been identified in wild-type GISTs [\\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e7\\u003c/span\\u003e]. The NTRK family includes three genes\\u0026mdash;\\u003cem\\u003eNTRK1\\u003c/em\\u003e, \\u003cem\\u003eNTRK2\\u003c/em\\u003e, and \\u003cem\\u003eNTRK3\\u003c/em\\u003e\\u0026mdash;located on chromosomes 1q22, 9q21, and 15q25, respectively. These genes encode the highly homologous TrkA, TrkB, and TrkC receptor tyrosine kinases. Under physiological conditions, neurotrophic factors bind to these receptors, activating downstream signaling pathways such as \\u003cem\\u003eRAS\\u003c/em\\u003e, which play critical roles in nervous system development and function [\\u003cspan citationid=\\\"CR8\\\" class=\\\"CitationRef\\\"\\u003e8\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR9\\\" class=\\\"CitationRef\\\"\\u003e9\\u003c/span\\u003e]. However, \\u003cem\\u003eNTRK\\u003c/em\\u003e gene fusions often lead to the expression of chimeric TRK proteins with constitutively active or overexpressed kinase activity, promoting oncogenesis. Such fusions have been implicated in a variety of cancers, including lung cancer, thyroid cancer, secretory breast cancer, and infantile fibrosarcoma [\\u003cspan additionalcitationids=\\\"CR11\\\" citationid=\\\"CR10\\\" class=\\\"CitationRef\\\"\\u003e10\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR12\\\" class=\\\"CitationRef\\\"\\u003e12\\u003c/span\\u003e]. Notably, TRK inhibitors such as Larotrectinib and Entrectinib have shown significant and durable antitumor efficacy in patients with \\u003cem\\u003eNTRK\\u003c/em\\u003e fusion-positive tumors, including GISTs [\\u003cspan citationid=\\\"CR13\\\" class=\\\"CitationRef\\\"\\u003e13\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR14\\\" class=\\\"CitationRef\\\"\\u003e14\\u003c/span\\u003e]. Therefore, detecting \\u003cem\\u003eNTRK\\u003c/em\\u003e fusions in GISTs is essential for enabling targeted therapeutic interventions and improving patient prognosis.\\u003c/p\\u003e\\u003cp\\u003eDespite the clinical significance of \\u003cem\\u003eNTRK\\u003c/em\\u003e fusion-positive GISTs, reported cases remain scarce, with only 22 documented in the literature to date [\\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e7\\u003c/span\\u003e, \\u003cspan additionalcitationids=\\\"CR15 CR16 CR17 CR18 CR19 CR20 CR21\\\" citationid=\\\"CR14\\\" class=\\\"CitationRef\\\"\\u003e14\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR22\\\" class=\\\"CitationRef\\\"\\u003e22\\u003c/span\\u003e]. The clinicopathological characteristics, prognosis and treatment of this rare GIST subtype are not yet well defined. In this study, we reviewed all wild-type GIST cases from our institution over the past five years. Using immunohistochemistry, fluorescence in situ hybridization (FISH), and next-generation sequencing (NGS), we identified five cases harboring \\u003cem\\u003eNTRK\\u003c/em\\u003e gene fusions. This study aims to further elucidate the clinicopathological and molecular features, diagnostic strategies, treatment options, and prognostic implications of this rare GIST subtype.\\u003c/p\\u003e\"},{\"header\":\"Materials and Methods\",\"content\":\"\\u003cdiv id=\\\"Sec3\\\" class=\\\"Section2\\\"\\u003e\\u003ch2\\u003ePatient Selection\\u003c/h2\\u003e\\u003cp\\u003eWe reviewed pathological records from 1,273 patients diagnosed with GISTs between January 2019 and December 2023 at Renji Hospital and Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, China. All cases were initially evaluated using immunohistochemical (IHC) staining for CD117 and DOG1, along with Sanger sequencing of \\u003cem\\u003eKIT\\u003c/em\\u003e (exons 9, 11, 13, and 17) and \\u003cem\\u003ePDGFRA\\u003c/em\\u003e (exons 12 and 18). Among these, 47 cases were initially classified as \\u003cem\\u003eKIT/PDGFRA\\u003c/em\\u003e wild-type (WT) GISTs. Subsequent IHC analysis for SDHB and BRAF V600E led to the exclusion of 12 \\u003cem\\u003eSDH\\u003c/em\\u003e-deficient and 9 \\u003cem\\u003eBRAF\\u003c/em\\u003e-mutant GISTs. Thus, 26 cases of \\u003cem\\u003eKIT/PDGFRA/SDH/BRAF\\u003c/em\\u003e wild-type GISTs were included for further evaluation. These cases underwent additional testing including IHC for TRK expression, fluorescence in situ hybridization (FISH) for \\u003cem\\u003eNTRK1\\u003c/em\\u003e, \\u003cem\\u003eNTRK2\\u003c/em\\u003e, and \\u003cem\\u003eNTRK3\\u003c/em\\u003e rearrangements, and next-generation sequencing (NGS).\\u003c/p\\u003e\\u003cp\\u003eThe screening workflow for GIST cases was summarized in Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e. Medical records and pathological slides were re-examined, and histologic features were independently evaluated by two pathologists. Clinical data were retrieved from an electronic medical database. Follow-up information was obtained via postoperative records and telephone interviews. Overall survival (OS) was defined as the interval from initial diagnosis to the last follow-up or death, and disease-free survival (DFS) as the period from surgery to tumor recurrence, metastasis, or last follow-up. Both OS and DFS were measured in months, with follow-up concluding in January 2025. This study was approved by the Ethics Committees of Renji Hospital and Ruijin Hospital, Shanghai Jiaotong University School of Medicine.\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003c/div\\u003e\\n\\u003ch3\\u003eIHC for TRK, SDHB, and BRAF V600E Expression\\u003c/h3\\u003e\\n\\u003cp\\u003eAll tissue specimens were fixed in 4% buffered formalin and embedded in paraffin. IHC was performed on 4-\\u0026micro;m-thick formalin-fixed, paraffin-embedded (FFPE) sections using a BenchMark XT automated stainer (Roche, Basel, Switzerland). The primary antibody against pan-TRK (clone EPR17341; rabbit monoclonal; Abcam, Cambridge, MA, USA) was applied at a dilution of 1:100. Antigen retrieval was conducted at 95\\u0026deg;C for 30 minutes, followed by incubation with the primary antibody at 37\\u0026deg;C for 30 minutes. A known positive control and a negative control (with phosphate-buffered saline replacing the primary antibody) were included in each run. Pan-TRK positivity was defined as immunoreactivity (nuclear, cytoplasmic, or membranous) in \\u0026ge;\\u0026thinsp;5% of tumor cells.\\u003c/p\\u003e\\n\\u003ch3\\u003eFluorescence In Situ Hybridization (FISH)\\u003c/h3\\u003e\\n\\u003cp\\u003eFISH was performed on 3-\\u0026micro;m FFPE sections using dual-color break-apart probes for \\u003cem\\u003eNTRK1\\u003c/em\\u003e, \\u003cem\\u003eNTRK2\\u003c/em\\u003e, and \\u003cem\\u003eNTRK3\\u003c/em\\u003e (LBP, Guangzhou, China), following the manufacturer\\u0026rsquo;s instructions. After denaturation at 95\\u0026deg;C for 5 minutes, hybridization was carried out at 37\\u0026deg;C for 16 hours in a ThermoBrite automated hybridizer (Abbott, Chicago, IL, USA). Signal interpretation was performed under a fluorescence microscope (Leica DM2500, Germany) at 1000\\u0026times; magnification using appropriate filters. A positive result was defined by either of the following patterns in \\u0026ge;\\u0026thinsp;30% of tumor cells: (1) a classic break-apart pattern, showing one fused signal plus separated 3\\u0026prime; (orange) and 5\\u0026prime; (green) signals (separation\\u0026thinsp;\\u0026ge;\\u0026thinsp;2 signal diameters); or (2) an atypical pattern, exhibiting one fused signal and an isolated orange signal without a corresponding green signal.\\u003c/p\\u003e\\n\\u003ch3\\u003eNext-Generation Target Sequencing\\u003c/h3\\u003e\\n\\u003cp\\u003eGenomic DNA and total RNA were extracted from formalin-fixed paraffin-embedded (FFPE) tumor samples using the QIAamp DNA FFPE Tissue Kit (Qiagen) and the RNeasy DSP FFPE Kit (Qiagen), respectively, following the manufacturers\\u0026rsquo; protocols. A comprehensive genomic profiling was performed using a hybrid capture-based targeted sequencing panel covering 506 cancer-related genes at the DNA level and 201 genes at the RNA level (Geneseeq, Nanjing, China). Libraries were prepared according to the manufacturer's instructions and sequenced on an Illumina NextSeq 550 platform with 2\\u0026times;150 bp paired-end reads (Illumina, San Diego, CA, USA).\\u003c/p\\u003e\\u003cp\\u003eFor DNA sequencing, raw reads were aligned to the human reference genome (hg19) using BWA (v0.7.12). PCR duplicates were removed with Picard (v2.5.0), and base quality recalibration was performed using BaseRecalibrator from GATK (v3.1.1). Somatic variants and indels were detected using an in-house developed pipeline. Gene fusions were identified from RNA-seq data via the STAR aligner (v2.5.3) and STAR-Fusion (v0.8). A fusion event was considered valid if supported by \\u0026ge;\\u0026thinsp;5 spanning reads.\\u003c/p\\u003e\\n\\u003ch3\\u003eTranscriptome sequencing and data analysis\\u003c/h3\\u003e\\n\\u003cp\\u003eRNA quantity was assessed using Qubit, with a minimum input requirement of 1 \\u0026micro;g. Quality was evaluated with the Bioanalyzer 2100, and samples with an RNA Integrity Number (RIN)\\u0026thinsp;\\u0026gt;\\u0026thinsp;7 were retained. Library preparation was conducted using the KAPA Library Preparation Kit, which included RNA fragmentation, reverse transcription, end repair, A-tailing, adapter ligation, and PCR enrichment.\\u003c/p\\u003e\\u003cp\\u003eLibraries were sequenced on the Illumina HiSeq platform with PE150 chemistry, yielding at least 30\\u0026nbsp;million reads per sample. Raw sequencing data in BCL format were converted to FASTQ using bcl2fastq (v2.17.1.14). Quality control and adapter trimming were performed with Trimmomatic. Clean reads were aligned to the transcriptome using STAR (v2.5.3a), and gene expression quantification was carried out with RSEM (v1.3.0). Differential expression analysis was conducted using DESeq2 (v1.16.1) and edgeR (v3.18.1), with significance thresholds set at |fold change| \\u0026gt;2 and adjusted p-value\\u0026thinsp;\\u0026lt;\\u0026thinsp;0.05. Functional enrichment analyses, including Gene Set Enrichment Analysis (GSEA), Gene Ontology (GO), and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses, were performed using the ClusterProfiler R package (v4.8.3).\\u003c/p\\u003e\"},{\"header\":\"Results\",\"content\":\"\\u003cdiv id=\\\"Sec9\\\" class=\\\"Section2\\\"\\u003e\\u003ch2\\u003eScreening of GISTs with NTRK Gene Fusions\\u003c/h2\\u003e\\u003cp\\u003ePan-TRK immunohistochemistry (IHC) was performed on 26 cases of KIT/PDGFRA/SDH/BRAF wild-type GISTs. TRK expression was negative in 14 cases and positive in 12 cases. Among the positive cases, three exhibited perinuclear punctate staining (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, A2, C2-\\u003cspan refid=\\\"Fig4\\\" class=\\\"InternalRef\\\"\\u003e4\\u003c/span\\u003e, D2), two showed a mixed nuclear and perinuclear punctate pattern (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, B2-\\u003cspan refid=\\\"Fig4\\\" class=\\\"InternalRef\\\"\\u003e4\\u003c/span\\u003e, E2), and seven displayed weak to moderate cytoplasmic staining (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, F1). Two case presented weak and focal staining that was only detectable under high magnification (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, C2 and E2).\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003cp\\u003e\\u003cdiv class=\\\"gridtable\\\"\\u003e\\u003ctable float=\\\"Yes\\\" id=\\\"Tab1\\\" border=\\\"1\\\"\\u003e\\u003ccaption language=\\\"En\\\"\\u003e\\u003cdiv class=\\\"CaptionNumber\\\"\\u003eTable 1\\u003c/div\\u003e\\u003cdiv class=\\\"CaptionContent\\\"\\u003e\\u003cp\\u003eCharacteristics of GIST patients harboring \\u003cem\\u003eETV6::NTRK3\\u003c/em\\u003e fusions.\\u003c/p\\u003e\\u003c/div\\u003e\\u003c/caption\\u003e\\u003ccolgroup cols=\\\"6\\\"\\u003e\\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c1\\\" colnum=\\\"1\\\"\\u003e\\u003c/div\\u003e\\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c2\\\" colnum=\\\"2\\\"\\u003e\\u003c/div\\u003e\\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c3\\\" colnum=\\\"3\\\"\\u003e\\u003c/div\\u003e\\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c4\\\" colnum=\\\"4\\\"\\u003e\\u003c/div\\u003e\\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c5\\\" colnum=\\\"5\\\"\\u003e\\u003c/div\\u003e\\u003cdiv align=\\\"left\\\" class=\\\"colspec\\\" colname=\\\"c6\\\" colnum=\\\"6\\\"\\u003e\\u003c/div\\u003e\\u003cthead\\u003e\\u003ctr\\u003e\\u003cth align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u0026nbsp;\\u003c/th\\u003e\\u003cth align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eCase 1\\u003c/p\\u003e\\u003c/th\\u003e\\u003cth align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eCase 2\\u003c/p\\u003e\\u003c/th\\u003e\\u003cth align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eCase \\u003cspan refid=\\\"FPar3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e\\u003c/p\\u003e\\u003c/th\\u003e\\u003cth align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eCase 4\\u003c/p\\u003e\\u003c/th\\u003e\\u003cth align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eCase 5\\u003c/p\\u003e\\u003c/th\\u003e\\u003c/tr\\u003e\\u003c/thead\\u003e\\u003ctbody\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eAge (years)\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003e64\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003e64\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003e60\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003e26\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003e77\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eSex\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eMale\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eMale\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eMale\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eMale\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eMale\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eLocation\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eRectum\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eSmall intestine\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eSmall intestine\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eStomach\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eSmall intestine\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eSize (cm)\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003e1.5\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003e6.5\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003e8\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003e12\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003e10\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eMitotic index (/50HPF)\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003e0\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003e2\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003e2\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003e\\u0026gt;\\u0026thinsp;10\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003e\\u0026gt;\\u0026thinsp;10\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eMorphology\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eSpindled\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eEpithelioid\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eSpindled\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eEpithelioid\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eSpindled\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eRisk assessment\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eVLR\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eHR\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eHR\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eHR\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eHR\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eIHC (Pan-TRK)\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003emoderate perinuclear punctate staining\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003emoderate\\u003c/p\\u003e\\u003cp\\u003enuclei positive perinuclear punctate staining\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003efocal and weak perinuclear punctate staining\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003emoderate perinuclear punctate staining\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eweak\\u003c/p\\u003e\\u003cp\\u003eperinuclear punctate staining\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eFISH (NTRK3)\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003etypical pattern\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eatypical pattern\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003etypical pattern\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003etypical pattern\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003etypical pattern\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eNTRK fusion\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eETV6::NTRK3\\u003c/em\\u003e\\u003c/p\\u003e\\u003cp\\u003e(E5::N14)\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eETV6::NTRK3\\u003c/em\\u003e\\u003c/p\\u003e\\u003cp\\u003e(E4::N14)\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eETV6::NTRK3\\u003c/em\\u003e\\u003c/p\\u003e\\u003cp\\u003e(E4::N15)\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eETV6::NTRK3\\u003c/em\\u003e\\u003c/p\\u003e\\u003cp\\u003e(E5::N14)\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eETV6::NTRK3\\u003c/em\\u003e\\u003c/p\\u003e\\u003cp\\u003e(E5::N15)\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003econcomitant mutation\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eCDKN2C\\u003c/em\\u003e p.L65*\\u003c/p\\u003e\\u003cp\\u003e\\u003cem\\u003eARID1A\\u003c/em\\u003e del \\u003cem\\u003eTNFRSF14\\u003c/em\\u003e del\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eCDKN2A\\u003c/em\\u003e del\\u003c/p\\u003e\\u003cp\\u003e\\u003cem\\u003eCDKN2B\\u003c/em\\u003e del\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eSETD2\\u003c/em\\u003e p.V2223Tfs*22\\u003c/p\\u003e\\u003cp\\u003e\\u003cem\\u003eARID1A\\u003c/em\\u003e del \\u003cem\\u003eTNFRSF14\\u003c/em\\u003e del\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003e\\u003cem\\u003eMDM2\\u003c/em\\u003e amp\\u003c/p\\u003e\\u003cp\\u003e\\u003cem\\u003eDDIT3\\u003c/em\\u003e amp\\u003c/p\\u003e\\u003cp\\u003e\\u003cem\\u003eCDK4\\u003c/em\\u003e amp\\u003c/p\\u003e\\u003cp\\u003e\\u003cem\\u003eHMGA2\\u003c/em\\u003e amp\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eCNV analysis\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u0026nbsp;\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003e1p del\\u003c/p\\u003e\\u003cp\\u003e1q and 7q amp\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003e1p del\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003e1q, 10q, 14q, 22q and 7p del\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003e1p, 22q,4p, 4q, 9p and 9q del\\u003c/p\\u003e\\u003cp\\u003e11p amp\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eMSI/TMB\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eMSS/TMB:3.9\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eMSS/TMB:4.0\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eMSS/TMB:1.94\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eMSS/TMB:1.94\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eMSS/TMB:3.87\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eSurgery\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eYes\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eYes\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eYes\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eYes\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eYes\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eDrugs\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eNo\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eNo\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eIM\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eIM, SU, Reg\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eIM, SU, Rip\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eTRK inhibitor\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eNo\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eNo\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eNo\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eLo\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eNo\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eRelapse/Time(month)\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eNED, 40\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eNED, 12\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eNED, 36\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003e42\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003e6\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003ctr\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c1\\\"\\u003e\\u003cp\\u003e\\u003cb\\u003eDeath/Time(month)\\u003c/b\\u003e\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c2\\\"\\u003e\\u003cp\\u003eNED, 40\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c3\\\"\\u003e\\u003cp\\u003eNED, 12\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c4\\\"\\u003e\\u003cp\\u003eNED, 36\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c5\\\"\\u003e\\u003cp\\u003eAWD, 63\\u003c/p\\u003e\\u003c/td\\u003e\\u003ctd align=\\\"left\\\" colname=\\\"c6\\\"\\u003e\\u003cp\\u003eAWD, 34\\u003c/p\\u003e\\u003c/td\\u003e\\u003c/tr\\u003e\\u003c/tbody\\u003e\\u003c/colgroup\\u003e\\u003ctfoot\\u003e\\u003ctr\\u003e\\u003ctd colspan=\\\"6\\\"\\u003eAbbreviation: VLR: Very low risk, HR: High risk, IHC: Immunohistochemistry, FISH: Fluorescence in Situ Hybridization, CNV: copy number variation, MSI: microsatellite instability, MSS: microsatellite stable, TMB: Tumor mutation burden, IM: Imatinib, SU: Sunitinib, Reg: Regorafenib, Rip: Ripretinib; Lo: Larotrectinib, NED: No evidence of disease, AWD: Alive with disease, del: deletion, amp: amplification; fs: frameshift\\u003c/td\\u003e\\u003c/tr\\u003e\\u003c/tfoot\\u003e\\u003c/table\\u003e\\u003c/div\\u003e\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003cp\\u003e\\u003c/p\\u003e\\u003cp\\u003eFluorescence in situ hybridization (FISH) was used to assess NTRK1, NTRK2, and NTRK3 fusions in all 26 cases. NTRK3 fusions were identified in five tumors, while no fusions were detected in NTRK1 or NTRK2. Among the NTRK3 fusion-positive cases, four displayed a classic break-apart signal pattern\\u0026mdash;one fused signal plus separated orange and green signals (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, A3, C5, D3)\\u0026mdash;and one exhibited an atypical pattern with one fused signal and a single orange signal (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, B5).\\u003c/p\\u003e\\u003cp\\u003eAll five cases with NTRK3 fusion were positive for pan-TRK IHC, showing either perinuclear punctate staining or a mixed nuclear and perinuclear pattern. The remaining seven pan-TRK-positive cases, which showed only weak to moderate cytoplasmic staining (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, F1\\u0026ndash;F4), were negative for NTRK fusions by FISH.\\u003c/p\\u003e\\u003cp\\u003eNext-generation sequencing (NGS) analysis confirmed the presence of ETV6::NTRK3 fusions in all five FISH-positive cases, with ETV6 identified as the fusion partner. Additionally, NGS detected NF1 mutations in 11 of the 26 wild-type GISTs. Among the seven pan-TRK IHC-positive/FISH-negative cases, five harbored NF1 mutations, while the remaining two showed no definitive or clinically significant mutations.\\u003c/p\\u003e\\u003cp\\u003eIn summary, five cases of ETV6::NTRK3 fusion-positive GISTs were identified among the 26 wild-type tumors. Pan-TRK IHC demonstrated 100% sensitivity and 66.7% specificity for detecting NTRK fusion in this cohort.\\u003c/p\\u003e\\u003cp\\u003e\\u003cb\\u003eClinicopathological Features of GISTs with\\u003c/b\\u003e \\u003cb\\u003eETV6::NTRK3\\u003c/b\\u003e \\u003cb\\u003eFusion\\u003c/b\\u003e\\u003c/p\\u003e\\u003cp\\u003eThe clinicopathological characteristics of the five identified GIST patients harboring the ETV6::NTRK3 fusion were summarized in Table\\u0026nbsp;\\u003cspan refid=\\\"Tab1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e. Histologic features, TRK expression patterns, and FISH results indicating NTRK fusion in these fusion-positive cases were presented in Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, A\\u0026ndash;E.\\u003c/p\\u003e\\u003cp\\u003eAll five patients were male. Four were elderly, with ages of 60, 64, 64, and 77 years, respectively, while one patient was younger, aged 26 years. Each underwent curative surgical resection. Tumor sites included the small intestine (three cases), rectum (one case), and stomach (one case). Tumor sizes ranged from 1.5 to 12 cm in diameter. Mitotic counts were \\u0026le;\\u0026thinsp;5 per 50 high-power fields (HPF) in three cases, and exceeded 10/50 HPF in the remaining two. Morphologically, three tumors were of the spindle cell type (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, A1, C1, E1), and two exhibited epithelioid morphology (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig2\\\" class=\\\"InternalRef\\\"\\u003e2\\u003c/span\\u003e, B1, D1). Based on the modified NIH risk classification, one tumor was categorized as very low risk and the other four as high risk for recurrence. No distinctive histological characteristics were specifically associated with NTRK fusion-positive GISTs.\\u003c/p\\u003e\\u003cp\\u003e\\u003cb\\u003eGenetic Features and Prognosis of GISTs with\\u003c/b\\u003e \\u003cb\\u003eETV6::NTRK3\\u003c/b\\u003e \\u003cb\\u003eFusion\\u003c/b\\u003e\\u003c/p\\u003e\\u003cp\\u003eThe next-generation sequencing (NGS) results for the five ETV6::NTRK3 fusion-positive GISTs were summarized in Table\\u0026nbsp;\\u003cspan refid=\\\"Tab1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e. Two cases harbored fusions between exons 1\\u0026ndash;5 of ETV6 and exons 14\\u0026ndash;19 of NTRK3. The remaining three cases exhibited the following fusion patterns: exons 1\\u0026ndash;4 of ETV6 with exons 15\\u0026ndash;19 of NTRK3; exons 1\\u0026ndash;4 of ETV6 with exons 14\\u0026ndash;19 of NTRK3; and exons 1\\u0026ndash;5 of ETV6 with exons 15\\u0026ndash;19 of NTRK3. In all cases, the kinase domain of NTRK3 remained intact. NGS analysis also revealed that all five tumors were microsatellite stable, with a tumor mutational burden (TMB) ranging from 1.25 to 4.0.\\u003c/p\\u003e\\u003cp\\u003e\\u003cstrong\\u003eCase 3\\u003c/strong\\u003e\\u003cp\\u003eshowed copy number deletions in ARID1A and TNFRSF14, along with a frameshift mutation in CDKN2C. Case 4 exhibited copy number deletions of CDKN2A and CDKN2B. Case 5 displayed more extensive genetic alterations, including a frameshift mutation in SETD2, copy number deletions of ARID1A and TNFRSF14, and amplifications of MDM2, CDK4, DDIT3, and HMGA2. No other clinically significant variants were detected in Cases 1 and 2.\\u003c/p\\u003e\\u003c/p\\u003e\\u003cp\\u003eArm-level chromosomal copy number variation (CNV) analysis across the five cases revealed recurrent deletions at 1p (60%), 22q (40%), 14q (20%), 4p (20%), 4q (20%), 9p (20%), 9q (20%), 1q (20%), 10q (20%), and 7p (20%), as well as amplifications at 11p (20%), 1q (20%), and 7q (20%). Overall, chromosomal deletions were more frequent than amplifications, particularly in Cases 4 and 5. The complete CNV profiles are summarized in Table\\u0026nbsp;\\u003cspan refid=\\\"Tab1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e and Supplementary Table\\u0026nbsp;1.\\u003c/p\\u003e\\u003cp\\u003eNotably, Cases 1 and 2, which lacked additional mutations, did not receive adjuvant targeted therapy after complete resection. Both patients showed no recurrence during follow-up periods of 40 and 12 months, respectively. Case \\u003cspan refid=\\\"FPar3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e received adjuvant imatinib (400 mg daily) and remained recurrence-free over 12 months of follow-up. CNV analysis indicated that these three fusion-positive GISTs had either no or minimal CNV alterations. In contrast, Cases 4 and 5, which carried more complex genomic profiles including additional mutations and CNVs, received adjuvant imatinib but experienced recurrence at 42 months and 6 months, respectively.\\u003c/p\\u003e\\u003cp\\u003e\\u003cb\\u003eComparison of Transcriptome Profiles Between\\u003c/b\\u003e \\u003cb\\u003eNTRK\\u003c/b\\u003e \\u003cb\\u003eFusion-Positive and KIT-Mutated GISTs\\u003c/b\\u003e\\u003c/p\\u003e\\u003cp\\u003eWe performed transcriptome analysis on five NTRK fusion-positive GISTs\\u0026mdash;to our knowledge for the first time\\u0026mdash;and compared their expression profiles with those of seven KIT-mutant GISTs (including three high-risk and four low-risk cases) that served as controls. The objective was to identify gene expression differences between NTRK fusion-driven GISTs and the more common KIT-mutant subtype.\\u003c/p\\u003e\\u003cp\\u003eContrary to expectations, principal component analysis (PCA) did not clearly segregate NTRK fusion-positive and KIT-mutant GISTs into two separate clusters. The five NTRK fusion positive GIST cases and seven KIT-mutant cases collectively formed three distinct groups in the PCA space (Fig.\\u0026nbsp;\\u003cspan refid=\\\"Fig3\\\" class=\\\"InternalRef\\\"\\u003e3\\u003c/span\\u003e). Although NTRK-related genes showed marked upregulation in the fusion-positive group, our observations indicated that there could be considerable transcriptomic heterogeneity within these tumors.\\u003c/p\\u003e\\u003cp\\u003eWe further examined the clinicopathological characteristics of these three groups and conducted differential gene expression analyses. Group 1, located in the lower-right quadrant of the PCA plot, comprised three low-risk gastric GISTs with KIT mutations. Group 2, situated in the upper-left quadrant, included six intestinal GISTs (four high-risk and two low-risk). Group 3, clustered in the upper-right quadrant, consisted of three high-risk gastric GISTs. Among these, two carried KIT mutations with higher variant allele frequencies (VAF), suggestive of loss of heterozygosity (LOH), and one case harbored an NTRK fusion along with CDKN2A/CDKN2B deletion. Detailed clinicopathological information for all 12 GIST cases across these groups is provided in Supplementary Table\\u0026nbsp;2.\\u003c/p\\u003e\\u003cp\\u003eWhen compared to Group 1, the differentially expressed genes in Group 2 were significantly enriched in pathways related to circulatory system processes. In contrast, Group 3 exhibited prominent enrichment in pathways involved in the mitotic cell cycle process. Additionally, genes associated with interferon signaling were significantly upregulated in Group 1.\\u003c/p\\u003e\\u003c/div\\u003e\\n\\u003ch3\\u003eTreatment with TRK Inhibitor in a GIST Patient Harboring NTRK Fusion\\u003c/h3\\u003e\\n\\u003cp\\u003eIn Case 4 (summarized in Table\\u0026nbsp;\\u003cspan refid=\\\"Tab1\\\" class=\\\"InternalRef\\\"\\u003e1\\u003c/span\\u003e), tumor recurrence occurred 42 months after initial surgery while the patient was on Imatinib therapy, presenting as multiple masses near the gastric anastomosis. The patient underwent a second surgical procedure, followed by initiation of Sunitinib. However, a new subcapsular nodule (2.9 cm) appeared three months later, prompting a switch to Regorafenib. After 15 months, disease progression was noted with an increase in tumor size to 6.0 cm. Treatment was subsequently changed to Larotrectinib. Within three months of starting this agent, the subcapsular splenic lesion showed significant regression. Figure\\u0026nbsp;\\u003cspan refid=\\\"Fig4\\\" class=\\\"InternalRef\\\"\\u003e4\\u003c/span\\u003e illustrates the radiographic changes of the lesion throughout the course of these TKI treatments.\\u003c/p\\u003e\"},{\"header\":\"Discussion\",\"content\":\"\\u003cp\\u003eAlthough NTRK gene fusions are exceptionally rare across all gastrointestinal stromal tumors (GISTs), their identification carries considerable clinical significance. A major diagnostic challenge lies in selecting an optimal screening strategy for NTRK fusions in wild-type (WT) GISTs\\u0026mdash;those lacking mutations in KIT, PDGFRA, RAS, and SDH. In this study, we identified the ETV6::NTRK3 fusion in five out of 26 WT GIST patients using next-generation sequencing (NGS), fluorescence in situ hybridization (FISH), and immunohistochemistry (IHC). Both NGS and FISH yielded consistent results in detecting NTRK fusions, confirming that either method was effective for identifying NTRK fusions in GIST. IHC, being more accessible, cost-effective, and rapid, serves as a practical initial screening tool in most clinical settings. The pan-TRK antibody used in IHC targeted the conserved C-terminal region common to all TRK proteins to detect TRK overexpression. Although all five NTRK3 fusion-positive cases in our study showed positive pan-TRK staining\\u0026mdash;exhibiting perinuclear punctate or nuclear patterns\\u0026mdash;with 100% sensitivity, reported sensitivity in the literature varies widely (0%\\u0026ndash;100%) [\\u003cspan additionalcitationids=\\\"CR19\\\" citationid=\\\"CR18\\\" class=\\\"CitationRef\\\"\\u003e18\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR20\\\" class=\\\"CitationRef\\\"\\u003e20\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR23\\\" class=\\\"CitationRef\\\"\\u003e23\\u003c/span\\u003e]. It is noteworthy that in our study, two cases exhibited very weak and focal pan-TRK immunostaining, which posed a risk of misinterpretation. Accurate identification required examination under high magnification, underscoring the need for experienced pathologists in the evaluation. Additionally, the specificity of pan-TRK IHC remained limited. Our data indicated that pan-TRK immunoreactivity occured in a notable subset of KIT/PDGFRA wild-type GISTs, particularly those associated with neurofibromatosis type 1 (NF-1). This was consistent with reports by Hung et al. and Solomon et al., who observed reduced specificity of pan-TRK IHC in sarcomas\\u0026mdash;especially tumors with neural or smooth muscle differentiation\\u0026mdash;due to physiological TRK expression in these tissues [\\u003cspan citationid=\\\"CR24\\\" class=\\\"CitationRef\\\"\\u003e24\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR25\\\" class=\\\"CitationRef\\\"\\u003e25\\u003c/span\\u003e]. Therefore, while pan-TRK IHC may be useful for initial screening, its reliability for definitively identifying NTRK fusions in WT GISTs is constrained by variable sensitivity and suboptimal specificity.\\u003c/p\\u003e\\u003cp\\u003eAll five patients with NTRK fusion-positive GIST in our cohort were male, with tumors primarily located in the small intestine, and most classified as high-risk. These findings imply that NTRK fusion-positive GISTs may possess distinct clinicopathological characteristics. Through a comprehensive PubMed review [\\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e7\\u003c/span\\u003e, \\u003cspan additionalcitationids=\\\"CR15 CR16 CR17 CR18 CR19 CR20 CR21\\\" citationid=\\\"CR14\\\" class=\\\"CitationRef\\\"\\u003e14\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR22\\\" class=\\\"CitationRef\\\"\\u003e22\\u003c/span\\u003e], we identified 22 previously reported cases of NTRK fusion-positive GISTs from 10 publications and two clinical studies. We summarized the clinicopathological, immunohistochemical, and genetic features of these 22 cases along with our five cases in Supplementary Table\\u0026nbsp;3. Among the 27 total cases, clinicopathological data were available for 23. Of these, seven patients were \\u0026le;\\u0026thinsp;45 years old and 16 were older; 16 were male and seven female. The most common tumor site was the intestine (69.57%, 16/23), followed by the stomach (26.09%, 6/23), with one case (4.35%, 1/23) of unknown origin. Morphological data were available for 16 cases: seven exhibited spindle cell morphology, six had epithelioid morphology (including one with small round cell features), and three showed mixed morphology. Among 21 patients with documented disease extent, only one had metastatic disease at diagnosis; the rest had localized tumors. Based on modified NIH risk criteria, 16 patients were high-risk, one was at least intermediate-risk, one was low-risk, and two were very low-risk. These results suggested that NTRK fusion-positive GISTs tend to occur at a younger age, show male predominance, often exhibit epithelioid morphology, and arise primarily in the intestine. Importantly, most were high-risk, indicating that NTRK fusion-positive GISTs may follow a more aggressive clinical course compared to non-NTRK fusion-positive GISTs.\\u003c/p\\u003e\\u003cp\\u003eWe performed the first comprehensive analysis of co-occurring mutations and copy number variations (CNVs) in these five NTRK fusion-positive GISTs. Ident alterations included frameshift mutations in CDKN2C and SETD2, deletion of CDKN2A/2B, and amplification of MDM2, DDIT3, CDK4, and HMGA2. CNV analysis revealed recurrent deletions at 1p, 22q, 14q, 4p, 4q, 9p, 9q, 1q, 10q, and 7p, as well as amplifications at 11p, 1q, and 7q. Overall, chromosomal deletions were more frequent than amplifications. Similar CNV patterns have been reported in common KIT- or PDGFRA-mutated GISTs. Our findings also suggest that aggressive NTRK fusion-positive GISTs harbor more genomic aberrations, a trait commonly seen in advanced KIT/PDGFRA-mutant GISTs [\\u003cspan additionalcitationids=\\\"CR27 CR28 CR29\\\" citationid=\\\"CR26\\\" class=\\\"CitationRef\\\"\\u003e26\\u003c/span\\u003e\\u0026ndash;\\u003cspan citationid=\\\"CR30\\\" class=\\\"CitationRef\\\"\\u003e30\\u003c/span\\u003e]. These shared molecular features imply that NTRK fusion GISTs may progress through mechanisms similar to those of conventional GIST subtypes. Therefore, established prognostic biomarkers\\u0026mdash;such as tumor site, size, mitotic rate, loss of chromosomes 1p and 22q, and cell cycle gene abnormalities\\u0026mdash;may also be relevant in NTRK fusion-positive GISTs.\\u003c/p\\u003e\\u003cp\\u003eInitial analyses from transcriptome sequencing and gene expression profiling lead us to tentatively hypothesize that NTRK fusion-positive GISTs might form a molecularly heterogeneous group with a profile that bears resemblance to KIT-mutated GISTs. Using PCA, we classified our cohort (five NTRK-fusion positive and seven KIT-mutated GISTs) into three groups, each with distinct clinicopathological features. In a related study, Xie et al. classified common KIT- and PDGFRA-mutated GISTs into four molecular subtypes (C1\\u0026ndash;C4): C1 (genome-stable): mainly low/intermediate-risk gastric GISTs with KIT exon 11 mutations and good prognosis after surgery; C2 (CD8\\u0026thinsp;+\\u0026thinsp;inflamed): primarily high-risk or metastatic intestinal GISTs, potentially responsive to TKIs plus immunotherapy; C3 (immune desert): mainly high-risk or metastatic gastric GISTs with KIT exon 11 mutations, unlikely to benefit from immunotherapy but potential candidates for CDK4/6 inhibitors and TKIs; C4 (PDGFRA-driven): all PDGFRA-mutated GISTs, suitable for Avapritinib [\\u003cspan citationid=\\\"CR30\\\" class=\\\"CitationRef\\\"\\u003e30\\u003c/span\\u003e]. Subtypes C1\\u0026ndash;C3 closely correspond to our groups 1\\u0026ndash;3. Our cohort did not include PDGFRA-mutated GISTs, so the C4 subtype was not represented. The five NTRK fusion-positive GISTs in our study fell into groups 2 and 3, aligning with Xie et al.\\u0026rsquo;s C2 and C3 subtypes. The comparison of the molecular typing in our study with that in Xie et al. 's study is summarized in Supplementary Table\\u0026nbsp;2. Whether the treatment strategies recommended for these subtypes apply to NTRK fusion GISTs merits further investigation. It is important to note that NTRK fusion-positive GISTs are exceedingly rare. Consequently, the small cohort size in this study, while valuable, limits the generalizability of our findings. Future studies with larger patient numbers are essential to validate the observations and hypotheses proposed here.\\u003c/p\\u003e\\u003cp\\u003eNTRK fusion-positive GISTs are exceedingly rare, and reports of treatment with TRK inhibitors are even scarcer. Here, we presented an additional case of NTRK fusion-positive GIST that responded markedly to Larotrectinib. To date, eight patients with NTRK fusion GISTs have received TRK inhibitors, with detailed clinical data available for seven, all of whom responded positively [\\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e7\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR14\\\" class=\\\"CitationRef\\\"\\u003e14\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR15\\\" class=\\\"CitationRef\\\"\\u003e15\\u003c/span\\u003e, \\u003cspan citationid=\\\"CR21\\\" class=\\\"CitationRef\\\"\\u003e21\\u003c/span\\u003e]. However, due to the high cost and limited availability of TRK inhibitors, as well as insufficient awareness of NTRK fusions during treatment planning, some patients still receive Imatinib as first-line therapy\\u0026mdash;even in neoadjuvant or adjuvant settings. Previous case reports indicated that Imatinib failed to control tumor growth in NTRK fusion-positive GISTs when used preoperatively [\\u003cspan citationid=\\\"CR7\\\" class=\\\"CitationRef\\\"\\u003e7\\u003c/span\\u003e]. In this study, we reported two patients with NTRK fusion-positive GIST who experienced recurrence during adjuvant imatinib therapy after complete resection. These cumulative findings suggest that patients with NTRK fusion-positive GISTs are unlikely to benefit from Imatinib or other non-TRK TKIs but may achieve significant clinical responses with TRK inhibitor therapy.\\u003c/p\\u003e\"},{\"header\":\"Declarations\",\"content\":\"\\u003cp\\u003e\\u003cb\\u003eHuman ethics and consent to participate\\u003c/b\\u003e This research involved the collection of FFPE sections from resection tissues. All related protocols were approved by the ethics committee of Renji Hospital (Ethics approval number: LY2024-264-B) and were carried out according to the Ethical Principles of the Declaration of Helsinki. All patients signed written informed consent forms.\\u003c/p\\u003e\\u003cp\\u003e\\u003cstrong\\u003eCompeting interests\\u003c/strong\\u003e\\u003cp\\u003eThe authors declare no competing interests.\\u003c/p\\u003e\\u003c/p\\u003e\\u003ch2\\u003eFunding\\u003c/h2\\u003e\\u003cp\\u003eThis work was supported by the National Natural Science Foundation of China (No. 82070207 and 82300212).\\u003c/p\\u003e\\u003ch2\\u003eAuthor Contribution\\u003c/h2\\u003e\\u003cp\\u003eY.S., A.L., Z.L., and Q.P., and J.W. participated in the literature search and in the writing. Y.S., L.Y., L.Z., and Y.C. participated in data analysis and visualization. Q.L., L.W., and J.Y. participated in data curation and methodology. A.L., M.W., and Z.L. supervised, coordinated the study, participated in the writing, and reviewed the final manuscript. All authors reviewed the manuscript.\\u003c/p\\u003e\\u003ch2\\u003eAcknowledgments\\u003c/h2\\u003e\\u003cp\\u003eWe gratefully acknowledge the support of Dr. Romel Somwar from Memorial Sloan Kettering Cancer Center for his support of this work.\\u003c/p\\u003e\"},{\"header\":\"References\",\"content\":\"\\u003col\\u003e\\u003cli\\u003e\\u003cspan\\u003eC.L. Corless, C.M. Barnett, M.C. Heinrich, Gastrointestinal stromal tumours: origin and molecular oncology. Nat. Rev. Cancer. \\u003cb\\u003e11\\u003c/b\\u003e(12), 865\\u0026ndash;878 (2011)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eH. Joensuu, E. Wardelmann, H. Sihto, M. Eriksson, K. Sundby Hall, A. Reichardt et al., Effect of KIT and PDGFRA mutations on survival in patients with gastrointestinal stromal tumors treated with adjuvant imatinib: an exploratory analysis of a randomized clinical trial. Jama Oncol. \\u003cb\\u003e3\\u003c/b\\u003e(5), 602\\u0026ndash;609 (2017)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eC.L. Corless, K.V. Ballman, C.R. Antonescu, V. Kolesnikova, R.G. Maki, P.W.T. Pisters et al., Pathologic and molecular features correlate with long-term outcome after adjuvant therapy of resected primary GI stromal tumor: the ACOSOG Z9001 trial. J. Clin. Oncol. \\u003cb\\u003e32\\u003c/b\\u003e(15), 1563\\u0026ndash;1570 (2014)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eM.A. Pantaleo, M. Nannini, C.L. Corless, M.C. Heinrich, Quadruple wild-type (WT) GIST: defining the subset of GIST that lacks abnormalities of KIT, PDGFRA, SDH, or RAS signaling pathways. Cancer Med. \\u003cb\\u003e4\\u003c/b\\u003e(1), 101\\u0026ndash;103 (2015)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eM. Nannini, G. Biasco, A. Astolfi, M.A. Pantaleo, An overview on molecular biology of KIT/PDGFRA wild type (WT) gastrointestinal stromal tumours (GIST). J. Med. Genet. \\u003cb\\u003e50\\u003c/b\\u003e(10), 653\\u0026ndash;661 (2013)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eS.A. Boikos, A.S. Pappo, J.K. Killian, M.P. LaQuaglia, C.B. Weldon, S. George et al., Molecular subtypes of KIT/PDGFRA wild-type gastrointestinal stromal tumors: a report from the national institutes of health gastrointestinal stromal tumor clinic. Jama Oncol. \\u003cb\\u003e2\\u003c/b\\u003e(7), 922\\u0026ndash;928 (2016)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eE. Shi, J. Chmielecki, C. Tang, K. Wang, M.C. Heinrich, G. Kang et al., FGFR1 and NTRK3 actionable alterations in Wild-Type gastrointestinal stromal tumors. J. Transl Med. \\u003cb\\u003e14\\u003c/b\\u003e(1), 339 (2016)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eE. Cocco, M. Scaltriti, A. Drilon, NTRK fusion-positive cancers and TRK inhibitor therapy. \\u003cb\\u003e15\\u003c/b\\u003e(12), 731\\u0026ndash;747 (2018)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eL.M. Weiss, V.A. Funari, NTRK fusions and Trk proteins: what are they and how to test for them. Hum. Pathol. \\u003cb\\u003e112\\u003c/b\\u003e, 59\\u0026ndash;69 (2021)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eY. Ma, Q. Zhang, K. Zhang, Y. Liang, F. Ren, J. Zhang et al., NTRK fusions in thyroid cancer: Pathology and clinical aspects. 184, 103957 (2023)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eF. Zito Marino, S. Buono, M. Montella, R. Giannatiempo, F. Messina, G. 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Farago et al., Entrectinib in patients with advanced or metastatic NTRK fusion-positive solid tumours: integrated analysis of three phase 1\\u0026ndash;2 trials. Lancet Oncol. \\u003cb\\u003e21\\u003c/b\\u003e(2), 271\\u0026ndash;282 (2020)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eD.S. Hong, S.G. DuBois, S. Kummar, A.F. Farago, C.M. Albert, K.S. Rohrberg et al., Larotrectinib in patients with TRK fusion-positive solid tumours: a pooled analysis of three phase 1/2 clinical trials. Lancet Oncol. \\u003cb\\u003e21\\u003c/b\\u003e(4), 531\\u0026ndash;540 (2020)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eM. Brenca, S. Rossi, M. Polano, D. Gasparotto, L. Zanatta, D. Racanelli et al., Transcriptome sequencing identifies ETV6-NTRK3 as a gene fusion involved in GIST. J. Pathol. \\u003cb\\u003e238\\u003c/b\\u003e(4), 543\\u0026ndash;549 (2016)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eR. 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Surg. \\u003cb\\u003e46\\u003c/b\\u003e(10), 4435\\u0026ndash;4436 (2023)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eI. Machado, R. Claramunt-Alonso, J. Lavernia, I. Romero, M. Barrios, M.J. Safont et al., Etv6::ntrk3 fusion-positive wild-type gastrointestinal stromal tumor (gist) with abundant lymphoid infiltration (tils and tertiary lymphoid structures): a report on a new case with therapeutic implications and a literature review. Int. J. Mol. Sci. \\u003cb\\u003e25\\u003c/b\\u003e(7), 3707 (2024)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eW. Xiang, W. Yuan, L. Ren, W. Huang, H. Liang, J. Huang et al., A case of quadruple wild-type gastrointestinal stromal tumor with CDC42BPB::NTRK3 fusion and abundant lymphoid infiltration. Diagn. Pathol. \\u003cb\\u003e20\\u003c/b\\u003e(1), 31 (2025)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eM. Castillon, S. Kammerer-Jacquet, M. Cariou, S. Costa, G. Conq, L. Samaison et al., Fluorescent in situ hybridization must be preferred to pan-trk immunohistochemistry to diagnose NTRK3-rearranged gastrointestinal stromal tumors (GIST). Appl. Immunohistochem. Mol. Morphol. \\u003cb\\u003e29\\u003c/b\\u003e(8), 626\\u0026ndash;634 (2021)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eY.P. Hung, C.D.M. Fletcher, Hornick. Evaluation of pan-trk immunohistochemistry in infantile fibrosarcoma, lipofibromatosis-like neural tumour and histological mimics. Histopathology. \\u003cb\\u003e73\\u003c/b\\u003e(4), 634\\u0026ndash;644 (2018)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eJ.P. Solomon, I. Linkov, A. Rosado, K. Mullaney, E.Y. Rosen, D. Frosina et al., NTRK fusion detection across multiple assays and 33,997 cases: diagnostic implications and pitfalls. Mod. Pathol. \\u003cb\\u003e33\\u003c/b\\u003e(1), 38\\u0026ndash;46 (2020)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eI.M. Schaefer, C. Delfs, S. Cameron, B. Gunawan, A. Agaimy, B.M. Ghadimi et al., Chromosomal aberrations in primary PDGFRA-mutated gastrointestinal stromal tumors. Hum. Pathol. \\u003cb\\u003e45\\u003c/b\\u003e(1), 85\\u0026ndash;97 (2014)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eM. Silva, I. Veiga, F.R. Ribeiro, J. Vieira, C. Pinto, M. Pinheiro et al., Chromosome copy number changes carry prognostic information independent of KIT/PDGFRA point mutations in gastrointestinal stromal tumors. BMC Med. \\u003cb\\u003e8\\u003c/b\\u003e, 26 (2010)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eW. El-Rifai, M. Sarlomo-Rikala, L.C. Andersson, S. Knuutila, M. Miettinen, Dna sequence copy number changes in gastrointestinal stromal tumors: tumor progression and prognostic significance. Cancer Res. \\u003cb\\u003e60\\u003c/b\\u003e(14), 3899\\u0026ndash;3903 (2000)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eT.G. Kleijn, B. Ameline, R.F. Bleckman, W. Kooistra, E. van den Broek, G. Diercks et al., Genome-wide dna methylation and copy number alterations in gastrointestinal stromal tumors. Chromosomes Cancer. \\u003cb\\u003e64\\u003c/b\\u003e(3), e70046 (2025)\\u003c/span\\u003e\\u003c/li\\u003e\\u003cli\\u003e\\u003cspan\\u003eF. Xie, S. Luo, D. Liu, X. Lu, M. Wang, X. Liu et al., Genomic and transcriptomic landscape of human gastrointestinal stromal tumors. Nat. Commun. \\u003cb\\u003e15\\u003c/b\\u003e(1), 9495 (2024)\\u003c/span\\u003e\\u003c/li\\u003e\\u003c/ol\\u003e\"}],\"fulltextSource\":\"\",\"fullText\":\"\",\"funders\":[],\"hasAdminPriorityOnWorkflow\":false,\"hasManuscriptDocX\":true,\"hasOptedInToPreprint\":true,\"hasPassedJournalQc\":\"\",\"hasAnyPriority\":false,\"hideJournal\":false,\"highlight\":\"\",\"institution\":\"\",\"isAcceptedByJournal\":true,\"isAuthorSuppliedPdf\":false,\"isDeskRejected\":\"\",\"isHiddenFromSearch\":false,\"isInQc\":false,\"isInWorkflow\":false,\"isPdf\":false,\"isPdfUpToDate\":true,\"isWithdrawnOrRetracted\":false,\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"cellular-oncology\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":false,\"externalIdentity\":\"ceon\",\"sideBox\":\"Learn more about [Cellular Oncology](http://link.springer.com/journal/13402)\",\"snPcode\":\"13402\",\"submissionUrl\":\"https://submission.nature.com/new-submission/13402/3\",\"title\":\"Cellular Oncology\",\"twitterHandle\":\"\",\"acdcEnabled\":true,\"dfaEnabled\":true,\"editorialSystem\":\"em\",\"reportingPortfolio\":\"Springer Hybrid\",\"inReviewEnabled\":true,\"inReviewRevisionsEnabled\":false},\"keywords\":\"gastrointestinal stromal tumor, Pan-TRK, NTRK3 fusion, NTRK inhibitors\",\"lastPublishedDoi\":\"10.21203/rs.3.rs-7615382/v1\",\"lastPublishedDoiUrl\":\"https://doi.org/10.21203/rs.3.rs-7615382/v1\",\"license\":{\"name\":\"CC BY 4.0\",\"url\":\"https://creativecommons.org/licenses/by/4.0/\"},\"manuscriptAbstract\":\"\\u003ch2\\u003ePurpose\\u003c/h2\\u003e\\u003cp\\u003eThis study aimed to characterize the clinicopathological, immunophenotypic, and molecular features of gastrointestinal stromal tumors (GISTs) harboring \\u003cem\\u003eNTRK\\u003c/em\\u003e fusions and to evaluate their diagnostic, prognostic, and therapeutic implications.\\u003c/p\\u003e\\u003ch2\\u003eMethods\\u003c/h2\\u003e\\u003cp\\u003eTwenty-six cases of \\u003cem\\u003eKIT/PDGFRA/SDH/BRAF\\u003c/em\\u003e wild-type GISTs were evaluated using pan-TRK immunohistochemistry (IHC), fluorescence in situ hybridization (FISH) for *NTRK1/2/3*, and next-generation sequencing (NGS). Transcriptome analysis was performed on all \\u003cem\\u003eNTRK\\u003c/em\\u003e fusion-positive cases. Seven \\u003cem\\u003eKIT\\u003c/em\\u003e-mutant GISTs served as controls. Clinicopathological parameters, IHC profiles, genetic alterations, and treatment responses were analyzed, supplemented by a literature review.\\u003c/p\\u003e\\u003ch2\\u003eResults\\u003c/h2\\u003e\\u003cp\\u003eFive of the 26 wild-type GISTs harbored \\u003cem\\u003eNTRK\\u003c/em\\u003e fusions, all confirmed by NGS as \\u003cem\\u003eETV6::NTRK3\\u003c/em\\u003e. Pan-TRK IHC showed 100% sensitivity and 66.7% specificity. All five patients were male; four tumors were intestinal and one gastric. Four cases were high-risk and one very low-risk. Two cases recurred post-resection, showing additional mutations and copy number variations (CNVs). Transcriptome analysis revealed molecular heterogeneity among \\u003cem\\u003eNTRK\\u003c/em\\u003e fusion-positive GISTs, with profiles overlapping those of \\u003cem\\u003eKIT\\u003c/em\\u003e-mutant GISTs. Both recurrent patients received multi-line TKI therapy (imatinib, sunitinib, regorafenib, ripretinib) with disease progression; one subsequently achieved remission with larotrectinib.\\u003c/p\\u003e\\u003ch2\\u003eConclusion\\u003c/h2\\u003e\\u003cp\\u003e\\u003cem\\u003eNTRK\\u003c/em\\u003e fusion-positive GISTs are rare and exhibit distinct clinicopathological characteristics. FISH and NGS are reliable detection methods, while pan-TRK IHC has limited specificity. Co-occurring genetic alterations may confer aggressive behavior. These tumors respond to TRK inhibition but are resistant to conventional TKIs, underscoring the need for molecularly guided therapy.\\u003c/p\\u003e\",\"manuscriptTitle\":\"Clinicopathologic and molecular spectrum of gastrointestinal stromal tumor (GIST) with NTRK fusion and marked response to Larotrectinib in GIST with NTRK fusion: a case report\",\"msid\":\"\",\"msnumber\":\"\",\"nonDraftVersions\":[{\"code\":1,\"date\":\"2025-09-30 07:11:04\",\"doi\":\"10.21203/rs.3.rs-7615382/v1\",\"editorialEvents\":[{\"type\":\"communityComments\",\"content\":0},{\"type\":\"decision\",\"content\":\"Revision requested\",\"date\":\"2025-10-24T02:10:07+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"editorInvitedReview\",\"content\":\"\",\"date\":\"2025-10-24T02:08:42+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"62976301326252898854147092801453400411\",\"date\":\"2025-10-24T01:30:29+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"editorInvitedReview\",\"content\":\"\",\"date\":\"2025-10-15T01:59:06+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"294211916689111366556932364154942542113\",\"date\":\"2025-10-14T10:07:37+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"331785745293919457869376528289118617344\",\"date\":\"2025-10-09T23:27:01+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"editorInvitedReview\",\"content\":\"\",\"date\":\"2025-10-07T13:34:53+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"227149402229104750547668982394370872769\",\"date\":\"2025-09-26T22:14:47+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewerAgreed\",\"content\":\"24428744014816429315125491252295924394\",\"date\":\"2025-09-24T07:08:03+00:00\",\"index\":\"hide\",\"fulltext\":\"\"},{\"type\":\"reviewersInvited\",\"content\":\"\",\"date\":\"2025-09-18T02:31:00+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"editorAssigned\",\"content\":\"\",\"date\":\"2025-09-17T07:18:16+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"checksComplete\",\"content\":\"\",\"date\":\"2025-09-17T07:17:19+00:00\",\"index\":\"\",\"fulltext\":\"\"},{\"type\":\"submitted\",\"content\":\"Cellular Oncology\",\"date\":\"2025-09-15T02:07:18+00:00\",\"index\":\"\",\"fulltext\":\"\"}],\"status\":\"published\",\"journal\":{\"display\":true,\"email\":\"info@researchsquare.com\",\"identity\":\"cellular-oncology\",\"isNatureJournal\":false,\"hasQc\":true,\"allowDirectSubmit\":false,\"externalIdentity\":\"ceon\",\"sideBox\":\"Learn more about [Cellular Oncology](http://link.springer.com/journal/13402)\",\"snPcode\":\"13402\",\"submissionUrl\":\"https://submission.nature.com/new-submission/13402/3\",\"title\":\"Cellular Oncology\",\"twitterHandle\":\"\",\"acdcEnabled\":true,\"dfaEnabled\":true,\"editorialSystem\":\"em\",\"reportingPortfolio\":\"Springer Hybrid\",\"inReviewEnabled\":true,\"inReviewRevisionsEnabled\":false}}],\"origin\":\"\",\"ownerIdentity\":\"3daf9f2b-a0d3-477e-bfa5-58ade07f9957\",\"owner\":[],\"postedDate\":\"September 30th, 2025\",\"published\":true,\"recentEditorialEvents\":[],\"rejectedJournal\":[],\"revision\":\"\",\"amendment\":\"\",\"status\":\"published-in-journal\",\"subjectAreas\":[],\"tags\":[],\"updatedAt\":\"2026-01-26T16:03:08+00:00\",\"versionOfRecord\":{\"articleIdentity\":\"rs-7615382\",\"link\":\"https://doi.org/10.1007/s13402-025-01146-6\",\"journal\":{\"identity\":\"cellular-oncology\",\"isVorOnly\":false,\"title\":\"Cellular Oncology\"},\"publishedOn\":\"2026-01-19 15:58:26\",\"publishedOnDateReadable\":\"January 19th, 2026\"},\"versionCreatedAt\":\"2025-09-30 07:11:04\",\"video\":\"\",\"vorDoi\":\"10.1007/s13402-025-01146-6\",\"vorDoiUrl\":\"https://doi.org/10.1007/s13402-025-01146-6\",\"workflowStages\":[]},\"version\":\"v1\",\"identity\":\"rs-7615382\",\"journalConfig\":\"researchsquare\"},\"__N_SSP\":true},\"page\":\"/article/[identity]/[[...version]]\",\"query\":{\"redirect\":\"/article/rs-7615382\",\"identity\":\"rs-7615382\",\"version\":[\"v1\"]},\"buildId\":\"XKTyCvWXoU3ODBz1xrDgd\",\"isFallback\":false,\"isExperimentalCompile\":false,\"dynamicIds\":[84888],\"gssp\":true,\"scriptLoader\":[]}","source_license":"CC-BY-4.0","license_restricted":false}