Tyrosine Kinase Inhibition Activates Intratumoral γδ T Cells in Gastrointestinal Stromal Tumor.

γδ T cells are a rare but potent subset of T cells with pleiotropic functions. They commonly reside within tumors but the response of γδ T cells to tyrosine kinase inhibition is unknown. To address this, we studied a genetically engineered mouse model of gastrointestinal stromal tumor (GIST) driven by oncogenic Kit signaling that responds to the Kit inhibitor imatinib. At baseline, γδ T cells were antitumoral, as blockade of either γδ T-cell receptor or IL17A increased tumor weight and decreased antitumor immunity. However, imatinib therapy further stimulated intratumoral γδ T cells, as determined by flow cytometry and single-cell RNA sequencing (scRNA-seq). Imatinib expanded a highly activated γδ T-cell subset with increased IL17A production and higher expression of immune checkpoints and cytolytic effector molecules. Consistent with the mouse model, γδ T cells produced IL17A in fresh human GIST specimens, and imatinib treatment increased γδ T-cell gene signatures, as measured by bulk tumor RNA-seq. Furthermore, tumor γδ T cells correlated with survival in patients with GIST. Our findings highlight the interplay between tumor cell oncogene signaling and antitumor immune responses and identify γδ T cells as targets for immunotherapy in GIST.

Combined hepatic resection and ablation for high burden of colorectal liver metastases demonstrates safety and durable survival.

BACKGROUND: Hepatic resection (HR) and thermal ablation of Colorectal Liver Metastases (CRLM) have each individually demonstrated safety and survival benefit. We sought to provide our experience with the combination of HR + ablation within one operation for patients with multiple CRLM. METHODS: Review of a single institution database of patients who underwent HR + ablation between 2010 and 2019. RESULTS: 161 patients were identified who underwent HR + ablation for isolated CRLM (mean age: 59, male 63.4%). 125 (77.6%) patients had bilobar disease and 92 (57.1%) patients had ≥5 tumors. 28 (17.4%) patients experienced minor (grade 1 or 2) complications while 20 (12.4%) had grade 3-5 complications. Patients who underwent simultaneous colon resection with HR + ablation had a higher complication rate (22 of 47, 46.8%) than those undergoing HR + ablation only (26 of 114, 22.8%, p = 0.002). Median and 5-year OS for all patients undergoing HR + ablation was 38.2 months and 33.2%, respectively. 5-year hepatic recurrence free survival was 23.5%. Patients with 5 or more tumors demonstrated no difference in median survival compared to those with fewer than 5 tumors (37.0 months vs 38.4 months, p = 0.326). CONCLUSIONS: In this population of CRLM patients with a relatively high burden of disease, HR + ablation demonstrated an acceptable safety profile as well as durable long-term survival.

Long-term overall survival in patients undergoing liver resection for metastatic anal squamous cell carcinoma.

BACKGROUND AND OBJECTIVES: There is limited research on management of metastatic anal canal squamous cell carcinoma (SCC) to the liver. This study aimed to describe outcomes for patients undergoing liver resection of anal SCC metastases. METHODS: A multicenter, retrospective cohort study was conducted by three tertiary-referral centers. Patients undergoing liver surgery between 2008 and 2022 were included. Cox regression analysis was performed to evaluate predictors of recurrence and survival and Kaplan-Meier analysis was performed for 1-, 3-, and 5-year survival. RESULTS: Twenty-one patients underwent liver resection and/or ablation. None were HIV positive and 24% had known HPV infection. 20/21(95%) patients had undergone Nigro protocol for management of the primary tumor with 12/21 (57%) patients experiencing complete response. 4/21 (19%) patients had synchronous liver metastases at time of diagnosis. Median tumor size was 5.0 cm and median tumor number was one. At analysis, 52% remained alive. Median overall survival was 32.2 months. 5-year overall survival was 50%. Median recurrence-free survival was 7.7 months and 5-year recurrence-free survival was 30%. Need for salvage abdominoperineal resection was negatively associated with recurrence-free survival. The most common site of recurrence was the liver. CONCLUSIONS: Liver resection for metastatic anal SCC can be beneficial for appropriately selected patients.

Multiple intratumoral sources of kit ligand promote gastrointestinal stromal tumor.

Gastrointestinal stromal tumor (GIST) is the most common human sarcoma and is typically driven by a single mutation in the Kit or PDGFRA receptor. While highly effective, tyrosine kinase inhibitors (TKIs) are not curative. The natural ligand for the Kit receptor is Kit ligand (KitL), which exists in both soluble and membrane-bound forms. While KitL is known to stimulate human GIST cell lines in vitro, we used a genetically engineered mouse model of GIST containing a common human KIT mutation to investigate the intratumoral sources of KitL, importance of KitL during GIST oncogenesis, and contribution of soluble KitL to tumor growth in vivo. We discovered that in addition to tumor cells, endothelia and smooth muscle cells produced KitL in KitV558Δ/+ tumors, even after imatinib therapy. Genetic reduction of total KitL in tumor cells of KitV558Δ/+ mice impaired tumor growth in vivo. Similarly, genetic reduction of tumor cell soluble KitL in KitV558Δ/+ mice decreased tumor size. By RNA sequencing, quantitative PCR, and immunohistochemistry, KitL expression was heterogeneous in human GIST specimens. In particular, PDGFRA-mutant tumors had much higher KitL expression than Kit-mutant tumors, suggesting the benefit of Kit activation in the absence of mutant KIT. Serum KitL was higher in GIST patients with tumors resistant to imatinib and in those with tumors expressing more KitL RNA. Overall, KitL supports the growth of GIST at baseline and after imatinib therapy and remains a potential biomarker and therapeutic target.

Myosin Light-Chain Kinase Inhibition Potentiates the Antitumor Effects of Avapritinib in PDGFRA D842V-Mutant Gastrointestinal Stromal Tumor.

PURPOSE: To create an in vivo model of PDGFRA D842V-mutant gastrointestinal stromal tumor (GIST) and identify the mechanism of tumor persistence following avapritinib therapy. EXPERIMENTAL DESIGN: We created a patient-derived xenograft (PDX) of PDGFRA D842V-mutant GIST and tested the effects of imatinib, avapritinib, and ML-7, an inhibitor of myosin light-chain kinase (MYLK). Bulk tumor RNA sequencing and oncogenic signaling were evaluated. Apoptosis, survival, and actin cytoskeleton were evaluated in GIST T1 cells and isolated PDX cells in vitro. Human GIST specimens were analyzed for MYLK expression. RESULTS: The PDX was minimally responsive to imatinib but sensitive to avapritinib. Avapritinib therapy increased tumor expression of genes related to the actin cytoskeleton, including MYLK. ML-7 induced apoptosis and disrupted actin filaments in short-term cultures of PDX cells and decreased survival in GIST T1 cells in combination with imatinib or avapritinib. Combined therapy with ML-7 improved the antitumor effects of low-dose avapritinib in vivo. Furthermore, MYLK was expressed in human GIST specimens. CONCLUSIONS: MYLK upregulation is a novel mechanism of tumor persistence after tyrosine kinase inhibition. Concomitant MYLK inhibition may enable the use of a lower dose of avapritinib, which is associated with dose-dependent cognitive side effects.

Tyrosine Kinase Inhibition Alters Intratumoral CD8+ T-cell Subtype Composition and Activity.

Targeted therapy with a tyrosine kinase inhibitor (TKI) such as imatinib is effective in treating gastrointestinal stromal tumor (GIST), but it is rarely curative. Despite the presence of a robust immune CD8+ T-cell infiltrate, combining a TKI with immune-checkpoint blockade (ICB) in advanced GIST has achieved only modest effects. To identify limitations imposed by imatinib on the antitumor immune response, we performed bulk RNA sequencing (RNA-seq), single-cell RNA-seq, and flow cytometry to phenotype CD8+ T-cell subsets in a genetically engineered mouse model of GIST. Imatinib reduced the frequency of effector CD8+ T cells and increased the frequency of naïve CD8+ T cells within mouse GIST, which coincided with altered tumor chemokine production, CD8+ T-cell recruitment, and reduced CD8+ T-cell intracellular PI3K signaling. Imatinib also failed to induce intratumoral T-cell receptor (TCR) clonal expansion. Consistent with these findings, human GISTs sensitive to imatinib harbored fewer effector CD8+ T cells but more naïve CD8+ T cells. Combining an IL15 superagonist (IL15SA) with imatinib restored intratumoral effector CD8+ T-cell function and CD8+ T-cell intracellular PI3K signaling, resulting in greater tumor destruction. Combination therapy with IL15SA and ICB resulted in the greatest tumor killing and maintained an effector CD8+ T-cell population in the presence of imatinib. Our findings highlight the impact of oncogene inhibition on intratumoral CD8+ T cells and support the use of agonistic T-cell therapy during TKI and/or ICB administration.

Oncogenic KIT Modulates Type I IFN-Mediated Antitumor Immunity in GIST.

Type I IFNs are implicated in tumor immunogenicity and response to systemic therapy, but their interaction with oncogene signaling is not well understood. Here, we studied oncogenic KIT, which drives gastrointestinal stromal tumor (GIST), the most common sarcoma. Using mouse models of GIST, we found that KIT inhibition reduced type I IFN production and signaling, which downregulated tumor MHC class I expression. Absence of type I IFN signaling increased tumor size, in part due to CD8+ T-cell impairment. Oncogenic KIT was required for GIST type I IFN signal transduction via STAT1. In human GIST cell lines and surgical specimens, type I IFN signaling contributed to human lymphocyte antigen class I expression and correlated with tumor immunogenicity. Augmenting the type I IFN response partially compensated for the immunosuppressive effects of KIT inhibition. Thus, KIT signaling contributes to type I IFN signaling, whereas KIT inhibition attenuates tumor immunogenicity and is partly rescued by innate immune stimulation.See related Spotlight on p. 489.

Tailored management of primary gastrointestinal stromal tumors.

Gastrointestinal stromal tumors (GISTs) are the most common human sarcoma and can form along the entire gastrointestinal tract. Over the last 20 years, considerable advances have been made in our understanding of the biology of GISTs. The advent of tyrosine kinase inhibitors has provided effective medical therapy for the first time. In fact, given that GIST typically is driven by either a KIT or PDGFRA gene mutation, it has become a paradigm of targeted molecular therapy. In addition, diagnostic and surgical techniques have been refined. Here, the critical aspects of primary GISTs and how they are now managed with an integrated approach are summarized. Treatment plans are developed based on specific pathologic and molecular features of the tumor. The authors outline the general principles of therapy and highlight some of the nuances. Particular focus is given to diagnosis, surgical considerations, and the use of preoperative and postoperative tyrosine kinase inhibitors.

Predictors of false negative sentinel lymph node biopsy in trunk and extremity melanoma.

BACKGROUND: Nodal recurrence following negative sentinel lymph node biopsy (SLNB) for melanoma is known as false-negative (FN) SLNB. Risk factors for FN SLNB among patients with trunk and extremity melanoma have not been well-defined. METHODS: After retrospective review, SLNB procedures were classified FN, true positive (TP; positive SLNB), or true negative (TN; negative SLNB without recurrence). Factors associated with high false negative rate (FNR) and low negative predictive value (NPV) were identified by comparing FNs to TPs and TNs, respectively. Survival was evaluated using Kaplan-Meier methods. RESULTS: Of 1728 patients, 234 were TP and 37 were FN for overall FNR of 14% and NPV of 97.5%. Age ≥65 years was independently associated with high FNR (FNR 20% in this group). Breslow thickness >1 mm and ulceration were independently associated with low NPV. Among patients with ulcerated tumors >4 mm, NPV was 88%. Median time to recurrence for FNs was 13 months. Among patients with primary melanomas ≤2 mm in depth, overall and distant disease-free survival were significantly shorter with FN SLNB than TP SLNB. CONCLUSIONS: Older age is associated with increased FNR; patients with thick, ulcerated lesions should be considered for increased nodal surveillance after negative SLNB given low NPV in this group.

Identification of Patients with Intermediate Thickness Melanoma at Low Risk for Sentinel Lymph Node Positivity.

INTRODUCTION: Sentinel lymph node (SLN) biopsy is recommended for all patients with intermediate-thickness melanomas. We sought to identify such patients at low risk of SLN positivity. METHODS: All patients with intermediate-thickness melanomas (1.01-4 mm) undergoing SLN biopsy at a single institution from 1995-2011 were included in this retrospective cohort study. Univariate and multivariate logistic regression determined factors associated with a low risk of SLN positivity. Classification and regression tree (CART) analysis was used to stratify groups based on risk of positivity. RESULTS: Of the 952 study patients, 157 (16.5 %) had a positive SLN. In the multivariate analysis, thickness <1.5 mm (odds ratio [OR] 0.29), age ≥60 (OR 0.69), present tumor-infiltrating lymphocytes (OR 0.60), absent lymphovascular invasion (OR 0.46), and absent satellitosis (OR 0.44) were significantly associated with a low risk of SLN positivity. CART analysis identified thickness of 1.5 mm as the primary cut point for risk of SLN metastasis. Patients with a thickness of <1.5 mm represented 36 % of the total cohort and had a SLN positivity rate of 6.6 % (95 % confidence interval 3.8-9.4 %). In patients with melanomas <1.5 mm in thickness, the presence of additional low risk factors identified 257 patients (75 % of patients with <1.5 mm melanomas) in which the rate of SLN positivity was <5 %. CONCLUSIONS: Despite a SLN positivity rate of 16.5 % overall, substantial heterogeneity of risk exists among patients with intermediate-thickness melanoma. Most patients with melanoma between 1.01 and 1.5 mm have a risk of SLN positivity similar to that in patients with thin melanomas.