Kyle Gowen

Activation of MET via Diverse Exon 14 Splicing Alterations Occurs in Multiple Tumor Types and Confers Clinical Sensitivity to MET Inhibitors

UNLABELLED Focal amplification and activating point mutation of the MET gene are well-characterized oncogenic drivers that confer susceptibility to targeted MET inhibitors. Recurrent somatic splice site alterations at MET exon 14 (METex14) that result in exon skipping and MET activation have been characterized, but their full diversity and prevalence across tumor types are unknown. Here, we report analysis of tumor genomic profiles from 38,028 patients to identify 221 cases with METex14 mutations (0.6%), including 126 distinct sequence variants. METex14 mutations are detected most frequently in lung adenocarcinoma (3%), but also frequently in other lung neoplasms (2.3%), brain glioma (0.4%), and tumors of unknown primary origin (0.4%). Further in vitro studies demonstrate sensitivity to MET inhibitors in cells harboring METex14 alterations. We also report three new patient cases with METex14 alterations in lung or histiocytic sarcoma tumors that showed durable response to two different MET-targeted therapies. The diversity of METex14 mutations indicates that diagnostic testing via comprehensive genomic profiling is necessary for detection in a clinical setting. SIGNIFICANCE Here we report the identification of diverse exon 14 splice site alterations in MET that result in constitutive activity of this receptor and oncogenic transformation in vitro. Patients whose tumors harbored these alterations derived meaningful clinical benefit from MET inhibitors. Collectively, these data support the role of METex14 alterations as drivers of tumorigenesis, and identify a unique subset of patients likely to derive benefit from MET inhibitors.

Inactivation of Capicua drives cancer metastasis

Metastasis is the leading cause of death in people with lung cancer, yet the molecular effectors underlying tumor dissemination remain poorly defined. Through the development of an in vivo spontaneous lung cancer metastasis model, we show that the developmentally regulated transcriptional repressor Capicua (CIC) suppresses invasion and metastasis. Inactivation of CIC relieves repression of its effector ETV4, driving ETV4-mediated upregulation of MMP24, which is necessary and sufficient for metastasis. Loss of CIC, or an increase in levels of its effectors ETV4 and MMP24, is a biomarker of tumor progression and worse outcomes in people with lung and/or gastric cancer. Our findings reveal CIC as a conserved metastasis suppressor, highlighting new anti-metastatic strategies that could potentially improve patient outcomes.

Comprehensive Genomic Profiling Identifies a Subset of Crizotinib-Responsive ALK-Rearranged Non-Small Cell Lung Cancer Not Detected by Fluorescence In Situ Hybridization

INTRODUCTION For patients with non-small cell lung cancer (NSCLC) to benefit from ALK inhibitors, sensitive and specific detection of ALK genomic rearrangements is needed. ALK break-apart fluorescence in situ hybridization (FISH) is the U.S. Food and Drug Administration approved and standard-of-care diagnostic assay, but identification of ALK rearrangements by other methods reported in NSCLC cases that tested negative for ALK rearrangements by FISH suggests a significant false-negative rate. We report here a large series of NSCLC cases assayed by hybrid-capture-based comprehensive genomic profiling (CGP) in the course of clinical care. MATERIALS AND METHODS Hybrid-capture-based CGP using next-generation sequencing was performed in the course of clinical care of 1,070 patients with advanced lung cancer. Each tumor sample was evaluated for all classes of genomic alterations, including base-pair substitutions, insertions/deletions, copy number alterations and rearrangements, as well as fusions/rearrangements. RESULTS A total of 47 patients (4.4%) were found to harbor ALK rearrangements, of whom 41 had an EML4-ALK fusion, and 6 had other fusion partners, including 3 previously unreported rearrangement events: EIF2AK-ALK, PPM1B-ALK, and PRKAR1A-ALK. Of 41 patients harboring ALK rearrangements, 31 had prior FISH testing results available. Of these, 20 were ALK FISH positive, and 11 (35%) were ALK FISH negative. Of the latter 11 patients, 9 received crizotinib based on the CGP results, and 7 achieved a response with median duration of 17 months. CONCLUSION Comprehensive genomic profiling detected canonical ALK rearrangements and ALK rearrangements with noncanonical fusion partners in a subset of patients with NSCLC with previously negative ALK FISH results. In this series, such patients had durable responses to ALK inhibitors, comparable to historical response rates for ALK FISH-positive cases. IMPLICATIONS FOR PRACTICE Comprehensive genomic profiling (CGP) that includes hybrid capture and specific baiting of intron 19 of ALK is a highly sensitive, alternative method for identification of drug-sensitive ALK fusions in patients with non-small cell lung cancer (NSCLC) who had previously tested negative using standard ALK fluorescence in situ hybridization (FISH) diagnostic assays. Given the proven benefit of treatment with crizotinib and second-generation ALK inhibitors in patients with ALK fusions, CGP should be considered in patients with NSCLC, including those who have tested negative for other alterations, including negative results using ALK FISH testing.

EGFR Fusions as Novel Therapeutic Targets in Lung Cancer

UNLABELLED Here, we report that novel epidermal growth factor receptor (EGFR) gene fusions comprising the N-terminal of EGFR linked to various fusion partners, most commonly RAD51, are recurrent in lung cancer. We describe five patients with metastatic lung cancer whose tumors harbored EGFR fusions, four of whom were treated with EGFR tyrosine kinase inhibitors (TKI) with documented antitumor responses. In vitro, EGFR-RAD51 fusions are oncogenic and can be therapeutically targeted with available EGFR TKIs and therapeutic antibodies. These results support the dependence of EGFR-rearranged tumors on EGFR-mediated signaling and suggest several therapeutic strategies for patients whose tumors harbor this novel alteration. SIGNIFICANCE We report for the first time the identification and therapeutic targeting of EGFR C-terminal fusions in patients with lung cancer and document responses to the EGFR inhibitor erlotinib in 4 patients whose tumors harbored EGFR fusions. Findings from these studies will be immediately translatable to the clinic, as there are already several approved EGFR inhibitors. Cancer Discov; 6(6); 601-11. ©2016 AACR.See related commentary by Paik, p. 574This article is highlighted in the In This Issue feature, p. 561.

Emergence of RET rearrangement co-existing with activated EGFR mutation in EGFR-mutated NSCLC patients who had progressed on first- or second-generation EGFR TKI

OBJECTIVES The gatekeeper mutation T790M mutation is the responsible for the majority of the resistance to epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) in patients with EGFR-mutated non-small cell lung cancer (NSCLC). Other previously described resistance mechanisms include HER2 amplification, MET amplification, PIK3CA mutation, epithelial-mesenchymal transition (EMT), small cell transformation have also been identified. However other resistance mechanisms remains to be discovered. MATERIALS AND METHODS Hybrid-capture based comprehensive genomic profiling (CGP) was performed on pre- and post-EGFR TKI progression EGFR-mutated NSCLC tumor samples during routine clinical care. We identify two paired pre- and post-EGFR TKI progression EGFR-mutated NSCLC patient tumor samples where both post EGFR TKI samples harbored in-frame CCDC6-RET rearrangements but not in the pre-EGFR TKI tumor samples. Furthermore analysis of the clinical database revealed one additional NCOA4-RET rearrangement co-existing with activated EGFR mutation in an EGFR-mutated NSCLC patient who had progressed on afatinib. None of the known resistance mechanisms to EGFR TKI including EGFR T790M, EGFR amplification, HER2 amplification, MET amplification, PIK3CA mutation, BRAF mutation, EMT or small cell transformation was identified in the three post progression samples that now harbored RET rearrangements. RESULTS AND CONCLUSIONS This is the first report of RET rearrangement co-existing with activated EGFR mutations in EGFR-mutated patients who had progressed on either first- or second generation EGFR TKI. As such, RET rearrangement may serve as a potential resistance mechanism to EGFR TKI in EGFR-mutated NSCLC.

Impact of EML4-ALK Variant on Resistance Mechanisms and Clinical Outcomes in ALK-Positive Lung Cancer

Purpose Advanced anaplastic lymphoma kinase ( ALK) fusion-positive non-small-cell lung cancers (NSCLCs) are effectively treated with ALK tyrosine kinase inhibitors (TKIs). However, clinical outcomes in these patients vary, and the benefit of TKIs is limited as a result of acquired resistance. Emerging data suggest that the ALK fusion variant may affect clinical outcome, but the molecular basis for this association is unknown. Patients and Methods We identified 129 patients with ALK-positive NSCLC with known ALK variants. ALK resistance mutations and clinical outcomes on ALK TKIs were retrospectively evaluated according to ALK variant. A Foundation Medicine data set of 577 patients with ALK-positive NSCLC was also examined. Results The most frequent ALK variants were EML4-ALK variant 1 in 55 patients (43%) and variant 3 in 51 patients (40%). We analyzed 77 tumor biopsy specimens from patients with variants 1 and 3 who had progressed on an ALK TKI. ALK resistance mutations were significantly more common in variant 3 than in variant 1 (57% v 30%; P = .023). In particular, ALK G1202R was more common in variant 3 than in variant 1 (32% v 0%; P < .001). Analysis of the Foundation Medicine database revealed similar associations of variant 3 with ALK resistance mutation and with G1202R ( P = .010 and .015, respectively). Among patients treated with the third-generation ALK TKI lorlatinib, variant 3 was associated with a significantly longer progression-free survival than variant 1 (hazard ratio, 0.31; 95% CI, 0.12 to 0.79; P = .011). Conclusion Specific ALK variants may be associated with the development of ALK resistance mutations, particularly G1202R, and provide a molecular link between variant and clinical outcome. ALK variant thus represents a potentially important factor in the selection of next-generation ALK inhibitors.

Oncogenic ALK Fusion in Rare and Aggressive Subtype of Colorectal Adenocarcinoma as a Potential Therapeutic Target

Purpose: Chromosomal translocations in the anaplastic lymphoma kinase (ALK) gene have been identified as oncogenic drivers in lung adenocarcinomas and other tumors, recently including rare cases of colorectal carcinoma. We identified a patient with refractory metastatic colorectal carcinoma harboring a STRN–ALK gene fusion who achieved an exceptional clinical benefit to the ALK inhibitor ceritinib. Our goal was to further define the clinicopathologic features of ALK-rearranged colorectal carcinoma in a large cohort. Experimental Design: Clinical cases of colorectal carcinoma evaluated by comprehensive genomic profiling (CGP) or by ALK immunohistochemistry (IHC) were reviewed retrospectively. FISH and microsatellite instability (MSI) analyses were performed. Results: Nine colorectal carcinoma cases harbored ALK gene fusions. Six cases were identified by CGP of 3,157 colorectal carcinoma (0.2%) and three by IHC of 2,980 colorectal carcinoma (0.1%). The ALK fusions involved known ALK partners EML4, C2orf44, CAD, and the novel STRN, PPP1R21, SENPF, MAPRE3, and PRKAP1B partners. These advanced-stage colorectal carcinomas lacked mutations in other oncogenic drivers, predominantly involved the proximal colon, and often exhibited MSI and mucinous phenotype. The index patient was treated with the ALK inhibitor ceritinib, resulting in a marked decrease in size of a skin metastasis, and resolution by computerized tomography of all contrast enhancing tumor. After 9 months of treatment, biopsy of progressive disease demonstrated a KRAS mutation, consistent with acquired resistance to ceritinib. Conclusions: Colorectal carcinoma harboring ALK fusions represent a rare aggressive subtype of colorectal carcinoma with distinct clinicopathologic features. This report provides the first clinical evidence that such patients may benefit from targeted monotherapy with ALK inhibitors. Clin Cancer Res; 22(15); 3831–40. ©2016 AACR.

BRAFV600E Mutations in High Grade Colorectal Neuroendocrine Tumors May Predict Responsiveness to BRAF-MEK Combination Therapy

UNLABELLED Neuroendocrine tumors comprise a heterogeneous group of malignancies with a broad spectrum of clinical behavior. Poorly differentiated tumors follow an aggressive course with limited treatment options, and new approaches are needed. Oncogenic BRAF V600E (BRAF(V600E)) substitutions are observed primarily in melanoma, colon cancer, and non-small cell lung cancer, but have been identified in multiple tumor types. Here, we describe the first reported recurrent BRAF(V600E) mutations in advanced high-grade colorectal neuroendocrine tumors and identify a BRAF alteration frequency of 9% in 108 cases. Among these BRAF alterations, 80% were BRAF(V600E) Dramatic response to BRAF-MEK combination therapy occurred in two cases of metastatic high-grade rectal neuroendocrine carcinoma refractory to standard therapy. Urinary BRAF(V600E) circulating tumor DNA monitoring paralleled disease response. Our series represents the largest study of genomic profiling in colorectal neuroendocrine tumors and provides strong evidence that BRAF(V600E) is an oncogenic driver responsive to BRAF-MEK combination therapy in this molecular subset. SIGNIFICANCE BRAF(V600E) is an established oncogenic driver, but significant disparities in response exist among tumor types. Two patients with treatment-refractory high-grade colorectal neuroendocrine tumors harboring BRAF(V600E) exhibited rapid and durable response to combined BRAF-MEK inhibition, providing the first clinical evidence of efficacy in this aggressive tumor type. Cancer Discov; 6(6); 594-600. ©2016 AACR.This article is highlighted in the In This Issue feature, p. 561.

TPD52L1-ROS1, a new ROS1 fusion variant in lung adenosquamous cell carcinoma identified by comprehensive genomic profiling

Crizotinib was approved for the treatment of ROS1-rearranged non-small cell lung cancer (NSCLC) patients in the US on 11 March, 2016. Interestingly no one companion diagnostic test (CDx) has been approved simultaneously with this approval of crizotinib. Hence, an ideal and adequate CDx will have to be able to identify ROS1 fusions without the knowledge of the fusion partners to ROS1, and as to date there are 13 fusion partners reported for ROS1 in NSCLC. Here we report a novel TPD52L1-ROS1 fusion variant in NSCLC. This novel TPD52L1-ROS1 fusion variant is generated by the fusion of exons 1-3 of TPD52L1 on chromosome 6q22-23 to the exons 33-43 of ROS1 on chromosome 6q22, likely from an intra-chromosomal deletion and subsequent fusion event similar to the generation of EML4-ALK. The predicted TPD52L1-ROS1 protein product contains 655 amino acids comprising of the N-terminal amino acids 1-95 of TPD52L1 and C-terminal amino acids of 1789-2348 of ROS1. In summary, TPD52L1-ROS1 is a novel ROS1 fusion variant in NSCLC identified by comprehensive genomic profiling and should be included in any ROS1 detecting assays that depend on identifying the corresponding fusion partners, such as reverse transcriptase-polymerase chain reaction (RT-PCR).

A recurrent endometrial stromal sarcoma harbors the novel fusion JAZF1-BCORL1

Highlights • Genomic alterations may improve diagnostic certainty and subsequent treatment of endometrial stromal sarcoma.• Novel JAZF1-BCORL1 mutation was identified.• Targeted therapeutics to down-stream targets may improve survival benefit in these patients.