Oncogenic switch and single-agent MET inhibitor sensitivity in a subset of EGFR-mutant lung cancer.

The clinical efficacy of epidermal growth factor receptor (EGFR)­targeted therapy in EGFR-mutant non­small cell lung cancer is limited by the development of drug resistance. One mechanism of EGFR inhibitor resistance occurs through amplification of the human growth factor receptor (MET) proto-oncogene, which bypasses EGFR to reactivate downstream signaling. Tumors exhibiting concurrent EGFR mutation and MET amplification are historically thought to be codependent on the activation of both oncogenes. Hence, patients whose tumors harbor both alterations are commonly treated with a combination of EGFR and MET tyrosine kinase inhibitors (TKIs). Here, we identify and characterize six patient-derived models of EGFR-mutant, MET-amplified lung cancer that have switched oncogene dependence to rely exclusively on MET activation for survival. We demonstrate in this MET-driven subset of EGFR TKI-refractory cancers that canonical EGFR downstream signaling was governed by MET, even in the presence of sustained mutant EGFR expression and activation. In these models, combined EGFR and MET inhibition did not result in greater efficacy in vitro or in vivo compared to single-agent MET inhibition. We further identified a reduced EGFR:MET mRNA expression stoichiometry as associated with MET oncogene dependence and single-agent MET TKI sensitivity. Tumors from 10 of 11 EGFR inhibitor­resistant EGFR-mutant, MET-amplified patients also exhibited a reduced EGFR:MET mRNA ratio. Our findings reveal that a subset of EGFR-mutant, MET-amplified lung cancers develop dependence on MET activation alone, suggesting that such patients could be treated with a single-agent MET TKI rather than the current standard-of-care EGFR and MET inhibitor combination regimens.

Discrimination of Germline EGFR T790M Mutations in Plasma Cell-Free DNA Allows Study of Prevalence Across 31,414 Cancer Patients.

Purpose: Plasma cell-free DNA (cfDNA) analysis is increasingly used clinically for cancer genotyping, but may lead to incidental identification of germline-risk alleles. We studied EGFR T790M mutations in non-small cell lung cancer (NSCLC) toward the aim of discriminating germline and cancer-derived variants within cfDNA.Experimental Design: Patients with EGFR-mutant NSCLC, some with known germline EGFR T790M, underwent plasma genotyping. Separately, deidentified genomic data and buffy coat specimens from a clinical plasma next-generation sequencing (NGS) laboratory were reviewed and tested.Results: In patients with germline T790M mutations, the T790M allelic fraction (AF) in cfDNA approximates 50%, higher than that of EGFR driver mutations. Review of plasma NGS results reveals three groups of variants: a low-AF tumor group, a heterozygous group (∼50% AF), and a homozygous group (∼100% AF). As the EGFR driver mutation AF increases, the distribution of the heterozygous group changes, suggesting increased copy number variation from increased tumor content. Excluding cases with high copy number variation, mutations can be differentiated into somatic variants and incidentally identified germline variants. We then developed a bioinformatic algorithm to distinguish germline and somatic mutations; blinded validation in 21 cases confirmed a 100% positive predictive value for predicting germline T790M. Querying a database of 31,414 patients with plasma NGS, we identified 48 with germline T790M, 43 with nonsquamous NSCLC (P < 0.0001).Conclusions: With appropriate bioinformatics, plasma genotyping can accurately predict the presence of incidentally detected germline risk alleles. This finding in patients indicates a need for genetic counseling and confirmatory germline testing. Clin Cancer Res; 23(23); 7351-9. ©2017 AACR.

Assigning clinical meaning to somatic and germ-line whole-exome sequencing data in a prospective cancer precision medicine study.

PURPOSE: Implementing cancer precision medicine in the clinic requires assessing the therapeutic relevance of genomic alterations. A main challenge is the systematic interpretation of whole-exome sequencing (WES) data for clinical care. METHODS: One hundred sixty-five adults with metastatic colorectal and lung adenocarcinomas were prospectively enrolled in the CanSeq study. WES was performed on DNA extracted from formalin-fixed paraffin-embedded tumor biopsy samples and matched blood samples. Somatic and germ-line alterations were ranked according to therapeutic or clinical relevance. Results were interpreted using an integrated somatic and germ-line framework and returned in accordance with patient preferences. RESULTS: At the time of this analysis, WES had been performed and results returned to the clinical team for 165 participants. Of 768 curated somatic alterations, only 31% were associated with clinical evidence and 69% with preclinical or inferential evidence. Of 806 curated germ-line variants, 5% were clinically relevant and 56% were classified as variants of unknown significance. The variant review and decision-making processes were effective when the process was changed from that of a Molecular Tumor Board to a protocol-based approach. CONCLUSION: The development of novel interpretive and decision-support tools that draw from scientific and clinical evidence will be crucial for the success of cancer precision medicine in WES studies.Genet Med advance online publication 26 January 2017.

Association Between Younger Age and Targetable Genomic Alterations and Prognosis in Non-Small-Cell Lung Cancer.

IMPORTANCE: Non-small-cell lung cancer (NSCLC) diagnosed in young patients is rare, and the genomics and clinical characteristics of this disease are poorly understood. In contrast, the diagnosis of other cancers at a young age has been demonstrated to define unique disease biology. Herein, we report on the association of young age with targetable genomic alterations and prognosis in a cohort of 2237 patients with NSCLC. OBJECTIVE: To determine the relationship between young age at diagnosis and the presence of a potentially targetable genomic alteration, disease prognosis, and natural history. DESIGN, SETTING, AND PARTICIPANTS: A cohort of all 2237 patients with NSCLC who were genotyped at the Dana-Farber Cancer Institute between January 2002 and December 2014 were identified. Tumor genotype, patient characteristics, and clinical outcomes were collected and studied at a National Cancer Institute-designated comprehensive cancer center. Multivariate logistic regression was used to analyze the relationship between age and mutation status, and multivariate Cox proportional hazard models were fitted for survival analysis. MAIN OUTCOMES AND MEASURES: The frequency of targetable genomic alterations by defined age categories as well as the association of these age groups with survival. Age categories used in this analysis were younger than 40, 40 to 49, 50 to 59, 60 to 69, and 70 years or older. RESULTS: A cohort of 2237 patients with NSCLC was studied. Of the 2237 participants, 1939 (87%) had histologically confirmed adenocarcinoma, 269 (12%) had NSCLC not otherwise specified, and 29 (1%) had squamous histologic findings; 1396 (63%) had either stage IIIB or IV cancers; and median (range) age was 62 (20-95) years. We found that gene mutations for EGFR (P = .02) and ALK (P < .001) were associated with cancer diagnosis at a younger age, and a similar trend existed for ERBB2 (P = .15) and ROS1 (P = .10) but not BRAF V600E (P = .43). Among patients tested for all 5 targetable genomic alterations (n = 1325), younger age was associated with an increased frequency of a targetable genotype (P < .001). Those diagnosed at 50 years or younger have a 59% increased likelihood of harboring a targetable genotype. While presence of a potentially targetable genomic alteration treated with a targeted agent was associated with improved survival, the youngest and oldest age groups had similarly poor outcomes even when a targetable genotype was present. CONCLUSIONS AND RELEVANCE: Younger age is associated with an increased likelihood of harboring a targetable genotype and is an underappreciated clinical biomarker in NSCLC. The survival of young patients with NCSLC is unexpectedly poor compared with other age groups, suggesting more aggressive disease biology. These findings underscore the importance of comprehensive genotyping, including next-generation sequencing, in younger patients with lung cancer.

MET Exon 14 Mutations in Non-Small-Cell Lung Cancer Are Associated With Advanced Age and Stage-Dependent MET Genomic Amplification and c-Met Overexpression.

PURPOSE: Non-small-cell lung cancers (NSCLCs) harboring mutations in MET exon 14 and its flanking introns may respond to c-Met inhibitors. We sought to describe the clinical, pathologic, and genomic characteristics of patients with cancer with MET exon 14 mutations. PATIENTS AND METHODS: We interrogated next-generation sequencing results from 6,376 cancers to identify those harboring MET exon 14 mutations. Clinical characteristics of MET exon 14 mutated NSCLCs were compared with those of NSCLCs with activating mutations in KRAS and EGFR. Co-occurring genomic mutations and copy number alterations were identified. c-Met immunohistochemistry and real-time polymerase chain reaction to detect exon 14 skipping were performed where sufficient tissue was available. RESULTS: MET exon 14 mutations were identified in 28 of 933 nonsquamous NSCLCs (3.0%) and were not seen in other cancer types in this study. Patients with MET exon 14-mutated NSCLC were significantly older (median age, 72.5 years) than patients with EGFR-mutant (median age, 61 years; P < .001) or KRAS-mutant NSCLC (median age, 65 years; P < .001). Among patients with MET exon 14 mutations, 68% were women, and 36% were never-smokers. Stage IV MET exon 14-mutated NSCLCs were significantly more likely to have concurrent MET genomic amplification (mean ratio of MET to chromosome 7, 4.3) and strong c-Met immunohistochemical expression (mean H score, 253) than stage IA to IIIB MET exon 14-mutated NSCLCs (mean ratio of MET to chromosome 7, 1.4; P = .007; mean H score, 155; P = .002) and stage IV MET exon 14-wild-type NSCLCs (mean ratio of MET to chromosome 7, 1.2; P < .001; mean H score, 142; P < .001). A patient whose lung cancer harbored a MET exon 14 mutation with concurrent genomic amplification of the mutated MET allele experienced a major partial response to the c-Met inhibitor crizotinib. CONCLUSION: MET exon 14 mutations represent a clinically unique molecular subtype of NSCLC. Prospective clinical trials with c-Met inhibitors will be necessary to validate MET exon 14 mutations as an important therapeutic target in NSCLC.

Clinical Implications of Variant ALK FISH Rearrangement Patterns.

INTRODUCTION: Break-apart fluorescence in situ hybridization (FISH) is the FDA-approved assay for detecting anaplastic lymphoma kinase (ALK) rearrangements in non-small-cell lung cancer (NSCLC), identifying patients who can gain dramatic benefit from ALK kinase inhibitors. Assay interpretation can be technically challenging, and either splitting of the 5' and 3' probes or loss of the 5' probe constitute rearrangement. We hypothesized that there may be clinical differences depending on rearrangement pattern on FISH. METHODS: An IRB-approved database of NSCLC patients at Dana-Farber Cancer Institute was queried for ALK rearrangement. Clinical characteristics and response to crizotinib were reviewed. Immunohistochemistry (IHC) and targeted next-generation sequencing (NGS) were obtained when available. RESULTS: Of 1614 NSCLC patients with ALK testing, 82 patients (5.1%) had ALK rearrangement by FISH: 30 patients with split signals, 25 patients with 5' deletion, and 27 patients with details unavailable. Patients with 5' deletion were older (p = 0.01) and tended to have more extensive smoking histories (p = 0.08). IHC was positive for ALK rearrangement in all 27 patients with FISH split signals, whereas three of 21 patients with FISH 5' deletion had negative IHC (p = 0.05). Targeted NGS on two of three cases with discordant FISH and IHC results did not identify ALK rearrangement, instead finding driver mutations in EGFR and KRAS. Patients with 5' deletion treated with crizotinib had a smaller magnitude of tumor response (p = 0.03). CONCLUSIONS: Patients with 5' deletion on ALK FISH harbor features less typical of ALK-rearranged tumors, potentially indicating that some cases with this variant are false positives. Corroborative testing with IHC or NGS may be beneficial.