Christine A. Lydon

Clinical, Pathologic, and Biologic Features Associated with BRAF Mutations in Non–Small Cell Lung Cancer

Purpose: BRAF mutations are found in a subset of non–small cell lung cancers (NSCLC). We examined the clinical characteristics and treatment outcomes of patients with NSCLC harboring BRAF mutations. Experimental Design: Using DNA sequencing, we successfully screened 883 patients with NSCLC for BRAF mutations between July 1, 2009 and July 16, 2012. Baseline characteristics and treatment outcomes were compared between patients with and without BRAF mutations. Wild-type controls consisted of patients with NSCLC without a somatic alteration in BRAF, KRAS, EGFR, and ALK. In vitro studies assessed the biologic properties of selected non-V600E BRAF mutations identified from patients with NSCLC. Results: Of 883 tumors screened, 36 (4%) harbored BRAF mutations (V600E, 18; non-V600E, 18) and 257 were wild-type for BRAF, EGFR, KRAS, and ALK negative. Twenty-nine of 36 patients with BRAF mutations were smokers. There were no distinguishing clinical features between BRAF-mutant and wild-type patients. Patients with advanced NSCLC with BRAF mutations and wild-type tumors showed similar response rates and progression-free survival (PFS) to platinum-based combination chemotherapy and no difference in overall survival. Within the BRAF cohort, patients with V600E-mutated tumors had a shorter PFS to platinum-based chemotherapy compared with those with non-V600E mutations, although this did not reach statistical significance (4.1 vs. 8.9 months; P = 0.297). We identified five BRAF mutations not previously reported in NSCLC; two of five were associated with increased BRAF kinase activity. Conclusions: BRAF mutations occur in 4% of NSCLCs and half are non-V600E. Prospective trials are ongoing to validate BRAF as a therapeutic target in NSCLC. Clin Cancer Res; 19(16); 4532–40. ©2013 AACR.

Expression of PD-1 and Its Ligands, PD-L1 and PD-L2, in Smokers and Never Smokers with KRAS-Mutant Lung Cancer

Introduction: Clinical responses to immune checkpoint blockade by anti-programmed cell death protein-1 (PD-1)/PD-L1 monoclonal antibodies in non–small-cell lung cancer (NSCLC) are associated with PD-L1 expression and smoking status. We aimed to determine the expression profile of PD-1 and its ligands, PD-L1 and PD-L2, in both smokers and never smokers patients with KRAS-mutant NSCLC. Methods: We retrospectively analyzed the clinical and molecular characteristics of 114 KRAS-mutant NSCLC patients (84 smokers and 30 never smokers) and their clinical tumor samples for the expression of PD-1, PD-L1, and PD-L2 by immunohistochemistry (IHC). We used murine monoclonal antibodies anti-PD-L1 (clone 9A11) and anti-PD-L2 (clone 9E5) to examine for tumor cell expression (0, negative; 1, weak; 2, moderate; 3, intense) and anti-PD-1 (clone EH33) for tumor-infiltrating lymphocytes. Results: PD-L1 expression was detected in 27 of 114 patients (24%; 95% confidence interval [CI], 16–33%) and associated with smoking status (current smokers, 44%; former smokers, 20%; never smokers, 13%; p = 0.03). Higher intensity of PD-L1 expression (IHC-2+/IHC-3+) was more frequently observed in smokers and associated with more pack-years. PD-L2 was positive in 54 of 114 patients (47%; 95% CI, 38–57%) and not related to smoking status. PD-1–positive tumor-infiltrating lymphocytes were present in 77 of 114 tumor specimens tested (68%; 95% CI, 59–77%) including 21 of 27 samples with PD-L1 expression and 39 of 54 samples with PD-L2–positive expression. We found that PD-L1 expression fades with the age of the specimens used for analyses decreasing beyond 3 years (p = 0.016). Conclusions: The expression of PD-1 and its ligands PD-L1 and PD-L2 is heterogeneous within KRAS-mutant NSCLC and suggests an inducible expression of PD-L1 by smoking.

Immunohistochemical loss of LKB1 is a biomarker for more aggressive biology in KRAS mutant lung adenocarcinoma.

Purpose: LKB1 loss is common in lung cancer, but no assay exists to efficiently evaluate the presence or absence of LKB1. We validated an IHC assay for LKB1 loss and determined the impact of LKB1 loss in KRAS-mutant non–small cell lung cancer (NSCLC). Experimental Design: We optimized and validated an IHC assay for LKB1 (clone Ley37D/G6) using a panel of lung cancer cell lines and tumors with known LKB1 mutations, including 2 patients with Peutz–Jeghers syndrome (PJS) who developed lung adenocarcinoma. We retrospectively analyzed tumors for LKB1 using IHC from 154 KRAS-mutant NSCLC patients, including 123 smokers and 31 never-smokers, and correlated the findings with patient and tumor characteristics and clinical outcome. Results: LKB1 expression was lost by IHC in 30% of KRAS-mutant NSCLC (smokers 35% vs. never-smokers 13%, P = 0.017). LKB1 loss did not correlate with a specific KRAS mutation but was more frequent in tumors with KRAS transversion mutations (P = 0.029). KRAS-mutant NSCLC patients with concurrent LKB1 loss had a higher number of metastatic sites at the time of diagnosis (median 2.5 vs. 2, P = 0.01), higher incidence of extrathoracic metastases (P = 0.01), and developed brain metastasis more frequently (48% vs. 25%, P = 0.02). There was a nonsignificant trend to worse survival in stage IV KRAS-mutant NSCLC patients with LKB1 loss. Conclusions: LKB1 IHC is a reliable and efficient assay to evaluate for loss of LKB1 in clinical samples of NSCLC. LKB1 loss is more common in smokers, and is associated with a more aggressive clinical phenotype in KRAS-mutant NSCLC patients, accordingly to preclinical models. Clin Cancer Res; 21(12); 2851–60. ©2015 AACR.

Long-term Benefit of PD-L1 Blockade in Lung Cancer Associated with JAK3 Activation.

Van Allen, Golay, Liu, and colleagues genomically profiled tumor and germline samples from a patient with activating JAK3 variants who achieved long-term clinical benefit from anti–PD-L1 therapy, suggesting that alterations in JAK signaling may be immunogenomic modulators of response to PD-L1 immunotherapy. PD-1 immune checkpoint blockade occasionally results in durable clinical responses in advanced metastatic cancers. However, mechanism-based predictors of response to this immunotherapy remain incompletely characterized. We performed comprehensive genomic profiling on a tumor and germline sample from a patient with refractory lung adenocarcinoma who achieved marked long-term clinical benefit from anti–PD-L1 therapy. We discovered activating somatic and germline amino acid variants in JAK3 that promoted PD-L1 induction in lung cancer cells and in the tumor immune microenvironment. These findings suggest that genomic alterations that deregulate cytokine receptor signal transduction could contribute to PD-L1 activation and engagement of the PD-1 immune checkpoint in lung cancer. Cancer Immunol Res; 3(8); 855–63. ©2015 AACR.

Five-Year Survival in EGFR-Mutant Metastatic Lung Adenocarcinoma Treated with EGFR-TKIs

Introduction: Activating mutations in the epidermal growth factor receptor gene (EGFR) predict for prolonged progression‐free survival in patients with advanced non–small cell lung cancer (NSCLC) treated with EGFR tyrosine kinase inhibitors (EGFR‐TKIs) versus chemotherapy. Long‐term survival outcomes, however, remain undefined. The objective of this study was to determine the 5‐year survival in these patients and identify clinical factors associated with overall survival (OS). Methods: Patients with EGFR‐mutant metastatic lung adenocarcinoma who had been treated with erlotinib or gefitinib at Dana‐Farber Cancer Institute between 2002 and 2009 were included. OS was analyzed. Results: Among 137 patients, median progression‐free survival and OS were 12.1 months (95% CI: 10.2–13.5) and 30.9 months (95% CI: 28.2–35.7), respectively. Twenty patients (14.6%) were 5‐year survivors. In multivariate analysis, exon 19 deletions (hazard ratio [HR] = 0.63, 95% CI: 0.44–0.91, p = 0.01), absence of extrathoracic (HR = 0.62, 95% CI: 0.41–0.93, p = 0.02) or brain metastasis (HR = 0.48, 95% CI: 0.30–0.77, p = 0.002), and not a current smoker (HR = 0.23, 95% CI: 0.09–0.59, p = 0.002) were associated with prolonged OS. Age; sex; stage at diagnosis; liver, bone, or adrenal metastasis; specific TKI; and line of TKI therapy were not associated with OS. Conclusions: Our data suggest that the rate of 5‐year survival among patients with EGFR‐mutant metastatic lung adenocarcinoma treated with erlotinib or gefitinib is 14.6%. Exon 19 deletions and absence of extrathoracic or brain metastasis are associated with prolonged survival. On the basis of our findings, clinicians can gain an enhanced estimation of long‐term outcomes in this population.

Acquired METD1228V Mutation and Resistance to MET Inhibition in Lung Cancer

Amplified and/or mutated MET can act as both a primary oncogenic driver and as a promoter of tyrosine kinase inhibitor (TKI) resistance in non-small cell lung cancer (NSCLC). However, the landscape of MET-specific targeting agents remains underdeveloped, and understanding of mechanisms of resistance to MET TKIs is limited. Here, we present a case of a patient with lung adenocarcinoma harboring both a mutation in EGFR and an amplification of MET, who after progression on erlotinib responded dramatically to combined MET and EGFR inhibition with savolitinib and osimertinib. When resistance developed to this combination, a new MET kinase domain mutation, D1228V, was detected. Our in vitro findings demonstrate that METD1228V induces resistance to type I MET TKIs through impaired drug binding, while sensitivity to type II MET TKIs is maintained. Based on these findings, the patient was treated with erlotinib combined with cabozantinib, a type II MET inhibitor, and exhibited a response. SIGNIFICANCE With several structurally distinct MET inhibitors undergoing development for treatment of NSCLC, it is critical to identify mechanism-based therapies for drug resistance. We demonstrate that an acquired METD1228V mutation mediates resistance to type I, but not type II, MET inhibitors, having therapeutic implications for the clinical use of sequential MET inhibitors. Cancer Discov; 6(12); 1334-41. ©2016 AACR.See related commentary by Trusolino, p. 1306This article is highlighted in the In This Issue feature, p. 1293.

Prognostic Impact of KRAS Mutation Subtypes in 677 Patients with Metastatic Lung Adenocarcinomas

Background: We previously demonstrated that patients with metastatic KRAS mutant lung cancers have a shorter survival compared with patients with KRAS wild-type cancers. Recent reports have suggested different clinical outcomes and distinct activated signaling pathways depending on KRAS mutation subtype. To better understand the impact of KRAS mutation subtype, we analyzed data from 677 patients with KRAS mutant metastatic lung cancer. Methods: We reviewed all patients with metastatic or recurrent lung cancers found to have KRAS mutations over a 6-year time period. We evaluated the associations among KRAS mutation type, clinical factors, and overall survival in univariate and multivariate analyses. Any significant findings were validated in an external multi-institution patient dataset. Results: Among 677 patients with KRAS mutant lung cancers (53 at codon 13, 624 at codon 12), there was no difference in overall survival for patients when comparing KRAS transition versus transversion mutations (p = 0.99), smoking status (p = 0.33), or when comparing specific amino acid substitutions (p = 0.20). In our dataset, patients with KRAS codon 13 mutant tumors (n = 53) had shorter overall survival compared with patients with codon 12 mutant tumors (n = 624) (1.1 versus 1.3 years, respectively; p = 0.009), and the findings were confirmed in a multivariate Cox model controlling for age, sex, and smoking status (hazard ratio: 1.52, 95% confidence interval: 1.11–2.08; p = 0.008). In an independent validation set of tumors from 682 patients with stage IV KRAS mutant lung cancers, there was no difference in survival between patients with KRAS codon 13 versus codon 12 mutations (1.0 versus 1.1 years, respectively; p = 0.41). Conclusions: Among individuals with KRAS mutant metastatic lung cancers treated with conventional therapy, there are no apparent differences in outcome based on KRAS mutation subtype.

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

Response heterogeneity of EGFR and HER2 exon 20 insertions to covalent EGFR and HER2 inhibitors

Insertion mutations in EGFR and HER2 both occur at analogous positions in exon 20. Non-small cell lung cancer (NSCLC) patients with tumors harboring these mutations seldom achieve clinical responses to dacomitinib and afatinib, two covalent quinazoline-based inhibitors of EGFR or HER2, respectively. In this study, we investigated the effects of specific EGFR and HER2 exon 20 insertion mutations from NSCLC patients that had clinically achieved a partial response after dacomitinib treatment. We identified Gly770 as a common feature among the drug-sensitive mutations. Structural modeling suggested that this mutation may facilitate inhibitor binding to EGFR. Introduction of Gly770 into two dacomitinib-resistant EGFR exon 20 insertion mutants restored sensitivity to dacomitinib. Based on these findings, we used afatinib to treat an NSCLC patient whose tumor harbored the HER2 V777_G778insGSP mutation and achieved a durable partial response. We further identified secondary mutations in EGFR (T790M or C797S) and HER2 (C805S) that mediated acquired drug resistance in drug-sensitive EGFR or HER2 exon 20 insertion models. Overall, our findings identified a subset of EGFR and HER2 exon 20 insertion mutations that are sensitive to existing covalent quinazoline-based EGFR/HER2 inhibitors, with implications for current clinical treatment and next-generation small-molecule inhibitors. Cancer Res; 77(10); 2712-21. ©2017 AACR.

Interstitial lung abnormalities in treatment-naïve advanced non-small-cell lung cancer patients are associated with shorter survival

OBJECTIVE Interstitial lung diseases are associated with increased risk of lung cancer. The prevalence of ILA at diagnosis of advanced non-small-cell lung cancer (NSCLC) and its impact on overall survival (OS) remain to be investigated. MATERIALS AND METHOD The study included 120 treatment-naïve stage IV NSCLC patients (53 males, 67 females). ILA was scored on CT prior to any systemic therapy using a 4-point scale [0=no evidence of ILA, 1=equivocal for ILA, 2=suspicious for ILA, 3=ILA] by a sequential reading method previously reported. ILA scores of 2 or 3 indicated the presence of ILA. RESULTS ILA was present in 17 patients (14%) with advanced NSCLC prior to any treatment (score3: n=2, score2: n=15). These 17 patients were significantly older (median age: 69 vs. 63, p=0.04) and had a heavier smoking history (median: 40 vs. 15.5 pack-year, p=0.003) than those with ILA score 0 or 1. Higher ILA scores were associated with shorter OS (p=0.001). Median OS of the 17 patients with ILA was 7.2 months [95%CI: 2.9-9.4] compared to 14.8 months [95%CI: 11.1-18.4] in patients with ILA score 0 or 1 (p=0.002). In a multivariate model, the presence of ILA remained significant for increased risk for death (HR=2.09, p=0.028) after adjusting for first-line systemic therapy (chemotherapy, p<0.001; TKI, p<0.001; each compared to no therapy) and pack years of smoking (p=0.40). CONCLUSION Radiographic ILA was present in 14% of treatment-naïve advanced NSCLC patients. Higher ILA scores were associated with shorter OS, indicating that ILA could be a marker of shorter survival in advanced NSCLC.