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Purpose Preclinical data and subset analyses from immunotherapy clinical trials indicate that prior radiation therapy was associated with better progression-free survival and overall survival when combined with immune checkpoint inhibitors in patients with non-small cell lung cancer. We present a prospective study of hypofractionated image guided radiation therapy (HIGRT) to a single site of metastatic disease concurrently with atezolizumab in patients with metastatic non-small cell lung cancer. Methods and Materials Patients meeting eligibility criteria received 1200 mg of atezolizumab intravenously every 3 weeks with concurrent 3- or 5-fraction HIGRT starting no later than the second cycle. The 3-fraction regimen employed a minimum of 8 Gy per fraction compared with 6 Gy for the 5-fraction regimen. Imaging was obtained every 12 weeks to assess response. Results From October 2015 to February 2017, 12 patients were enrolled in the study (median age 64; range, 55-77 years). The best response by the Response Evaluation in Solid Tumors criteria was partial response in 3 and stable disease in 3, for a disease control rate of 50%. Five patients had a grade 3 immune-related adverse event, including choreoretinitis (n = 1), pneumonitis (n = 1), transaminitis (n = 1), fatigue (n = 1), and peripheral neuropathy (n = 1). The median progression-free survival was 2.3 months, and the median overall survival was 6.9 months (range, 0.4-not reached). There was no clear association between peripheral blood T cell repertoire characteristics at baseline, PD-L1, or tumor mutations and response or outcome. One long-term survivor exhibited oligoclonal T cell populations in a baseline tumor biopsy that were consistently detected in peripheral blood over the entire course of the study. Conclusions HIGRT plus atezolizumab resulted in an overall response rate of 25% and disease control rate of 50% in this pilot study. The incidence of grade 3 adverse events was similar to that of atezolizumab alone. Alhough it was a pilot study with limited sample size, the results generated hypotheses worthy of further investigation.

Small cell lung cancer (SCLC) is an aggressive malignancy that accounts for 15% of all lung cancers. It is characterized by initial responsiveness to therapy followed by rapid disease progression that is relatively resistant to further treatment. Recently, the addition of an immune checkpoint inhibitor (ICI) to chemotherapy has improved survival in patients with advanced disease, the first advance in systemic therapy in SCLC in over 30 years.In this review, we present an overview of SCLC with a focus on the scope of the problem and standard treatment, followed by a critical assessment of scientific rationale for immunotherapy in SCLC and the clinical trials that have been performed with ICIs in SCLC. Finally, we address ongoing hurdles for the development of ICIs in SCLC and potential avenues for further study.Despite solid biological rationale, the results of clinical trials of ICIs in SCLC have yielded modest benefits. A small subset of patients does achieve long-term benefit, but further development of ICIs in SCLC will depend on the identification of predictive biomarkers and the design of combination regimens that take advantage of the molecular alterations that drive the immune-avoidance mechanisms and survival of SCLC cells.

Purpose Immunotherapy is a relatively new treatment modality for advanced non-small cell lung cancer following platinum-based chemotherapy. Nivolumab, pembrolizumab, and atezolizumab demonstrated superior outcomes and improved tolerability compared to standard treatment in randomized controlled trials; however, these studies vary significantly in inclusion criteria and study design. To our knowledge, the efficacy and safety of nivolumab and atezolizumab following platinum-based chemotherapy have not been directly compared to one another in a real-world clinic setting. Methods We retrospectively compared immunotherapy response rates and toxicity in patients with stage IV or recurrent non-small cell lung cancer following progression during or after platinum-based chemotherapy. Results Among 124 eligible patients, the objective response rate was 14.8% in the nivolumab group (n = 81) vs. 13.9% in the atezolizumab group (n = 43) (p = 0.897). Median overall survival was 8.4 months with nivolumab (95% confidence interval (CI), 6.3 to 11.2) vs. 6.5 months with atezolizumab (95% CI, 4.7 to not reached). Median progression free survival was 2.2 months (95% CI, 1.7 to 2.8) and 2.0 months (95% CI, 1.8 to 2.7) in the nivolumab and atezolizumab groups, respectively. Treatment-related adverse events occurred in 70.4% of patients in the nivolumab group and 65.1% in the atezolizumab group. Conclusions There was no statistically significant difference in efficacy outcomes in patients with non-small cell lung cancer who received atezolizumab or nivolumab after progression during or after platinum-based chemotherapy. Response rates in this study were numerically lower than response rates observed in the landmark randomized controlled trials leading to approval of immunotherapy in this setting. Rates of treatment-related adverse events were similar between groups.

3079 Background: RET fusions are targetable oncogenic drivers in multiple solid tumor types. ARROW study (NCT03037385) data supported the US FDA approval of pralsetinib, a once-daily (QD) oral highly potent and selective RET inhibitor, for RET-altered metastatic non-small cell lung cancer (NSCLC) and advanced/metastatic thyroid cancer. Here we provide an update on the clinical activity of pralsetinib in patients (pts) with advanced RET fusion-positive solid tumors other than NSCLC and thyroid cancer (“other” RET fusion–positive solid tumors). Methods: The global ongoing ARROW study (84 sites in 13 countries) includes phase 1 dose-escalation (30–600 mg [QD or twice daily]) and phase 2 expansion cohorts (400 mg QD) defined by tumor type and RET alteration status. Primary objectives are overall response rate (ORR; blinded independent central review per RECIST v1.1) and safety. Results: Updated analyses were completed as of Nov 6, 2020 (data cut-off) for 21 pts with other RET fusion–positive solid tumors enrolled by May 22, 2020 (enrollment cut-off) (lung other than NSCLC, n = 4; pancreatic, n = 3; colon, n = 3; cholangiocarcinoma, n = 3; unknown primary [UP], n = 2; other, n = 6). Overall, 11 (52%) pts received ≥2 prior lines of therapy for metastatic disease. The most common RET fusion partners were CCDC6 and KIF5B (24% each), NCOA4 (19%), other (10%), and unknown (24%). Two pts with colon cancer were excluded from efficacy analyses due to other driver mutations ( KRAS, PIK3CB). In 19 evaluable pts, ORR was 53% (95% CI, 29–76) with 2 (11%) complete responses (CR) and 8 (42%) partial responses (PR). Responses occurred across multiple tumor types including 3/3 pts with pancreatic cancer (including a CR ongoing at 20.8 months on treatment), 2/2 pts with UP, 2/3 pts with cholangiocarcinoma, and in pts with mesenchymal, salivary duct, and lung carcinoid tumors. Median duration of response was 19.0 months (95% CI, 5.5–not estimable). Clinical benefit rate (proportion with CR, PR, or stable disease persisting ≥16 weeks) was 68% (95% CI, 43–87). Tumor shrinkage was observed in 89% of 18 evaluable pts with post-baseline tumor assessment. In all pts enrolled in ARROW who received pralsetinib 400 mg QD irrespective of tumor type (n = 471) the most common (≥25%) treatment-related adverse events (TRAEs) were increased aspartate aminotransferase (39%), anemia (35%), increased alanine aminotransferase (28%), constipation (26%), and hypertension (25%). Overall, 6% of pts discontinued treatment due to TRAEs. Conclusions: Pralsetinib showed robust, durable antitumor activity in patients with multiple RET fusion‒positive, heavily pre-treated, advanced solid tumors, and was well tolerated. These data highlight the need for broad RET testing to identify candidates who could benefit from treatment with pralsetinib. Enrollment of patients with other RET fusion–positive solid tumors in ARROW is ongoing. Clinical trial information: NCT03037385.

PURPOSE Pembrolizumab monotherapy has shown antitumor activity in patients with small-cell lung cancer (SCLC). The randomized, double-blind, phase III KEYNOTE-604 study compared pembrolizumab plus etoposide and platinum (EP) with placebo plus EP for patients with previously untreated extensive-stage (ES) SCLC. METHODS Eligible patients were randomly assigned 1:1 to pembrolizumab 200 mg once every 3 weeks or saline placebo for up to 35 cycles plus 4 cycles of EP. Primary end points were progression-free survival (PFS; RECIST version 1.1, blinded central review) and overall survival (OS) in the intention-to-treat population. Objective response rate (ORR) and duration of response were secondary end points. Prespecified efficacy boundaries were one-sided P = .0048 for PFS and .0128 for OS. RESULTS Of the 453 participants, 228 were randomly assigned to pembrolizumab plus EP and 225 to placebo plus EP. Pembrolizumab plus EP significantly improved PFS (hazard ratio [HR], 0.75; 95% CI, 0.61 to 0.91; P = .0023). Twelve-month PFS estimates were 13.6% with pembrolizumab plus EP and 3.1% with placebo plus EP. Although pembrolizumab plus EP prolonged OS, the significance threshold was not met (HR, 0.80; 95% CI, 0.64 to 0.98; P = .0164). Twenty-four-month OS estimates were 22.5% and 11.2%, respectively. ORR was 70.6% in the pembrolizumab plus EP group and 61.8% in the placebo plus EP group; the estimated proportion of responders remaining in response at 12 months was 19.3% and 3.3%, respectively. In the pembrolizumab plus EP and placebo plus EP groups, respectively, any-cause adverse events were grade 3-4 in 76.7% and 74.9%, grade 5 in 6.3% and 5.4%, and led to discontinuation of any drug in 14.8% and 6.3%. CONCLUSION Pembrolizumab plus EP significantly improved PFS compared with placebo plus EP as first-line therapy for patients with ES-SCLC. No unexpected toxicities were seen with pembrolizumab plus EP. These data support the benefit of pembrolizumab in ES-SCLC.

Radiation esophagitis (RE) is a common adverse event associated with radiotherapy for non–small cell lung cancer (NSCLC). While plasma cytokine levels have been correlated with other forms of radiation-induced toxicity, their association with RE has been less well studied. We analyzed data from 126 patients treated on 4 prospective clinical trials. Logistic regression models based on combinations of dosimetric factors [maximum dose to 2 cubic cm (D2cc) and generalized equivalent uniform dose (gEUD)], clinical variables, and pretreatment plasma levels of 30 cytokines were developed. Cross-validated estimates of area under the receiver operating characteristic curve (AUC) and log likelihood were used to assess prediction accuracy. Dose-only models predicted grade 3 RE with AUC values of 0.750 (D2cc) and 0.727 (gEUD). Combining clinical factors with D2cc increased the AUC to 0.779. Incorporating pretreatment cytokine measurements, modeled as direct associations with RE and as potential interactions with the dose-esophagitis association, produced AUC values of 0.758 and 0.773, respectively. D2cc and gEUD correlated with grade 3 RE with odds ratios (ORs) of 1.094/Gy and 1.096/Gy, respectively. Female gender was associated with a higher risk of RE, with ORs of 1.09 and 1.112 in the D2cc and gEUD models, respectively. Older age was associated with decreased risk of RE, with ORs of 0.992/year and 0.991/year in the D2cc and gEUD models, respectively. Combining clinical with dosimetric factors but not pretreatment cytokine levels yielded improved prediction of grade 3 RE compared to prediction by dose alone. Such multifactorial modeling may prove useful in directing radiation treatment planning.

Small-cell lung cancer (SCLC) is an aggressive tumor that accounts for 13–15% of all lung cancer cases and is the seventh most common cause of cancer-related death in the U.S. with a 5-year overall survival rate of 6.3% (1). At initial diagnosis, about 2/3 of patients have extensive-stage disease (ES-SCLC), for which all treatment is given with palliative intent. Platinum-based, two-drug chemotherapy has been the standard first-line treatment for patients with ES-SCLC for over 20 years. In the U.S., the most common regimens utilized are carboplatin plus etoposide (CE) or cisplatin plus etoposide (PE), which yield a response rate of 50–70% and a median overall survival time of 8–11 months (2). Despite the remarkable activity of initial chemotherapy, nearly all patients relapse within months and the benefits of second-line therapy are limited. Thus, there is a desperate need for more effective first-line therapy.

Introduction Most patients with lung cancer are elderly and poorly represented in randomized clinical trials. They are often undertreated because of concerns about their ability to tolerate aggressive treatment. We tested the hypothesis that elderly patients undergoing definitive lung radiation might tolerate treatment differently than younger patients. Methods A total of 125 patients who underwent definitive lung radiotherapy were identified from a prospective institutional database (University of Michigan cohort). Logistic regression modeling was performed to assess the impact of age on esophagitis grade 2 or higher or grade 2 or higher and pneumonitis grade 3 or higher or grade 2 or higher, with adjustment for esophageal and lung dose, respectively, as well as for chemotherapy utilization, smoking status, and performance status. The analysis was validated in a large cohort of 691 patients from the Michigan Radiation Oncology Quality Consortium registry, an independent statewide prospective database. Results In the University of Michigan cohort, multivariable regression models revealed a significant inverse correlation between age and rate of esophagitis for both toxicity levels, (adjusted OR = 0.93 for both models and 95% confidence intervals of 0.88–0.98 and 0.87–0.99), with areas under the curve of 0.747 and 0.721, respectively, demonstrating good fit. This same association was noted in the Michigan Radiation Oncology Quality Consortium cohort. There was no significant association between age and pneumonitis. Conclusions There is a lower incidence of esophagitis with increasing age even after adjustment for use of chemotherapy. This is a novel finding in thoracic oncology. No age dependence was noted for pulmonary toxicity. The elderly are able to tolerate definitive thoracic radiation well and should be offered this option when clinically warranted.

(1) Background: Circulating tumor cell (CTC) clusters are emerging as clinically significant harbingers of metastases in solid organ cancers. Prior to engaging these CTC clusters in animal models of metastases, it is imperative for technology to identify them with high sensitivity. These clusters often present heterogeneous surface markers and current methods for isolation of clusters may fall short. (2) Methods: We applied an inertial microfluidic Labyrinth device for high-throughput, biomarker-independent, size-based isolation of CTCs/CTC clusters from patients with metastatic non-small-cell lung cancer (NSCLC). (3) Results: Using Labyrinth, CTCs (PanCK+/DAPI+/CD45&minus, ) were isolated from patients (n = 25). Heterogeneous CTC populations, including CTCs expressing epithelial (EpCAM), mesenchymal (Vimentin) or both markers were detected. CTCs were isolated from 100% of patients (417 &plusmn, 1023 CTCs/mL). EpCAM&minus, CTCs were significantly greater than EpCAM+ CTCs. Cell clusters of &ge, 2 CTCs were observed in 96% of patients&mdash, of which, 75% were EpCAM&minus, CTCs revealed identical genetic aberrations as the primary tumor for RET, ROS1 and ALK genes using fluorescence in situ hybridization (FISH) analysis. (4) Conclusions: The Labyrinth device recovered heterogeneous CTCs in 100% and CTC clusters in 96% of patients with metastatic NSCLC. The majority of recovered CTCs/clusters were EpCAM&minus, suggesting that these would have been missed using traditional antibody-based capture methods.

TPS8589 Background: Increased expression of BCL-2, BCL-xL, and MCL-1 allows certain tumors to evade apoptosis. Pelcitoclax is a novel, dual BCL-2/BCL-xL inhibitor with strong single-agent antitumor activity against tumor cells addicted to BCL-2, BCL-xL, and BCL-w, and exhibits even broader antitumor activity when administered with chemotherapy. Pelcitoclax reduces tumor growth in SCLC and other human cancer xenograft models, with manageable effects on platelet counts. Preliminary findings from the first-in-human study suggested promising antitumor activity and a favorable safety profile. Methods: This open-label study is evaluating the safety and preliminary efficacy of pelcitoclax combined with paclitaxel in adults with R/R SCLC that has progressed on or after initial treatment. Prior treatments may include platinum-based therapy (± thoracic radiation), immunotherapy, or chemotherapeutic agents other than paclitaxel. Eligible patients have an ECOG performance status of 0-2; adequate organ function; no known bleeding diathesis, immune thrombocytopenic purpura, autoimmune hemolytic anemia, serious gastrointestinal bleeding, or concomitant use of most anticoagulants; and no residual grade ≥ 2 adverse events from previous treatment. In the phase Ib study, the pelcitoclax maximum tolerated dose is being determined using a time-to-event continual reassessment method. In this phase, pelcitoclax is administered by IV infusion over 30 minutes on Days 1, 8, and 15 at dose levels of 80, 160, and 240 mg per week, with fixed-dose paclitaxel 80 mg/m2 on Days 1 and 8 of a 21-day cycle. In addition to a baseline scan within 4 weeks before study entry, computed tomography will be performed every two cycles to evaluate antitumor response. Treatment will continue until disease progression, unacceptable toxicity, consent withdrawal, or administrative discontinuation. The primary endpoint of this phase includes dose-limiting toxicity by NCI CTCAE v5.0 over 21 days. After determination of the recommended phase II dose of pelcitoclax in the phase Ib study, the efficacy of pelcitoclax with paclitaxel will be determined in the phase II study using a Simon two-stage design, with overall response rate as the primary endpoint. Other study endpoints in the phase II study include pharmacokinetics of pelcitoclax with paclitaxel, as well as progression-free and overall survival. As of February 8, 2021, 15 of 58 patients had been enrolled. Internal study identifier APG1252SU101. Clinical trial information: NCT04210037.

9089 Background: RET fusions are targetable oncogenic drivers in 1–2% of non-small cell lung cancer (NSCLC). ARROW (NCT03037385) supported the US FDA approval of pralsetinib, a highly potent oral selective RET inhibitor for RET-altered NSCLC and thyroid cancer. Here, we present updated results for a larger population of patients with RET fusion–positive NSCLC enrolled in ARROW. Methods: ARROW is a phase 1/2 open-label study conducted at 84 sites in 13 countries. Phase 2 expansion cohorts included patients with RET fusion–positive NSCLC. Initially, all treatment-naïve patients were not candidates for platinum-based therapy, a requirement removed by protocol amendment in July 2019. Primary objectives are overall response rate (ORR; blinded independent central review [BICR] per RECIST v1.1), assessed for patients with baseline measurable disease, and safety. Results: Updated analyses were completed as of Nov 6, 2020 (data cut-off), for patients who initiated pralsetinib 400 mg QD by May 22, 2020 (enrollment cut-off). Efficacy results, including analyses for treatment-naïve patients enrolled after eligibility criteria were revised to allow candidates for platinum-based therapy, are shown in the Table. Conclusions: Pralsetinib showed rapid, potent, and durable clinical activity in patients with RET fusion-positive NSCLC (regardless of prior therapies), including poor prognosis patients not eligible for platinum-based therapy. Overall, pralsetinib was well-tolerated. These data highlight the need for RET testing early in the course of disease to identify candidates who may benefit from treatment with pralsetinib. Clinical trial information: NCT03037385. [Table: see text]

Background: Circulating tumor cell (CTC) clusters are emerging as clinically significant harbingers of metastases in solid organ cancers. Prior to engaging these CTC clusters in animal models of metastases, it is imperative for technology to identify them with high sensitivity. These clusters often present heterogeneous surface markers, and current methods for isolation of clusters may fall short. Methods: We have applied the inertial microfluidic Labyrinth device for high-throughput, biomarker-independent, size-based isolation of CTCs and CTC clusters from patients with metastatic non-small cell lung cancer (NSCLC). Results: Using the Labyrinth device, CTCs (PanCK+/DAPI+/CD45-) were isolated from metastatic NSCLC patients (n=25). Heterogeneous CTC populations were detected, including CTCs expressing epithelial (EpCAM), mesenchymal (vimentin), or both markers. CTCs were isolated from 100% of patients (417±1023 CTCs/mL), and CTCs that were EpCAM negative were significantly higher in numbers than EpCAM+ CTCs. Cell clusters of ≥2 CTCs were observed in 96% of patients, of which 75% were negative for EpCAM. Patients with higher number of CTC clusters than single CTCs had worse progression-free survival (PFS) (p=0.05). Recovered CTCs from patients with RET, ROS1, and ALK-rearranged tumors revealed identical genetic aberrations as the primary tumor for each gene using FISH analysis. We have successfully expanded the recovered CTCs from 2 patients and screened for therapeutic targeting. We have found that TPX-0005 might be effective in these patients and would direct them to a clinical trial using this compound. Conclusions: The label-free Labyrinth device demonstrated the capability of collecting recovered CTCs from the device using a continuous processing technique while in a suspension state. This advantage opens the opportunities not only for CTC expansion off-chip, but also for ex vivo drug testing to direct patient-specific therapies. Citation Format: Mina Zeinali, Wei Huang, Maggie Lee, Arthi Nadhan, Anvya Mathur, Casey Hedman, Eric Lin, Ramdane Harouaka, Max S. Wicha, Lili Zhao, Nallasivam Palanisamy, Mathias Hafner, Rishindra Reddy, Gregory P. Kalemkerian, Bryan J. Schneider, Khaled A. Hassan, Nithya Ramnath, Sunitha Nagrath. High-throughput label-free isolation and expansion of circulating tumor cells (CTCs) from non-small cell lung cancer (NSCLC) patients for personalized treatments [abstract]. In: Proceedings of the AACR Special Conference on Advances in Liquid Biopsies; Jan 13-16, 2020; Miami, FL. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(11_Suppl):Abstract nr B45.

PURPOSE: Radiation-induced cardiac toxicity (RICT) is an increasingly well-appreciated source of morbidity and mortality in patients receiving thoracic radiotherapy (RT). Currently available methods to predict RICT are suboptimal. We investigated circulating microRNAs (c-miRNAs) as potential biomarkers of RICT in patients undergoing definitive RT for non-small cell lung cancer (NSCLC). METHODS: Data from 63 patients treated on institutional trials were analyzed. Prognostic models of grade 3 or greater (G3+) RICT based on pre-treatment c-miRNA levels (‘c-miRNA’), mean heart dose (MHD) and pre-existing cardiac disease (PCD) (‘clinical’), and a combination of these (‘c-miRNA + clinical’) were developed. Elastic net Cox regression and full cross-validation were used for variable selection, model building, and model evaluation. Concordance statistic (c-index) and integrated Brier score (IBS) were used to evaluate model performance. RESULTS: MHD, PCD, and serum levels of 14 c-miRNA species were identified as jointly prognostic for G3+ RICT. The ‘c-miRNA and ‘clinical’ models yielded similar cross-validated c-indices (0.70 and 0.72, respectively) and IBSs (0.26 and 0.28, respectively). However, prognostication was not improved by combining c-miRNA and clinical factors (c-index 0.70, IBS 0.28). The ‘c-miRNA’ and ‘clinical’ models were able to significantly stratify patients into high- and low-risk groups of developing G3+ RICT. Chi-square testing demonstrated a marginally significantly higher prevalence of PCD in patients with high- compared to low-risk c-miRNA profile (p=0.09), suggesting an association between some c-miRNAs and PCD. CONCLUSIONS: We identified a pre-treatment c-miRNA signature prognostic for G3+ RICT. With further development, pre- and mid-treatment c-miRNA profiling could contribute to multiparametric, patient-specific dose-selection and treatment-adaptation.

9001 Background: Pembro monotherapy showed durable antitumor activity as third-line or later therapy for metastatic SCLC, leading to FDA approval in that setting. KEYNOTE-604 was a double-blind, phase 3 study of pembro + EP vs placebo + EP as first-line therapy for ES-SCLC (NCT03066778). Methods: Eligible patients (pts) with previously untreated ES-SCLC and no untreated CNS metastases were randomized 1:1 to pembro 200 mg Q3W or saline placebo for up to 35 cycles plus 4 cycles of standard-dose EP. Pts with CR or PR after cycle 4 could receive PCI at investigator discretion. Randomization was stratified by platinum choice (carboplatin vs cisplatin), ECOG PS (0 vs 1), and LDH (≤ULN vs > ULN). Primary endpoints were OS and PFS (RECIST v1.1, blinded central review) in the ITT population. ORR, DOR, and safety were secondary endpoints. OS and PFS treatment differences were assessed by the stratified log-rank test. The protocol specified 2 interim analyses (IAs) and a final analysis (FA). Prespecified efficacy boundaries were one-sided P = 0.0048 for PFS at IA2 (prespecified final PFS analysis) and 0.0128 for OS at FA. Results: 453 pts were randomized. 223/228 pts assigned to pembro + EP and 222/225 assigned to placebo + EP received ≥1 dose of assigned treatment; 1 pt assigned to pembro + EP received placebo + EP in error. Median age was 65 y, 74% had ECOG PS 1, and 57% had LDH > ULN; more pts in the pembro + EP arm had baseline brain metastases (14% vs 10%). At FA (median follow-up, 21.6 mo), 9% of pts in the pembro + EP arm and 1% in the placebo + EP arm remained on study treatment; 12% and 14% received PCI. At IA2 (median follow-up, 13.5 mo), pembro + EP significantly improved PFS in the ITT population (HR 0.75 [95% CI 0.61-0.91], P = 0.0023; median 4.5 vs 4.3 mo). At FA, pembro + EP prolonged OS in the ITT population, but the significance threshold was not met (HR 0.80 [95% CI 0.64-0.98], P = 0.0164; median 10.8 vs 9.7 mo). In a post hoc analysis of OS in the as-treated population, the nominal P value was smaller than the significance threshold (HR 0.78 [95% CI 0.63-0.97], P = 0.0124). ORR at FA was 71% for pembro + EP vs 62% for placebo + EP; median DOR was 4.2 vs 3.7 mo. Observed AEs were as expected; any-cause AEs were grade 3-4 in 77% vs 75%, grade 5 in 6% vs 5%, and led to discontinuation in 15% vs 6%. Conclusions: Pembro + EP significantly improved PFS and prolonged OS compared with placebo + EP as first-line therapy for pts with ES-SCLC. No unexpected toxicities were seen with pembro + EP. These data support the benefit of pembro-containing regimens for ES-SCLC. Clinical trial information: NCT03066778.

Almost all patients with EGFR-driven lung cancer who are treated with EGFR tyrosine kinase inhibitors (TKI) develop resistance to treatment. A single base (c.2369C>T) transition mutation, EGFR T790M, is the most frequent resistance event after first-generation exposure to EGFR TKIs. Whether T790M mutation is acquired or is selected from a preexisting clone has been a matter of significant debate. In this study, we show that treatment with EGFR TKIs leads to activation of the NFκB pathway, which in turn induces expression of activation-induced cytidine deaminase (AICDA). In turn, AICDA causes deamination of 5-methylcytosine to thymine at position c.2369 to generate the T790M mutation. Pharmacologic inhibition of the NFκB pathway or knockout of AICDA decreased the frequency or prevented the development of T790M mutation, respectively. In addition, patients treated with first-line EGFR TKI displayed increased expression of AICDA and detection of the T790M mutation upon progression. These results identify the mechanism of T790M acquisition and present an opportunity to target the process to delay or prevent it. Significance: These findings identify the mechanism behind acquisition of a common resistance mutation to TKI treatment in lung cancer.

Purpose To assess the utility of circulating serum microRNAs (c-miRNAs) to predict response to high-dose radiation therapy for locally advanced non-small cell lung cancer (NSCLC). Methods and Materials Data from 80 patients treated from 2004 to 2013 with definitive standard- or high-dose radiation therapy for stages II-III NSCLC as part of 4 prospective institutional clinical trials were evaluated. Pretreatment serum levels of 62 miRNAs were measured by quantitative reverse transcription–polymerase chain reaction array. We combined miRNA data and clinical factors to generate a dose–response score (DRS) for predicting overall survival (OS) after high-dose versus standard-dose radiation therapy. Elastic net Cox regression was used for variable selection and parameter estimation. Model assessment and tuning parameter selection were performed through full cross-validation. The DRS was also correlated with local progression, distant metastasis, and grade 3 or higher cardiac toxicity using Cox regression, and grade 2 or higher esophageal and pulmonary toxicity using logistic regression. Results Eleven predictive miRNAs were combined with clinical factors to generate a DRS for each patient. In patients with low DRS, high-dose radiation therapy was associated with significantly improved OS compared to treatment with standard-dose radiation therapy (hazard ratio 0.22). In these patients, high-dose radiation also conferred lower risk of distant metastasis and local progression, although the latter association was not statistically significant. Patients with high DRS exhibited similar rates of OS regardless of dose (hazard ratio 0.78). The DRS did not correlate with treatment-related toxicity. Conclusions Using c-miRNA signature and clinical factors, we developed a DRS that identified a subset of patients with locally advanced NSCLC who derive an OS benefit from high-dose radiation therapy. This DRS may guide dose escalation in a patient-specific manner.

Background: Circulating tumor cells (CTCs) have emerged as important blood-based surrogate markers of primary tumors. Current methods for isolation of lung CTCs mostly rely on biomarker dependent antibody-based capture, missing populations that may be stem-like in nature. Results: We have applied the microfluidic Labyrinth device for high throughput, label-free, size-based isolation of CTCs from non-small cell lung cancer patients (NSCLC). The Labyrinth device was optimized and tested for inertial separation of cancer cells using the human lung cancer cell line H1650. The recovery and purity were >82% and >78%, respectively, operating at a flow rate of 2.5 mL/min. Using the biomarker-independent Labyrinth separation device, heterogeneous CTC populations were isolated from metastatic NSCLC patients (n=21). Heterogeneous CTC populations were detected, including CTCs (PanCK+ and CD45-), CTCs expressing EpCAM or Vimentin, and CTCs expressing both markers representing an EMT-like population of CTCs. Using Labyrinth, we were able to isolate CTCs from 100% of patients with an average yield of 180±168 CTCs/mL. Among the captured CTCs, EpCAM- CTCs were significantly more common than EpCAM+ CTCs (115.7 vs. 39.1 CTCs/mL respectively). Cell clusters of 2 or more CTCs were also observed in 95% of patients; 79% of these clusters were negative for EpCAM expression, whereas 35% expressed Vimentin, suggestive of an EMT phenotype. Recovered CTCs from patients with RET, ROS1 and ALK rearranged tumors showed aberrations matching with the primary tumor for each gene using FISH analysis. We have successfully expanded the recovered CTCs from 2 patients and screened for therapeutic targeting. We have found that TPX-0005 might be effective in these patients and would direct them to a clinical trial using this compound. Conclusion: The label-free Labyrinth device demonstrated the capability of collecting recovered CTCs from the device using a continuous processing technique while in a suspension state. This advantage opens the opportunities not only for CTC expansion off-chip, but also for ex-vivo drug testing to direct patient-specific therapies. Citation Format: Mina Zeinali, Maggie Lee, Arthi Nadhan, Anvya Mathur, Wei Huang, Eric Lin, Ramdane Harouaka, Max S. Wicha, Nallasivam Palanisamy, Mathias Hafner, Rishindra Reddy, Gregory P. Kalemkerian, Bryan J. Schneider, Khaled A. Hassan, Nithya Ramnath, Sunitha Nagrath. High Throughput isolation and expansion of circulating tumor cells (CTCs) from Non-small cell lung cancer (NSCLC) patients for personalized treatments [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1332.

Background: Epidermal growth factor receptor (EGFR) activation mutations occur in 10-50% of lung adenocarcinomas. EGFR tyrosine kinase inhibitors (TKIs) are the mainstay of treatment for stage IV non-small cell lung cancer (NSCLC) patients with EGFR mutations. However, acquired EGFR mutations are the main mechanism of on-target resistance to TKIs. T790M mutation that occurs after first generation TKI treatment, is a cytosine to thymine (C>T) single nucleotide transition leading to a threonine to methionine amino acid change at position 790 (i.e. T790M). Our data suggest that resistant mutations are acquired events secondary to cytosine deamination through Activation Induced Cytosine Deamination enzyme (AICDA). Interestingly, treatment with Osimertinib, that overcomes the T790M mutation, leads to other acquired resistant mutations (C797S, G796S/R and L792F/H) that are mostly cytosine based. Results: Sub clones of the lung adenocarcinoma cell line PC9 with no evidence of T790M mutation by digital droplet PCR (ddPCR) at baseline, were treated with EGFR TKI. After serially increasing the treatment dose, T790M mutation was detected by ddPCR associated with a significant increase in AICDA expression. Similarly, when the resistant T790M PC9 clones were treated with Osimertinib, the expression of AICDA was also induced. Knocking down AICDA by shRNA or CRISPR-Cas9, decreases the development of T790M in PC9 cell lines after TKI exposure. Using mass spectrometry, we established that cytosine at codon 790 is methylated; thus, deamination of 5-methylcytosine leads to thymine directly, explaining the T790M C>T mutation. In addition, using ChIP assay and pharmacological inhibition we confirm that upon TKI exposure, NFκB binds AICDA promoter and induces its expression. In a mouse xenograft model, the induction of NFκB and AICDA after EGFR TKI exposure is abrogated by concurrent use of an NFκB inhibitor. Finally, patients treated with EFR TKI had an increased expression of AICDA upon progression. Conclusion: In EGFR driven lung adenocarcinoma, NFκB pathway is activated upon exposure to EGFR TKIs which induces AICDA expression. AICDA deaminates cytosine into other nucleotides leading to treatment resistance. Citation Format: Najwa El Kadi, Luo Wang, April Davis, Alexander Cooke, Varun Vadnala, Hasan Korkaya, Gregory P. Kalemkerian, Khaled A. Hassan. The T790M mutation is acquired through 5-methylcytosine deamination after EGFR TKI treatment in lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 931.

e20061 Background: Malignant pleural mesothelioma (MPM) is a disease that is resistant to chemotherapy and there remains an unmet need for better therapeutic options. Nintedanib (BIBF 1120) is an oral multikinase inhibitor impacting VEGFR, FGFR, PDGFR, and other kinase activity such as TGFß signaling pathways. VEGF, FGF, and TGFβ are commonly expressed in MPM. We conducted a phase II trial in patients with recurrent MPM after platinum-based chemotherapy. Methods: Patients (pts) with MPM previously treated with platinum-based chemotherapy, performance status (PS) 0-1, adequate organ function, and no contraindications to anti-angiogenic therapy were eligible for treatment. Nintedanib 200 mg twice per day was administered until disease progression or unacceptable toxicity. The primary endpoint was the 4-month progression-free survival (PFS). A two-stage design was used and > 4 pts had to have a PFS of ≥4 months to proceed to the second stage. Results: Twenty pts. were enrolled. The median age was 70 yrs. (32-81), 90% were male, and 80% were PS = 1. The histology was 70% epithelioidal, 5% sarcomatoid, 10% biphasic, and 15% unknown. 15% had prior bevacizumab. The median follow-up is 16.4 mo. A median of 2 treatment cycles (range 1-18) were delivered. There were no responses but 40% had stable disease. The median PFS was 1.8 mo. (95% CI: 1.68, 3.55) and the PFS rate at 4 mo. was 13%. The median OS was 4.2 mo. (95% CI: 2.53, 8.74) and the OS rate at 4 mo. was 55%. Toxicities were usually grade 1-2 and included diarrhea, fatigue, edema, transaminase elevation, anorexia, nausea, vomiting and dyspnea. Conclusions: The activity of nintedanib in previously treated MPM pts. was modest. The trial did not meet the primary PFS endpoint. However, there was a small subset of pts. that had prolonged stable disease for > 4 months thus potentially deriving some clinical benefit from treatment. Supported by Boehringer Ingelheim. Clinical trial information: NCT02568449.

Purpose: The cancer stem cell theory postulates that tumors contain a subset of cells with stem cell properties of self-renewal, differentiation, and tumor initiation. The purpose of this study is to determine the role of Notch activity in identifying lung cancer stem cells. Experimental Design: We investigated the role of Notch activity in lung adenocarcinoma using a Notch GFP reporter construct and a γ-secretase inhibitor (GSI), which inhibits Notch pathway activity. Results: Transduction of lung cancer cells with Notch GFP reporter construct identified a subset of cells with high Notch activity (GFP-bright). GFP-bright cells had the ability to form more tumor spheres in serum-free media and were able to generate both GFP-bright and GFP-dim (lower Notch activity) cell populations. GFP-bright cells were resistant to chemotherapy and were tumorigenic in serial xenotransplantation assays. Tumor xenografts of mice treated with GSI had decreased expression of downstream effectors of Notch pathway and failed to regenerate tumors upon reimplantation in NOD/SCID mice. Using multivariate analysis, we detected a statistically significant correlation between poor clinical outcome and Notch activity (reflected in increased Notch ligand expression or decreased expression of the negative modulators), in a group of 443 patients with lung adenocarcinoma. This correlation was further confirmed in an independent group of 89 patients with adenocarcinoma in which Hes-1 overexpression correlated with poor overall survival. Conclusions: Notch activity can identify lung cancer stem cell–like population and its inhibition may be an appropriate target for treating lung adenocarcinoma. Clin Cancer Res; 19(8); 1972–80. ©2013 AACR.

Preclinical studies demonstrated antiproliferative synergy of 1,25-D3 (calcitriol) with cisplatin. The goals of this phase I/II study were to determine the recommended phase II dose (RP2D) of 1,25-D3 with cisplatin and docetaxel and its efficacy in metastatic non-small-cell lung cancer.Patients were ≥18 years, PS 0-1 with normal organ function. In the phase I portion, patients received escalating doses of 1,25-D3 intravenously every 21 days prior to docetaxel 75 mg/m(2) and cisplatin 75 mg/m(2) using standard 3 + 3 design, targeting dose-limiting toxicity (DLT) rate33 %. Dose levels of 1,25-D3 were 30, 45, 60, and 80 mcg/m(2). A two-stage design was employed for phase II portion. We correlated CYP24A1 tagSNPs with clinical outcome and 1,25-D3 pharmacokinetics (PK).34 patients were enrolled. At 80 mcg/m(2), 2/4 patients had DLTs of grade 4 neutropenia. Hypercalcemia was not observed. The RP2D of 1,25-D3 was 60 mcg/m(2). Among 20 evaluable phase II patients, there were 2 confirmed, 4 unconfirmed partial responses (PR), and 9 stable disease (SD). Median time to progression was 5.8 months (95 % CI 3.4, 6.5), and median overall survival 8.7 months (95 % CI 7.6, 39.4). CYP24A1 SNP rs3787554 (CT) correlated with disease progression (P = 0.03) and CYP24A1 SNP rs2762939 (CG) trended toward PR/SD (P = 0.08). There was no association between 1,25-D3 PK and CYP24A1 SNPs.The RP2D of 1,25-D3 with docetaxel and cisplatin was 60 mcg/m(2) every 21 days. Pre-specified endpoint of 50 % confirmed RR was not met in the phase II study. Functional SNPs in CYP24A1 may inform future studies individualizing 1,25-D3.

We investigated the novel combination of dose-dense pemetrexed, gemcitabine and bevacizumab as frontline treatment for advanced non–small cell lung cancer. Thirty-nine patients received pemetrexed (400 mg/m(2)), gemcitabine (1200 mg/m(2)), and bevacizumab (10 mg/kg), given every 14 days. Median progression-free survival was 6.1 months (95% confidence interval [CI], 4.2–7.9) and median overall survival was 18.4 months (95% CI, 13.1–29.5). Treatment met the primary endpoint and represents a reasonable therapeutic option. INTRODUCTION: Platinum-based chemotherapy is standard for untreated, advanced non-small-cell lung cancer (NSCLC). We investigated the activity and tolerability of the novel combination of dose-dense pemetrexed, gemcitabine, and bevacizumab in patients with advanced NSCLC. METHODS: This multicenter phase II trial evaluated the safety and efficacy of the combination of pemetrexed (400 mg/m(2)), gemcitabine (1200 mg/m(2)), and bevacizumab (10 mg/kg), given every 14 days in patients with untreated, advanced NSCLC. The primary endpoint was progression-free survival with secondary endpoints of response rate and overall survival. RESULTS: Thirty-nine patients were enrolled. Treatment was well tolerated; the most common grade 3–4 toxicities were neutropenia and fatigue. Of the 38 patients evaluable for tumor response, 1 (3%) had complete response, 15 (39%) had partial response, 12 (31%) had stable disease, and 10 (26%) had progressive disease. Median progression-free survival was 6.1 months (95% confidence interval [CI], 4.2–7.9) and median overall survival was 18.4 months (95% CI, 13.1–29.5). The 1-year overall survival rate was 64% (95% CI, 51%−81%) and the 2-year overall survival rate was 41% (95% CI, 28%−60%). CONCLUSIONS: Treatment with dose-dense pemetrexed, gemcitabine, and bevacizumab met the primary endpoint with promising efficacy and a manageable safety profile in patients with untreated advanced NSCLC. This regimen represents a reasonable therapeutic option.

Etoposide and irinotecan are key drugs in the treatment of small-cell lung cancer. We did this study to investigate whether combined chemotherapy with cisplatin, etoposide, and irinotecan was superior to topotecan monotherapy as second-line chemotherapy in patients with sensitive relapsed small-cell lung cancer.We did this open-label, multicentre, randomised phase 3 trial at 29 institutions in Japan. Patients with small-cell lung cancer that responded to first-line treatment but showed evidence of disease relapse or progression at least 90 days after completion of the first-line treatment were eligible to participate. Enrolled patients were randomly assigned (1:1) to receive combination chemotherapy with cisplatin plus etoposide plus irinotecan or topotecan alone. Randomisation was done via the minimisation method with biased-coin balancing for Eastern Cooperative Oncology Group performance status, disease stage at enrolment, and institution. Combination chemotherapy consisted of five 2-week courses of intravenous cisplatin 25 mg/m(2) on days 1 and 8, intravenous etoposide 60 mg/m(2) on days 1-3, and intravenous irinotecan 90 mg/m(2) on day 8, with granulocyte colony-stimulating factor given by hypodermic injection every day starting from day 9 of the first course (except on the days anticancer drugs were given). Topotecan therapy consisted of four courses of intravenous topotecan 1·0 mg/m(2) on days 1-5, every 3 weeks. The primary endpoint was overall survival in the intention-to-treat population, which was analysed with a one-sided α of 5%, and safety was assessed in all patients who received at least one dose of study drug. The trial is registered with University Hospital Medical Information Network Clinical Trials Registry, number UMIN000000828.Between Sept 20, 2007, and Nov 30, 2012, 180 patients were enrolled, with 90 assigned to each treatment group. The median follow-up for censored patients was 22·7 months (IQR 20·0-35·3). Overall survival was significantly longer in the combination chemotherapy group (median 18·2 months, 95% CI 15·7-20·6) than in the topotecan group (12·5 months, 10·8-14·9; hazard ratio 0·67, 90% CI 0·51-0·88; p=0·0079). The most common grade 3 or 4 adverse events were neutropenia (75 [83%] patients in the combination chemotherapy group vs 77 [86%] patients in the topotecan group), anaemia (76 [84%] vs 25 [28%]), and leucopenia (72 [80%] vs 46 [51%]). Grade 3 or 4 febrile neutropenia was more common in the combination chemotherapy group than in the topotecan group (28 [31%] vs six [7%]), as was grade 3 or 4 thrombocytopenia (37 [41%] vs 25 [28%]). Serious adverse events were reported in four (4%) patients in the topotecan group and nine (10%) in the combination chemotherapy group. Two treatment-related deaths (one each of pneumonitis and pulmonary infection) occurred in the topotecan group and one (febrile neutropenia with sepsis) occurred in the combination chemotherapy group.Combination chemotherapy with cisplatin plus etoposide plus irinotecan could be considered the standard second-line chemotherapy for selected patients with sensitive relapsed small-cell lung cancer.National Cancer Center and the Ministry of Health, Labour and Welfare of Japan.

BACKGROUND: The objectives of this study were to evaluate survival among current smokers, former smokers, and never smokers who are diagnosed with non–small cell lung cancer (NSCLC). METHODS: The study included patients who participated in the National Comprehensive Cancer Network's NSCLC Database Project. Current, former, and never smokers were compared with respect to overall survival by fitting Cox regression models. RESULTS: Data from 4200 patients were examined, including 618 never smokers, 1483 current smokers, 380 former smokers who quit 1 to 12 months before diagnosis, and 1719 former smokers who quit >12 months before diagnosis. Among patients with stage I, II, and III disease, only never smokers had better survival than current smokers (hazard ratio, 0.47 [95% confidence interval, 0.26-0.85] vs 0.51 [95% confidence interval, 0.38-0.68], respectively). Among patients with stage IV disease, the impact of smoking depended on age: Among younger patients (aged ≤55 years), being a never smoker and a former smoker for ≥12 months increased survival. After age 85 years, smoking status did not have a significant impact on overall survival. CONCLUSIONS: Patients who were smoking at the time of diagnosis had worse survival compared with never smokers. Among younger patients with stage IV disease, current smokers also had worse survival compared with former smokers who quit >12 months before diagnosis. It is likely that tumor biology plays a major role in the differences observed; however, to improve survival, it is prudent to encourage all smokers to quit smoking if they are diagnosed with NSCLC. Cancer 2013. © 2012 American Cancer Society.

Background: Epidermal growth factor receptor (EGFR) activation mutations occur in 10-50% of lung adenocarcinomas. This leads to constitutive activation of EGFR, which triggers multiple downstream survival and proliferation pathways. EGFR tyrosine kinase inhibitors (TKIs) are the mainstay of treatment for stage IV non-small cell lung cancer (NSCLC) patients with EGFR mutations. Acquired EGFR mutations are the main mechanism of on-target resistance to TKIs. T790M mutation that occurs after first line TKI treatment, is a cytosine to thymine (C>T) single nucleotide transition leading to a threonine to methionine amino acid change at position 790 (i.e. T790M). Interestingly, treatment with Osimertinib, that overcomes the T790M mutation, leads to other acquired resistant mutations, C797S, G796S/R and L792F/H. Our data suggest that resistant mutations are acquired events secondary to cytosine deamination through Activation Induced Cytosine Deamination enzyme (AICDA). Results: Sub clones of the lung adenocarcinoma cell line PC9 with no evidence of T790M mutation by droplet digital PCR (ddPCR) at baseline, were treated with EGFR TKI. After serially increasing the treatment dose, T790M mutation was detected by ddPCR associated with a significant increase in AICDA expression. Knocking down AICDA by shRNA, decreases the development of T790M in PC9 cell lines after TKI exposure. Similarly, when the resistant T790M PC9 clones were treated with Osimertinib, the expression of AICDA was also induced. Using mass spectrometry, we established that cytosine at codon 790 is methylated; thus, deamination of 5-methylcytosine leads to thymine directly, explaining the T790M C>T mutation. In addition, using ChIP assay and pharmacological inhibition we confirm that upon TKI exposure, NFĸB binds AICDA promoter and induces its expression. In a mouse xenograft model, the induction of NFĸB and AICDA after EGFR TKI exposure is abrogated by concurrent use of an NFĸB inhibitor. Finally, patients treated with EFR TKI had an increased expression of AICDA upon progression. Conclusion: In EGFR driven lung adenocarcinoma, NFĸB pathway is activated upon exposure to EGFR TKIs which induces AICDA expression. AICDA deaminates cytosine into other nucleotides leading to treatment resistance. Citation Format: Najwa El Kadi, April Davis, Alexander Cooke, Luo Wang, Hasan Korkaya, Gregory Kalemkerian, Khaled Hassan. Activation Induced Cytosine Deamination, AICDA, is induced after EGFR TKI exposure leading to secondary resistant mutations in lung adenocarcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5836.

Background The prognosis for patients with extensive-stage small-cell lung cancer remains poor. This trial was designed to evaluate irinotecan/cisplatin plus maintenance imatinib in patients with c-Kit–positive disease (the transmembrane receptor c-Kit is the product of the c-KIT protooncogene). Patients and Methods Immunohistochemistry for c-Kit was performed before enrollment. Treatment consisted of irinotecan 65 mg/m 2 on days 1 and 8 plus cisplatin 60 mg/m 2 on day 1 and every 21 days for 4 cycles. Imatinib was administered at 400 mg twice a day until progression or unacceptable toxicity occurred. Results Fourteen patients were enrolled. Slow accrual led to early study termination. Six patients did not begin treatment with imatinib because of disease progression, persistent toxicity, or referral for radiation therapy. Eight patients had a partial response with irinotecan/cisplatin and received imatinib. The median number of weeks on imatinib was 6.1 (range, 4.1-25.1 weeks). Reasons for imatinib discontinuation included disease progression (n = 7) and persistent neutropenia (n = 1). No objective responses to imatinib were evident, but 3 patients (21%) exhibited stable disease for 12, 15, and 25 weeks. The median progression-free survival was 4.3 months (95% CI, 2.9-4.8 months). The median overall survival was 7.8 months (95% CI, 5.7-10.0 months). The irinotecan/cisplatin regimen was well tolerated (grade 1/2 neutropenia, 29%; anemia, 43%; thrombocytopenia, 14%; and diarrhea, 29%), except in 1 patient with grade 3 vomiting. Imatinib toxicity included grade 1/2 nausea in 50% of the patients, peripheral edema in 75% of the patients, grade 3 fatigue in 13% of the patients, and neutropenia in 13% of the patients. Conclusion Despite the selection of tumors expressing c-Kit, imatinib did not appear to delay disease progression after response to chemotherapy. However, this trial was underpowered because of its early termination. Although disease stability with imatinib was evident in 3 patients and the therapy was well tolerated, this approach does not appear to warrant further clinical study.

e21175Background: Three immunotherapy agents are currently FDA-approved for the treatment of NSCLC after platinum-based chemotherapy. These agents have demonstrated superior outcomes and tolerabili...

Purpose: An embryonic stem cell (ESC) profile correlates with poorly differentiated breast, bladder, and glioma cancers. In this article, we assess the correlation between the ESC profile and clinical variables in lung cancer. Experimental Design: Microarray gene expression analysis was done using Affymetrix Human Genome U133A on 443 samples of human lung adenocarcinoma and 130 samples of squamous cell carcinoma (SCC). To identify gene set enrichment patterns, we used the Genomica software. Results: Our analysis showed that an increased expression of the ESC gene set and a decreased expression of the Polycomb target gene set identified poorly differentiated lung adenocarcinoma. In addition, this gene expression signature was associated with markers of poor prognosis and worse overall survival in lung adenocarcinoma. However, there was no correlation between this ESC gene signature and any histologic or clinical variable assessed in lung SCC. Conclusions: This work suggests that not all poorly differentiated non–small cell lung cancers exhibit a gene expression profile similar to that of ESC, and that other characteristics may play a more important role in the determination of differentiation and survival in SCC of the lung. (Clin Cancer Res 2009;15(20):6386–90)

Small cell lung cancer (SCLC) is an aggressive, poorly differentiated neuroendocrine carcinoma with distinct clinical, pathological and molecular characteristics. Despite robust responses to initial chemotherapy and radiation, the prognosis of patients with SCLC remains poor with an overall 5-year survival rate of less than 10 %. Despite the fact that numerous molecularly targeted approaches have thus far failed to demonstrate clinical utility in SCLC, further advances will rely on better definition of the biological pathways that drive survival, proliferation and metastasis. Recent next-generation, molecular profiling studies have identified many new therapeutic targets in SCLC, as well as extreme genomic instability which explains the high degree of resistance. A wide variety of anti-angiogenic agents, growth factor inhibitors, pro-apoptotic agents, and epigenetic modulators have been evaluated in SCLC and many studies of these strategies are on-going. Perhaps the most promising approaches involve agents targeting cancer stem cell pathways and immunomodulatory drugs that interfere with the PD1 and CTLA-4 pathways. SCLC offers many barriers to the development of successful therapy, including limited tumor samples, inadequate preclinical models, high mutational burden, and aggressive tumor growth which impairs functional status and hampers enrollment on clinical trials.

We aimed to report the final toxicity results on a radiation-dose escalation trial designed to test a hypothesis that very high doses of radiation could be safely administered to patients with non-small-cell lung cancer (NSCLC) by quantifying the dose-volume toxicity relationship of the lung.A total of 109 patients with unresectable or medically inoperable NSCLC were enrolled and treated with radiation-dose escalation (on the basis of predicted normal-lung toxicity) either alone or with neoadjuvant chemotherapy by use of 3D conformal techniques. Eighty-four patients (77%) received more than 69 Gy, the trial was stopped after the dose reached 103 Gy. Estimated median follow-up was 110 months.There were 17 (14.6%) Grade 2 to 3 pneumonitis and 15 (13.8%) Grade 2 to 3 fibrosis and no Grade 4 to 5 lung toxicity. Multivariate analyses showed them to be (1) not associated with the dose prescribed to the tumor, and (2) significantly (p0.001) associated with lung-dosimetric parameters such as the mean lung dose (MLD), volume of lung that received at least 20 Gy (V20), and the normal-tissue complication probability (NTCP) of the lung. If cutoffs are 30% for V20, 20 Gy for MLD, and 10% for NTCP, these factors have positive predictive values of 50% to 71% and negative predictive value of 85% to 89%.With long-term follow-up for toxicity, we have demonstrated that much higher doses of radiation than are traditionally administered can be safely delivered to a majority of patients with NSCLC. Quantitative lung dose-volume toxicity-based dose escalation can form the basis for individualized high-dose radiation treatment to maximize the therapeutic ratio in these patients.

The peroxisome proliferator-activated receptor-gamma (PPAR-gamma) is a member of the nuclear hormone receptor superfamily of ligand-activated transcription factors and a crucial regulator of cellular differentiation. Differentiation-inducing and antiproliferative effects of PPAR-gamma suggest that PPAR-gamma agonists might be useful as effective anticancer agents. Few studies have examined the efficacy of these agonists in animal models of tumorigenesis, and their mechanism(s) of action are still not clear. Our studies indicate higher PPAR-gamma expression in primary tumors from non-small-cell lung cancer (NSCLC) patients when compared to normal surrounding tissue. The expression of PPAR-gamma was also observed in several NSCLC lines. The treatment of lung adenocarcinoma cells (A549) with troglitazone (Tro), a PPAR-gamma ligand, enhanced PPAR-gamma transcriptional activity and induced a dose-dependent inhibition of A549 cell growth. The observed growth arrest was predominantly due to the inhibition of cell proliferation without significant induction of apoptosis. Cell cycle analysis of Tro-treated cells revealed a cell cycle arrest at G(0)/G(1) with concomitant downregulation of G(0)/G(1) cyclins D and E. In addition, Tro treatment stimulated sustained Erk1/2 activation in A549 cells, suggesting the activation of a differentiation-inducing pathway. Furthermore, treatment of A549 tumor-bearing SCID mice with Tro or Pio inhibited primary tumor growth by 66.7% and significantly inhibited the number of spontaneous lung metastatic lesions. Collectively, our data demonstrate that activation of PPAR-gamma impedes lung tumor progression and suggest that PPAR-gamma ligands may serve as potential therapeutic agents for NSCLC.

Although race, in and of itself, is not a relevant biologic variable, racial differences in disease characteristics and outcomes have been reported in many malignancies, including lung cancer. The lung cancer incidence rate in blacks has been consistently higher than that in whites for many years. This racial disparity is seen primarily in men and is significantly greater in younger age groups. The reason for higher lung cancer incidence rates in blacks remains unclear, but racial differences in smoking habits, socioeconomic variables, and the metabolism of tobacco carcinogens may all play an important role. Blacks are also more likely than whites to present with squamous cell carcinoma and with advanced-stage disease. A significant racial difference in survival rates has developed over the past 30 years, with a poorer prognosis noted in black patients, particularly those with local- and regional-stage disease. This disparity appears to be due to a lack of improvement in the survival of black patients with lung cancer, but the biological and/or societal basis for racial variations in survival have not been determined. In summary, significant racial differences exist in lung cancer incidence and survival rates. Further research is required to determine the factors responsible for these differences and to develop effective preventative and therapeutic interventions that will impact favorably on the incidence and prognosis of this disease.

In phase I clinical trial design, it has become common to extend the protocol beyond the standard dose escalation phase (eg, the cohorts-of-three “3+3” algorithm, accelerated titration, or continual reassessment method [CRM] [1,2]), which estimates a maximum tolerated dose (MTD). This protocol extension, called a dose expansion cohort (DEC), treats additional patients at the estimated MTD, sometimes stratifying enrollment of the DEC by multiple disease sites. The typical aim is to ensure that a sufficient number of patients has been treated at the estimated MTD by the trial’s completion to begin to characterize efficacy, tolerability, and pharmacokinetic or pharmacodynamic (PK/PD) endpoints (3–11). Recent discussions have highlighted the increasing size and complexity of DECs as well as the need to prospectively justify corresponding aims and analyses, which may be lacking (9–11). This paper provides a statistically based assessment of DECs and offers recommendations based on these results. A systematic review of 611 single-agent phase I trials published from 2006 to 2011 found that 149 (24%) included a DEC, with the use of DECs increasing over time (9). Thirty-eight of these trials did not include any specific objectives for the DEC; among those that did, safety was most common (89 trials, 60%), followed by efficacy (50 trials, 34%). There is also heterogeneity in the size of DECs. Among the 149 trials identified by Manji, et al., the median DEC size was 17, and the range was two to 271 (9). In 10 case studies from Iasonos and O’Quigley, seven had a DEC of size less than 25, and the remaining three were 43, 97, and 100 (10). Dahlberg, et al. report a median DEC size of 27 from 60 phase I trials submitted at Harvard in 2011 (11). Reviewing all phase I trials at our institution (University of Michigan Comprehensive Cancer Center) currently enrolling patients (as of June, 2014), we found that 54 have a planned DEC in the trial protocol, with a median size of 19 and range of four to 105. Trials with a small DEC, eg, fewer than 25 patients, are of greatest relevance in a phase I context, where safety remains of primary importance and preliminary evidence of efficacy is secondary. Because these studies are not statistically powered to evaluate efficacy, they fall short of being true phase I/II trials. Recent papers have proposed dose-finding designs based on efficacy and toxicity (12–14). Although DECs may have additional endpoints beyond toxicity, a DEC strategy should first be rigorously evaluated with regard to patient safety, both to future patients, that is, finding the true MTD, as well as patients in the current trial, that is, treating many current patients at the true MTD (15–17). Thus, our focus here is on the role of DECs in traditional phase I designs based on toxicity considerations. A principled dose escalation mechanism should operate for the duration of the trial to satisfy the safety metrics just described. We support this assertion with a simulation study of trial designs with a 10 to 20 patient DEC. The benefit conferred by a DEC may depend on the trial design, and we evaluate two common dose escalation designs: the 3+3 algorithm and the CRM (18). Because the final sample size of a 3+3 is not predetermined, adding a DEC increases the size of a trial that might otherwise be small. In contrast, the sample size of the CRM design may be predetermined, leading to a decision between inclusion of a DEC following completion of the CRM or a larger CRM in which the dose assignment mechanism operates throughout the entire trial. In investigator-initiated trials at our institution, the CRM design is used exclusively, making this decision crucial and recurrent. To the best of our knowledge, however, the strategy of appending a DEC to a CRM trial has not been statistically justified. Thus, we consider a small CRM trial paired with a DEC and an equivalently sized “large” CRM trial with no DEC. Iasonos and O’Quigley provide guidance for safety monitoring during the DEC and present four strategies for reevaluating estimated dose-toxicity rates during and after the DEC based on safety or efficacy data (10). Also included was a simulation study of the 3+3 design to assess how frequently the final MTD estimate, after analyzing the DEC data, differs from the initial MTD estimate. Extending rule-based designs like the 3+3 with DECs is natural because such trials may otherwise terminate before enrolling enough patients to precisely evaluate secondary objectives such as efficacy or PK/PD (eg, 3–6). The authors found that the post-DEC MTD estimate from a complete analysis was more accurate than the pre-DEC estimate but did not report the frequency in which patients in the trial were assigned the true MTD. A final important and distinguishing feature of this paper is that we differentiate between the final estimated, or selected, MTD and the true, unknown MTD, the latter being the dose level that induces dose-limiting toxicity (DLT) in x% of patients, where x may be selected based on characteristics of a particular therapy. Treating more than six patients at the estimated MTD is the motivation for using DECs and drives how precisely efficacy endpoints such as response rates may be estimated. However, although it is often implicitly assumed that the MTD determined from the dose-escalation phase is the true MTD, eg, “[t]he MTD was defined as the highest dose level with DLTs in

Importance Our previous studies demonstrated that tumors significantly decrease in size and metabolic activity after delivery of 45 Gy of fractionated radiatiotherapy (RT), and that metabolic shrinkage is greater than anatomic shrinkage. This study aimed to determine whether 18 F-fludeoxyglucose–positron emission tomography/computed tomography (FDG-PET/CT) acquired during the course of treatment provides an opportunity to deliver higher-dose radiation to the more aggressive areas of the tumor to improve local tumor control without increasing RT-induced lung toxicity (RILT), and possibly improve survival. Objective To determine whether adaptive RT can target high-dose radiation to the FDG-avid tumor on midtreatment FDG-PET to improve local tumor control of locally advanced non–small-cell lung cancer (NSCLC). Design, Setting, and Participants A phase 2 clinical trial conducted at 2 academic medical centers with 42 patients who had inoperable or unresectable stage II to stage III NSCLC enrolled from November 2008, to May 2012. Patients with poor performance, more than 10% weight loss, poor lung function, and/or oxygen dependence were included, providing that the patients could tolerate the procedures of PET scanning and RT. Intervention Conformal RT was individualized to a fixed risk of RILT (grade >2) and adaptively escalated to the residual tumor defined on midtreatment FDG-PET up to a total dose of 86 Gy in 30 daily fractions. Medically fit patients received concurrent weekly carboplatin plus paclitaxel followed by 3 cycles of consolidation. Main Outcomes and Measures The primary end point was local tumor control. The trial was designed to achieve a 20% improvement in 2-year control from 34% of our prior clinical trial experience with 63 to 69 Gy in a similar patient population. Results The trial reached its accrual goal of 42 patients: median age, 63 years (range, 45-83 years); male, 28 (67%); smoker or former smoker, 39 (93%); stage III, 38 (90%). Median tumor dose delivered was 83 Gy (range, 63-86 Gy) in 30 daily fractions. Median follow-up for surviving patients was 47 months. The 2-year rates of infield and overall local regional tumor controls (ie, including isolated nodal failure) were 82% (95% CI, 62%-92%) and 62% (95% CI, 43%-77%), respectively. Median overall survival was 25 months (95% CI, 12-32 months). The 2-year and 5-year overall survival rates were 52% (95% CI, 36%-66%) and 30% (95% CI, 16%-45%), respectively. Conclusions and Relevance Adapting RT-escalated radiation dose to the FDG-avid tumor detected by midtreatment PET provided a favorable local-regional tumor control. The RTOG 1106 trial is an ongoing clinical trial to validate this finding in a randomized fashion. Trial Registration clinicaltrials.gov Identifier:NCT01190527

EGFR resistant mutation, T790M, in lung adenocarcinoma is acquired through a specific cytosine deamination mechanism. Background: Epidermal growth factor receptor (EGFR) activation mutations occur in 15% of lung adenocarcinomas. This leads to constitutive activation of EGFR, which triggers multiple downstream survival and proliferation pathways. Currently, EGFR tyrosine kinase inhibitors (TKIs) are first line therapy for stage IV non-small cell lung cancer (NSCLC) patients with EGFR mutations. Despite initial significant response to TKIs, most tumors develop resistance. The main mechanism of resistance detected in 50-60% of cases is a cytosine to thymine (C>T) single nucleotide transition mutation at position 2369. This causes a threonine to methionine amino acid change at position 790 (i.e. T790M). Our data suggests that the C>T mutation is an acquired event secondary to cytosine deamination by Activation Induced Cytosine Deamination enzyme (AICDA). Results: Single cell clones of lung adenocarcinoma cell line, PC9, were treated with EGFR TKI. At baseline, these clones have EGFR exon 19 deletion but no evidence of T790M mutation by digital droplet PCR (ddPCR). However, after treatment with a serial increasing dose of EGFR TKI, T790M mutation was detected by ddPCR. Assessing whether cytosine deamination enzymes were altered by this treatment, a significant increase in AICDA expression was seen. Furthermore, recombinant AICDA protein could deaminate cytosine at position 2369 in vitro. In addition, using mass spectrometry and methylation specific primers, we determined that cytosine at position 2369 is in fact methylated. This further supports our hypothesis since 5-methyl cytosine is deaminated into thymine directly. Since in germinal center B-lymphocytes, AICDA is activated through a non-canonical NFkB mechanism, we assessed NFkB pathway in PC9 cell line. RelB and p52 expression were significantly increased after TKI treatment. In addition direct interaction between RelB and AICDA promoter was confirmed by ChIP Assay. These findings were also seen in a mouse PC9 xenograft model. Daily oral gavage of EGFR TKI caused significant increase in the expression of RelB as well as AICDA. Adding NFkB inhibitor twice weekly inhibited the expression of RelB and AICDA. Finally, knocking down AICDA by shRNA prevented the development of T790M mutation in PC9 cell lines after TKI exposure. Conclusion: Our data suggest that the T790M mutation could be actively acquired after TKI treatment through a cytosine deamination process by AICDA. This would have significant implications for treatment with targeted therapy. In fact, Imatinib resistance in CML and GIST tumors have a similar C>T single nucleotide transition mutation. Citation Format: Khaled Hassan, Najwa El Kadi, April Davis, Gregory Kalemkerian, Luo Wang, Hasan Korkaya. The development of EGFR resistant mutation, T790M, in lung adenocarcinoma is acquired through a specific cytosine deamination mechanism [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4119. doi:10.1158/1538-7445.AM2017-4119

e20607 Background: The PD-1 inhibitors nivolumab and pembrolizumab are approved for the treatment of advanced non-small cell lung cancer (NSCLC), but little is known about patterns of disease progression when these agents fail. Understanding patterns of failure is key to developing strategies to improve duration of response to these agents. Methods: We gathered clinical and radiographic data on patients treated at the University of Michigan and the Ann Arbor VA for advanced NSCLC with standard-dose nivolumab (n = 68) or pembrolizumab (n = 23) after progression on platinum-based chemotherapy. Sites of disease progression were described as local (within the same lobe as primary disease), nodal (thoracic), or distant. Results: 91 patients were evaluable, of whom 56 (61.5%) had progression of disease after an average duration of therapy of 3.2 months (95%CI, 2.6-3.8). 10 (17.9%) patients had progression at local sites alone, 3 (5.4%) at nodal sites alone, 12 (21.4%) at distant sites alone, and 31 (55.4%) at a combination of sites. Overall, 41 (73.2%) had progression at distant sites. There was no statistically significant difference in clinical factors ( i.e. age, histology, comorbidity index) between progressors vs. non-progressors or distant-progressors vs. non-distant progressors. Of 37 patients who had prior radiation, 17 (45.9%) had progression at an irradiated site, with patients who had local and/or nodal progression more likely to have received radiation at site of progression (p = 0.01). The most common distant sites of progression were liver (13), bone (10), and brain (9). Conclusions: The most common site of disease progression was distant, with the liver being the most commonly involved site. This may be due to the immunotolerogenic environment of the liver, which is characterized by high IL-10 concentration and propagation of suppressive regulatory T cells, which can dampen activation of cytotoxic T cells. Prior irradiation did not seem to prevent disease progression. Our preliminary analysis suggests that the efficacy of checkpoint inhibitors is hindered in immune-privileged sites. Strategies to overcome immune tolerance should be investigated to improve duration of response to these agents.

8504 Background: The median progression free survival (PFS) and overall survival (OS) following initial chemotherapy in ES-SCLC pts are 2 and 7 months, respectively (Ready N, J Clin Oncol 2015). We evaluated the benefits of maintenance pembro in ES-SCLC pts who had response/stable disease after 4-6 cycles of platinum/etoposide. Methods: Pts were required to begin pembro within 8 weeks of completion of chemotherapy, with restaging scans no more than 3 weeks prior to start of pembro. Prophylactic cranial radiation was permitted. Pts were treated with pembro 200 mg I.V. every 3 weeks for a maximum of 2 years. Disease assessment was done every 2 cycles for the first 6 cycles and then as per investigator discretion. Primary end point of the study was PFS. PFS according to immune related response criteria (irPFS) and OS were also assessed. Tumor tissue was analyzed for PD-L1 expression by the DAKO 22C3 antibody. Any level of expression was considered as positive for PDL1. Blood for circulating tumor cells (CTCs) was collected prior to first, second and third cycle of pembro. Results: Of the 45 pts enrolled, 55% were males and 22% had brain metastases. Median age was 66 years. The median time from end of chemotherapy to start of pembro was 5 weeks. Median number of pembro cycles was 4 (1-20 cycles). 35 pts had measurable disease at study entry. The disease control rate with pembro was 42% (1 CR, 3 PR, 15 SD). At a median follow up of 6 months, the median PFS was 1.4 months (90% CI-1.3-4.0) and the irPFS was 4.7 months (90% CI- 1.8-6.7). The median OS was 9.2 months (90% CI-6.1-15.2). 11 pts are still on therapy (3-20 cycles). The median CTC prior to pembro was 1 (0-256, n=37 pts). Each unit increase in baseline CTC correlated with worse PFS (p = 0.052; adjusted for brain mets, age and sex). PDL1 could be assessed in 35 pts and was positive in 1 pt. Most common adverse events were fatigue, nausea, cough and dyspnea. One pt developed atrio-ventricular conduction block and 1 pt type 1 diabetes. Conclusions: Maintenance pembro did not improve PFS in these patients but favorable OS suggests that some SCLC patients can benefit from maintenance pembro. Biomarkers to identify patients most likely to benefit from pembro need to be defined. Clinical trial information: NCT02359019.

Standard chemotherapeutic regimens, such as cisplatin and etoposide, may improve quality of life and prolong survival in patients with incurable non-small cell lung cancer (NSCLC). This trial was designed to evaluate the activity and toxicity of a regimen combining three of the most active agents against advanced-stage NSCLC: mitomycin C, etoposide, and cisplatin (MEP). Sixty-eight patients with stage IIIB (pleural effusion) or IV NSCLC received cisplatin 80 mg/m2 i.v. on day 1 and etoposide 80 mg/m2 i.v. on days 1, 2, and 3 every 3 weeks along with mitomycin C 10 mg/m2 i.v. on day 1 of the first and third cycles for a median of four cycles (range, 1-11). Median age was 59 years, and nine patients were enrolled after relapse from previously treated early-stage NSCLC. Eighty-eight percent of patients had stage IV disease, and 14 (21%) had brain metastases at diagnosis. Palliative radiotherapy was given to 10 patients (15%) before MEP and to 17 (25%) concurrent with MEP. The major toxicity of MEP was myelosuppression, with grade 3-4 neutropenia in 74% of patients. Sixteen patients (24%) had documented infections, and there were eight (12%) treatment-related deaths. Partial response was observed in 24 patients (35%) with a median duration of 4.4 months, (range 1.4-13 months). Median survival was 8.1 months (range, 1-34 months), and 1-year survival was 32%. The addition of mitomycin C to cisplatin and etoposide resulted in response and survival rates comparable with those achieved with standard regimens in patients with advanced NSCLC but was associated with substantial hematologic toxicity and unacceptable treatment-related mortality.