Late-stage Non-small Cell Lung Cancer Patients Research Articles

Longitudinal tracking of circulating rare events in the liquid biopsy of stage III–IV non-small cell lung cancer patients

In the United States, lung cancer is the second most common type of cancer with non-small cell lung cancer (NSCLC) encompassing around 85% of total lung cancer cases. Late-stage patients with metastatic disease have worsening prognosis, highlighting the importance of longitudinal disease monitoring. Liquid biopsy (LBx) represents a way for physicians to non-invasively track tumor analytes, such as circulating tumor cells (CTCs), and understand tumor progression in real-time through analyzing longitudinal blood samples. CTCs have been shown to be effective predictive biomarkers in measuring treatment efficacy and survival outcomes. We used the third-generation High-Definition Single Cell Assay (HDSCA3.0) workflow to analyze circulating rare events longitudinally during treatment in a cohort of 10 late-stage NSCLC patients, identifying rare events including circulating cancer cells (i.e., CTCs), and oncosomes. Here, we show (1) that there is a cancer specific LBx profile, (2) there is considerable heterogeneity of rare cells and oncosomes, and (3) that LBx data elements correlated with patient survival outcomes. Additional studies are warranted to understand the biological significance of the rare events detected, and the clinical potential of the LBx to monitor and predict response to treatment in NSCLC patient care.

Therapeutic efficacy of rare earth carbonate with chemoradiotherapy in late-stage non-small cell lung cancer: a cohort prospective study.

To compare the therapeutic effects and adverse reactions of sterilizing rare earth carbonate combined with concurrent chemoradiotherapy and simple concurrent chemoradiotherapy in the treatment of late-stage non-small cell lung cancer (NSCLC), and to analyze the reasons for the differences. A total of 817 patients with pathologically diagnosed late-stage NSCLC from June 1, 2021 to December 30, 2022, in the affiliated hospital of Kunming University of Science and Technology, were selected. They were randomly divided into a control group of 394 people and an experimental group of 423 people. The control group was given concurrent chemoradiotherapy (cisplatin + etoposide), while the experimental group simultaneously took a low dose of sterilized rare earth carbonate (0.05mg/Kg). The χ² test and Fisher's test were used to compare the clinical pathological features, objective response rate (ORR), ECOG score, and adverse reactions of the two groups of patients, while survival analysis was used to compare the progression-free survival (PFS) of the two groups. Cox regression analysis was used to test factors related to prognosis. The differences in clinical pathological features between the two groups of patients were not statistically significant, with all P>0.05. The ORR of the control group was 45.18% (178/394), and the experimental group was 89.83% (380/423), with a statistically significant difference (P=0.001). After treatment, the ECOG score of the experimental group was lower than that of the control group, P<0.001. The adverse reaction grading of patients in both groups was below level 3 after treatment, and no treatment-related fatalities occurred. The risk of pulmonary infection and bone marrow suppression in the experimental group was lower than that in the control group. In late-stage NSCLC patients, compared with simple concurrent chemoradiotherapy, the combination of concurrent chemoradiotherapy and sterilizing rare earth carbonate can significantly improve the short-term therapeutic effect and prognosis of patients, with good safety.

Abstract 6687: Prevalence of four major gene mutation classes in a survey of ctDNA recovery from early and advanced stage NSCLC

Abstract Introduction: Circulating tumor DNA (ctDNA) has emerged as a powerful biomarker with many applications in oncology including early and late-stage disease prognosis and in treatment decision making. Liquid biopsies have become critical in non-small cell lung cancer (NSCLC) given the patient risks associated with biopsies. It is well established that circulating nucleic acid levels representative of all solid tumors are increased in advanced stages of the disease. Less well characterized is the status of cfDNA levels in the earliest stages of solid tumor progression. We used validated methods for quantitative fluorometry and a high sensitivity amplicon based NGS method to survey cfDNA recovery and prevalence for the four major mutation classes (copy number amplifications (CNAs), single nucleotide variations (SNVs), fusions, and insertions/deletions (INDELs)) from patients previously diagnosed with NSCLC. Methods: Cell-free nucleic acid was extracted from 1,139 late-stage and 185 early-stage NSCLC plasma specimens. cfDNA was quantified using fluorometry, and the 52 gene GeneStrat NGS Test was used for the detection of CNAs, SNVs, fusions and INDELs. We evaluated cfDNA concentration (normalized to plasma input volume), quantity in circulation, and the prevalence of the four classes of mutations in early-stage (I-IIIA) versus late-stage (IIIB-IV) NSCLC. Results: We observed significantly lower average concentrations of cf nucleic acids in plasma recovered from early versus late-stage NSCLC specimens (early-stage: 13.7 ng/mL; range (3.14, 118.65); n = 185; late-stage: 18.3 ng/mL; range (0.67, 187.2); n =1,139; p=0.00176). We also evaluated total cfDNA recovered for the sample set and observed significantly lower average measurements for early-stage specimens versus late-stage (early-stage: 101.4 ng; range (18.38, 652.6); n=185; late-stage: 140.5 ng; range (9.802, 1534); n=1,139; p=0.00692). We do note that there were overlaps at the low end of the ranges. Also significant was the finding that all mutation classes were more prevalent in late versus early-stage NSCLC. Specifically, 56 of 58 CNAs (96.6%); 737 of 805 SNVs (91.6%); 42 of 43 INDELs (97.3%) and 12 fusions (100%) were associated with late-stage specimens. Conclusion: In this observational study cohort, cfDNA yields were significantly higher in late-stage NSCLC patients compared to early-stage. We also observed significantly more and varied mutations in plasma from late-stage patients. These initial findings need to be extended to additional cancers and refined by stage at diagnosis. We are currently expanding our study to include additional technologies and panels to assess ctDNA mutations especially for the early-stage patients. Quantitative cfDNA data may be of utility in diagnosis, monitoring and recurrence of cancer. Citation Format: Claire Gould, Leisa Jackson, Colin Cochran, Audrey Audetat, Brittany D'Alessio, Amanda Weaver, Janice Riley, Gary A. Pestano. Prevalence of four major gene mutation classes in a survey of ctDNA recovery from early and advanced stage NSCLC [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6687.

365 Computational pathology delivers objective and quantitative PD-L1 expression analysis for enrichment of responders to durvalumab in non-small cell lung cancer (NSCLC)

BackgroundThe pathologist’s visual assessment of tumor proportion score (TPS) with 25% cutoff on PD-L1 stained tissue samples is an established method to select metastatic NSCLC patients that are likely to respond to an anti-PD-L1 monotherapy.1 However, manual scoring is often subject to subjectivity in human perception2 and there remains a critical need for more objective and quantitative methods to assess PD-L1 expression in immuno-oncology.MethodsWe used deep learning (DL) based image analysis (IA) to generate a novel PD-L1 Quantitative Continuous Score (QCS)3 in tumor cells. PD-L1 QCS consists of two DL models to first segment epithelial regions and second detect membranes, cytoplasm and nuclei of each tumor cell in PD-L1 immunohistochemically (IHC) stained tissue slides. The PD-L1 expression of each tumor cell compartment was estimated by the respective optical density (OD) of DAB, and tumor cells with a membrane OD greater than ODmin are considered as PD-L1-positive. A slide comprising at greater percentage of PD-L1-positive tumor cells than a cutoff value (CoV) is considered QCS-positive. The ODmin and CoV parameters were linked to patient overall survival (OS), by minimizing the Kaplan Meier log-rank p-values and keeping at least 50% prevalence in the QCS-positive subgroup.Fully supervised QCS-IA models were extensively trained using pathologists’ annotations and the performance was validated on unseen data to ensure its generalization and robustness.3 4 The QCS IA was locked and blindly applied on clinical trial data (NCT01693562, durvalumab-treated late-stage NSCLC cohort) without further refinement.ResultsData analytics techniques were used to determine optimal PD-L1 QCS parameters for the clinical trial cohort of N=162 late-stage NSCLC patients. A PD-L1 QCS algorithm (ODmin=8, CoV=57%) is able to stratify durvalumab-treated NSCLC patients at a higher prevalence and more significant log rank p-value (64%, p=0.0001) for OS (figure 1) compared to pathologist TPS (59%, p=0.01). Median OS times of (19.2 months vs 7.9 months) was observed in the QCS-positive vs negative subgroups, respectively. The box plots (figure 2) indicate an overall good agreement (72% concordance) of the fully automated QCS with the pathologists TPS, which quantitatively supports the positive visual assessment of the cell segmentation accuracy.Abstract 365 Figure 1Kaplan Meier (KM) curves for OS stratification. KM curves for Overall Survival (OS) stratification with (left) manual PD-L1 TPS score (25% cutoff), and (right) automated QCS (57% cutoff).Abstract 365 Figure 2QCS scores within TPS positive and negative groups. Box plot indicating percent positive cells (OD≥8) as measured by PD-L1 QCS within the PD-L1 high (red) and low (blue) groups as per pathologist assessment by TPS.ConclusionsThe novel Quantitative Continuous Scoring (QCS) provides an objective way of correlating a quantitative estimate of PD-L1 IHC expression on tumor cells with survival of durvalumab-treated NSCLC patients. This data establishes a first proof-of-concept demonstrating the potential utility of PD-L1 QCS towards precision medicine in immuno-oncology.ReferencesRebelatto M, et al. Development of a programmed cell death ligand-1 immunohistochemical assay validated for analysis of non-small cell lung cancer and head and neck squamous cell carcinoma. Diagnostic Pathology 2016.Tsao M S, et al. PD-L1 immunohistochemistry comparability study in real-life clinical samples: results of blueprint phase 2 project. Journal of Thoracic Oncology 2018.Gustavson M, et al. Novel approach to HER2 quantification: digital pathology coupled with AI-based image and data analysis delivers objective and quantitative HER2 expression analysis for enrichment of responders to trastuzumab deruxtecan (T-DXd; DS-8201), specifically in HER2-low patients. (2021) DOI: 10.1158/1538-7445.SABCS20-PD6-01Kapil A, et al. Domain adaptation-based deep learning for automated tumor cell (TC) scoring and survival analysis on PD-L1 stained tissue images. IEEE Transactions on Medical Imaging DOI: 10.1109/TMI.2021.3081396Ethics ApprovalClinical study NCT01693562, from which data in this report were obtained, was carried out in accordance with the Declaration of Helsinki and Good Clinical Practice guidelines. The study protocol, amendments, and participant informed consent document were approved by the appropriate institutional review boards.

Body Mass Index, Weight Loss, and Mortality Risk in Advanced-Stage Non-Small Cell Lung Cancer Patients: A Focus on EGFR Mutation

Body mass index (BMI) influences the prognosis of patients with non-small cell lung cancer (NSCLC), including both early-stage and late-stage NSCLC patients that are undergoing chemotherapies. However, earlier research on the relationship between BMI and survival in patients taking epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) yielded contradictory results. These publications either had a limited number of patients or were getting TKIs in various lines of therapy, which might explain why the outcomes were contradictory. As a result, we undertook retrospective study to examine the effect of BMI on survival outcomes in patients with advanced EGFR mutant NSCLC receiving first-line EGFR-TKIs. We also compared the findings to those with wild-type EGFR. Between November 2010 and March 2014, 513 patients with advanced NSCLC were enrolled in the study. According to the adjusted BMI cut-off point for Asia, 35 out of 513 (6.8%) patients were underweight (BMI < 18.5 kg/m2), whereas 197 (38.4%) were overweight (BMI > 24 kg/m2). Overweight patients with wild-type EGFR exhibited longer progression-free survival (4.6 vs. 2.1 months, p = 0.003) and overall survival (OS) (8.9 vs. 4.3 months, p = 0.003) than underweight patients. Overweight patients with EGFR mutations had a longer OS than normal-weight patients (23.0 vs. 20.2 months, p = 0.025). Bodyweight reduction was related to a shorter OS in both the mutant EGFR patients (17.1 vs. 30.5 months, p < 0.001) and the wild-type EGFR patients (7.8 vs. 18.7 months, p < 0.001). In conclusion, advanced stages NSCLC patients with a lower BMI and early weight loss had a worse outcome that was independent of EGFR mutation status.

Abstract 1615: Immune-related adverse effects (irAEs) associated proteomic profile in late-stage NSCLC patients after PD-1 blockade

Abstract Immune checkpoint inhibitors (ICIs) in cancer therapy have improved clinical responses and overall survival for patients with non-small cell lung cancer (NSCLC). However, their action is quite often correlated with increased amount of immune related adverse effects (irAEs). In severe cases, strict control and prediction of such events is of high importance for proper patient management. Interestingly, low grade irAEs, observed after anti-PD-1 treatment are associated with better overall response rate (ORR) or overall higher survival benefit as previously reported. Therefore, defining good predictors that could address the occurrence of irAEs has been addressed by many researchers and a potential use of neutrophil-to-lymphocyte ratio (NLR) and platelet-to-lymphocyte ratio (PLR) has been already reported. In this study we have focused on the discovery of plasma based proteomic profiles associated with irAEs occurrence in two cohorts of late stage NSCLC patients receiving PD-1 blockade. Deep, unbiased plasma proteome profiling was achieved using LC-MS/MS in DIA mode (HRM). Proteins from plasma samples were denatured, digested, and injected into Thermo-Fisher instrument. High depth library for DIA approach was generated for protein identification and quantification using Spectronaut 14 (Biognosys). Protein panel selection was done using sPLS-DA approach. Gene ontology enrichment analysis for biological processes was performed on selected proteins. Unbiased analysis of 125 plasma samples from late-stage NSCLC patients treated with anti- PD1 (75 baseline and 50 after 8-weeks treatment) resulted in identification and quantifications of 853 proteins. Progression free survival analysis using Kaplan-Meier estimators showed significant difference between subjects with reported irAEs against those without, within and between both cohorts. Classification analysis using sPLS-DA on 43 subjects (86 of 125 samples) after optimization provided proteomic signature which completely separated patients with and without reported irAEs. Identified proteomic panel was used for hierarchical clustering, confirming separation across all subjects (irrespective of cohort allocation) into two main blocks. Interestingly, subjects divided into both groups showed lack of significance with regards to T cell-monocyte ratio. However, analysis of biological processes present in the panel revealed strong enrichment of proteins involved in immune responses, especially related to B cell activation, regulation and signaling. In this study we confirm prior observations of survival benefit related to irAEs after treatment with PD-1 blockade in late stage NSCLC patients. We also demonstrate the power of deep proteomic profiling in biomarker selection associated to irAEs, allowing for further development of new tools for improved patient management and risk-benefit assessment. Citation Format: Kamil Sklodowski, Vito Dozio, Roberta Poli, Andrés Lanzós, Silvia Lopez-Lastra, Kristina Beeler, Emanuela Romano. Immune-related adverse effects (irAEs) associated proteomic profile in late-stage NSCLC patients after PD-1 blockade [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1615.

Dynamics of Serum Tumor Markers Can Serve as a Prognostic Biomarker for Chinese Advanced Non-small Cell Lung Cancer Patients Treated With Immune Checkpoint Inhibitors.

Background: Serum tumor markers carcinoembryonic antigen (CEA), cancer antigen 125 (CA125), cytokeratin 19 fragment (CYFRA21-1) and squamous-cell carcinoma-related antigen (SCC-Ag) are routinely used for monitoring the response to chemotherapy or targeted therapy in advanced-stage non-small cell lung cancer (NSCLC), however their role in immunotherapy remains unclear. The aim of this study was to investigate whether dynamics of these serum markers were associated with the efficacy and prognosis of Chinese late-stage NSCLC patients treated with programmed cell death-1/programmed cell death ligand-1 (PD-1/PD-L1) inhibitors.Methods: We initiated a longitudinal prospective study on advanced NSCLC patients treated with PD-1/PD-L1 inhibitors in Chinese PLA general hospital (Beijing, China). Blood samples of baseline and after 6 weeks' treatment were collected. CT scan were used by all patients to evaluate treatment efficacy according to RECIST 1.1. Serum tumor markers levels were measured with an electrochemical luminescence for SCC-Ag and with a chemiluminescent microparticle immunoassay for serum CEA, CA125, and CYFRA21-1. At least 20% decreases of the biomarkers from baseline were considered as meaningful improvements after 6 weeks of treatment with immune checkpoint inhibitors (ICIs). Optimization-based method was used to balance baseline covariates between different groups. Associations between serum tumor biomarker improvements and objective response rate (ORR), progression-free survival (PFS), and overall survival (OS) were analyzed.Results: A total of 308 Chinese patients with advanced NSCLC were enrolled in the study. After balancing baseline covariates, patients with meaningful improvements in <2 out of 4 biomarkers (CEA, CA125, CYFRA21-1, and SCC-Ag) was ended up with lower ORR (0.08 vs. 0.35, p < 0.001), shorten PFS (median: 5.4 vs. 12.5 months, p < 0.001), and OS (median: 11.7 vs. 25.6 months, p < 0.001) in the total population. Subgroup analysis of patients with adenocarcinoma revealed that patients with meaningful improvements in <2 out of 4 biomarkers had significant lower ORR (0.06 vs. 0.36, p < 0.001), shorten PFS (median: 4.1 vs. 11.9 months, p < 0.001), and OS (median: 11.9 vs. 24.2 months, p < 0.001). So as in patients with squamous cell carcinoma, meaningful improvements in at least 2 out of 4 biomarkers were linked to better ORR (0.42 vs. 0.08, p = 0.014), longer PFS (median: 13.1 vs. 5.6 months, p = 0.001), and OS (median: 25.6 vs. 10.9 months, p = 0.06).Conclusions: The dynamic change of CEA, CA125, CYFRA21-1, and SCC-Ag from baseline have prognostic value for late-stage NSCLC patients treated with PD-1/PD-L1 inhibitors. Decrease of associated biomarkers serum levels were associated with favorable clinical outcomes.

Pretreatment hemoglobin level as a predictor to evaluate the efficacy of immune checkpoint inhibitors in patients with advanced non-small cell lung cancer.

Background:Targeting immune checkpoints represents an immense breakthrough in cancer therapeutics. The prognostic value of hemoglobin (Hb) has been investigated in many malignancies including non-small cell lung cancer (NSCLC). However, the prognostic impact of pretreatment Hb count for immune checkpoint inhibitors (ICIs) in advanced NSCLC patients remains unclear.Methods:A total of 310 late-stage NSCLC patients who received ICI therapies between January 2015 and March 2019 were prospectively enrolled. We used a propensity score-matched cohort analysis for this study. Patients’ clinicopathological characteristics and pretreatment Hb concentration were assessed against the progression-free survival (PFS) and overall survival (OS) using the Kaplan–Meier method and Cox proportional hazards regression.Results:A propensity score (PS)-matched cohort analysis was applied to adjust for potential bias and to create two comparable groups according to patients’ clinicopathological characteristics. The patients with normal baseline Hb levels (⩾110 g/L) had significantly longer PFS [median: 10.0 versus 4.0 months, hazard ratio (HR): 0.63, 95% confidence interval (CI): 0.46−0.86; p = 0.001] and OS [median: 17.6 versus 10.5 months, HR (95% CI): 0.56 (0.40−0.79); p < 0.001] than those with decreased Hb count (<110 g/L) in a PS-matched cohort (n = 255). For patients with normal pretreatment Hb levels, ICI combination therapy was significantly associated with better PFS [median: 11.1 versus 8.0 months, HR (95% CI): 0.74 (0.50−1.06); p = 0.09] and OS [median: 26.0 versus 12.9 months, HR (95% CI): 0.56 (0.37−0.86); p = 0.008] than monotherapy, but there was no such trend for patients with decreased baseline Hb levels.Conclusion:Our findings showed that normal pretreatment Hb count served as a favorable prognostic marker in advanced NSCLC patients treated with ICIs, representing an economical biomarker with readily measuring performance among all reported ones.

Circulating tumor DNA as prognostic markers for late stage NSCLC with bone metastasis.

Bone metastases from non-small cell lung cancer (NSCLC) are common, and current prognostic stratification methods are challenging to predict outcomes. The aim of the study is to examine circulating tumor DNA as a potential biomarker to gauge overall survival. Late stage NSCLC patients associated with bone metastasis were recruited for the study. Circulating tumor DNA was quantified using droplet digital polymerase chain reaction, a sensitive assay that is capable to pick up low mutant DNA frequencies. KRAS and EGFR genes were serially profiled at monthly intervals to ascertain their variations in different patients during monitoring. These were correlated to overall survival as an endpoint measure. Analysis of circulating tumor DNA and tumor tissues samples yielded a 94.7% overall agreement for KRAS and EGFR mutations. Higher circulating tumor DNA quantities of more than 1.4-fold were also detected in patients with bone metastases. To gauge the prognostic utility of circulating tumor DNA for these patient groups, circulating tumor DNA quantities as well as the patients' genotypes were used to stratify patients in the survival analysis. Hazard ratios for patient groups with and without bone metastasis was 1.63. Patients groups separated by circulating tumor DNA quantity and molecular profile were 1.51 and 1.58, respectively. The results showed that circulating tumor DNA was a useful marker to identify patients with better survival outcomes. The good overall concordance in genetic profiling and circulating tumor DNA measurements associated with NSCLC patients with bone metastases supports plasma-based testing. This study presents exploratory evidence of the prognostic role of circulating tumor DNA that can be of value in the management of the disease.

Sensitivity of TargetSelector in clinical experience in ctDNA profiling of NSCLC 2000 cases.

e23033 Background: Targeted cancer therapy relies on identifying specific DNA mutations from a patient’s tumor. Tyrosine kinase inhibitors (TKIs) tend to be effective for non-small cell lung cancer (NSCLC) with epidermal growth factor receptor (EGFR) activating mutations, of which exon 19 deletions (Del19) and L858R are most common. Acquired resistance to TKI therapy is associated with a T790M mutation. Standard biomarker analyses may not reflect tumor heterogeneity; they entail tissue biopsies often with surgical complications. To address these limitations, Biocept developed a minimally invasive method to characterize cancer biomarkers in blood. Biocept's proprietary TargetSelector assays selectively amplify relevant mutations from circulating tumor DNA (ctDNA). Clinical validations demonstrated high concordances between molecular tests in blood vs tissue. As further validation, EGFR mutation detection frequencies were compared to US averages (mycancergenome.org). Here we analyze 2000 blood samples received at Biocept from 1Mar 2016 to 4Jan 2017 from late stage NSCLC patients. Methods: Blood was collected in Biocept OncoCEE BCT validated to preserve DNA ≤ 8 days. TargetSelector was used to detect ctDNA L858R, Del19 and T790M.EGFR allele copy numbers for wild type and each mutant were calculated. The prevalence of each mutation was compared to US averages. Results: Del19, L858R, and T790M mutations were detected in 12.9%, 8.5%, and 9.9% of the analyzed blood samples, respectively. This is concordant with US averages, which are 10% for each mutation. Median copy numbers/ml of blood were 30 for Del19 (range: 1 – 91974), 15 for L858R (range: 1 – 91200), and 10 for T790M (range: 1 – 137360). The median wild type EGFR copy number detected/ml blood was 2304 (range: 8 – 2498725). In ~80% of T790M cases, ≥ 1 concomitant activating mutation was detected. Conclusions: Biocept's TargetSelector detects EGFR mutations (Del19, L585R, and T790M) at a very high level of sensitivity down to 1 mutant copy/ml in advanced NSCLC patients at frequencies consistent with cited US rates. Moreover, the underlying activating mutation was detected in ~80% of T790M cases.

Utility of urinary circulating tumor DNA for EGFR mutation detection in different stages of non-small cell lung cancer patients.

Non-invasive methods of molecular profiling for non-small cell lung cancer (NSCLC) are useful for monitoring disease progression. The aim of the current study was to ascertain if transrenal DNA is sensitive for clinical correlation and EGFR detection in NSCLC patients. 160 patients at various stages of the disease participated and samples were collected prospectively at 2-month intervals. A baseline sample was taken before treatment commencement. To ascertain the sensitivity of transrenal DNA, we compared its results with plasma DNA. ddPCR was used to profile the urine and blood samples for key EGFR mutations. Using tumor tissues as references, our study showed good concordance in EGFR mutations with transrenal DNA before treatment. Results were highly matching in late-stage NSCLC patients, with stage III/IV patients yielding an agreement of more than 90%. The assay was also sensitive to detect early-stage patients after surgical procedures. Profiles were highly concordant with results derived from plasma DNA, demonstrating the specificity of transrenal DNA assays. Serial monitoring of these patients showed stable molecular signatures and correlated to different treatments. Survival analysis showed good prognostic utility for late-stage patients with high transrenal DNA variations and patients that acquired T790M mutation. The study demonstrated the feasibility of using transrenal DNA in mutation profiling for different stages of NSCLC patients. It highlights the importance of continual monitoring and has potential clinical utility in the clinical management of NSCLC.

Investigation of transrenal KRAS mutation in late stage NSCLC patients correlates to disease progression

Purpose: Using transrenal DNA to detect KRAS mutations in non-small cell lung cancer (NSCLC), the study addressed the clinical impact for longitudinal monitoring and prognostic value for disease outcome.Methods: Digital droplet PCR was used to detect the mutant DNA. A total of 200 NSCLC patients were recruited with varying molecular profiles. To ascertain the specificity of transrenal DNA to accurately profile the disease, primary tissues were compared. Subsequently, serial samplings were performed at different treatment cycles to gauge the predictive value.Results: Transrenal DNA was successfully detected in all 200 patients. Overall concordance rate for mutant KRAS DNA within urine specimens and primary tissue biopsies was 95% (k = 0.87; 95% CI: 0.82–0.95). Patients with positive results at baseline had lower median overall survival (OS) than the wildtype group. More importantly, longitudinal monitoring of urine specimens showed an increase in the quantity of transrenal DNA, which were highly associated with disease progression and outcome.Conclusions: Our study showed a highly associative link to the patient’s tumor KRAS profile. Monitoring its variations aided in stratifying patients with worse outcome. Urinary specimens that can be extracted non-invasively presents new opportunities to track patients with KRAS mutation undergoing therapy.

Highlights of This Issue

Disrupting BRD4 activity through the use of small molecule probes has proven to be a widely effective preclinical anticancer strategy. To date, all reported BRD4 probes and clinical drug candidates operate through a monovalent binding mode. In this report, Rhyasen, Hattersley, and colleagues describe the pharmacology of AZD5153, a bivalent BRD4 inhibitor. The unique bivalent binding mode affords AZD5153 enhanced binding affinity, cell potency, and in vivo efficacy against hematologic cancer models. Targeting BRD4 through a bivalent binding mode therefore represents an attractive anticancer strategy.While most anticancer drugs have a single target, SF1126 inhibits two key signaling proteins within the liver cancer cell, PI3K and BRD4. Inhibition of PI3K enhances MYC degradation and BRD4 inhibition and prevents MYC transcription, providing a “first-in-class” approach to maximally block MYC-driven cancers. Singh, Joshi, Burgoyne and colleagues report this novel strategy of targeting two central signaling nodes with one agent that is antitumorigenic in vitro and in vivo and synergistic with the multikinase inhibitor Sorafenib. SF1126 is expected to enter HCC Phase I trials later this year.Chondrosarcoma is a real challenge in orthopedic oncology and is notoriously chemoresistant. In this study, Peyrode, Weber, and colleagues develop an attractive proteoglycans-targeting chemotherapy approach using a quaternary ammonium conjugate of melphalan (Mel-QA). This work demonstrated the crucial role of the QA carrier for in vitro binding to aggrecan. In vivo, in chondrosarcoma-bearing rats, radiolabeled [3H]-Mel-QA was shown to accumulate in tumor tissue respectively to melphalan, which was associated with tumor growth inhibition and improvement of the therapeutic index. These results open new perspectives in the therapeutic management of chondrosarcoma.AC0010 is a novel, irreversible, epidermal growth factor receptor (EGFR) inhibitor designed to overcome T790M-induced resistance in non–small cell lung cancer (NSCLC) patients. Preclinical studies demonstrate that AC0010 is a selective and potent inhibitor of EGFR activation and T790M mutations. A Phase 1 dose-escalation clinical study indicates that AC0010 is safe and effective in late-stage NSCLC patients. In this study, no pneumonitis, hyperglycemia, or grade 3 prolongation of the corrected QT interval event were observed. The preclinical and clinical evidence warranted further development of AC0010 as an alternative therapeutic agent for NSCLC patients who acquire resistance to first-generation EGFR TKIs.

Consequences of Late-Stage Non–Small-Cell Lung Cancer Cachexia on Muscle Metabolic Processes

IntroductionThe loss of muscle is common in patients with advanced non–small-cell lung cancer (NSCLC) and contributes to the high morbidity and mortality of this group. The exact mechanisms behind the muscle loss are unclear. Patients and MethodsTo investigate this, 4 patients with stage IV NSCLC who met the clinical definitions for sarcopenia and cachexia were recruited, along with 4 age-matched healthy volunteers. After an overnight fast, biopsy specimens were obtained from the vastus lateralis, and the key components associated with inflammation and the control of muscle protein, carbohydrate, and fat metabolism were assessed. ResultsCompared with the healthy volunteers, significant increases in mRNA levels for interleukin-6 and NF-κB signaling and lower intramyocellular lipid content in slow-twitch fibers were observed in NSCLC patients. Although a significant decrease in phosphorylation of the mechanistic target of rapamycin (mTOR) signaling protein 4E-BP1 (Ser65) was observed, along with a trend toward reduced p70 S6K (Thr389) phosphorylation (P = .06), no difference was found between groups for the mRNA levels of MAFbx (muscle atrophy F box) and MuRF1 (muscle ring finger protein 1), chymotrypsin-like activity of the proteasome, or protein levels of multiple proteasome subunits. Moreover, despite decreases in intramyocellular lipid content, no robust changes in mRNA levels for key proteins involved in insulin signaling, glycolysis, oxidative metabolism, or fat metabolism were observed. ConclusionThese findings suggest that examining the contribution of suppressed mTOR signaling in the loss of muscle mass in late-stage NSCLC patients is warranted and reinforces our need to understand the potential contribution of impaired fat metabolism and muscle protein synthesis in the etiology of cancer cachexia.

Socioeconomic Status Is Associated with Depressive Severity Among Patients with Advanced Non–Small-Cell Lung Cancer: Treatment Setting and Minority Status Do Not Make a Difference

Non-small-cell lung cancer (NSCLC) is the leading cause of cancer-related morbidity and mortality. Unfortunately, patients with NSCLC have relatively poor survival rates compared with patients diagnosed with most other types of cancer. Accordingly, managing physical and mental health symptoms are important treatment goals. In the current investigation, we sought to determine whether individual socioeconomic status (SES; as indexed by level of education), racial/ethnic minority status, and hospital type (public versus tertiary care center) were associated with NSCLC cancer patients' depressive severity. Importantly, we investigated whether NSCLC patients' individual SES was more or less prognostic of their depressive severity compared with minority status and the hospital context where they received treatment. Patients scheduled for chemotherapy were assessed for depressed mood by the Beck Depression Inventory-II (BDI-II). Data were collected at baseline and at approximately 6, 12, and 18 weeks. NSCLC patients with less education had more depressive severity than those with more education. Treatment setting and minority status were not associated with depressive severity. The interaction between education level and treatment setting predicting depressive severity was not significant, suggesting that the association between education level and depressive severity did not differ by treatment setting. Our study brings heightened awareness to the substantial, persistent SES differences that exist in depressive severity among late-stage NSCLC patients. Furthermore, these findings seem to persist, regardless of minority status and whether the patient is treated at a public hospital or tertiary cancer center.

Peripheral Blood miR-328 Expression as a Potential Biomarker for the Early Diagnosis of NSCLC

Lung cancer is often diagnosed at an advanced stage, with subsequently poor prognosis. There are no biomarkers available to facilitate early diagnosis or to discriminate between benign and malignant nodules. MicroRNAs (miRNAs) are stable molecules that can be found and measured in peripheral blood, thus representing potential diagnostic biomarkers. We evaluated 100 individuals comprising 86 patients with predominantly early-stage non-small cell lung cancer (NSCLC) and 24 healthy donors. RNA was extracted from peripheral blood samples and the expression of a panel of miRNAs was analyzed by Real-Time PCR method. Expression levels of miR-328, miR-18a, miR-339 and miR-140 were significantly higher in NSCLC patients than in healthy donors (p < 0.05). In particular, miR-328 showed good diagnostic accuracy in discriminating between patients with early NSCLC and healthy donors (AUC ROC 0.82, 95% CI 0.72–0.92), with 70% sensitivity and 83% specificity at the best relative expression cut-off of 300. Moreover, miR-339 was a good discriminant between healthy donors and late-stage NSCLC patients (AUC ROC 0.79, 95% CI 0.68–0.91). In conclusion, miR-328 represents a potential diagnostic biomarker of NSCLC, especially for the identification of early-stage tumors. Its role in discriminating between benign and malignant nodules detected by spiral CT warrants further investigation.

Abstract 1350: Genetic variations in apoptosis pathway are associated with survival in late stage non-small cell lung cancer.

Abstract Dysregulation of apoptosis pathway plays an essential role in carcinogenesis, tumor progression, and resistance to chemotherapy. We hypothesized that genetic variations in genes involved in the apoptosis pathway may affect the survival in late stage non-small cell lung cancer (NSCLC) patients. To test this hypothesis, we genotyped 583 tagging SNPs in 598 late stage NSCLC patients and analyzed their associations with survival individually and jointly. Seven SNPs were statistically significantly associated with overall survival after correction for multiple comparisons (q&amp;lt;0.1). The most significant SNP was rs17592236 on FOXO1 gene under the dominant model (OR: 2.54, 95%CI: 1.71-3.78, p=4.15x10−6). We also observed a significant cumulative effect. Compared to the reference group (patients with no more than two risk alleles), those having five or more risk alleles were much more likely to die within seven months (HR: 3.58, 95% CI: 2.55-5.04, p&amp;lt;0.001). Notably, the median survival time (MST) was significantly reduced by up to 15 months in high risk patients when compared to the reference group (MST: 22.2 months, p&amp;lt;0.001). Survival tree analysis revealed potential higher order gene-gene interactions among these SNPs. Our results provided evidence that genetic variations in the apoptosis pathway modulate NSCLC survival and may serve as biomarkers to differentiate NSCLC patients with different prognosis. Citation Format: Xiang Shu, Charles Lu, Joe Y. Chang, Jack A. Roth, Xifeng Wu. Genetic variations in apoptosis pathway are associated with survival in late stage non-small cell lung cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1350. doi:10.1158/1538-7445.AM2013-1350

In vivo isolation of circulating tumor cells in non-small cell lung cancer (NSCLC) patients by a medical device.

10536 Background: Currently, circulating tumor cells (CTCs) are isolated in vitro from small limited volumes of blood samples. Furthermore, CTC results for NSCLC as a prognostic and stratification biomarker are scarce. The aim of the study was to assess a functionalized and structured medical wire (FSMW) for in vivo isolation of CTCs directly from the blood of NSCLC patients, and to compare it to the Cell Search method. Methods: The device was inserted in a cubital vein through a standard cannula for thirty minutes. The interaction of target CTCs with the FSMW was mediated by an antibody directed against the epithelial cell adhesion molecule (EpCAM). To confirm the CTCs binding to the wire, the immunohistochemical staining against EpCAM as well as CD45 for negative cell selection was performed. There were 72 applications of the wire in 48 stage I-IIIB NSCLC patients (12 double applications) and 12 non cancer patients. Enumeration data was available for 34 NSCLC patients and 8 non cancer patients. For 34 patients, samples were also tested in the Cell Search system. Results: The device was well tolerated in all 72 applications without side effects. We obtained in vivo isolation of CTCs in 32 of 34 NSCLC patients (94.1%) with a median (range) of 13 (0-300) CTCs. The sensitivity was similar for early and late stage NSCLC patients. In the non-cancer patients, no CTCs were detected (100 % specificity). In 2 of 34 samples (5.8%), CTCs were detected by the Cell Search method. In all matched pairs, the FSMW detected the same number or more CTCs compared to the Cell Search method. Conclusions: Whilst well tolerated without side effects, the CTC detection rate of the FSMW in NSCLC patients was &gt;90%. In contrast, &lt;10% detection was obtained using the Cell Search analysis. No CTCs were detected in non-cancer patients. This proof of concept study may have important clinical implications, as the implementation of the device into clinical practice may improve early detection, prognosis and therapy monitoring of NSCLC patients. The method may also allow the molecular analysis of the CTCs, with the possibility of establishing more personalized treatment regiments.

Genetic Variations in the Regulator of G-Protein Signaling Genes Are Associated with Survival in Late-Stage Non-Small Cell Lung Cancer

The regulator of G-protein signaling (RGS) pathway plays an important role in signaling transduction, cellular activities, and carcinogenesis. We hypothesized that genetic variations in RGS gene family may be associated with the response of late-stage non-small cell lung cancer (NSCLC) patients to chemotherapy or chemoradiotherapy. We selected 95 tagging single nucleotide polymorphisms (SNPs) in 17 RGS genes and genotyped them in 598 late-stage NSCLC patients. Thirteen SNPs were significantly associated with overall survival. Among them, rs2749786 of RGS12 was most significant. Stratified analysis by chemotherapy or chemoradiation further identified SNPs that were associated with overall survival in subgroups. Rs2816312 of RGS1 and rs6689169 of RGS7 were most significant in chemotherapy group and chemoradiotherapy group, respectively. A significant cumulative effect was observed when these SNPs were combined. Survival tree analyses identified potential interactions between rs944343, rs2816312, and rs1122794 in affecting survival time in patients treated with chemotherapy, while the genotype of rs6429264 affected survival in chemoradiation-treated patients. To our knowledge, this is the first study to reveal the importance of RGS gene family in the survival of late-stage NSCLC patients.

Increased numbers but functional defects of CD56+CD3+ cells in lung cancer

CD56+ T cells were studied in samples of peripheral blood from small-cell lung cancer (SCLC) and non-small-cell lung cancer (NSCLC) patients compared with healthy controls. Relative numbers of CD56+CD3+ cells were increased in NSCLC (P = 0.001) and SCLC (P = 0.002) compared with normal subjects but their ability to respond to activation by up-regulating CD25 or producing IFN-γ were both significantly impaired. Expression of the killer-immunoglobulin-like receptor CD158a was significantly lower on CD56+CD3+ cells in SCLC than controls and also in early stage compared with late stage NSCLC patients. Mean levels of CD158e were higher in NSCLC patients than controls. CD158e levels on CD56+CD3+ cells were increased in the presence of its ligand HLA-Bw4 compared with controls. Although the precise role of CD56+CD3+ cells is not clear, they appear to be functionally impaired in lung cancer, which may have implications for a reduction of direct or indirect anti-tumour responses.