Small Cell Lung Cancer Biomarker Research Articles

Current and future trends in non-small cell lung cancer biomarker testing: The American experience.

Biomarker testing in patients with advanced stage non-small cell lung cancer provides essential information that can be used to select the most appropriate therapy. The regular updates of guideline recommendations reflect the growing number of biomarkers that must be assessed, and as such signal the shift from single-gene assays to more comprehensive genomic profiling using next-generation sequencing modalities. Cytology and small biopsy specimens have proven to be more than adequate substrates for these types of ancillary molecular testing; however, other alternative testing substrates are beginning to emerge. These include so-called liquid biopsies as well the supernatant fluid from cytology specimens, both of which have demonstrated promise for use in the clinical realm. This review will briefly cover the current state of non-small cell lung cancer biomarker testing in the United States, with a focus on these novel nonconventional substrates that are increasingly being incorporated into testing paradigms.

Potential prognostic value of delta-like protein 3 in small cell lung cancer: a meta-analysis

BackgroundCurrent researches have revealed that delta-like protein 3 (DLL3) may be related with prognosis in patients with small cell lung cancer (SCLC). However, this finding remains controversial in small cell lung cancer. This meta-analysis was systematically performed to evaluate the prognostic value of DLL3 in SCLC.MethodsThe PubMed, EMBASE and Web of Science databases were retrieved to collect the eligible references. Through Stata 15.0 software, we pooled hazard ratios (HR) with 95% confidence intervals (CI) by using random or fixed-effects models to evaluate the association between DLL3 and SCLC survival results.ResultsA total of 6 inter-related studies including 645 patients were qualified. After we removed 1 study, the remaining 5 studies including 601 patients were pooled to testify that high expression of DLL3 was an inferior prognostic for patients with SCLC in Asian populations (HR = 1.37, 95% CI = 1.05, 1.69; I2 = 0.0%, p = 0.000). The pooled results showed that DLL3 might be higher expression in advanced metastasis SCLC in Asian populations (RR = 0.84, 95% CI = 0.71, 0.99; I2 = 44.7%, p = 0.039). But the expression of DLL3 was not correlated with sex (RR = 1.33, 95% CI = 0.98, 1.80; I2 = 0.0%, p = 0.064), smoking history (RR = 1.01, 95% CI = 0.58, 1.75; I2 = 72.1%, p = 0.967) and tumour stage (RR = 0.68, 95% CI = 0.44, 1.05; I2 = 66.6%, p = 0.081).ConclusionsOur meta-analysis confirms that in Asian populations, high expression of DLL3 was a potential poor prognostic biomarker for SCLC and DLL3 highly expressed in advanced stage SCLC in Asian populations.

Selective application of neuroendocrine markers in the diagnosis and treatment of small cell lung cancer

AimsHere, we explored the potential application and selection of neuroendocrine biomarkers in the diagnosis and treatment of small cell lung cancer. MethodsWe retrospectively analyzed 118 patients with small cell lung cancer (SCLC), 166 patients with non-small cell lung cancer (NSCLC), 33 patients with benign lung disease (BLD), and 200 healthy individuals admitted to Zhejiang Provincial People’s Hospital between January 1, 2015 and May 31, 2019. All the patients were newly diagnosed with either SCLC, NSCLC, or BLD and previously untreated. Peripheral blood levels of ProGRP, NSE, CEA, and CYFRA21-1 were analyzed during the follow-up treatment, and 2-fold upper limit of reference intervals were defined as effective elevation. We used paired results to analyze the diagnostic efficiency of proGRP and NSE on SCLC. ResultsIn the 118 SCLC patients, proGRP levels were significantly higher compared with NSE levels. The diagnostic efficiencies of NSE and ProGRP for SCLC were 0.8554 and 0.9053, respectively. The combined diagnostic efficiency (0.9426) was higher relative to NSE, but there was no significant difference compared with proGRP. The effective elevation rate of proGRP was 45.3% higher than that of NSE in the limited stage of SCLC. In the extensive disease of SCLC patients, 70.7% cases had more than 10-fold increase in proGRP value, whereas 56.9% cases had less than 5-fold increase in NSE value. Compared with pre-treatment, the median concentrations of proGRP increased by 204.0% higher than that of NSE (71.3%) in the progressive group. Besides, the dynamic change in imaging characteristics and tumor size had a strong correlation with the levels of proGRP. SignificanceProGRP is a reliable neuroendocrine biomarker in SCLC. The effective elevation of proGRP has a potential diagnostic and efficacy value in the evaluation of SCLC. However, the combined detection of proGRP and NSE does not significantly improve the diagnosis of lung cancer.

Biofunctionalized Graphene Quantum Dots Based Fluorescent Biosensor toward Efficient Detection of Small Cell Lung Cancer.

Quantitative detection of cancer biomarkers with higher accuracy and sensitivity provides an effective platform for screening, monitoring, early diagnosis, and disease surveillance. The present work demonstrates the fabrication and application of fluorescent turn-on biosensor for ultrasensitive detection of small cell lung cancer biomarker utilizing biofunctionalized graphene quantum dots as the energy donor and gold nanoparticles (AuNPs) as the energy acceptor. One-pot and the bottom-up hydrothermal route have been employed for the synthesis of in situ amine-functionalized and nitrogen-doped graphene quantum dots (amine-N-GQDs) and further characterized experimentally by different analytical techniques. The molecular simulation studies were performed using the Material Studio software for optimizing the possible chemical structure of synthesized amine-N-GQDs, a comprehensive analysis of experimental results to validate the presence of potential N-doping and amine functionalization sites. Then monoclonal neuron-specific enolase antibodies (anti-NSE) were covalently immobilized to amine-N-GQDs to provide the biofunctionalized GQDs (anti-NSE/amine-N-GQDs). A label-free and efficient fluorescent biosensor based on nanosurface energy transfer (NSET) between anti-NSE/amine-N-GQDs and AuNPs has been developed for neuron-specific enolase (NSE) detection. The fluorescence response studies of anti-NSE/amine-N-GQDs@AuNPs nanoprobe conducted as a function of NSE antigen exhibited fast response time (16 min), broader linear detection range (0.1 pg mL-1 to 1000 ng mL-1), and remarkably low detection limit (0.09 pg mL-1). Additionally, the fluorescent biosensor exhibited excellent performance in real samples, with an average recovery value of 94.69%.

JCSE01.21 Changes of Peripheral Blood sPD-L1 in Patients with Small Cell Lung Cancer During Chemotherapy and Its Clinical Significance

As a new immunotherapeutic target, the inhibitors of programmed death 1 (PD-1) and programmed death ligand-1 (PD-L1) pathway have been used to treat a variety of tumors including small cell lung cancer (SCLC). However, the biomarkers now used to predict the efficacy of SCLC immunological checkpoint inhibitors are still in the exploratory phase. The aim of this prospective study was to investigate the prevalence and prognostic roles of soluble PD-L1(sPD-L1 ) protein in the blood of patients with lung cancer. A total of 94 patients with SCLC who were diagnosed by histopathology or cytopathology between March 2018 to November 2018 were enrolled. Blood samples plasma were collected at the time of diagnosis. 17 samples of healthy subjects matching in sex and age from the Health care Center of the hospital were also studied as control. The level of sPD-L1 protein in the blood was measured using an enzyme-linked immunosorbent assay (ELISA). And the correlation of sPD-L1 expression with tumor stage, distant metastasis, and pro gastrin releasing peptide (ProGRP) was analyzed. Expression of sPD-L1 in SCLC patients was significantly higher than healthy people(P<0.05).A cut-off value of 1.362ng/ml was distinguished in patients according to Receiver operating characteristic curve (ROC). Dynamic changes of sPD-L1 are associated with progressive disease（PD）a, partial response（PR）a and stable disease（SD）b in SCLC patients（a P<0.01，b P>0.05）.The expression of sPD-L1 in serum was positively correlated with ProGRP.View Large Image Figure ViewerDownload Hi-res image Download (PPT) Our results indicated that changes of plasma SPD-L1 levels in SCLC patients are associated with prognosis. Plasma sPD-L1 protein is a great biomarker in SCLC and may play an important role in sifting the beneficiaries of immunotherapy.

A new small cell lung cancer biomarker identified by Cell-SELEX generated aptamers

Small cell lung cancer (SCLC) has been a recalcitrant cancer without significant breakthroughs in clinical treatment during the past three decades. As there is a lack of effective protein inhibitor for SCLC targeted therapy, the discovery of new druggable SCLC biomarkers is a pressing work. Here we identified a new protein biomarker of SCLC, which is high density lipoprotein binding protein (HDLBP), through the aptamer generated by cell-SELEX against SCLC cells. Immunohistochemistry results showed an elevated HDLBP level in SCLC tissues from clinical samples. Attenuating HDLBP expression with siRNA inhibited proliferation and metastasis of SCLC cells in vitro and tumor formation in vivo. Mechanism study revealed the new function of HDLBP in promoting G1/S cell cycle transition for tumor progression. While the inhibitor of HDLBP has been reported, our work suggested a promising potential of targeting HDLBP to improve the treatment of fatal SCLC and a powerful tool of using cell-SELEX in cancer medicine.

Selective Fishing for Peptides with Antibody-Immobilized Acrylate Monoliths, Coupled Online with NanoLC-MS.

An online microfluidics-mass spectrometry platform was developed for determining proteotypic peptides from in-solution digested samples. Accelerated and selective sample cleanup was achieved by integrating proteotypic epitope peptide immunoextraction with nano liquid chromatography-tandem mass spectrometry (online IE-nanoLC-MS/MS). Ten individually prepared 180 μm inner diameter capillaries with ethylene glycol dimethacrylate- co-vinyl azlactone (EDMA- co-VDM) monoliths were immobilized with anti-protein antibodies that are used in routine immunoassays of the intact small cell lung cancer biomarker ProGRP. The resulting AB columns provided linearity correlation coefficients of 0.96-0.99 for protein amounts and concentrations of 10 pg to 5 ng and 0.5-250 ng/mL, respectively. The columns/platform gave relative peak area RSDs below 15%. The IE-nanoLC-MS/MS platform provided a limit of detection (LOD) of 520 pg/mL of ProGRP in human serum. The approach was applicable for other matrixes and proteins, i.e., primary glioblastoma cells and endogenous αV integrin chain. Thus, EDMA- co-VDM monoliths immobilized with antibodies are suited for automated peptide capture in microfluidic formats.

Transbronchoscopic patient biopsy-derived xenografts as a preclinical model to explore chemorefractory-associated pathways and biomarkers for small-cell lung cancer

Insufficient tumor tissue is a major barrier for cancer biology research in small-cell lung cancer (SCLC) and has driven the development of patient-derived xenografts (PDXs) from biopsy tumor tissues. Here, we utilized transbronchoscopic biopsy specimens from SCLC tumors to establish PDXs and evaluated the genomic profile using next-generation sequencing and an RNA sequencing platform. The PDX establishment rate was 54.1% (40/74). PDXs largely recapitulated the major characteristics of their corresponding primary tumors, such as histopathology, genetic profile, and chemo-responsiveness. Compared with chemosensitive (chemo-S) PDXs, chemorefractory (chemo-R) PDXs demonstrated significant gene aberrances in the mitogen-activated protein kinase (MAPK) pathway and a higher frequency of receptor tyrosine kinase (RTK)-related genes. Phosphorylated ERK (pERK) was associated with chemo-R status. Patients with positive pERK expression demonstrated significantly inferior progression-free survival after first-line chemotherapy compared with that of patients who were negative for pERK (p < 0.001). Collectively, transbronchoscopic biopsy SCLC PDXs can serve as a model for genomic profiling and identifying biomarkers predictive of chemo-R status. Using PDXs, RTK-related gene aberrances and pERK expression were found to be associated with chemo-R SCLC.

Circulating endothelial cells and microparticles as diagnostic and prognostic biomarkers in small-cell lung cancer

ObjectivesIt has been proposed that circulating endothelial cells (CECs) and microparticles (MPs) may be useful for the assessment of patients with non-small-cell lung cancer (NSCLC). However, little is known about the potential clinical relevance of these biomarkers in small-cell lung cancer (SCLC). Therefore, we investigated the utility of baseline levels of CECs and MPs in SCLC patients. Materials and methodsAn immunomagnetic separation (IMS) technique was used to isolate and quantify CECs in the peripheral blood, while plasma samples were analyzed using flow cytometry for the measurement of circulating MPs. ResultsWe prospectively collected data from 56 patients and 41 healthy individuals. Forty-three patients presented at initial diagnosis and 13 patients presented at relapse. Baseline levels of CECs and MPs were significantly higher in SCLC patients either at initial diagnosis or at relapse than in healthy subjects (p < 0.0002 and p < 0.007, respectively). However, estimated tumor volume (ETV) was significantly correlated with basal MP values (p < 0.0001) but not with pretreatment CECs (p = 0.57). The amount of baseline CECs and MPs was significantly lower in patients with an objective response (OR, n = 23) than in those with progressive disease (PD, n = 15) after treatment (p = 0.016 and 0.05, respectively). With cut-off values of 110 cells/mL for CECs and 1257 events/μL for MPs according to receiver operating characteristics (ROC) analysis, baseline levels of these biomarkers were not significantly correlated with either progression-free survival (PFS) or overall survival (OS). However, patients with 6-month PFS displayed significantly decreased pretreatment CEC counts (p = 0.042), whereas basal MP values significantly increased in 1-year survivors compared with those in non-survivors (p = 0.05). ConclusionOur results suggest that baseline CECs and MPs may be predictive biomarkers of tumor response and long-term survival in SCLC patients.

On-Chip Spyhole Nanoelectrospray Ionization Mass Spectrometry for Sensitive Biomarker Detection in Small Volumes.

A polyimide microfluidic chip with a microhole emitter (Ø 10-12μm) created on top of a microchannel by scanning laser ablation has been designed for nanoelectrospray ionization (spyhole-nanoESI) to couple microfluidics with mass spectrometry. The spyhole-nanoESI showed higher sensitivity compared to standard ESI and microESI from the end of the microchannel. The limits of detection (LOD) for peptide with the spyhole-nanoESI MS reached 50pM, which was 600 times lower than that with standard ESI. The present microchip emitter allows the analysis of small volumes of samples. As an example, a small cell lung cancer biomarker, neuron-specific enolase (NSE), was detected by monitoring the transition of its unique peptide with the spyhole-nanoESI MS/MS. NSE at 0.2nM could be well identified with a signal to noise ratio (S/N) of 50, and thereby its LOD was estimated to be 12pM. The potential application of the spyhole-nanoESI MS/MS in cancer diagnosis was further demonstrated with the successful detection of 2nM NSE from 1μL of human serum. Before the detection, the serum sample spiked with NSE was first depleted with immune spin column, then desalted by centrifugal filter device, and finally digested by trypsin, without any other complicated preparation steps. The concentration matched the real condition of clinical samples. In addition, the microchips can be disposable to avoid any cross contamination. The present technique provides a highly efficient way to couple microfluidics with MS, which brings additional values to various microfluidics and MS-based analysis. Graphical Abstract ᅟ.

Self-assembled selenium nanoparticles and their application in the rapid diagnostic detection of small cell lung cancer biomarkers.

By coupling molecular imprinting, chitosan biosorption and TiO2 photocatalysis, selenium nanoparticles (Se NPs) were self-assembled in a controlled manner on the molecular imprinting sites of zeolite-chitosan-TiO2 microspheres. Se NPs with different sizes and areal densities were individually synthesized by controlling the rapid adsorption of molecular-imprinted nanocomposites and photocatalytic reaction of TiO2 nanoparticles. In order to improve the sensitivity and specificity of rapid diagnostic detection, Se NPs were self-assembled again into high-order and spherically stable structures with an average size of 80 nm by well-defined monomer units, after separation from zeolite-chitosan-TiO2 microspheres with a stabilizer of 0.3% (v/v) bovine serum albumin. Due to their biological activity, spherical-shaped Se NPs were used for dot-blot immunoassays with multiple native antigens for rapid serodiagnosis of human lung cancer. The sensitivity of the dot immunoassays for detecting progastrin-releasing peptide (ProGRP) was 75 pg mL-1. The detection time of colloidal Se dot immunoassays for ProGRP was only 5 min. No positive results were observed with other commonly potential interfering substances, including carcinoembryonic antigen, α-fetoprotein antigen and BSA. The research presents a simple and green method for the reuse of SeO32- and the controlled synthesis of Se NPs for biological and medical applications by bioaffinity adsorption and photoreduction.

P1.05-024 Preoperative Neuron-Specific Enolase to Albumin Ratio is a Prognostic Biomarker for Patients with Operable Non-Small-Cell Lung Cancer

Neuron-specific Enolase (NSE) is a widely used tumor biomarker in small-cell lung cancer (SCLC) diagnosis, and serum albumin (Alb) levels are commonly used as indicators of the nutritional status of cancer patients. However, the prognostic value of these markers in combination has not been examined. This study was designed to explore the value of the combination between NSE and Alb in non-small-cell lung cancer (NSCLC). We retrospectively evaluated the prognostic value of the preoperative NSE to albumin ratio (NAR) in 319 patients with operable NSCLC. We analyzed associations among the NAR, clinicopathological characteristics, and inflammatory biomarkers. Univariate and multivariate analyses were performed to identify the clinicopathological characteristics associated with OS. Furthermore, we compared the prognostic value of the NAR with other established prognostic indexes by evaluating the area under the curves (AUC). The optimal NAR cutoff level was found to be 3.2×10-7. We found that a higher NAR was associated with more advanced TNM staged cancers (P=0.041) and higher tumor stage (P=0.011).The NAR was also associated with the inflammatory biomarker albumin/globulin ratio (AGR, P=0.032), but not the neutrophil/lymphocyte ratio (NLR, P=0.295) or platelet/lymphocyte ratio (PLR, P=0.260). In multivariate analyses, the NAR was an independent prognostic factor for NSCLC patients (P<0.001). The AUC of the NAR was higher than the NLR, PLR or AGR at 24 and 36 months of follow-up. The preoperative NAR might be an independent prognostic factor for patients with operable NSCLC, and a higher NAR indicates a poorer prognosis.

Abstract 3756: PARP inhibitor sensitivity in small cell lung cancer cell lines and patient-derived xenografts correlates with SLFN11 expression but not with structural homologous recombination deficiency

Abstract Poly-(ADP)-ribose polymerase (PARP) was identified as a potential novel therapeutic target for small cell lung cancer (SCLC) in two key proteomic studies. These findings led to the inclusion of a SCLC cohort in a Phase I study of talazoparib (BMN 673) - currently the most potent PARP inhibitor available - with promising efficacy signals. Identifying predictors of response to PARP inhibition would bring a needed therapeutic advance in this recalcitrant disease. PARP inhibition was first demonstrated to have marked efficacy in BRCA1/2 mutant cancers that are deficient in homologous recombination (HR). Large chromosomal structural alterations characteristic of these tumors can be quantitated by three correlated HR deficiency (HRD) metrics. We sought to determine if HRD would be a relevant predictive biomarker in SCLC. Thirty-six SCLC cell lines represented in the Cancer Cell Line Encyclopedia (CCLE) were interrogated for sensitivity to talazoparib, olaparib and cisplatin. We found significant correlation between these three pharmacologic agents and sought to determine response predictors. Three different HRD metrics were computed in these 36 SCLC cell lines using publically available CCLE Affymetrix SNP 6.0 array datasets. Surprisingly, none of these three measures of HRD were discriminatory for response to talazoparib in vitro. However, we found that after controlling for HRD and PARP1 target expression, expression of a single gene, SLFN11, was significantly correlated with talazoparib sensitivity as measured by the CCLE Affymetrix U133+2 arrays (q value = 0.008). After deriving SLFN11 knock-out SCLC cell lines by CRISPR/Cas9 gene editing, we found that loss of SLFN11 conferred resistance to talazoparib (IC50 &amp;gt;log difference) as compared to parental SLFN11 proficient cells. We then treated several SCLC patient-derived xenograft (PDX) models with talazoparib daily and found that SLFN11 expression in vivo was necessary but not sufficient for response. None of the SLFN11 low expressing PDXs responded to therapy, whereas 3/5 (60%) of SLFN11 high expressing PDXs demonstrated significant tumor growth inhibition of 54-75% after 21-28 days of daily treatment with talazoparib 0.3mg/kg. In conclusion, we demonstrate that SLFN11 expression is correlated with response to PARP inhibition in SCLC in vitro and in vivo. HRD structural aberration scores do not predict for in vitro PARP inhibitor sensitivity, however functional HR interrogation (such as RAD51 foci recruitment) may still be a relevant discriminator in addition to ongoing in vivo analysis. SLFN11 may be a pertinent predictive biomarker in SCLC for therapeutic response to PARP inhibition with considerable clinical implications. Citation Format: Benjamin H. Lok, Ying Feng, G. Karen Yu, Eric E. Gardner, Nadeem Riaz, Yuanbin (Kevin) Ru, Elisa de Stanchina, Simon N. Powell, Jerry Shen, John T. Poirier, Charles M. Rudin. PARP inhibitor sensitivity in small cell lung cancer cell lines and patient-derived xenografts correlates with SLFN11 expression but not with structural homologous recombination deficiency. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3756.

Genomic Landscape Survey Identifies SRSF1 as a Key Oncodriver in Small Cell Lung Cancer.

Small cell lung cancer (SCLC) is an aggressive disease with poor survival. A few sequencing studies performed on limited number of samples have revealed potential disease-driving genes in SCLC, however, much still remains unknown, particularly in the Asian patient population. Here we conducted whole exome sequencing (WES) and transcriptomic sequencing of primary tumors from 99 Chinese SCLC patients. Dysregulation of tumor suppressor genes TP53 and RB1 was observed in 82% and 62% of SCLC patients, respectively, and more than half of the SCLC patients (62%) harbored TP53 and RB1 mutation and/or copy number loss. Additionally, Serine/Arginine Splicing Factor 1 (SRSF1) DNA copy number gain and mRNA over-expression was strongly associated with poor survival using both discovery and validation patient cohorts. Functional studies in vitro and in vivo demonstrate that SRSF1 is important for tumorigenicity of SCLC and may play a key role in DNA repair and chemo-sensitivity. These results strongly support SRSF1 as a prognostic biomarker in SCLC and provide a rationale for personalized therapy in SCLC.

Clinical evaluation and therapeutic monitoring value of serum tumor markers in lung cancer.

Tumor markers CYFRA21-1, CEA, NSE, CA125, pro-GRP and SCC are routinely used for lung cancer. However, there has been no systematic evaluation of these markers in the same cohort. The aim of this study was to evaluate the diagnostic and therapeutic monitoring value of these markers. The levels of 6 serum tumor markers were measured in 392 patients, including 308 patients with non-small cell lung cancer (NSCLC) and 84 with small cell lung cancer (SCLC), and 116 patients with benign lung diseases and 144 healthy controls. 34 patients were followed up after operation and chemotherapy. Multiple logistic models and receiver operating characteristic (ROC) curves were used to evaluate their diagnostic value. CEA, NSE, CA125 and pro-GRP in SCLC, and CYFRA21-1 as well as CEA in NSCLC, were higher than those in control groups. The level of CEA and CA125 were related to the clinical stages of NSCLC. Pro-GRP was significantly increased in extensive disease (ED) compared with limited disease (LD) in SCLC. CYFRA21-1 was reduced after the third and fifth treatment cycle respectively in patients who undergoing operation and without operation. NSE and pro-GRP were reduced significantly after the second and third treatment cycles, respectively. CEA, NSE, CA125 and pro-GRP could serve as biomarkers for SCLC, and CEA and CYFRA21-1 could serve as biomarkers for NSCLC. Pro-GRP, CA125 and CEA were related to the clinical stages of lung cancer. CYFRA21-1, NSE and pro-GRP could be used for monitoring the effect of chemotherapy.

Abstract C153: Investigation of Chk1 as a novel therapeutic target for small cell lung cancer (SCLC)

Abstract Background: Small cell lung cancer (SCLC) is the most lethal form of lung cancer, accounting for around 15% of lung cancers in the United States, currently having no targeted therapies with established efficacy. Using a high-throughput proteomic screen, we demonstrated overexpression of checkpoint kinase 1 (Chk1) in SCLC. Chk1 being the master regulator of the cell cycle in absence of p53, we hypothesize that Chk1 targeting may be effective SCLC because of near ubiquitous loss of p53. In the present study, we tested the effect of Chk1 targeting by shRNA-mediated knockdown and pharmacologic inhibition in an extensive panel of SCLC cell lines and in animal models. Experimental procedures: shRNA-mediated knockdown of Chk1 was done in select human SCLC lines and confirmed at the mRNA and protein level. The effect of Chk1 inhibition by LY2606368 alone and in combination with cisplatin or the PARP1/2 inhibitor talazoparib was tested in 39 SCLC cell lines. Proliferation was assessed by cell titer glo analysis and effect on cell cycle was tested by flow cytometry. Drug sensitivity (IC50) was then correlated with baseline expression level of &amp;gt;190 total or phosphorylated proteins measured by reverse phase protein array (RPPA) to identify potential predictive biomarkers. In vivo effect of the Chk1 inhibitor with or without cisplatin or talazoparib was tested in syngeneic and xenograft model of SCLC. Results: shRNA-mediated targeted Chk1 knockdown significantly decreased the proliferation of SCLC lines confirming Chk1 as a viable target (p&amp;lt;0.01). Targeted Chk1 knockdown also increased the expression of pγH2Ax indicating double-stranded DNA damage upon Chk1 inhibition. Chk1 inhibition by LY2606368 had striking single-agent activity in a majority of SCLC lines (median IC50 2-50nM) and the combination of LY2606368 with either cisplatin or talazoparib was additive in a majority of cell lines. Proteomic analysis revealed association between Chk1 inhibitor sensitivity and elevated basal expression of total and phospho cMyc (p = 0.01), 14-3-3 protein (p = 0.003), pro-apoptotic protein Bax (p&amp;lt;0.05); in contrast, resistance to LY2606368 at multiple doses was correlated to with higher levels of PARP, SGK and p90RSK (p&amp;lt;0.05). In syngeneic in vivo model, single agent LY2606368 was well tolerated and resulted in near-total regression of tumor (&amp;gt;90% reduction in volume) which was durable (ongoing response &amp;gt;50 days in 7/9 animals). The combination with cisplatin or talazoparib also resulted in tumor regression and was significantly more effective than either cisplatin or talazoparib alone. Tumor regression following LY2606368 treatment was also observed in cis-resistant H69 xenograft model of SCLC. Conclusion: The study established checkpoint kinase 1 (Chk1) targeting as a novel and active therapeutic strategy for SCLC, a disease for which the standard of care has remained unchanged for &amp;gt;30 years. Chk1 knockdown was sufficient to inhibit proliferation in SCLC cell lines and LY2606368, a Chk1 inhibitor in current clinical trials, showed activity as a single agent and in combination with cisplatin and PARP inhibition in vitro and in vivo. TP53-mutant cells (hallmark of SCLC) rely on Chk1 for cell cycle arrest; hence, inhibition of Chk1 in SCLC likely works by driving cells to mitotic catastrophe and apoptosis. These findings support the further investigation of Chk1 targeting and Chk1 predictive biomarkers in SCLC. Citation Format: Triparna Sen, Pan Tong, C. Allison Stewart, You Hong-Fan, Ying Feng, Jing Wang, Lauren A. Byers. Investigation of Chk1 as a novel therapeutic target for small cell lung cancer (SCLC). [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr C153.

Challenges and Opportunities for Identifying Non–Small Cell Lung Cancer Biomarkers Using Bronchoscopy

Non–small cell lung cancer is no longer viewed as a single disease, but a constellation of cancer types that benefit from different treatments. Advancements in the identification of driver mutations with specific Food and Drug Administration–regulated molecular tests have been approved for matching the right treatment to the right patient. These biomarkers, referred to as theranostic biomarkers, because of their ability to support treatment decisions and predict a clinical response are changing the way oncologists formulate non–small cell lung cancer treatment plans. Currently approved biomarker tests for lung cancer include fluorescence in situ hybridization testing for genetic rearrangements and reverse-transcriptase polymerase chain reaction testing for genetic mutations. As the use of molecular medicine progresses, an emerging treatment challenge in thoracic oncology is using limited cytologic material to reliably measure the essential theranostic biomarkers. The majority of newly diagnosed lung cancer patients have advanced disease at the time of presentation, and often undergo the least invasive biopsy possible to ascertain a diagnosis and disease stage. As less invasive diagnostic techniques become more common, it is critical to optimize collection and minimize loss of cancer tissue during processing. The physicians performing these procedures must be knowledgeable about tissue requirements for molecular testing based on tumor type, and should develop collection strategies to optimize molecular biomarker testing. This review explores current literature recommendations to maximize the collection and processing of cytology material for biomarker evaluation and evaluate which cytology samples provide the best yields. As our ability to truly individualize lung cancer therapy evolves, it will be critical to reliably evaluate and treat based on biomarker analysis.

Abstract 1546: The research of the mechanism of lung cancer carcinogenesis and the target gene of microRNA-20a in lung cancer small cells

Abstract Purpose: Recent studies have suggested that microRNA biomarkers could be useful for stratifying lung cancer subtypes, but microRNA signatures varied between different populations. Small cell lung cancer (SCLC) is the lowest differentiation degree and the highest malignant subtype of lung cancer that urgently needs biomarkers to aid patient management. Here, we undertook the first comprehensive investigation on microRNA in Chinese SCLC patients of limited stage. Experimental Design: MicroRNA expression was measured in cancerous and noncancerous tissue pairs strictly collected from Chinese SCLC patients of limited stage, who had not been treated with chemotherapy or radiotherapy prior to surgery. The molecular targets of proposed microRNA were further examined. Results: We identified that high expression of hsa-miR-20a was associated with poor survival in these 20 SCLC patients by Kaplan-Meier analysis (P=0.03), This association was independently validated in a separate cohort of 38 SCLC patients in Kaplan-Meier analysis(P=0.046). We then determined that the tumor suppressor PTEN is a target of hsa-miR-20a. Expression of hsa-miR-20a in a human lung cancer cell line repressed PTEN activity. Conclusions: Our results identified a new prognostic biomarker of SCLC, hsa-miR-20a. Note: This abstract was not presented at the meeting. Citation Format: Tan Xiaogang, Hu Mu, Zhi Xiuyi, Jin Shimeng. The research of the mechanism of lung cancer carcinogenesis and the target gene of microRNA-20a in lung cancer small cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1546. doi:10.1158/1538-7445.AM2014-1546

Multiplexing Determination of Small Cell Lung Cancer Biomarkers and Their Isovariants in Serum by Immunocapture LC-MS/MS

A multiplex method for the determination of the small cell lung cancer (SCLC) markers progastrin releasing peptide (ProGRP) and neuron specific enolase (NSE) is presented, which involves coextraction by immunoaffinity (IA) beads and codetermination by selected reaction monitoring (SRM). The performance was compared with two IA SRM methods which were recently validated for individual marker determination. The multiplexing method reduces sample volume, handling time per sample, and reagent consumption and shows good linearity, recovery, quantitative measurements, and sensitivity with lower limit of detection (LLOD) values of 7.2 pM (=90 pg/mL) and 4.5 pM (=210 pg/mL) and lower limit of quantitation (LLOQ) values of 24 pM (=300 pg/mL) and 15 pM (=700 pg/mL), for total ProGRP and γ-NSE, respectively. The novel aspect of this approach is the multiplexing of ProGRP and NSE with the additional ability to perform fingerprinting by the selective determination of ProGRP isoform 1, ProGRP isoform 3, and total ProGRP, as well as the α- and the γ-subunit of NSE isoenzymes. Six serum samples from patients with SCLC were analyzed to demonstrate the methods feasibility to simultaneously differ between and individually quantify ProGRP, NSE, and their isoform and isoenzyme variants, respectively. Both the presence of and variation between all the isoforms and isoenzymes, as well as covarying results with the conventional immunometric assays for total ProGRP and γ-NSE, were seen in the analyses of patient serum samples.

Evaluation of serum cytokines in small cell lung cancer and its clinical significance

背景与目的已有的研究表明：小细胞肺癌（small cell lung cancer, SCLC）患者可出现血清细胞因子表达水平的变化，本研究通过检测SCLC患者血清中细胞因子的差异改变，探讨其在SCLC中的诊断价值。方法首先使用Reybiotech G6/G7细胞因子芯片对4例SCLC患者、4例健康人和4例炎症患者血清进行细胞因子差异表达筛查，进一步应用酶联免疫吸附法（ELISA试剂盒）对197例SCLC患者、180例正常对照血清以及97例炎性病变患者血清进行验证。结果芯片检测120种细胞因子，从有明显差异的细胞因子中选出最有研究价值的4种进行验证，包括尿激酶型纤溶酶原激活剂受体（urokinase plasminogen activator receptor, uPAR）、人瘦素（Leptin）、巨噬细胞刺激蛋白（macrophage stimulating protein α, MSP-α）和巨噬细胞炎症蛋白1β（macrophage inflammatory protein 1β, MIP-1β）。采用ELISA法验证上述结果，SCLC患者血清uPAR较健康人群以及炎症患者增高（P < 0.05），诊断的敏感度为52.93%，特异度为83.36%。Leptin在无体重变化SCLC组较健康人群以及炎症患者增高，诊断的敏感度为50.11%，特异度86.77%；而Leptin在体重下降组较对照组无明显差异。此外，血清MSP-α、MIP-1β水平在三组之间比较无明显差异。结论血清uPAR升高在SCLC中具有一定的诊断价值，而Leptin在无体重变化的SCLC中可能具有诊断意义。