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The tumorous niche may drive the plasticity of heterogeneity and cancer stemness, leading to drug resistance and metastasis, which is the main reason of treatment failure in most cancer patients. The aim of this study was to establish a tumor microenvironment (TME)-based screening to identify drugs that can specifically target cancer stem cells (CSCs) and cancer-associated fibroblasts (CAFs) in the TME. Methods: Lung cancer patient-derived cancer cell and CAFs were utilized to mimic the TME and reproduce the stemness properties of CSCs in vitro and develop a high-throughput drug screening platform with phenotypical parameters. Limiting dilution assay, sphere-forming and ALDH activity assay were utilized to measure the cancer stemness characteristics. In vivo patient-derived xenograft (PDX) models and single-cell RNA sequencing were used to evaluate the mechanisms of the compounds in CSCs and CAFs. Results: The TME-based drug screening platform could comprehensively evaluate the response of cancer cells, CSCs and CAFs to different treatments. Among the 1,524 compounds tested, several drugs were identified to have anti-CAFs, anticancer and anti-CSCs activities. Aloe-emodin and digoxin both show anticancer and anti-CSCs activity in vitro and in vivo, which was further confirmed in the lung cancer PDX model. The combination of digoxin and chemotherapy improved therapeutic efficacy. The single-cell transcriptomics analysis revealed that digoxin could suppress the CSCs subpopulation in CAFs-cocultured cancer cells and cytokine production in CAFs. Conclusions: The TME-based drug screening platform provides a tool to identify and repurpose compounds targeting cancer cells, CSCs and CAFs, which may accelerate drug development and therapeutic application for lung cancer patients.

Background Malignant pleural effusion (MPE)-macrophage (Mφ) of lung cancer patients within unique M1/M2 spectrum showed plasticity in M1–M2 transition. The M1/M2 features of MPE-Mφ and their significance to patient outcomes need to be clarified; furthermore, whether M1-repolarization could benefit treatment remains unclear. Methods Total 147 stage-IV lung adenocarcinoma patients undergoing MPE drainage were enrolled for profiling and validation of their M1/M2 spectrum. In addition, the MPE-Mφ signature on overall patient survival was analyzed. The impact of the M1-polarization strategy of patient-derived MPE-Mφ on anti-cancer activity was examined. Results We found that MPE-Mφ expressed both traditional M1 (HLA-DRA) and M2 (CD163) markers and showed a wide range of M1/M2 spectrum. Most of the MPE-Mφ displayed diverse PD-L1 expression patterns, while the low PD-L1 expression group was correlated with higher levels of IL-10. Among these markers, we identified a novel two-gene MPE-Mφ signature, IL-1β and TGF-β1, representing the M1/M2 tendency, which showed a strong predictive power in patient outcomes in our MPE-Mφ patient cohort (N = 60, p = 0.013) and The Cancer Genome Atlas Lung Adenocarcinoma dataset (N = 478, p Conclusions We identified MPE-Mφ on the M1/M2 spectrum and plasticity and described a two-gene M1/M2 signature that could predict the outcome of late-stage lung cancer patients. In addition, we found that “re-education” of these MPE-Mφ toward anti-cancer M1 macrophages using clinically applicable strategies may overcome tumor immune escape and benefit anti-cancer therapies.

Lung cancer is the leading cause of cancer death worldwide and cancer relapse accounts for the majority of cancer mortality. The mechanism is still unknown, especially in hereditary lung cancer without known actionable mutations. To identify genetic alternations involved in hereditary lung cancer and relapse is urgently needed. We collected genetic materials from a unique hereditary lung cancer patient's blood, first cancer tissue (T1), adjacent normal tissue (N1), relapse cancer tissue (T2), and adjacent normal tissue (N2) for whole genome sequencing. We identified specific mutations in T1 and T2, and attributed them to tumorigenesis and recurrence. These tumor specific variants were enriched in antigen presentation pathway. In addition, a lung adenocarcinoma cohort from the TCGA dataset was used to confirm our findings. Patients with high mutation burdens in tumor specific genes had decreased relapse‐free survival (P = 0.017, n = 186). Our study may provide important insight for designing immunotherapeutic treatment for hereditary lung cancer.

PURPOSE Epidermal growth factor receptor (EGFR)-tyrosine kinase inhibitors (TKIs) show efficacy in treating patients with lung adenocarcinoma with EGFR-activating mutations. However, a significant subset of targeted patients fail to respond. Unlike acquired resistance (AR), intrinsic resistance (IR) remains poorly understood. We investigated whether epigenomic factors contribute to patient-to-patient heterogeneity in the EGFR-TKI response and aimed to characterize the IR subpopulation that obtains no benefit from EGFR-TKIs. PATIENTS AND METHODS We conducted genome-wide DNA methylation profiling of 79 tumors sampled from patients with advanced lung adenocarcinoma before they received EGFR-TKI treatment and analyzed the patient responses. Pyrosequencing was performed in a validation cohort of 163 patients with EGFR-activating mutations. RESULTS A DNA methylation landscape of 216 CpG sites with differential methylation was established to elucidate the association of DNA methylation with the characteristics and EGFR-TKI response status of the patients. Functional analysis of 37 transcription-repressive sites identified the enrichment of transcription factors, notably homeobox ( HOX) genes. DNA methylation of HOXB9 (cg13643585) in the enhancer region yielded 88% sensitivity for predicting drug response (odds ratio [OR], 6.64; 95% CI, 1.98 to 25.23; P = .0009). Pyrosequencing validated that HOXB9 gained methylation in patients with a poor EGFR-TKI response (OR, 3.06; 95% CI, 1.13 to 8.19; P = .019). CONCLUSION Our data suggest that homeobox DNA methylation could be a novel tumor cellular state that can aid the precise categorization of tumor heterogeneity in the study of IR to EGFR-TKIs. We identified, for the first time, an epigenomic factor that can potentially complement DNA mutation status in discriminating patients with lung adenocarcinoma who are less likely to benefit from EGFR-TKI treatment, thereby leading to improved patient management in precision medicine.

// Hsuan-Yu Chen 1, 2, 3 , Chia-Hsin Liu 1, 4, 5, * , Ya-Hsuan Chang 1, * , Sung-Liang Yu 6 , Bing-Ching Ho 6 , Chung-Ping Hsu 7 , Tsung-Ying Yang 7 , Kun-Chieh Chen 7 , Kuo-Hsuan Hsu 7 , Jeng-Sen Tseng 7 , Jiun-Yi Hsia 7 , Cheng-Yen Chuang 7 , Chi-Sheng Chang 6 , Yu-Cheng Li 1 , Ker-Chau Li 1 , Gee-Chen Chang 7, 8, 9 , Pan-Chyr Yang 6, 10, 11 1 Institute of Statistical Sciences, Academia Sinica, Taipei, Taiwan 2 College of Medicine, National Taiwan University, Taipei, Taiwan 3 College of Life Science, National Taiwan University, Taipei, Taiwan 4 Bioinformatics Program, Taiwan International Graduate Program, Academia Sinica, Taipei, Taiwan 5 Institute of Biomedical Informatics, National Yang-Ming University, Taipei, Taiwan 6 Center of Genomic Medicine, National Taiwan University, Taipei, Taiwan 7 Taichung Veterans General Hospital, Taichung, Taiwan 8 Faculty of Medicine, School of Medicine, National Yang-Ming University, Taipei, Taiwan 9 Comprehensive Cancer Center, Taichung Veterans General Hospital, Taichung, Taiwan 10 Department of Internal Medicine, National Taiwan University Hospital, Taipei, Taiwan 11 Institute of Biomedical Sciences, Academia Sinica, Taipei, Taiwan * Chia-Hsin Liu and Ya-Hsuan Chang contributed equally as the first author Correspondence to: Hsuan-Yu Chen, e-mail: hychen@stat.sinica.edu.tw Gee-Chen Chang, e-mail: august@vghtc.gov.tw Keywords: lung adenocarcinoma, DNA copy number abundance, ErbB family, EGFR-activating mutation, prognosis Received: July 07, 2015 Accepted: January 12, 2016 Published: January 27, 2016 ABSTRACT In this study, EGFR -activating mutation status and DNA copy number abundances of members of ErbB family were measured in 261 lung adenocarcinomas. The associations between DNA copy number abundances of ErbB family, EGFR -activating mutation status, and prognosis were explored. Results showed that DNA copy number abundances of EGFR , ERBB2 , ERBB3 , and ERBB4 had associations with overall survival in lung adenocarcinoma with EGFR -activating mutations. In the stratification analysis, only ERBB2 showed significant discrepancy in patients carrying wild type EGFR and other members of ErbB family in patients carrying EGFR -activating mutation. This indicated that CNAs of ErbB family had effect modifications of EGFR -activating mutation status. Findings of this study demonstrate potential molecular guidance of patient management of lung adenocarcinoma with or without EGFR -activating mutations.

e17512 Background: Very-small-nuclear circulating tumor cells (vsnCTCs) as a subset of CTCs with nuclear size < 8.5 μm associated with visceral metastases (VM) in advanced metastatic, castration-resistant prostate cancer (mCRPC). As VM predicts foreshortened survival, we hypothesized that vsnCTCs would be directly associated with poor overall survival (OS). Methods: Clinically annotated, blood samples from patients with mCRPC with survival follow-up within the Translational Oncology Program Blood & Biospecimen Bank were selected for analysis. CTCs were isolated and analyzed using the NanoVelcro assay. The nuclear size from all CTCs from each patient sample was compared to OS and progression-free survival (PFS) from the time of the blood draw. Results: A total of 76 patients had samples suitable for analysis. Sixty-six (87%) had CTCs; 49 (64%) were vsnCTC+ (≥1 vsnCTC). vsnCTCs were more common in mCRPC patients with previous androgen receptor signaling inhibitor (ARSI) therapy and/or 2 or more lines of treatment. OS was significantly shorter for vsnCTC+ than vsnCTC- patients (median 34 vs. 149 weeks; log-rank HR = 2.6; 95% CI = 1.5 to 4.5; P = 0.0006). Fifty patient samples were available for PFS analysis (i.e. drawn within 4 weeks of starting therapy). vsnCTC+ patients experienced more rapid progression than vsnCTC- patients (median 12 vs. 26 weeks, log-rank HR = 2.2, 95% CI = 1.3 to 4.0; P = 0.004). Multivariable Cox regression revealed that vsnCTC status was independently associated with OS and PFS. P-spline plot analysis showed that the HR of OS increased as the minimum CTC nuclear size decreased. The minimum CTC nuclear size was also independently associated with OS and PFS in a multivariable Cox regression analysis. Patients with prior use of androgen receptor signaling inhibitor (ARSI) therapy had significantly smaller minimum CTC nuclear size compared to those without prior use of ARSI. Average and median nuclear size did not strongly associate with OS or PFS. Conclusions: vsnCTC+ patients are at risk for more rapid clinical progression and have greater risk of death than vsnCTC-. We posit that the vsnCTC may be a biomarker of aggressive mCRPC with foreshortened survival. Interestingly, the CTCs with the smallest nuclei appear to best reflect the observed clinical behavior. This has potential importance in optimizing therapeutic choices and may point toward a unique biology relating nuclear size to aggressive molecular features. Our group continues to explore the biology underlying the vsnCTC.

Predicting a patient’s risk of recurrence after the resection of liver metastases from colorectal cancer is critical for evaluating and selecting therapeutic approaches. Clinical and pathologic parameters have shown limited accuracy thus far. Therefore, we combined the clinical status with a genomic approach to stratify relapse-free survival in colorectal cancer liver metastases patients. To identify new molecular and genetic signatures specific to colorectal cancer with liver metastasis (CRCLM) patients, we conducted DNA copy number profiling on a cohort of 21 Taiwanese CRCLM patients using a comparative genomic hybridization (CGH) array. We identified a three-gene signature based on differential copy number alteration between patients with different statuses of (1) recurrence and (2) synchronous metastasis. In relapse hotspot regions, only three genes (S100PBP, CSMD2, and TGFBI) were significantly associated with the synchronous liver metastasis factor. A final set of three genes—S100PBP, CSMD2, TGFBI—significantly predicted relapse-free survival in our cohort (p = 0.04) and another CRCLM cohort (p = 0.02). This three-gene signature is the first genomic signature validated for relapse-free survival in post-hepatectomy CRCLM patients. Our three-gene signature was developed using a whole-genome CGH array and has a good prognostic position for the relapse-free survival of CRCLM patients after hepatectomy.

BACKGROUND Although enumeration of circulating tumor cells (CTCs) has shown some clinical value, the pool of CTCs contains a mixture of cells that contains additional information that can be extracted. The authors subclassified CTCs by shape features focusing on nuclear size and related this with clinical information. METHODS A total of 148 blood samples were obtained from 57 patients with prostate cancer across the spectrum of metastatic states: no metastasis, nonvisceral metastasis, and visceral metastasis. CTCs captured and enumerated on NanoVelcro Chips (CytoLumina, Los Angeles, Calif) were subjected to pathologic review including nuclear size. The distribution of nuclear size was analyzed using a Gaussian mixture model. Correlations were made between CTC subpopulations and metastatic status. RESULTS Statistical modeling of nuclear size distribution revealed 3 distinct subpopulations: large nuclear CTCs, small nuclear CTCs, and very small nuclear CTCs (vsnCTCs). Small nuclear CTCs and vsnCTC identified those patients with metastatic disease. However, vsnCTC counts alone were found to be elevated in patients with visceral metastases when compared with those without (0.36 ± 0.69 vs 1.95 ± 3.77 cells/mL blood; P

α-Catulin is an oncoprotein that helps sustain proliferation by preventing cellular senescence. Here, we report that α-catulin also drives malignant invasion and metastasis. α-Catulin was upregulated in highly invasive non–small cell lung cancer (NSCLC) cell lines, where its ectopic expression or short-hairpin RNA–mediated attenuation enhanced or limited invasion or metastasis, respectively. α-Catulin interacted with integrin-linked kinase (ILK), a serine/threonine protein kinase implicated in cancer cell proliferation, antiapoptosis, invasion, and angiogenesis. Attenuation of ILK or α-catulin reciprocally blocked cell migration and invasion induced by the other protein. Mechanistic investigations revealed that α-catulin activated Akt-NF-κB signaling downstream of ILK, which in turn led to increased expression of fibronectin and integrin αvβ3. Pharmacologic or antibody-mediated blockade of NF-κB or αvβ3 was sufficient to inhibit α-catulin–induced cell migration and invasion. Clinically, high levels of expression of α-catulin and ILK were associated with poor overall survival in patients with NSCLC. Taken together, our study shows that α-catulin plays a critical role in cancer metastasis by activating the ILK-mediated Akt-NF-κB-αvβ3 signaling axis. Cancer Res; 73(1); 428–38. ©2012 AACR.

INTRODUCTION AND OBJECTIVES: The relation of serum androgens and the development of prostate cancer (PCa) is subject of debate. Lower total testosterone (TT) levels have been associated with increased PCa detection and worse pathological features after treatment. The aim of this study is to investigate the association of serum androgens levels and PCa detection in a prospective screening study of men at higher genetic risk due to BRCA mutation METHODS: Six hundred and fifty seven participants in the IMPACT study had sequential TT and sexual hormone binding globulin (SHBG) measurements in serum samples at annual visits, giving 2783 prospective androgen levels, from 234 BRCA1 carriers, 268 BRCA2 carriers and 155 mutation negative controls. Free and bioavailable testosterone were calculated from TT and SHBG using the Sordergaard equation. Age, BMI, PSA and hormonal concentrations were compared between men affected and unaffected with PCa within each genetic cohort using unpaired t-test. The Fisher exact test was used to correlate hypogonadic levels of testosterone and genetic status with PCa diagnosis within each genetic category RESULTS: There were no significant differences in the PSA levels and BMI at study entry. Mean ages were 53.52, 52.17, and 51.17 years, for the control, BRCA1 and BRCA2 groups, respectively (BRCA1 p1⁄40.24 and BRCA2 p1⁄40.01). In the BRCA2 cohort, serum TT levels were lower in carriers diagnosed with PCa, compared with unaffected carriers and controls (11.2, 13.78, 13.4 nmol/L, mean values per group, respectively, p1⁄40.02 and p1⁄40.009). In the BRCA1 cohort, no significant difference was observed between affected and unaffected carriers and controls (13.6, 13.5, 13.4, nmol/L, mean values per group respectively, p1⁄40.82 and p-0.69). SHBG levels were significantly higher in BRCA1 carriers with PCa than unaffected carriers and controls (47.62, 37.97, 40.34 nmol/L, respectively; p1⁄40.002 and p1⁄40.03). TT levels

Lung cancer is the leading cause of cancer-related deaths worldwide. High-throughput technologies such as microarrays provide an opportunity to explore biomarkers for cancer prevention, prognosis and treatment guidance. Recent studies have revealed many biomarkers with the potential for clinical application. However, major limitations still exist. Although useful data on cancer genomics has accumulated rapidly, there has also been a simultaneous tendency for amplification of the complex relationships among the enormous number of variables that need to be considered. Disentangling these complex gene–gene interactions requires new approaches to data analysis to reveal information that has been obscured by traditional methods. Here, we review the current findings on biomarker identification in lung cancer, address their limitations and discuss some future directions for improvements in this area of research.

Purpose Patients with non–small-cell lung cancer (NSCLC) with epidermal growth factor receptor (EGFR)–activating mutations have excellent response to EGFR tyrosine kinase inhibitors (TKIs), but T790M mutation accounts for most TKI drug resistance. This study used highly sensitive methods to detect T790M before and after TKI therapy and investigated the association of T790M and its mutation frequencies with clinical outcome. Patients and Methods Direct sequencing, matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) and next-generation sequencing (NGS) were used to assess T790M in the following two cohorts of patients with NSCLC: TKI-naive patients (n = 107) and TKI-treated patients (n = 85). Results were correlated with TKI treatment response and survival. Results MALDI-TOF MS was highly sensitive in detecting and quantifying the frequency of EGFR-activating mutations and T790M (detection limits, 0.4% to 2.2%). MALDI-TOF MS identified more T790M than direct sequencing in TKI-naive patients with NSCLC (27 of 107 patients, 25.2% v three of 107 patients, 2.8%, respectively; P < .001) and in TKI-treated patients (before TKI: 23 of 73 patients, 31.5% v two of 73 patients, 2.7%, respectively; P < .001; and after TKI: 10 of 12 patients, 83.3% v four of 12 patients, 33.3%, respectively; P = .0143). The EGFR mutations and their frequencies were confirmed by NGS. T790M was an independent predictor of decreased progression-free survival (PFS) in patients with NSCLC who received TKI treatment (P < .05, multivariate Cox regression). Conclusion T790M may not be a rare event before or after TKI therapy in patients with NSCLC with EGFR-activating mutations. The pretreatment T790M mutation was associated with shorter PFS with EGFR TKI therapy in patients with NSCLC.

Purpose Although epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have been proven more effective for patients with lung adenocarcinoma with EGFR-activating mutation rather than wild type, the former group still includes approximately 30% nonresponders. The molecular basis of this substantial response heterogeneity is unknown. Our purpose was to seek molecular aberrations contributing to disease progression at the genome-wide level and identify the prognostic signature unique to patients with EGFR-activating mutation. Patients and Methods We first investigated the molecular differences between tumors with EGFR-activating mutation and wild-type tumors by conducting high-density array comparative genomic hybridization on a collection of 138 adenocarcinoma tissues. We then used an independent group of 114 patients to validate the clinical relevance of copy-number alterations (CNAs) in predicting overall and disease-free survival. Finally, focusing on 23 patients with EGFR mutation receiving EGFR-TKI treatment, we investigated the association between CNAs and response to EGFR-TKIs. Results We identified chromosome regions with differential CNAs between tumors with EGFR-activating mutation and wild-type tumors and found the aberration sites to cluster highly on chromosome 7p. A cluster of six representative chromosome 7p genes predicted overall and disease-free survival for patients with EGFR-activating mutation but not for those with wild type. Importantly, simultaneous presence of more genes with increased CNAs in this cluster correlated with less favorable response to EGFR-TKIs in patients with EGFR-activating mutation. Conclusion Our results shed light on why responses to EGFR-TKIs are heterogeneous among patients with EGFR-activating mutation. They may lead to better patient management in this population.

Previous studies have demonstrated focal but limited molecular similarities between circulating tumor cells (CTCs) and biopsies using isolated genetic assays. We hypothesized that molecular similarity between CTCs and tissue exists at the single cell level when characterized by whole genome sequencing (WGS). By combining the NanoVelcro CTC Chip with laser capture microdissection (LCM), we developed a platform for single-CTC WGS. We performed this procedure on CTCs and tissue samples from a patient with advanced prostate cancer who had serial biopsies over the course of his clinical history. We achieved 30X depth and ≥ 95% coverage. Twenty-nine percent of the somatic single nucleotide variations (SSNVs) identified were founder mutations that were also identified in CTCs. In addition, 86% of the clonal mutations identified in CTCs could be traced back to either the primary or metastatic tumors. In this patient, we identified structural variations (SVs) including an intrachromosomal rearrangement in chr3 and an interchromosomal rearrangement between chr13 and chr15. These rearrangements were shared between tumor tissues and CTCs. At the same time, highly heterogeneous short structural variants were discovered in PTEN, RB1, and BRCA2 in all tumor and CTC samples. Using high-quality WGS on single-CTCs, we identified the shared genomic alterations between CTCs and tumor tissues. This approach yielded insight into the heterogeneity of the mutational landscape of SSNVs and SVs. It may be possible to use this approach to study heterogeneity and characterize the biological evolution of a cancer during the course of its natural history.

One method for characterizing spatiotemporal variation in brain activity levels is based on the use of statistical dimension reduction. This reduction finds temporal components in data that best preserve the spatiotemporal regression structure. The method does this by suppressing more prominent waveforms that do not vary in a spatially predictable pattern.

Introduction and Objective: Patients with metastatic castration-resistant prostate cancer (mCRPC) who develop visceral metastases (VM) have poorer clinical outcomes in comparison to those without VM. Currently, VM are discovered late in the clinical course of mCRPC-VM and this aggressive natural history typically culminates in organ failure. There are no existing tests that identify men at risk for VM other than radiography. Our team performed circulating tumor cell (CTC) enumeration using NanoVelcro CTC Assay on prostate cancer patients across the spectrum of metastatic states: no metastasis, non-visceral metastasis, and VM. We identified an association between the presence of very-small-nuclear CTCs (vsnCTCs, DAPI+/Cytokeratin+/CD45- with nuclear size < 8.5μm) and VM. Serial enumeration studies suggested the emergence of vsnCTCs occurred before the radiographic detection of VM, and the change of vsnCTC counts reflected the patients’ disease progression. We then hypothesized that presence of vsnCTC signals the presence of VM and has predictive and prognostic value with respect to VM. Methods: We identified mCRPC patients who had progressed through next generation hormonal maneuvers such as abiraterone, enzalutamide, or an equivalent drug. Serial blood specimens were used for vsnCTC enumeration using NanoVelcro CTC Assay as previously published. The vsnCTC counts were related to the presence and development of VM (evaluated by radiography) as well as the response to anti-cancer treatment. Results: Blood specimens were identified from 28 patients who met the eligibility criteria; 15/28 patients presented with VM and 13/28 had bone-only disease at their first CTC enumeration. Five out of 13 non-VM patients developed VM during follow-up, and vsnCTCs were detected 86-196 days prior to radiographic detection of VM (true positive); 4/13 had vsnCTCs detected but no VM was found by the time of analysis (false positive). None of the vsnCTC(-) patients developed VM. vsnCTCs were detected in 20/20 VM patients compared to 4/8 non-VM patients. Reduction of vsnCTC count occurred at initiation of anti-cancer treatment; transition from vsnCTC(-) to vsnCTC(+) was seen prior to progression under the treatment. Of the patients who have VM, 14 passed away at the time of this abstract submission including all the patients converting from non-VM to VM during the time of follow-up. Two out of 8 non-VM patients passed away including one patient who had vsnCTCs detected around 6 months prior to death. Conclusions: vsnCTCs are associated with the presence of VM. The vsnCTC is a potential biomarker for predicting the development of VM and monitoring the treatment response in mCRPC. Transition from vsnCTC(-) to vsnCTC(+) was associated with the development of VM and progression under the treatment. Citation Format: Jie-Fu Chen, Hao Ho, Elisabeth Hodara, Alexander Ureno, Ann Go, Elizabeth Kaufman, Margarit Sievert, Daniel J. Luthringer, Jiaoti Huang, Leland Chung, Zunfu Ke, Ker-Chau Li, Hsian-Rong Tseng, Edwin M. Posadas. Very-small-nuclear circulating tumor cell (vsnCTC) as a putative biomarker for visceral metastasis in metastatic castration-resistant prostate cancer (mCRPC). [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 4962.

Modern graphical tools have enhanced our ability to learn many things from data directly. With much user-friendly graphical software available, we are encouraged to plot a lot more often than before. The benefits from direct interaction with graphics have been enormous. But trailing behind these high-tech advances is the issue of appropriate guidance on what to plot. There are too many directions to project a high-dimensional data set and unguided plotting can be time-consuming and fruitless. In a recent article, Li set up a statistical framework for study on this issue, based on a notion of effective dimension reduction (edr) directions. They are the directions to project a high dimensional input variable for the purpose of effectively viewing and studying its relationship with an output variable. A methodology, sliced inverse regression, was introduced and shown to be useful in finding edr directions. This article introduces another method for finding edr directions. It begins with the observat...

To many biomedical researchers, effective tumor classification methods such as the support vector machine often appear like a black box not only because the procedures are complex but also because the required specifications, such as the choice of a kernel function, suffer from a clear guidance either mathematically or biologically. As commonly observed, samples within the same tumor class tend to be more similar in gene expression than samples from different tumor classes. But can this well-received observation lead to a useful procedure of classification and prediction? To address this issue, we first conceived a statistical framework and derived general conditions to serve as the theoretical foundation that supported the aforementioned empirical observation. Then we constructed a classification procedure that fully utilized the information obtained by comparing the distributions of within-class correlations with between-class correlations via Kullback–Leibler divergence. We compared our approach with many machine-learning techniques by applying to 22 binary- and multiclass gene-expression datasets involving human cancers. The results showed that our method performed as efficiently as support vector machine and Naïve Bayesian and outperformed other learning methods (decision trees, linear discriminate analysis, and k -nearest neighbor). In addition, we conducted a simulation study and showed that our method would be more effective if the arriving new samples are subject to the often-encountered baseline shift or increased noise level problems. Our method can be extended for general classification problems when only the similarity scores between samples are available.

Purpose: Metastasis is the main cause of mortality in non–small cell lung cancer (NSCLC) patients. Genes that can discriminate the invasion ability of cancer cells may become useful candidates for clinical outcome prediction. We identify invasion-associated genes through computational and laboratorial approach that supported this idea in NSCLC. Experimental Design: We first conducted invasion assay to characterize the invasion abilities of NCI-60 lung cancer cell lines. We then systematically exploited NCI-60 microarray databases to identify invasion-associated genes that showed differential expression between the high and the low invasion cell line groups. Furthermore, using the microarray data of Duke lung cancer cohort (GSE 3141), invasion-associated genes with good survival prediction potentials were obtained. Finally, we validated the findings by conducting quantitative PCR assay on an in-house collected patient group (n = 69) and by using microarray data from two public western cohorts (n = 257 and 186). Results: The invasion-associated four-gene signature (ANKRD49, LPHN1, RABAC1, and EGLN2) had significant prediction in three validation cohorts (P = 0.0184, 0.002, and 0.017, log-rank test). Moreover, we showed that four-gene signature was an independent prognostic factor (hazard ratio, 2.354, 1.480, and 1.670; P = 0.028, 0.014, and 0.033), independent of other clinical covariates, such as age, gender, and stage. Conclusion: The invasion-associated four-gene signature derived from NCI-60 lung cancer cell lines had good survival prediction power for NSCLC patients. (Clin Cancer Res 2009;15(23):7309–15)

SummaryWe investigated whether microRNA expression profiles can predict clinical outcome of NSCLC patients. Using real-time RT-PCR, we obtained microRNA expressions in 112 NSCLC patients, which were divided into the training and testing sets. Using Cox regression and risk-score analysis, we identified a five-microRNA signature for the prediction of treatment outcome of NSCLC in the training set. This microRNA signature was validated by the testing set and an independent cohort. Patients with high-risk scores in their microRNA signatures had poor overall and disease-free survivals compared to the low-risk-score patients. This microRNA signature is an independent predictor of the cancer relapse and survival of NSCLC patients.

Background Circulating tumor cells (CTCs) are an emerging biomarker in prostate cancer (PC). Most efforts have focused on enumeration, we and others have identified subsets within this pool of cells which may be clinically informative. Using the highly sensitive NanoVelcro technology for CTC isolation and fluorescence microscopy, our group sub-classified CTCs by nuclear size and focused on the correlation with sites of metastatic disease in advanced PC patients. Methods Blood samples were obtained from PC patients across the spectrum of metastatic states, i.e. no metastasis, non-visceral (osseous/lymph node) metastasis, and visceral metastasis. The patients were divided into training and validation cohorts. CTCs were captured from the blood and enumerated on NanoVelcro Chips. These were then subjected to pathologic review for cellular morphology and nuclear size. The distribution of nuclear sizes was analyzed using a Gaussian Mixture Model to sub-classify the CTCs. Correlations were made between CTC subpopulations and metastatic status. Individual CTCs were isolated by laser-capture microdissection for subsequent characterization focusing on cellular markers and transcriptomic signatures in comparison between vsnCTC and other CTC subpopulations. Results Statistical analysis and modeling of nuclear size distribution revealed 3 distinct subpopulations in the training cohort: large-nuclear CTCs (lnCTC), small-nuclear CTCs (snCTC), and very-small-nuclear CTCs (vsnCTCs). While the lnCTC subpopulation alone failed to distinguish metastatic disease from non-metastatic disease, snCTC + vsnCTC counts could make that distinction. Furthermore, vsnCTC counts were elevated in PC patients with visceral metastases when compared to those without visceral metastases (0.5 ± 0.5 versus 3.1 ± 5.2 cells/mL of blood, p = 0.005). This difference remained statistically significant in the validation cohort (0.6 ± 1.0 versus 3.5 ± 4.0 cells/mL of blood, p = 0.007). Serial enumerations for individual patients showed the emergence of vsnCTCs prior to detection of newly developed visceral lesions. The recurrence of vsnCTC in patients under treatment is also associated with radiographic progression of existing visceral lesions through therapy. Molecular characterization of vsnCTCs reveals differential expression of genes related to neuroendocrine, stem cell, and androgen biology. Conclusions Subsets of CTCs contain information on PC disease status when analyzed with pathologic approaches, such as nuclear size measurement. In particular, the detection of vsnCTCs correlated with the presence of visceral metastatic lesions and should be explored as a potential biomarker to identify patients at risk for developing this more aggressive form of PC. Citation Format: Jie-Fu Chen, Hao Ho, Jake Lichterman, Yi-Tsung Lu, Yang Zhang, Mitch A. Garcia, Shang-Fu Chen, An-Jou Liang, Haiyen E. Zhau, Shuang Hou, Rafi S. Ahmed, Daniel J. Luthringer, Jiaoti Huang, Ker-Chau Li, Leland WK Chung, Zunfu Ke, Hsian-Rong Tseng, Edwin M. Posadas. Sub-classification of prostate cancer circulating tumor cells (CTCs) by nuclear size reveals very-small nuclear CTCs in patients with visceral metastases. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3473. doi:10.1158/1538-7445.AM2015-3473

11027 Background: In prostate cancer (PCa), morphologic classification remains a standard clinical practice in pathology. It has been shown that nuclear size and shape in tumor sections correlate w...

We investigated how different the patients of lung adenocarcinoma with and without EGFR activating mutation are in the genomes of their cancer cells. We conducted EGFR mutation test for 141 patients and used high-density even-coverage CGH arrays to chart the sites of DNA copy number alteration (CNA) in primary adenocarcinomas of each patient. We analyzed the site distribution of the most notable alterations from various perspectives and revealed several leading characteristics that converged on chromosome 7p. The clinical relevance of our findings was confirmed by conducting a survival study on an independent group of 114 patients, wherein a gene-signature that used the average copy numbers of 6 genes on 7p as measured by genomic real-time rt-PCR, showed statistical significance in predicting the overall survival and the relapse-free survival for the EGFR-mutation patients but not for the EGFR-wild-type patients. Our findings suggest that chromosome 7p contains very rich information for deciphering the pivotal genomic component contributing to the heterogeneity of lung adenocarcinoma. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3882. doi:10.1158/1538-7445.AM2011-3882