Gene Drug Sensitivity Research Articles

Construction of artificial neural network model for predicting the efficacy of first-line FOLFOX chemotherapy for metastatic colorectal cancer

Objective: To explore and establish an artificial neural network (ANN) model for predicting the efficacy of first-line FOLFOX chemotherapy for metastatic colorectal cancer. Methods: A set of FOLFOX chemotherapy data from a group of patients with metastatic colorectal cancer (mCRC) (GSE104645) was downloaded from the GEO database as a training set. According to the FOLFOX protocol, the efficacy was divided into two groups: the chemo-sensitive group (including complete response and partial response) and the chemo-resistant group (including stable disease and progressive disease), including 31 cases in the sensitive group and 23 in the resistant group. Then, chip data (accessible number: GSE69657) from Fujian Medical University Union Hospital were chosen as a test set. A total of 30 patients were enrolled in the study, including 13 in the sensitive group and 17 in the resistant group. The batch effect correction was performed on the expression values of the two sets of matrices using the R 3.5.1 software Combat package. The gene expression difference of sensitive and resistant group in GSE104645 was analyzed by the GEO2R platform. P<0.05 and the absolute value of log(2)FC>0.33 (FC abbreviation of fold change) were used as the threshold value to screen the drug resistance and sensitive genes of the FOLFOX regimen. An ANN was constructed using the multi-layer perceptron (MLP) to perform the FOLFOX regimen on the GSE104645 dataset. The GSE69657 expression matrix and clinical efficacy parameters were then used for retrospective verification. Receiver operating characteristic(ROC) curves were used to evaluate the test results and predictive power. Results: A total of 2, 076 differentially expressed genes in GSE104645 were selected, of which 822 genes were up-regulated and 1, 254 genes were down-regulated in the chemo-resistance group. The down-regulated genes were sensitive genes. GO analysis of the biological processes in which the differentially expressed genes were involved, revealed that they were mainly involved in the regulation of substance metabolism. A total of 39 genes were included in the final model construction. This was a neural network model with two hidden layers. The accuracy of predicting training samples and test samples was 75.7% and 76.5%, respectively, and the area under the ROC curve was 0.875. The chip data set of our department (GSE69657) was set as the test set, and the area under the ROC curve was 0.778. Conclusions: In this study, an artificial neural network model is successfully constructed to predict the efficacy of first-line FOLFOX regimen for metastatic colorectal cancer based on the microarray, and an independent external verification is also conducted. The model has good stability and well prediction efficiency. Besides, the results of this study suggest that the gene functions related to oxaliplatin resistance are mainly enriched in the regulation process of substance metabolism.

Markov Neighborhood Regression for High-Dimensional Inference

This article proposes an innovative method for constructing confidence intervals and assessing p-values in statistical inference for high-dimensional linear models. The proposed method has successfully broken the high-dimensional inference problem into a series of low-dimensional inference problems: For each regression coefficient βi , the confidence interval and p-value are computed by regressing on a subset of variables selected according to the conditional independence relations between the corresponding variable Xi and other variables. Since the subset of variables forms a Markov neighborhood of Xi in the Markov network formed by all the variables , the proposed method is coined as Markov neighborhood regression (MNR). The proposed method is tested on high-dimensional linear, logistic, and Cox regression. The numerical results indicate that the proposed method significantly outperforms the existing ones. Based on the MNR, a method of learning causal structures for high-dimensional linear models is proposed and applied to identification of drug sensitive genes and cancer driver genes. The idea of using conditional independence relations for dimension reduction is general and potentially can be extended to other high-dimensional or big data problems as well. Supplementary materials for this article are available online.

Analysis of antibiotic sensitivity and resistance genes of Bordetella pertussis in Chinese children.

To understood the pathogen detection status and clinical characteristics of suspected pertussis in children and to observe the drug sensitivity and drug resistance genes of Bordetella pertussis (B. pertussis). Three hundred fifty-one cases were collected and their nasopharyngeal swab samples were analyzed by culture and fluorescent quantitative polymerase chain reaction. The susceptibility to erythromycin, clindamycin, ampicillin, levofloxacin, and sulfamethoxazole-trimethoprim were tested by E-test for the positive strains, and the susceptibility to erythromycin was also tested for the KB disk diffusion method. The 23S rRNA gene of the positive strains was amplified and sequenced, and statistical analysis was performed in conjunction with clinical data. The positive rate of bacterial culture was 16.8% (59/351), and the positive rate of PCR was 62.4% (219/351). Two cases were positive about bacterial culture and negative for PCR. There were 221 confirmed cases of pertussis. The E-test results showed that the rate of the sensitivity of the 55 strains of pertussis to erythromycin and clindamycin was 50.9% (28/55), the minimum antibiotic concentration50 (MIC50) and MIC90 values were 0.094/>256 and 0.75/>256 mg/L, respectively, and the MIC50/MIC90 to ampicillin, levofloxacin, and sulfamethoxazole were 0.125/0.19, 0.38/0.5, and 0.125/0.25 mg/L, respectively. The KB disk diffusion method showed 27 of the 55 strains 49.1% (27/55) was resistant to erythromycin; all of the resistant strains had the 23S rRNA gene A2047G mutation, and their MIC of erythromycin was >256 mg/L. The diagnosis of pertussis by a fluorescent quantitative polymerase chain reaction method is more sensitive than that of bacterial culture. The resistance of B. pertussis to erythromycin was prominent. All of the strains of B. pertussis resistant to erythromycin in our center had the A2047G mutation of the 23S rRNA gene.

Clinical application of drug sensitive gene detection in postoperative instillation for non-muscle invasive bladder cancer

BackgroundBladder cancer is the most common malignant tumor of the urinary system. One of the biological characteristics of NMIBC is the high recurrence rate after surgery. The implementation of this project aimed to investigate the role of pharmacogenomic testing-guided intravesical perfusion of chemotherapeutic agents in the postoperative perfusion therapy for non-muscle invasive bladder cancer.MethodFrom January 2015 to December 2016, 298 patients with non-muscle-invasive bladder cancer were enrolled in this prospective study. These patients received chemotherapy drugs after electrotherapy. According to the presence or absence of tumor susceptibility gene detection after surgery, they were divided into two groups, including the drug sensitive group(N = 44) and the control group(N = 254). The drug sensitive group received bladder infusion therapy with sensitive chemotherapy drugs based on drug sensitivity gene detection results. The control group received intravesical instillation of pirarubicin. The preoperative general data and tumor grade of patients were recorded. Cystoscopy was performed before and every 3 months after surgery. The chest CT, upper abdomen CT, renal function, and urinary routine tests were performed. Tumor recurrence, metastasis and tumor-related death were recorded and evaluated during follow-up.ResultsThe drug sensitive group, which selected high-sensitivity drugs for intravesical instillation therapy based on gene expression, has a significantly lower relapse rate (11.36% vs 37.40%, P < 0.05) and a significantly longer time to relapse (17.80 ± 7.20 month vs11.20 ± 6.10 month, P < 0.05) compared with the control group. There were no significant differences in the time of mortality and death time between two groups.ConclusionThe pharmacogenomic testing-directed bladder instillation of chemotherapeutic drugs may be more effective than empiric drug administration in reducing the recurrence rate of non-muscle-invasive bladder cancer.

Patient-Derived Organoids from Multiple Colorectal Cancer Liver Metastases Reveal Moderate Intra-patient Pharmacotranscriptomic Heterogeneity.

Molecular tumor heterogeneity may have important implications for the efficacy of targeted therapies in metastatic cancers. Inter-metastatic heterogeneity of sensitivity to anticancer agents has not been well explored in colorectal cancer. We established a platform for ex vivo pharmacogenomic profiling of patient-derived organoids (PDO) from resected colorectal cancer liver metastases. Drug sensitivity testing (n = 40 clinically relevant agents) and gene expression profiling were performed on 39 metastases from 22 patients. Three drug-response clusters were identified among the colorectal cancer metastases, based primarily on sensitivities to EGFR and/or MDM2 inhibition, and corresponding with RAS mutations and TP53 activity. Potentially effective therapies, including off-label use of drugs approved for other cancer types, could be nominated for eighteen patients (82%). Antimetabolites and targeted agents lacking a decisive genomic marker had stronger differential activity than most approved chemotherapies. We found limited intra-patient drug sensitivity heterogeneity between PDOs from multiple (2-5) liver metastases from each of ten patients. This was recapitulated at the gene expression level, with a highly proportional degree of transcriptomic and pharmacological variation. One PDO with a multi-drug resistance profile, including resistance to EGFR inhibition in a RAS-mutant background, showed sensitivity to MEK plus mTOR/AKT inhibition, corresponding with low-level PTEN expression. Intra-patient inter-metastatic pharmacological heterogeneity was not pronounced and ex vivo drug screening may identify novel treatment options for metastatic colorectal cancer. Variation in drug sensitivities was reflected at the transcriptomic level, suggesting potential to develop gene expression-based predictive signatures to guide experimental therapies.

Cytological and Gene Profile Expression Analysis Reveals Modification in Metabolic Pathways and Catalytic Activities Induce Resistance in Botrytis cinerea Against Iprodione Isolated From Tomato

Grey mold is one of the most serious and catastrophic diseases, causing significant yield losses in fruits and vegetables worldwide. Iprodione is a broad spectrum agrochemical used as a foliar application as well as a seed protectant against many fungal and nematode diseases of fruits and vegetables from the last thirty years. The extensive use of agrochemicals produces resistance in plant pathogens and is the most devastating issue in food and agriculture. However, the molecular mechanism (whole transcriptomic analysis) of a resistant mutant of B. cinerea against iprodione is still unknown. In the present study, mycelial growth, sporulation, virulence, osmotic potential, cell membrane permeability, enzymatic activity, and whole transcriptomic analysis of UV (ultraviolet) mutagenic mutant and its wild type were performed to compare the fitness. The EC50 (half maximal effective concentration that inhibits the growth of mycelium) value of iprodione for 112 isolates of B. cinerea ranged from 0.07 to 0.87 µg/mL with an average (0.47 µg/mL) collected from tomato field of Guangxi Province China. Results also revealed that, among iprodione sensitive strains, only B67 strain induced two mutants, M0 and M1 after UV application. The EC50 of these induced mutants were 1025.74 μg/mL and 674.48 μg/mL, respectively, as compared to its wild type 1.12 μg/mL. Furthermore, mutant M0 showed higher mycelial growth sclerotia formation, virulence, and enzymatic activity than wild type W0 and M1 on potato dextrose agar (PDA) medium. The bctubA gene in the mutant M0 replaced TTC and GAT codon at position 593 and 599 by TTA and GAA, resulting in replacement of phenyl alanine into leucine (transversion C/A) and aspartic acid into glutamic acid (transversion T/C) respectively. In contrast, in bctubB gene, GAT codon at position 646 is replaced by AAT and aspartic acid converted into asparagine (transition G/A). RNA sequencing of the mutant and its wild type was performed without (M0, W0) and with iprodione treatment (M-ipro, W-ipro). The differential gene expression (DEG) identified 720 unigenes in mutant M-ipro than W-ipro after iprodione treatment (FDR ≤ 0.05 and log2FC ≥ 1). Seven DEGs were randomly selected for quantitative real time polymerase chain reaction to validate the RNA sequencing genes expression (log fold 2 value). The gene ontology (GO) enrichment and Kyoto encyclopedia genes and genomes (KEGG) pathway functional analyses indicated that DEG’s mainly associated with lysophopholipase, carbohydrate metabolism, amino acid metabolism, catalytic activity, multifunctional genes (MFO), glutathione-S transferase (GST), drug sensitivity, and cytochrome P450 related genes are upregulated in mutant type (M0, M-ipro) as compared to its wild type (W0, W-ipro), may be related to induce resistant in mutants of B. cinerea against iprodione.

Diagnostic accuracy of line probe assays for drug-resistant tuberculosis: a Meta-analysis

Objective: To evaluate the diagnostic accuracy of line probe assays for drug- resistant tuberculosis (TB) in China. Methods: Chinese databases (CNKI, Wanfang, SinoMed, VIP Information) and English databases (PubMed, Embase, Cochrane Library) were used to retrieve the literatures regarding the accuracy of line probe assays in the diagnosis of drug-resistant tuberculosis in China between January 1, 2000 and September 1, 2017. Quality Assessment of Diagnostic Accuracy Studies-2 was used to evaluate the quality of the included studies. Sensitivity and specificity in different studies (using drug sensitivity test or gene sequencing as gold standard) were combined by Meta-analysis using bivariate or univariate model. In addition, subgroup analysis (GenoType MTBDRplus, GenoType MTBDRsl and Reverse dot blot hybridization) and sensitivity analysis were also carried out. Results: A total of 24 literatures involving 82 studies were included in the final analysis. The sensitivity and specificity of line probe assays for rifampicin resistant TB were 0.91(0.88-0.94) and 0.98 (0.97-0.99), respectively. The sensitivity and specificity of line probe assays for isoniazid resistant TB were 0.80 (0.77-0.83) and 0.98 (0.96-0.99), respectively. The sensitivity and specificity of line probe assays for multidrug-resistant TB were 0.81 (0.76-0.85) and 0.99 (0.99-1.00), respectively. The sensitivity and specificity of line probe assays for quinolone resistant TB were 0.92(0.88-0.95) and 0.94 (0.91-0.97), respectively. The sensitivity and specificity of line probe assays for second-line injectable drug resistant TB (including kanamycin, Capreomycin, amikacin) were 0.79(0.58-0.91) and 0.98 (0.90-1.00), respectively. The sensitivity and specificity of line probe assays for extensively drug-resistant TB were 0.46 (0.19-0.75) and 1.00 (0.98-1.00), respectively. Subgroup analysis showed that the overall diagnostic accuracy of GenoType MTBDRplus and GenoType MTBDRsl was higher than that of Reverse dot blot hybridization. According to the results of sensitivity analysis, the results of this study were robust. Conclusion: The diagnostic accuracy of line probe assays for drug-resistant TB is high.

Bayesian Neural Networks for Selection of Drug Sensitive Genes

ABSTRACTRecent advances in high-throughput biotechnologies have provided an unprecedented opportunity for biomarker discovery, which, from a statistical point of view, can be cast as a variable selection problem. This problem is challenging due to the high-dimensional and nonlinear nature of omics data and, in general, it suffers three difficulties: (i) an unknown functional form of the nonlinear system, (ii) variable selection consistency, and (iii) high-demanding computation. To circumvent the first difficulty, we employ a feed-forward neural network to approximate the unknown nonlinear function motivated by its universal approximation ability. To circumvent the second difficulty, we conduct structure selection for the neural network, which induces variable selection, by choosing appropriate prior distributions that lead to the consistency of variable selection. To circumvent the third difficulty, we implement the population stochastic approximation Monte Carlo algorithm, a parallel adaptive Markov Chain Monte Carlo algorithm, on the OpenMP platform that provides a linear speedup for the simulation with the number of cores of the computer. The numerical results indicate that the proposed method can work very well for identification of relevant variables for high-dimensional nonlinear systems. The proposed method is successfully applied to identification of the genes that are associated with anticancer drug sensitivities based on the data collected in the cancer cell line encyclopedia study. Supplementary materials for this article are available online.

Global view of a drug-sensitivity gene network.

An important challenge in drug development is to gain insight into the mechanism of drug sensitivity. Looking for insights into the global relationships between drugs and their sensitivity genes would be expected to reveal mechanism of drug sensitivity. Here we constructed a drug-sensitivity gene network (DSGN) based on the relationships between drugs and their sensitivity genes, using drug screened genomic data from the NCI-60 cell line panel, including 181 drugs and 1057 sensitivity genes, and 1646 associations between them. Through network analysis, we found that two drugs that share the same sensitivity genes tend to share the same Anatomical Therapeutic Chemical classification and side effects. We then found that the sensitivity genes of same drugs tend to cluster together in the human interactome and participate in the same biological function modules (pathways). Finally, we noticed that the sensitivity genes and target genes of the same drug have a significant dense distance in the human interactome network and they were functionally related. For example, target genes such as epidermal growth factor receptor gene can activate downstream sensitivity genes of the same drug in the PI3K/Akt pathway. Thus, the DSGN would provide great insights into the mechanism of drug sensitivity.

DBGC: A Database of Human Gastric Cancer.

The Database of Human Gastric Cancer (DBGC) is a comprehensive database that integrates various human gastric cancer-related data resources. Human gastric cancer-related transcriptomics projects, proteomics projects, mutations, biomarkers and drug-sensitive genes from different sources were collected and unified in this database. Moreover, epidemiological statistics of gastric cancer patients in China and clinicopathological information annotated with gastric cancer cases were also integrated into the DBGC. We believe that this database will greatly facilitate research regarding human gastric cancer in many fields. DBGC is freely available at http://bminfor.tongji.edu.cn/dbgc/index.do

Abstract 4855: DBGC: database of gastric cancer

Abstract DBGC is a comprehensive database, integrating various gastric cancer related data resources. In this database, gastric cancer related transcriptomics projects, proteomics projects, mutations, biomarkers and drug sensitive genes from different sources were collected and unified. Besides, follow-up information and clinicopathological information of gastric cancer patients in Shanghai, China, were also integrated through ID-mapping. We believe that this database will greatly facilitate the research work of gastric cancer researchers. Database URL: http://bminfor.tongji.edu.cn/dbgc/index.do Note: This abstract was not presented at the meeting. Citation Format: Yingyan Yu. DBGC: database of gastric cancer. [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 4855. doi:10.1158/1538-7445.AM2015-4855

The significance of copy number variation in multiple osteosarcoma's malignance grade, drug resistance and classification

Objective To study the clinical significance of copy number variation (CNV) and driver gene in multiple osteosarcoma (MOS) for the diagnosis, classification, clinical features and difference of chemo-drug sensitivity, and to discuss the applications of exons group sequencing technology in tumor research. Methods 2 cases of multiple osteosarcoma patients diagnosed from January 2014 to October 2014 were collected. The exome of each patient's normal tissue and multiple lesions was sequenced in accordance with the unified standards. After filtering the sequencing data, the sequence of mutation gene and its type was confirmed. Highlight the single nucleotide mutation/SNV and CNV. Analyze the similarities and differences of genetic variations between multiple lesions of each patient, and the gene variants that are shared by the all four tumor lesions. Finally, analyze the molecular targeted drug sensitive gene about the similarities and differences between different lesions. Results The sequencing results showed that all specimens of each patient had vast majority of the same part, which confirmed they were obtained from a single patient at the genetic level firstly. Then, through the SNV analysis of both tumor tissues, the final data showed that there was highly overlap between them. We found PIK3 and HNRNPA2B1, both drive gene variants played an important role in the process of tumor when analyzing one patient's SNV. The anti-oncogene TP53 was found with CNV between multiple lesions of the 2 cases, while the CNV mutation got the same variation trend but different range. The CNV mutation of the molecular targeted drug sensitive gene was found to have significant differences between the tumor sites of both patients. By analyzing the patients' somatic SNV and CNV globally, the results indicated that the enrolled two patients were all metastasis MOS that with only one primary lesion. Conclusion CNV is one of the significant features of genetic abnormality of MOS. Key words: Osteosarcoma; Exome; DNA copy number variations

A New Drug Combinatory Effect Prediction Algorithm on the Cancer Cell Based on Gene Expression and Dose-Response Curve.

Gene expression data before and after treatment with an individual drug and the IC20 of dose–response data were utilized to predict two drugs' interaction effects on a diffuse large B-cell lymphoma (DLBCL) cancer cell. A novel drug interaction scoring algorithm was developed to account for either synergistic or antagonistic effects between drug combinations. Different core gene selection schemes were investigated, which included the whole gene set, the drug-sensitive gene set, the drug-sensitive minus drug-resistant gene set, and the known drug target gene set. The prediction scores were compared with the observed drug interaction data at 6, 12, and 24 hours with a probability concordance (PC) index. The test result shows the concordance between observed and predicted drug interaction ranking reaches a PC index of 0.605. The scoring reliability and efficiency was further confirmed in five drug interaction studies published in the GEO database.

Restoring expression of miR-16: a novel approach to therapy for malignant pleural mesothelioma

BackgroundMalignant pleural mesothelioma (MPM) is recalcitrant to treatment and new approaches to therapy are needed. Reduced expression of miR-15/16 in a range of cancer types has suggested a tumour suppressor function for these microRNAs, and re-expression has been shown to inhibit tumour cell proliferation. The miR-15/16 status in MPM is largely unknown. Materials and methodsMicroRNA expression was analysed by TaqMan-based RT-qPCR in MPM tumour specimens and cell lines. MicroRNA expression was restored in vitro using microRNA mimics, and effects on proliferation, drug sensitivity and target gene expression were assessed. Xenograft-bearing mice were treated with miR-16 mimic packaged in minicells targeted with epidermal growth factor receptor (EGFR)-specific antibodies. ResultsExpression of the miR-15 family was consistently downregulated in MPM tumour specimens and cell lines. A decrease of 4- to 22-fold was found when tumour specimens were compared with normal pleura. When MPM cell lines were compared with the normal mesothelial cell line MeT-5A, the downregulation of miR-15/16 was 2- to 10-fold. Using synthetic mimics to restore miR-15/16 expression led to growth inhibition in MPM cell lines but not in MeT-5A cells. Growth inhibition caused by miR-16 correlated with downregulation of target genes including Bcl-2 and CCND1, and miR-16 re-expression sensitised MPM cells to pemetrexed and gemcitabine. In xenograft-bearing nude mice, intravenous administration of miR-16 mimics packaged in minicells led to consistent and dose-dependent inhibition of MPM tumour growth. ConclusionsThe miR-15/16 family is downregulated and has tumour suppressor function in MPM. Restoring miR-16 expression represents a novel therapeutic approach for MPM.

In vitroantimalarial susceptibility and molecular markers of drug resistance in Franceville, Gabon

BackgroundMalaria remains a major public health problem, due largely to emergence and widespread P. falciparum drug resistance. WHO recommends artemisinine combination based therapy (ACT) to overcome P. falciparum drug resistance, but reports of declining ACT efficacy have been published. A thorough understanding of the molecular bases of P. falciparum resistance to existing drugs is therefore needed. The aims of this study were to analyze the in vitro sensitivity of P. falciparum field isolates from Franceville, Gabon, to chloroquine (CQ), mefloquine (MF), dihydroartemisinine (DHA) and monodesethylamodiaquine (MDAQ), and to investigate polymorphisms associated with drug resistance.MethodsWe conducted a cross-sectional study of 53 field isolates. Field isolates sensitivity to CQ, MF, DHA and MDAQ was assessed using the colorimetric DELI test. The Pfmdr1 codons 86 and 1246, Pfcrt (haplotype codon 72 to 76) and the PfATPAse6 codons 110 and 2694 were analysed by PCR-RFLP. Associations between drug sensitivity and parasite gene polymorphisms were evaluated with the Chi square test, and routine hematological parameters were analyzed with Fisher’s exact test implemented with Epinfo software. In all statistical tests, significance was assumed at p<0.05.ResultsA total of 46 P. falciparum isolates were successfully cultured in vitro and their sensitivity was tested. The proportions of isolates resistant to CQ, MF and MDAQ were 43.5%, 23.4% and 56.5%, respectively. Some isolates (23.9%) had DHA IC50 values higher than 10 nM. The median IC50 values were 71.67 (interquartile range (IQR, 1–438.2), 6.59 (IQR, 0.08-96), 64.79 (IQR, 0.09-448) and 6.45 nM (IQR, 0.09-23) for CQ, MF, MDAQ and DHA, respectively. The strongest correlation between diminished DHA sensitivity and MF resistance was observed (r2=0.73), followed by correlation between diminished DHA sensitivity and CQ resistance. Cross-resistance between CQ and MF was also observed. The prevalence of the 86Y and 1246Y mutations in Pfmdr1, 76T in Pfcrt, and 110A and 2694T in PfATPase6 was respectively 42% and 17.1%, 97.8%, and 0% and 22.2%.ConclusionThese high levels of antimalarial drug resistance in Franceville, Gabon, call for reinforced surveillance of drug efficacy.

Establishment and characterization of six human lung cancer cell lines: EGFR, p53 gene mutations and expressions of drug sensitivity genes

Six human lung cancer cell lines (SNU-371, SNU-963, SNU-1327, SNU-1330, SNU-2292 and SNU-2315) were newly established through primary cell cultures. These cell lines were derived from a pulmonary blastoma, a small cell lung cancer, three adenocarcinomas and a squamous cell carcinoma of the lung of six Korean lung cancer patients. The histopathology of the primary tumors and their in vitro growth characteristics were described. DNA fingerprinting analysis and genetic alterations in the p53, β-catenin, TGFβRII, K-ras and EGFR genes were conducted. mRNA expressions levels of E-cadherin, COX-2, MDR1, MXR, CGA, synatophysin and TTF1 genes were investigated and sensitivity to anticancer drugs was screened. Five cell lines grew as adherent cells and one cell line grew as floating aggregates. All lines were free of mycoplasma or bacteria and were proven unique by DNA fingerprinting analysis. A significant polymorphism at codon 72 (Arg to Pro) of the p53 gene was found in one line (SNU-1327) and a mutation at codon 176 was found in SNU-2292. No mutations in the K-ras, β-catenin and TGF-βRII genes were observed. E-cadherin was not expressed in SNU-371 and COX-2 was overexpressed in SNU-1330, SNU-2292 and SNU-2315 cell lines. MDR1 was overexpressed in SNU-371 and SNU-2292 cell lines and MXR was overexpressed in SNU-1327 cell line. Interestingly, the SNU-371 cell line derived from a pulmonary blastoma and which overexpressed MDR1 displayed cross resistance for several anticancer drugs. Neuroendocrine markers, chromogranin A and synaptophysin, were overexpressed in the small cell lung cancer cell line, SNU-963 and thyroid transcription factor-1 was also over expressed in this cell line. Two mutations (p.Glu746_Ser752delinsVal and p.Glu746_Ala750del) in exon 19 of EGFR were found in SNU-1330 and SNU-2315 cell lines, respectively. These well-characterized lung cancer cell lines may be useful tools for investigations of the biological characteristics of lung cancers, particularly for investigations related to mutations of EGFR.

Effects of Bone Marrow Stromal Cells and Umbilical Cord Blood-Derived Stromal Cells on Daunorubicin-Resistant Residual Jurkat Cells

Objective To observe the effects of the hematopoietic inductive microenvironment (HIM) simulated by stromal cells of different origins on daunorubicin-resistant residual Jurkat cells (Jurkat/DNR cells). Methods Jurkat/DNR cells were cultured and identified. Human umbilical cord blood-derived stromal cells (UCBDSCs) and normal human bone marrow stromal cells (BMSCs) were isolated and cocultured with Jurkat/DNR cells. Jurkat/DNR cells were collected after 14 days of coculture and analyzed with regard to cell proliferation and differentiation abilities, apoptosis, drug sensitivity, and MRD1 multidrug resistance gene mRNA expression. Results UCBDSC-simulated HIM suppressed proliferation and promoted apoptosis, differentiation, and drug sensitivity of Jurkat/DNR cells more significantly than BMSC-simulated HIM. Conclusions Both BMSCs and UCBDSCs reconstruct the leukemic HIM and reverse drug resistance in Jurkat/DNR cells. UCBDSCs reconstruct the leukemic HIM and reverse drug resistance more significantly than BMSCs.

Abstract 3606: Gene and miRNA expression profiles in lung cancer cells cooperated with drug sensitivity to PKC inhibitor enzastaurin

Abstract PKC-beta inhibitor, Enzastaurin, is being evaluated in a Phase III trial as a maintenance therapy for the treatment of diffuse large B-cell lymphoma (DLBCL), as well as being evaluated in several Phase II studies across a variety of more common tumor types including: breast, colon, lung, ovarian and prostate cancers. Sequence-dependent, synergistic antiproliferative and proapoptotic effects of the combination of cytotoxic drugs and enzastaurin in non small cell lung cancer cells (NSCLC). To ascertain the potential for PKC-beta inhibitor-based treatment in NSCLC, we analyzed the antitumor effects of enzastaurin in 22 NSCLC cell lines via MTT assay. To identify a molecular model of sensitivity to PKC-beta inhibitor treatment in NSCLC, we conducted a gene and miRNA expression profiling study using cDNA arrays and Taqman MicroRNA Arrays on the same set of cell lines and related the cytotoxic activity of enzastaurin to corresponding gene and miRNA expression pattern. In addition, pathway analysis was done with Pathway Architect software. We used eight genes, which were identified by gene-drug sensitivity correlation and pathway analysis, to build a support vector machine algorithm model by which sensitive cell lines were distinguished from resistant cell lines. We also identified unique miRNA expression patterns correlated with enzastaurin sensitivity. Our results suggested that PKCβ inhibitor may be promising anticancer drugs to NSCLC and the eight-gene classifier is useful in predicting drug sensitivity to PKC-beta inhibitors and may contribute to achieving individualized therapy for NSCLC patients. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3606.

Establishment and characterization of 13 human colorectal carcinoma cell lines: mutations of genes and expressions of drug-sensitivity genes and cancer stem cell markers

Thirteen human colorectal cancer (CRC) cell lines were established from 10 primary tumors and 3 metastatic tumors obtained from 13 Korean patients. Characteristics of the cell lines including morphology in vivo and in vitro; mutations of the K-ras, p53, APC and MMR genes and microsatellite instability (MSI) status in vitro were determined. Expression of drug-sensitivity genes including MDR1, MXR, MRP1 and COX2 was also analyzed. The cell lines were unique as judged by DNA fingerprinting using 16 short tandem repeats. Eleven of the cell lines grew as adherent populations and the remaining two as floating aggregates. None of the cell lines were contaminated with Mycoplasma or bacteria. All cell lines showed high viability with relatively long doubling times. Six cell lines contained mutations at K-ras. Seven cell lines displayed p53 gene missense, nonsense and frameshift mutations. MSI was found in three cell lines and two cell lines with an MSI-high phenotype-possessed hMLH1 mutations. Nine cell lines had an APC mutation. MRP1 was highly expressed in all cell lines, and high expression of MDR1, MXR and COX2 evident in eight, six and six cell lines, respectively. Embryonal stem cell markers (MELK, SOX4 and OCT4) were expressed in most of cell lines. The cancer stem cell biomarkers CD133, CD44 and Lgr5 were expressed in 12, 13 and 13 cell lines, respectively. The presently well-characterized CRC cell lines should be useful in investigations of the biological characteristics of CRC, particularly for investigations related to gene alterations associated with CRC and biology of cancer stem cells.

Screening the Drug Sensitivity Genes Related to GEM and CDDP in the Lung Cancer Cell-lines.

Background and objective Screening of small-cell lung cancer (SCLC) and non-small cell lung cancer (NSCLC) cell lines with gemcitabine hydrochloride (GEM) and cisplatin (CDDP) related to drug sensitivity gene might clarify the action mechanism of anti-cancer drugs and provide a new clue for overcoming drug resistance and the development of new anti-cancer drugs, and also provide theoretical basis for the clinical treatment of individual. The drug sensitivity of CDDP and GEM in 4 SCLC cell lines and 6 NSCLC cell lines was determined using MTT colorimetric assay, while the cDNA macroarray was applied to detect the gene expression state related to drug sensitivity of 10 lung cancer cell line in 1 291, and the correlation between the two was analysized. There were 6 genes showing significant positive correlation (r>=0.632, P<0.05) with GEM sensitivity; 45 genes positively related to CDDP; another 41 genes related to both GEM and CDDP (r>=+/- 0.4). Lung cancer with GEM and CDDP sensitivity of two types of drugs significantly related genes were Metallothinein (Signal transduction molecules), Cathepsin B (Organization protease B) and TIMP1 (Growth factor); the GEM, CDDP sensitivity associated genes of lung cancer cell lines mainly distributed in Metallothinein, Cathepsin B, growth factor TIMP1 categories. There existed drug-related sensitive genes of GEM, CDDP in SCLC and NSCLC cell lines; of these genes, Metallothinein, Cathepsin B and TIMP1 genes presented a significant positive correlation with GEM drug sensitivity, a significant negative correlation with CDDP drug sensitivity.