Correction: Combined poly-ADP ribose polymerase and ataxia-telangiectasia mutated/Rad3-related inhibition targets ataxia-telangiectasia mutated-deficient lung cancer cells.

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Combined poly-ADP ribose polymerase and ataxia-telangiectasia mutated/Rad3-related inhibition targets ataxia-telangiectasia mutated-deficient lung cancer cells.

BACKGROUND: Up to 40% of lung adenocarcinoma have been reported to lack ataxia-telangiectasia mutated (ATM) protein expression. We asked whether ATM-deficient lung cancer cell lines are sensitive to poly-ADP ribose polymerase (PARP) inhibitors and determined the mechanism of action of olaparib in ATM-deficient A549 cells. METHODS: We analysed drug sensitivity data for olaparib and talazoparib in lung adenocarcinoma cell lines from the Genomics of Drug Sensitivity in Cancer (GDSC) project. We deleted ATM from A549 lung adenocarcinoma cells using CRISPR/Cas9 and determined the effects of olaparib and the ATM/Rad3-related (ATR) inhibitor VE-821 on cell viability. RESULTS: IC50 values for both olaparib and talazoparib positively correlated with ATM mRNA levels and gene amplification status in lung adenocarcinoma cell lines. ATM mutation was associated with a significant decrease in the IC50 for olaparib while a similar trend was observed for talazoparib. A549 cells with deletion of ATM were sensitive to ionising radiation and olaparib. Olaparib induced phosphorylation of DNA damage markers and reversible G2 arrest in ATM-deficient cells, while the combination of olaparib and VE-821 induced cell death. CONCLUSIONS: Patients with tumours characterised by ATM-deficiency may benefit from treatment with a PARP inhibitor in combination with an ATR inhibitor.

Cav3.1 overexpression is associated with negative characteristics and prognosis in non-small cell lung cancer.

INTRODUCTION: Voltage-gated calcium channels (VGCC) have been found to be differentially expressed in several different tumor types, but their role in tumor growth, malignant invasion, metastases and impact on clinical outcomes has not been clarified. MATERIALS AND METHODS: From a cohort database of 193 patients with early-stage NSCLC, 163 formalin-fixed paraffin-embedded specimens were available for analysis to construct tissue microarrays. Cav3.1 protein expression was detected using fluorescence immunohistochemistry, and quantified using automated image acquisition and analysis. RESULTS: Among the cohort of 193 NSCLC patients, adenocarcinoma (53.9%) and squamous cell carcinoma (SCC) (30.1%) were the most common histologies. There was no difference between SCC and non-SCC subtypes in overall survival (OS) or relapse-free survival (RFS); 74.2 vs 90.1 months (p = 0.543) and 48.8 vs 52.6 months (p = 0.766), respectively. T-type VGCC 3.1 (Cav3.1) overexpression was assessed by tissue microarray immunohistochemistry analysis from 163 available patient samples. Eighteen (11.0%) NSCLC primaries were found to have Cav3.1 overexpression levels, and were significantly associated with SCC histology (p < 0.001), larger tumor size (p < 0.001) and later stage disease at diagnosis (p = 0.019). Median OS was 48.6 vs 106.7 months for Cav3.1 overexpressing and non-overexpressing patients, respectively (p = 0.032). Regression analysis revealed a significantly negative effect for Cav3.1 overexpression on RFS (Hazard ratio [HR] = 2.02, p = 0.048). CONCLUSIONS: Cav3.1 overexpression is a potential biomarker for poorer patient outcomes. These results bring supportive evidence for calcium channels inducing an aggressive phenotype in NSCLC and potentially may serve as a therapeutic target in overexpressing tumors.

Loss of tumour-specific ATM protein expression is an independent prognostic factor in early resected NSCLC.

Ataxia-telangiectasia mutated (ATM) is critical in maintaining genomic integrity. In response to DNA double-strand breaks, ATM phosphorylates downstream proteins involved in cell-cycle checkpoint arrest, DNA repair, and apoptosis. Here we investigate the frequency, and influence of ATM deficiency on outcome, in early-resected non-small cell lung cancer (NSCLC). Tissue microarrays, containing 165 formalin-fixed, paraffin-embedded resected NSCLC tumours from patients diagnosed at the Tom Baker Cancer Centre, Calgary, Canada, between 2003 and 2006, were analyzed for ATM expression using quantitative fluorescence immunohistochemistry. Both malignant cell-specific ATM expression and the ratio of ATM expression within malignant tumour cells compared to that in the surrounding tumour stroma, defined as the ATM expression index (ATM-EI), were measured and correlated with clinical outcome. ATM loss was identified in 21.8% of patients, and was unaffected by clinical pathological variables. Patients with low ATM-EI tumours had worse survival outcomes compared to those with high ATM-EI (p < 0.01). This effect was pronounced in stage II/III patients, even after adjusting for other clinical co-variates (p < 0.001). Additionally, we provide evidence that ATM-deficient patients may derive greater benefit from guideline-recommended adjuvant chemotherapy following surgical resection. Taken together, these results indicate that ATM loss seems to be an early event in NSCLC carcinogenesis and is an independent prognostic factor associated with worse survival in stage II/III patients.

microRNA 338-3p exhibits tumor suppressor role and its down-regulation is associated with adverse clinical outcome in prostate cancer patients.

MicroRNAs (miRNAs) are small non-coding RNAs that function in transcriptional and post-transcriptional regulation of gene expression. Several miRNAs have been implicated in regulating prostate cancer (PCa) progression. Deregulations of miRNA regulatory networks have been reported in ERG positive PCa, which accounts for ~50 % of PCa and have been suggested to affect tumor aggressiveness. The function of miR338-3p, its prognostic significance, and its association with ERG positive PCa has not been fully investigated. Using microarray expression profiling, we identified miRNA338-3p as among the top deregulated miRNAs associated with ERG status in PCa. We investigated miR338-3p function using in vitro and in vivo experimental models and its expression was assessed and validated in clinical samples and a public cohort of localized and metastatic prostate cancer. miR338-3p was significantly down-regulated with disease progression from benign prostate tissue to primary and metastatic lesions. In localized disease, patients with lower miR338-3p expression levels showed increased association to biochemical recurrence and several adverse pathological parameters compared to patients with higher miRNA338-3p tissue expression levels. Using in vitro PCa cell models, overexpression of miR338-3p resulted in a decrease in cell invasion and expression of chemokine signalling genes CXCL12, CXCR4, and CXCR7. In vivo, orthotropic implantation of PC3 cells stably expressing miR338-3p was associated with a significant decrease in tumor weights compared to control cells. miR338-3p has anti-proliferative and anti-invasive properties. It affects CXCR4 axis, and its down-regulation is associated with adverse clinical outcomes in PCa patients.

Interrogation of ERG gene rearrangements in prostate cancer identifies a prognostic 10-gene signature with relevant implication to patients' clinical outcome.

OBJECTIVES: ERG-gene rearrangement defines a distinct molecular subtype of PCA with potential biological and clinical implications. To identify a molecular signature reflective of the downstream effects of ERG-mediated transcriptional regulation with prognostic implication in patients with prostate cancer (PCA). MATERIAL AND METHODS: We used a singular value decomposition (SVD) bioinformatics approach to re-analyse gene expression data previously generated from 46 prostate tumours, and identified an ERG-like gene signature. The signature was validated on several patient cohorts and individual genes were correlated to ERG expression and PCA progression. RESULTS: An ERG-like 10-gene signature was identified and validated in PCA cohorts of the physician health study (p115) (n = 110) in addition to three independent public datasets, and was significantly associated with disease progression, biochemical recurrence and PCA-specific mortality. Patients with the ERG-like signature were significantly associated with disease recurrence on univariate (hazard ratio [HR] 2.6; 95% confidence interval [CI]:1.3-5.2; P = 0.004) and multivariate analysis (HR 2.3; 95% CI:1.1-4.6, P = 0.016) compared with patients without this signature. Within the group of patients with Gleason score (GS) 6 and 7 PCA, the signature added prognostic value beyond GS and identified patients at higher risk of cancer deaths more accurately than GS alone or in combination with ERG status. Protein expression of the 10 genes were significantly associated with ERG and disease progression regardless of ERG status. CONCLUSION: The characterized ERG-like signature was reflective of aggressive features of ERG-mediated transcription and was prognostically robust. The combination of this signature with clinicopathological variables should be validated prospectively to explore its clinical utility in stratifying patients with PCA and in identifying those at higher risk of metastatic and lethal disease.

Elevated physiological levels of folic acid can increase in vitro growth and invasiveness of prostate cancer cells.

OBJECTIVES: • To investigate the effects of different folic acid concentrations on the growth and invasiveness of prostate cancer cell lines. • To determine if observed changes are correlated with changes in levels of the potential prostate cancer biomarker, sarcosine, a byproduct of folate metabolism. MATERIALS AND METHODS: • The prostate cancer cell lines PC-3, LNCaP and DU145 were cultured in media containing 4, 20 or 100 nm of folic acid and assayed for growth over 9 days by counting viable cells at 3-day intervals, or for invasion by passage through a Matrigel-coated transwell membrane. • Cells grown in the different folic acid media were collected and subjected to metabolomic analysis by gas chromatography and mass spectrometry to measure levels of intracellular sarcosine. RESULTS: • The results show that higher levels of folic acid can increase cell growth in PC-3 and LNCaP prostate cancer cell lines, and may also increase the invasive capacity of PC-3, LNCaP and DU145 cells. • We did not observe a correlation between increased invasion from higher folic acid concentrations and levels of sarcosine, but there were significant changes in other metabolites in cells grown in higher levels of folic acid. CONCLUSION: • These findings suggest that folic acid has an important and potentially negative role in prostate cancer progression.

Characterization of an unusual tropoelastin with truncated C-terminus in the frog.

Tropoelastin is the monomeric form of elastin, a major polymeric protein of the extracellular elastic matrix of vertebrate tissues with properties of extensibility and elastic recoil. Mammalian and avian species contain a single gene for tropoelastin. A tropoelastin gene has also previously been identified in amphibians. In contrast, two tropoelastin genes with different tissue expression patterns have been described in teleosts. While general characteristics of tropoelastins, such as alternating arrangements of hydrophobic and crosslinking domains, are conserved across a wide phylogenetic range, sequences of these domains are highly variable, particularly when amphibian and teleost tropoelastins are included. For this reason exon-to-exon correspondence is not clear, and overall alignment of tropoelastin sequences across all species is not possible. An exception to this is the C-terminal exon, whose coding sequence has been very well-conserved across all species described to date. In mammalians this C-terminal domain has been shown to be important for interactions with cells and other matrix-associated proteins involved in matrix assembly. Here we identify and characterize a second tropoelastin gene in the frog with several unusual characteristics, the most striking of which is truncation of the C-terminal domain, deleting normally conserved sequence motifs. We demonstrate that, in spite of the absence of these motifs, both frog tropoelastin genes are expressed and incorporated into the elastic matrix, although with differential tissue localizations.