Sample type bias in the analysis of cancer genomes

Abstract There is widespread agreement that cancer gene discovery requires high-quality tumor samples

NOTCH3 Is a Prognostic Factor That Promotes Glioma Cell Proliferation, Migration and Invasion via Activation of CCND1 and EGFR

<div><p>Using a GWA analysis of a comprehensive glioma specimen population, we identified whole gain of chromosome 19 as one of the major chromosomal aberrations that correlates to patients’ outcomes. Our analysis of significant loci revealed for the first time NOTCH3 as one of the most significant amplification. NOTCH3 amplification is associated with worse outcome compared to tumors with non-amplified locus. NOTCH receptors (NOTCH1-4) are key positive regulators of cell-cell interactions, angiogenesis, cell adhesion and stem cell niche development which have been shown to play critical roles in several human cancers. Our objective is to determine the molecular roles of NOTCH3 in glioma pathogenesis and aggressiveness. Here we show for the first time that NOTCH3 plays a major role in glioma cell proliferation, cell migration, invasion and apoptosis. Therefore, our study uncovers the prognostic value and the oncogenic function of NOTCH3 in gliomagenesis and supports NOTCH3 as a promising target of therapy in high grade glioma. Our studies allowed the identification of a subset of population that may benefit from GSI- or anti-NOTCH3- based therapies. This may lead to the design of novel strategies to improve therapeutic outcome of patients with glioma by establishing medical and scientific basis for personalized chemotherapies.</p></div

Tumor Derived Mutations of Protein Tyrosine Phosphatase Receptor Type K Affect Its Function and Alter Sensitivity to Chemotherapeutics in Glioma

<div><p>Poor prognosis and resistance to therapy in malignant gliomas is mainly due to the highly dispersive nature of glioma cells. This dispersive characteristic results from genetic alterations in key regulators of cell migration and diffusion. A better understanding of these regulatory signals holds promise to improve overall survival and response to therapy. Using mapping arrays to screen for genomic alterations in gliomas, we recently identified alterations of the protein tyrosine phosphatase receptor type kappa gene (PTPRK) that correlate to patient outcomes. These PTPRK alterations are very relevant to glioma biology as PTPRK can directly sense cell–cell contact and is a dephosphorylation regulator of tyrosine phosphorylation signaling, which is a major driving force behind tumor development and progression. Subsequent sequencing of the full length PTPRK transcripts revealed novel PTPRK gene deletion and missense mutations in numerous glioma biopsies. PTPRK mutations were cloned and expressed in PTPRK-null malignant glioma cells. The effect of these mutations on PTPRK anti-oncogenic function and their association with response to anti-glioma therapeutics, such as temozolomide and tyrosine kinase inhibitors, was subsequently analyzed using <i>in vitro</i> cell-based assays. These genetic variations altered PTPRK activity and its post-translational processing. Reconstitution of wild-type PTPRK in malignant glioma cell lines suppressed cell growth and migration by inhibiting EGFR and β-catenin signaling and improved the effect of conventional therapies for glioma. However, PTPRK mutations abrogated tumor suppressive effects of wild-type PTPRK and altered sensitivity of glioma cells to chemotherapy.</p></div

Redox active metals and H2O2 mediate the increased efficacy of pharmacological ascorbate in combination with gemcitabine or radiation in pre-clinical sarcoma models

Soft tissue sarcomas are a histologically heterogeneous group of rare mesenchymal cancers for which treatment options leading to increased overall survival have not improved in over two decades. The current study shows that pharmacological ascorbate (systemic high dose vitamin C achieving ⥠20 mM plasma levels) is a potentially efficacious and easily integrable addition to current standard of care treatment strategies in preclinical models of fibrosarcoma and liposarcoma both in vitro and in vivo. Furthermore, enhanced ascorbate-mediated toxicity and DNA damage in these sarcoma models were found to be dependent upon H2O2 and intracellular labile iron. Together, these data support the hypothesis that pharmacological ascorbate may represent an easily implementable and non-toxic addition to conventional sarcoma therapies based on taking advantage of fundamental differences in cancer cell oxidative metabolism. Keywords: Sarcoma, Pharmacological ascorbate, Radiation sensitization, Chemotherapy sensitization, Labile iro

NOTCH3 knockdown in U87-MG and U251-MG cells reduced expression of NOTCH3 and its target.

<p>(<b>A</b>) Western blot of U87-MG and U251-MG cells showing silencing of NOTCH3 protein expression at both full length and cleaved forms following shRNA treatment. GAPDH was used as loading control. (<b>B</b>) Quantitative-real time PCR showing downregulation of NOTCH3 transcripts and several of its known targets in U87-MG and U251-MG cells following shRNA knockdown. *indicates p<0.05, **indicates p<0.01 and **indicates p<0.001 (ANOVA and Tukey’s multiple comparison test).</p

NOTCH3 knockdown in U87-MG cells represses glioma cells growth and hinders cell migration and invasion.

<p>(<b>A</b>) Cell viabilities of U87-MG and U251-MG cells following NOTCH3 knockdown were determined using MTT assay. The data represent mean ± SE of three independent experiments performed in triplicate. (<b>B</b>) Growth curves U87-MG and (<b>C</b>) U251-MG cells resulting from specific NOTCH3 knockdown were monitored with crystal violet staining at indicated time points. The results are plotted as the average growth (A_540nm) ± SE of three independent experiments. (<b>D</b>) Colony formation ability of glioma cells was determined with soft agar colony formation assay. Data represent mean ± SE of three independent experiments. (<b>E</b>) Wound healing assay for U87-MG cells. ImageJ software was used to measure changes in cell migration per time in U87-MG cells transduced with sh235 (N = 20) or control (N = 23) lentivirus. Results are expressed as relative migration to the baseline. (<b>F</b>) Matrigel cell invasion assay. U87-MG cells expressing sh235 or scrambled control lentivirus were seeded into the upper chamber of the matrigel coated transwells. Invasive cells were stained with crystal violet, photographed under an inverted light microscope and quantified using ImageJ software in four representative fields. *indicates p<0.05, **indicates p<0.01 and **indicates p<0.001 (ANOVA and Tukey’s multiple comparison test).</p

Ketogenic diets as an adjuvant cancer therapy: History and potential mechanism

Cancer cells, relative to normal cells, demonstrate significant alterations in metabolism that are proposed to result in increased steady-state levels of mitochondrial-derived reactive oxygen species (ROS) such as O2•−and H2O2. It has also been proposed that cancer cells increase glucose and hydroperoxide metabolism to compensate for increased levels of ROS. Given this theoretical construct, it is reasonable to propose that forcing cancer cells to use mitochondrial oxidative metabolism by feeding ketogenic diets that are high in fats and low in glucose and other carbohydrates, would selectively cause metabolic oxidative stress in cancer versus normal cells. Increased metabolic oxidative stress in cancer cells would in turn be predicted to selectively sensitize cancer cells to conventional radiation and chemotherapies. This review summarizes the evidence supporting the hypothesis that ketogenic diets may be safely used as an adjuvant therapy to conventional radiation and chemotherapies and discusses the proposed mechanisms by which ketogenic diets may enhance cancer cell therapeutic responses