A Single Nucleotide Polymorphism in Catalase Is Strongly Associated with Ovarian Cancer Survival

<div><p>Ovarian cancer is the deadliest of all gynecologic cancers. Recent evidence demonstrates an association between enzymatic activity altering single nucleotide polymorphisms (SNP) with human cancer susceptibility. We sought to evaluate the association of SNPs in key oxidant and antioxidant enzymes with increased risk and survival in epithelial ovarian cancer. Individuals (n = 143) recruited were divided into controls, (n = 94): healthy volunteers, (n = 18), high-risk <i>BRCA1/2</i> negative (n = 53), high-risk <i>BRCA1/2</i> positive (n = 23) and ovarian cancer cases (n = 49). DNA was subjected to TaqMan SNP genotype analysis for selected oxidant and antioxidant enzymes. Of the seven selected SNP studied, no association with ovarian cancer risk (Pearson Chi-square) was found. However, a catalase SNP was identified as a predictor of ovarian cancer survival by the Cox regression model. The presence of this SNP was associated with a higher likelihood of death (hazard ratio (HR) of 3.68 (95% confidence interval (CI): 1.149–11.836)) for ovarian cancer patients. Kaplan-Meier survival analysis demonstrated a significant median overall survival difference (108 versus 60 months, p<0.05) for those without the catalase SNP as compared to those with the SNP. Additionally, age at diagnosis greater than the median was found to be a significant predictor of death (HR of 2.78 (95% CI: 1.022–7.578)). This study indicates a strong association with the catalase SNP and survival of ovarian cancer patients, and thus may serve as a prognosticator.</p></div

ALKBH7 Variant Related to Prostate Cancer Exhibits Altered Substrate Binding

<div><p>The search for prostate cancer biomarkers has received increased attention and several DNA repair related enzymes have been linked to this dysfunction. Here we report a targeted search for single nucleotide polymorphisms (SNPs) and functional impact characterization of human ALKBH family dioxygenases related to prostate cancer. Our results uncovered a SNP of <i>ALKBH7</i>, rs7540, which is associated with prostate cancer disease in a statistically significantly manner in two separate cohorts, and maintained in African American men. Comparisons of molecular dynamics (MD) simulations on the wild-type and variant protein structures indicate that the resulting alteration in the enzyme induces a significant structural change that reduces ALKBH7’s ability to bind its cosubstrate. Experimental spectroscopy studies with purified proteins validate our MD predictions and corroborate the conclusion that this cancer-associated mutation affects productive cosubstrate binding in ALKBH7.</p></div

Kaplan-Meier overall survival curves for in ovarian cancer utilizing a specific catalase SNP.

<p>The solid curve represents cases with (CC) homozygous wild-type genotype as compared to the dashed curve, which represents cases with homozygous mutant plus heterozygous mutant (CT+TT) genotypes. The X-axis represents patient survival in months; the Y-axis represents cumulative survival percentage. Chi-square p-value 0<0.05 is considered statistically significant.</p

Cox regression analysis for selected SNPs in key oxidants and antioxidants genes in ovarian cancer.

<p>Adjusted Hazard Ratio (HR) for the variables included in the model.</p><p>* p<0.05, degrees of freedom = 1 for all analyses. CYBA; NAD(P)H oxidase subunit (NOX4), GPX; glutathione peroxidase, GSR; glutathione reductase, MnSOD; manganese superoxide dismutase, MPO; myeloperoxidase, NOS2; inducible nitric oxide synthase. For this analysis, several “Method”simulations were performed such as: forced entry (ENTER), forward LR (likelihood ratio), etc. The forward LR was chosen for the final analysis. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0135739#pone.0135739.t004" target="_blank">Table 4</a> includes the strongest predictors kept by the model as well as those rejected by the model. The Cox regression model generated the scores in the table. The P-values are noted in the column significance; *p<0.05, is considered statistically significant.</p><p>Cox regression analysis for selected SNPs in key oxidants and antioxidants genes in ovarian cancer.</p

Comparison of cases and controls based on demographic, personal or family history of cancer, and genotypic characteristics.

<p><i>*p< 0</i>.<i>05</i>, <i>CAT; catalase</i>, <i>CYBA; NAD(P)H oxidase subunit (NOX4)</i>, <i>GPX; glutathione peroxidase</i>, <i>GSR; glutathione reductase</i>, <i>MnSOD; manganese superoxide dismutase</i>, <i>MPO; myeloperoxidase</i>, <i>NOS2; inducible nitric oxide synthase</i>.</p><p>Comparison of cases and controls based on demographic, personal or family history of cancer, and genotypic characteristics.</p

SNPs of <i>ALKBH</i> family genes with significant association (p ≤ 0.05) with prostate cancer phenotype.

<p>SNPs of <i>ALKBH</i> family genes with significant association (p ≤ 0.05) with prostate cancer phenotype.</p

Structural and dynamic comparison between WT and R191Q ALKBH7 with bound α-kg.

<p><b>a,</b> Overlay of representative structures for WT (gray) and R191Q mutant (yellow) forms of ALKBH7. Active site residues and α-kg as well as the site undergoing substitution are displayed (licorice). <b>b,</b> 180 degree rotation and close-up of the substituted site. <b>c,</b> 90 degree rotation and close-up of the active site, with each relevant active site residue and α-kg labeled. Dashed lines in gray represent the original bonds to the metal ion in the crystal structure, and dashed lines in orange represent the new bonds to the metal ion near the end of the trajectory for the variant protein. <b>d,</b> Correlation difference for each residue in the WT protein with respect to the R191Q variant mapped onto the protein structure using the mutation site as the reference. <b>e,</b> Distance analysis for key residues in the SNP variant and active sites (with respect to their centers of mass) throughout the simulation trajectory.</p

Average binding enthalpies for cosubstrate α-kg and Fe at the active site (in kcal/mol) for the simulation.

<p>Results for the product (succinate) are reported in the Supplementary Information.</p

Hydrogen bond analysis for the WT/R191Q variant with α-kg.

<p>Residues colored in red denote amino acids involved in H-bonds for over 30% of the WT trajectory and broken for over 90% of the R191Q variant trajectory. Residues colored in orange are involved in hydrogen bonds for both trajectories, but are present for at least 30% less of the time in the variant trajectory. The hydrogen bonds between these residues are displayed in blue. The corresponding analysis for the WT/SNP variant with succinate are given in the Supplementary Information.</p

Difference absorption spectra of WT ALKBH7 and its R191Q variant.

<p>The spectra of the anaerobic proteins (0.3 mM) were recorded in the presence of 2 mM α-kg and 100 μM Fe(II). The difference spectra were obtained by subtracting the spectra for proteins with α-kg, but without the metal. <b>A</b>, WT ALKBH7; <b>B</b>, R191Q ALKBH7.</p