Karina L. Mine

Gene network reconstruction reveals cell cycle and antiviral genes as major drivers of cervical cancer

Although human papillomavirus (HPV) was identified as an etiological factor in cervical cancer, the key human gene drivers of this disease remain unknown. Here we apply an unbiased approach integrating gene expression and chromosomal aberration data. In an independent group of patients, we reconstruct and validate a gene regulatory meta-network, and identify cell cycle and antiviral genes that constitute two major sub-networks up-regulated in tumour samples. These genes are located within the same regions as chromosomal amplifications, most frequently on 3q. We propose a model in which selected chromosomal gains drive activation of antiviral genes contributing to episomal virus elimination, which synergizes with cell cycle dysregulation. These findings may help to explain the paradox of episomal HPV decline in women with invasive cancer who were previously unable to clear the virus.

Construct and Compare Gene Coexpression Networks with DAPfinder and DAPview.

BackgroundDAPfinder and DAPview are novel BRB-ArrayTools plug-ins to construct gene coexpression networks and identify significant differences in pairwise gene-gene coexpression between two phenotypes.ResultsEach significant difference in gene-gene association represents a Differentially Associated Pair (DAP). Our tools include several choices of filtering methods, gene-gene association metrics, statistical testing methods and multiple comparison adjustments. Network results are easily displayed in Cytoscape. Analyses of glioma experiments and microarray simulations demonstrate the utility of these tools.ConclusionsDAPfinder is a new friendly-user tool for reconstruction and comparison of biological networks.

A novel strategy for defining haplotypes by selective depletion using restriction enzymes

Various single nucleotide polymorphisms (SNPs) have been investigated regarding association with gene expression levels or human diseases. Although different SNPs within one gene are frequently analyzed individually, it is highly probable that in the majority of the cases, a precise combination of SNP alleles, i.e., haplotype, determines a functional trait. Methods commonly used for haplotype determination, involving studies in families, cloning, or somatic cell hybrids, are expensive and time-consuming. We herein suggest a novel and simple strategy for haplotype determination, involving selective haplotype depletion with a restriction enzyme, followed by sequencing. We studied 11 LTA gene polymorphisms in 102 Brazilian individuals, and we applied this novel methodology for haplotyping 67 out of 70 LTA heterozygous individuals. We concluded that the method is rapid and efficient, and, as it includes only simple and widespread-used techniques, it could be used in most of the laboratories without further investment in equipments. The wider usage of haplotyping could be important to clarify contradictory results frequently observed among studies that focus on a single SNP.

Heightened expression of HLA-DQB1 and HLA-DQB2 in pre-implantation biopsies predicts poor late kidney graft function

BACKGROUND Accurate pre-transplant prediction of late graft function remains an unmet need in kidney transplantation. The aim of this study was to evaluate HLA genes expression levels in pre-implantation biopsies (PIB) of deceased donor kidneys as markers for long-term graft outcome. METHODS HLA genes expression analysis was initially performed using microarray data of 53 PIB, previously generated by our laboratory. The validation analysis was performed by real-time PCR in 116 PIB from an independent cohort. RESULTS The microarray data showed association between high expression levels of HLA class II genes, especially HLA-DQB1 and -DQB2, in kidneys from young (18 to 49-year-old) donors and poor (eGFR < 45 mL/min/1.73 m2) 1- and 5-year graft function. A subsequent study in an independent cohort, in which only HLA-DQB2 expression was evaluated, validated the association between increased HLA-DQB2 expression in PIB of kidneys from young donors and poor 1-year graft function: expression levels ≥0.0025 relative units conferred an odds ratio of 22.5, with positive and negative predictive values of 71.4% and 90.0%, respectively. CONCLUSION Heightened expression of HLA-DQB1 and -DQB2 in PIB are promising tools for pre-transplant risk assessment of poor late graft function in transplants with kidneys from 18 to 49-year-old donors.

Predicting delayed kidney graft function with gene expression in preimplantation biopsies and first-day posttransplant blood

The purpose of this study was to investigate possible markers for predicting delayed graft function (DGF). To this end we analyzed, in pre-implantation biopsies (PIB) and in first-day post-Tx peripheral blood mononuclear cells (PBMC), the expression of five genes (ACSL4, CUBN, DEFB1, FABP3, GK) through real-time TaqMan PCR assays. These genes were selected from a large scale gene expression study in PIB. DEFB1, FABP3 and GK expression levels in PIB were lower in cases with DGF and, in a multivariate analysis which included these genes and clinical variables, only FABP3 expression remained independently associated with DGF. FABP3 expression lower than -1.32 units of relative expression conferred an odds ratio for DGF of 41.1. Compared to the PBMC of recipients without DGF, recipients with prolonged DGF (pDGF) had lower ACSL4 and higher DEFB1 expression levels. In a multivariate analysis, including PBMC gene expression levels of ACSL4, DEFB1 and TLR4 (data from a previous study with the same patients) and clinical variables, only TLR4 remained independently associated with pDGF. In summary, this study revealed FABP3 expression in PIB as a marker for DGF and disclosed new genes possibly involved in the pathogenesis of DGF.