Survival Analysis with Large Dimensional Covariates: An Application in Microarray Studies

Use of microarray technology often leads to high-dimensional and low- sample size data settings. Over the past several years, a variety of novel approaches have been proposed for variable selection in this context. However, only a small number of these have been adapted for time-to-event data where censoring is present. Among standard variable selection methods shown both to have good predictive accuracy and to be computationally efficient is the elastic net penalization approach. In this paper, adaptation of the elastic net approach is presented for variable selection both under the Cox proportional hazards model and under an accelerated failure time (AFT) model. Assessment of the two methods is conducted through simulation studies and through analysis of microarray data obtained from a set of patients with diffuse large B-cell lymphoma where time to survival is of interest. The approaches are shown to match or exceed the predictive performance of a Cox-based and an AFT-based variable selection method. The methods are moreover shown to be much more computationally efficient than their respective Cox- and AFT- based counterparts.

QUANTIFYING UNCERTAINTY IN GENOTYPE CALLS

Genome-wide association studies (GWAS) are used to discover genes underlying complex, heritable disorders for which less powerful study designs have failed in the past. The number of GWAS has skyrocketed recently with findings reported in top journals and the mainstream media. Mircorarrays are the genotype calling technology of choice in GWAS as they permit exploration of more than a million single nucleotide polymorphisms (SNPs)simultaneously. The starting point for the statistical analyses used by GWAS, to determine association between loci and disease, are genotype calls (AA, AB, or BB). However, the raw data, microarray probe intensities, are heavily processed before arriving at these calls. Various sophisticated statistical procedures have been proposed for transforming raw data into genotype calls. We find that variability in microarray output quality across different SNPs, different arrays, and different sample batches has substantial inuence on the accuracy of genotype calls made by existing algorithms. Failure to account for these sources of variability, GWAS run the risk of adversely affecting the quality of reported findings. In this paper we present solutions based on a multi-level mixed model. Software implementation of the method described in this paper is available as free and open source code in the crlmm R/BioConductor.

A MULTILEVEL MODEL TO ADDRESS BATCH EFFECTS IN COPY NUMBER ESTIMATION USING SNP ARRAYS

Submicroscopic changes in chromosomal DNA copy number dosage are common and have been implicated in many heritable diseases and cancers. Recent high-throughput technologies have a resolution that permits the detection of segmental changes in DNA copy number that span thousands of basepairs across the genome. Genome-wide association studies (GWAS) may simultaneously screen for copy number-phenotype and SNP-phenotype associations as part of the analytic strategy. However, genome-wide array analyses are particularly susceptible to batch effects as the logistics of preparing DNA and processing thousands of arrays often involves multiple laboratories and technicians, or changes over calendar time to the reagents and laboratory equipment. Failure to adjust for batch effects can lead to incorrect inference and requires inefficient post-hoc quality control procedures that exclude regions that are associated with batch. Our work extends previous model-based approaches for copy number estimation by explicitly modeling batch effects and using shrinkage to improve locus-specific estimates of copy number uncertainty. Key features of this approach include the use of diallelic genotype calls from experimental data to estimate batch- and locus-specific parameters of background and signal without the requirement of training data. We illustrate these ideas using a study of bipolar disease and a study of chromosome 21 trisomy. The former has batch effects that dominate much of the observed variation in quantile-normalized intensities, while the latter illustrates the robustness of our approach to datasets where as many as 25% of the samples have altered copy number. Locus-specific estimates of copy number can be plotted on the copy-number scale to investigate mosaicism and guide the choice of appropriate downstream approaches for smoothing the copy number as a function of physical position. The software is open source and implemented in the R package CRLMM available at Bioconductor (http:www.bioconductor.org).

EXPLORATION, NORMALIZATION, AND GENOTYPE CALLS OF HIGH DENSITY OLIGONUCLEOTIDE SNP ARRAY DATA

In most microarray technologies, a number of critical steps are required to convert raw intensity measurements into the data relied upon by data analysts, biologists and clinicians. These data manipulations, referred to as preprocessing, can influence the quality of the ultimate measurements. In the last few years, the high-throughput measurement of gene expression is the most popular application of microarray technology. For this application, various groups have demonstrated that the use of modern statistical methodology can substantially improve accuracy and precision of gene expression measurements, relative to ad-hoc procedures introduced by designers and manufacturers of the technology. Currently, other applications of microarrays are becoming more and more popular. In this paper we describe a preprocessing methodology for a technology designed for the identification of DNA sequence variants in specific genes or regions of the human genome that are associated with phenotypes of interest such as disease. In particular we describe methodology useful for preprocessing Affymetrix SNP chips and obtaining genotype calls with the preprocessed data. We demonstrate how our procedure improves existing approaches using data from three relatively large studies including one in which large number independent calls are available. Software implementing these ideas are avialble from the Bioconductor oligo package.

REMOVING TECHNICAL VARIABILITY IN RNA-SEQ DATA USING CONDITIONAL QUANTILE NORMALIZATION

The ability to measure gene expression on a genome-wide scale is one of the most promising accomplishments in molecular biology. Microarrays, the technology that first permitted this, were riddled with problems due to unwanted sources of variability. Many of these problems are now mitigated, after a decade’s worth of statistical methodology development. The recently developed RNA sequencing (RNA-seq) technology has generated much excitement in part due to claims of reduced variability in comparison to microarrays. However, we show RNA-seq data demonstrates unwanted and obscuring variability similar to what was first observed in microarrays. In particular, we find GC-content has a strong sample specific effect on gene expression measurements that, if left uncorrected, leads to false positives in downstream results. We also report on commonly observed data distortions that demonstrate the need for data normalization. Here we describe statistical methodology that improves precision by 42% without loss of accuracy. Our resulting conditional quantile normalization (CQN) algorithm combines robust generalized regression to remove systematic bias introduced by deterministic features such as GC-content, and quantile normalization to correct for global distortions.