Microsatellite genotyping reveals a signature in breast cancer exomes.

Genomic instability at microsatellite loci is a hallmark of many cancers, including breast cancer. However, much of the genomic variation and many of the hereditary components responsible for breast cancer remain undetected. We hypothesized that variation at microsatellites could provide additional genomic markers for breast cancer risk assessment. A total of 1,345 germline and tumor DNA samples from individuals diagnosed with breast cancer, exome sequenced as part of The Cancer Genome Atlas, were analyzed for microsatellite variation. The comparison group for our analysis, representing healthy individuals, consisted of 249 females which were exome sequenced as part of the 1,000 Genomes Project. We applied our microsatellite-based genotyping pipeline to identify 55 microsatellite loci that can distinguish between the germline of individuals diagnosed with breast cancer and healthy individuals with a sensitivity of 88.4 % and a specificity of 77.1 %. Further, we identified additional microsatellite loci that are potentially useful for distinguishing between breast cancer subtypes, revealing a possible fifth subtype. These findings are of clinical interest as possible risk diagnostics and reveal genes that may be of potential therapeutic value, including genes previously not associated with breast cancer.

Population-scale analysis of human microsatellites reveals novel sources of exonic variation.

Using our microsatellite specific genotyping method, we analyzed tandem repeats, which are known to be highly variable with some recognized as biomarkers causative of disease, in over 500 individuals who were exon sequenced in a 1000 Genomes Project pilot study. We were able to genotype over 97% of the microsatellite loci in the targeted regions. A total of 25,115 variations were observed, including repeat length and single nucleotide polymorphisms, corresponding to an average of 45.6 variations per individual and a density of 1.1 variations per kilobase. Standard variant detection did not report 94.2% of the exonic repeat length variations in part because the alignment techniques are not ideal for repetitive regions. Additionally some standard variation detection tools rely on a database of known variations, making them less likely to call repeat length variations as only a small percent of these loci (~6000) have been accurately characterized. A subset of the hundreds of non-synonymous variations we identified was experimentally validated, indicating an accuracy of 96.5% for our microsatellite-based genotyping method, with some novel variants identified in genes associated with cancer. We propose that microsatellite-based genotyping be used as a part of large scale sequencing studies to identify novel variants.

Evaluation of microsatellite variation in the 1000 Genomes Project pilot studies is indicative of the quality and utility of the raw data and alignments.

We performed an analysis of global microsatellite variation on the two kindreds sequenced at high depth (~20×-60×) in the 1000 Genomes Project pilot studies because alterations in these highly mutable repetitive sequences have been linked with many phenotypes and disease risks. The standard alignment technique performs poorly in microsatellite regions as a consequence of low effective coverage (~1×-5×) resulting in 79% of the informative loci exhibiting non-Mendelian inheritance patterns. We used a more stringent approach in computing robust allelotypes resulting in 94.4% of the 1095 informative repeats conforming to traditional inheritance. The high-confidence allelotypes were analyzed to obtain an estimate of the minimum polymorphism rate as a function of motif length, motif sequence, and distribution within the genome.

A long AAAG repeat allele in the 5' UTR of the ERR-γ gene is correlated with breast cancer predisposition and drives promoter activity in MCF-7 breast cancer cells.

We sequenced the 5' UTR of the estrogen-related receptor gamma gene (ERR-γ) in ~500 patient and volunteer samples and found that longer alleles of the (AAAG)(n) microsatellite were statistically and significantly more likely to exist in the germlines of breast cancer patients when compared to healthy volunteers. This microsatellite region contains multiple binding sites for a number of transcription factors, and we hypothesized that the polymorphic AAAG-containing sequence in the 5' UTR region of ERR-γ might modulate expression of ERR-γ. We found that the 369 bp PCR product containing the AAAG repeat drove expression of a reporter gene in estrogen receptor positive breast cancer cells. Our results support a role for the 5' UTR region in ERR-γ expression, which is potentially mediated via binding to the variable tandem AAAG repeat, the length of which correlates with breast cancer pre-disposition. Our study indicates that the AAAG tetranucleotide repeat polymorphism in ERR-γ gene 5' UTR region may be a new biomarker for genetic susceptibility to breast cancer.

Status asthmaticus complicated by cardiac arrest.

In the setting of severe acute asthma, electrocardiographic abnormalities are not uncommon and some patients will develop reversible systolic dysfunction. However acute myocardial infarction and potentially fatal arrhythmias are rare. We report the case of a middle-aged indigenous male who suffered an ST-elevation myocardial infarction and then pulseless ventricular tachycardia arrest while still in the acute phase of treatment for status asthmaticus.

Global microsatellite content distinguishes humans, primates, animals, and plants.

Microsatellites are highly mutable, repetitive sequences commonly used as genetic markers, but they have never been studied en masse. Using a custom microarray to measure hybridization intensities of every possible repetitive nucleotide motif from 1-mers to 6-mers, we examined 25 genomes. Here, we show that global microsatellite content varies predictably by species, as measured by array hybridization signal intensities, correlating with established taxonomic relationships, and particular motifs are characteristic of one species versus another. For instance, hominid-specific microsatellite motifs were identified despite alignment of the human reference, Celera, and Venter genomic sequences indicating substantial variation (30-50%) among individuals. Differential microsatellite motifs were mainly associated with genes involved in developmental processes, whereas those found in intergenic regions exhibited no discernible pattern. This is the first description of a method for evaluating microsatellite content to classify individual genomes.