Summary We analyzed primary breast cancers by genomic DNA copy number arrays, DNA methylation, exome sequencing, mRNA arrays, microRNA sequencing and reverse phase protein arrays. Our ability to integrate information across platforms provided key insights into previously-defined gene expression subtypes and demonstrated the existence of four main breast cancer classes when combining data from five platforms, each of which shows significant molecular heterogeneity. Somatic mutations in only three genes (TP53, PIK3CA and GATA3) occurred at > 10% incidence across all breast cancers; however, there were numerous subtype-associated and novel gene mutations including the enrichment of specific mutations in GATA3, PIK3CA and MAP3K1 with the Luminal A subtype. We identified two novel protein expression-defined subgroups, possibly contributed by stromal/microenvironmental elements, and integrated analyses identified specific signaling pathways dominant in each molecular subtype including a HER2/p-HER2/HER1/p-HER1 signature within the HER2-Enriched expression subtype. Comparison of Basal-like breast tumors with high-grade Serous Ovarian tumors showed many molecular commonalities, suggesting a related etiology and similar therapeutic opportunities. The biologic finding of the four main breast cancer subtypes caused by different subsets of genetic and epigenetic abnormalities raises the hypothesis that much of the clinically observable plasticity and heterogeneity occurs within, and not across, these major biologic subtypes of breast cancer.

Genetic information storage and processing rely on just two polymers, DNA and RNA. Whether their role reflects evolutionary history or fundamental functional constraints is unknown. Using polymerase evolution and design, we show that genetic information can be stored in and recovered from six alternative genetic polymers (XNAs) based on simple nucleic acid architectures not found in nature. We also select XNA aptamers, which bind their targets with high affinity and specificity, demonstrating that beyond heredity, specific XNAs have the capacity for Darwinian evolution and for folding into defined structures. Thus, heredity and evolution, two hallmarks of life, are not limited to DNA and RNA but are likely to be emergent properties of polymers capable of information storage.The nucleic acids DNA and RNA provide the molecular basis for all life through their unique ability to store and propagate information. To better understand these singular properties and discover relevant parameters for the chemical basis of molecular information encoding, nucleic acid structure has been dissected by systematic variation of nucleobase, sugar and backbone moieties (1-7).These studies have revealed the profound influence of backbone, sugar and base chemistry on nucleic acid properties and function. Crucially, only a small subset of chemistries allows information transfer through base pairing with DNA or RNA, a prerequisite for crosstalk with extant biology. However, base pairing alone cannot conclusively determine the capacity of a given chemistry to serve as a genetic system, as hybridization need not preserve information content (8). A more thorough examination of candidate genetic polymers' potential for information storage, propagation and evolution requires a system for replication which would allow a systematic exploration of the informational, evolutionary and functional potential of synthetic genetic polymers and open up applications ranging from biotechnology to material science.In principle, informational polymers can be synthesized and replicated chemically (9) with advances in the non-enzymatic polymerization of mononucleotides (10) (11, 12) enabling model selection experiments (13). Nevertheless, chemical polymerization remains relatively inefficient. On the other hand, enzymatic polymerization has been hindered by the stringent substrate selectivity of polymerases. Despite progress in understanding the determinants of polymerase substrate specificity and in engineering polymerases with expanded substrate spectra (7), most unnatural nucleotide analogues are poor polymerase substrates at full substitution, both as nucleotides for polymer synthesis and as templates for reverse transcription. Notable exceptions are 2′OMe-DNA and TNA. 2′OMe-DNA is present in eukaryotic rRNAs, is well-tolerated by natural reverse transcriptases (RTs) and has been shown to support heredity and evolution at near full substitution (14). TNA allowed polymer synthesis and evolution in a three letter system (15) but only limited re...

The use of chemically synthesized short interfering RNAs (siRNAs) is currently the method of choice to manipulate gene expression in mammalian cell culture, yet improvements of siRNA design is expectably required for successful application in vivo. Several studies have aimed at improving siRNA performance through the introduction of chemical modifications but a direct comparison of these results is difficult. We have directly compared the effect of 21 types of chemical modifications on siRNA activity and toxicity in a total of 2160 siRNA duplexes. We demonstrate that siRNA activity is primarily enhanced by favouring the incorporation of the intended antisense strand during RNA-induced silencing complex (RISC) loading by modulation of siRNA thermodynamic asymmetry and engineering of siRNA 3′-overhangs. Collectively, our results provide unique insights into the tolerance for chemical modifications and provide a simple guide to successful chemical modification of siRNAs with improved activity, stability and low toxicity.