DNA methylation patterns in luminal breast cancers differ from non-luminal subtypes and can identify relapse risk independent of other clinical variables.

The diversity of breast cancers reflects variations in underlying biology and affects the clinical implications for patients. Gene expression studies have identified five major subtypes- Luminal A, Luminal B, basal-like, ErbB2+ and Normal-Like. We set out to determine the role of DNA methylation in subtypes by performing genome-wide scans of CpG methylation in breast cancer samples with known expression-based subtypes. Unsupervised hierarchical clustering using a set of most varying loci clustered the tumors into a Luminal A majority (82%) cluster, Basal-like/ErbB2+ majority (86%) cluster and a non-specific cluster with samples that were also inconclusive in their expression-based subtype correlations. Contributing methylation loci were both gene associated loci (30%) and non-gene associated (70%), suggesting subtype dependant genome-wide alterations in the methylation landscape. The methylation patterns of significant differentially methylated genes in luminal A tumors are similar to those identified in CD24 + luminal epithelial cells and the patterns in basal-like tumors similar to CD44 + breast progenitor cells. CpG islands in the HOXA cluster and other homeobox (IRX2, DLX2, NKX2-2) genes were significantly more methylated in Luminal A tumors. A significant number of genes (2853, p < 0.05) exhibited expression-methylation correlation, implying possible functional effects of methylation on gene expression. Furthermore, analysis of these tumors by using follow-up survival data identified differential methylation of islands proximal to genes involved in Cell Cycle and Proliferation (Ki-67, UBE2C, KIF2C, HDAC4), angiogenesis (VEGF, BTG1, KLF5), cell fate commitment (SPRY1, OLIG2, LHX2 and LHX5) as having prognostic value independent of subtypes and other clinical factors.

Complex landscapes of somatic rearrangement in human breast cancer genomes.

Multiple somatic rearrangements are often found in cancer genomes; however, the underlying processes of rearrangement and their contribution to cancer development are poorly characterized. Here we use a paired-end sequencing strategy to identify somatic rearrangements in breast cancer genomes. There are more rearrangements in some breast cancers than previously appreciated. Rearrangements are more frequent over gene footprints and most are intrachromosomal. Multiple rearrangement architectures are present, but tandem duplications are particularly common in some cancers, perhaps reflecting a specific defect in DNA maintenance. Short overlapping sequences at most rearrangement junctions indicate that these have been mediated by non-homologous end-joining DNA repair, although varying sequence patterns indicate that multiple processes of this type are operative. Several expressed in-frame fusion genes were identified but none was recurrent. The study provides a new perspective on cancer genomes, highlighting the diversity of somatic rearrangements and their potential contribution to cancer development.

The estrogen receptor pathway in rhabdomyosarcoma: a role for estrogen receptor-beta in proliferation and response to the antiestrogen 4'OH-tamoxifen.

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children. Highly malignant, RMS frequently fails to respond to conventional aggressive multimodal radiation, surgery, and chemotherapy treatment protocols that also cause significant sequelae in the growing child. Other tumors of mesenchymal origin, such as locally aggressive fibromatoses and desmoid tumors, have been successfully treated with a selective estrogen receptor (ER) modulator, tamoxifen. In an effort to identify new targets for RMS therapy, our group investigated the previously uncharacterized ER pathway in RMS cell culture and primary tumors. We detected ER isoform beta (ER beta), but not isoform alpha, RNA, and protein in five RMS cell lines. Immunohistochemical staining of primary RMS tumor sections confirmed high levels of ER beta but not ER alpha protein. RMS cell growth was dramatically inhibited in steroid-free conditions, and this growth inhibition was rescued with 17-beta-estradiol (E2) supplementation. Exposure of RMS cells to 4'OH-tamoxifen (4OHT) decreased cell viability and inhibited colony formation as detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and colony-forming assays. 4OHT also induced apoptotic signaling in RMS cells as detected by cleavage of caspase-3 and poly(ADP)ribose polymerase. This effect increased 3- to 8-fold in steroid-deprived conditions but was rescued by supplementation with E2. Immunofluorescence studies detected a change in the subcellular localization of ER beta in response to 4OHT. Together, these data suggest an active ER beta-mediated signal transduction pathway in RMS. The ability of 4OHT to induce apoptotic signaling and disrupt estradiol-mediated proliferation provides a rationale to explore a role for selective ER modulators in the treatment of RMS.