SUMMARY Sarcomas are a broad family of mesenchymal malignancies exhibiting remarkable histologic diversity. We describe the multi-platform molecular landscape of 206 adult soft tissue sarcomas representing 6 major types. Along with novel insights into the biology of individual sarcoma types, we report three overarching findings: 1) unlike most epithelial malignancies, these sarcomas (excepting synovial sarcoma) are characterized predominantly by copy number changes, with low mutational loads and only a few genes (TP53, ATRX, RB1) highly recurrently mutated across sarcoma types, 2) within sarcoma types, genomic and regulomic diversity of driver pathways defines molecular subtypes associated with patient outcome, and 3) the immune microenvironment, inferred from DNA methylation and mRNA profiles, associates with outcome and may inform clinical trials of immune checkpoint inhibitors. Overall, this large-scale analysis reveals previously unappreciated sarcoma type-specific changes in copy number, methylation, RNA, and protein, providing insights into refining sarcoma therapy and relationships to other cancer types.

Angiosarcoma is an aggressive malignancy that arises spontaneously or secondarily to ionising radiation or chronic lymphoedema1. Previous work has identified aberrant angiogenesis, including occasional somatic mutations in angiogenesis signalling genes, as a key driver of angiosarcoma1. Here, we employed whole genome, exome, and targeted sequencing to study the somatic changes underpinning primary and secondary angiosarcoma. We identified recurrent mutations in two genes, PTPRB and PLCG1, which are intimately linked to angiogenesis. The endothelial phosphatase PTPRB, a negative regulator of vascular growth factor tyrosine kinases, harboured predominantly truncating mutations in 10/39 (26%) tumours. PLCG1, a signal transducer of tyrosine kinases, presented with a recurrent, likely activating R707Q missense variant in 3/34 cases (9%). Overall, 15/39 (38%) tumours harboured at least one driver mutation in angiogenesis signalling genes. Our findings inform and reinforce current therapeutic efforts to target angiogenesis signalling in angiosarcoma.

The ability of 20-50 nm nanoparticles to target and modulate the biology of specific types of cells will enable major advancements in cellular imaging and therapy in cancer and atherosclerosis. A key challenge is to load an extremely high degree of targeting, imaging, and therapeutic functionality into small, yet stable particles. Herein we report ~30 nm stable uniformly sized near-infrared (NIR) active, superparamagnetic nanoclusters formed by kinetically controlled self-assembly of goldcoated iron oxide nanoparticles. The controlled assembly of nanocomposite particles into clusters with small primary particle spacings produces collective responses of the electrons that shift the absorbance into the NIR region. The nanoclusters of ~70 iron oxide primary particles with thin gold coatings display intense NIR (700-850 nm) absorbance with a cross section of ~10 −14 m 2 . Because of the thin gold shells with an average thickness of only 2 nm, the r 2 spin-spin magnetic relaxivity is 219 mM −1 s −1 , an order of magnitude larger than observed for typical iron oxide particles with thicker gold shells. Despite only 12% by weight polymeric stabilizer, the particle size and NIR absorbance change very little in deionized water over 8 months. High uptake of the nanoclusters by macrophages is facilitated by the dextran coating, producing intense NIR contrast in dark field and hyperspectral microscopy, both in cell culture and an in vivo rabbit model of atherosclerosis. Small nanoclusters with optical, magnetic, and therapeutic functionality, designed by assembly of *Address correspondence to: kpj@che.utexas.edu, FELDMANM@uthscsa.edu. Supporting Information Available: Reproducibility in nanorose size distribution; porosity of dextran in the shells about the iron oxide particle; estimation of number of particles per nanocluster; average optical density spectra in macrophages labeled with nanorose by hyperspectral microscopy; and laser vaporization of macrophages in vitro. This material is available free of charge via the Internet at http://pubs.acs.org. NIH Public AccessAuthor Manuscript ACS Nano. Author manuscript; available in PMC 2010 September 22. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript nanoparticle building blocks, offer broad opportunities for targeted cellular imaging, therapy, and combined imaging and therapy. Keywordsgold; iron oxide; nanocluster; near-infrared; macrophage targeted imaging; MRI; atherosclerosis; cancer Clinical imaging and/or therapy with multifunctional nanoparticles that target specific types of cells has the potential to transform health care in cancer, atherosclerosis, and other diseases. When the nanoparticle diameters are reduced to 20-50 nm, the biological pathways in targeted cells can undergo profound changes. [1][2][3][4][5] Small nanoparticles, the size of small viruses, permeate barriers more rapidly including cell membranes and leaky vasculature in cancers. The efficacy of vaccines may be enhanced with ultrasmall 20 nm nanoparticles that can dif...

Metal nanoparticles with surface plasmon resonance (SPR) in the near-infrared region (NIR) are of great interest for imaging and therapy. Presently, gold nanoparticles with NIR absorbance are typically larger than 50nm, above the threshold size of ~5 nm required for efficient renal clearance. As these nanoparticles are not biodegradable, concerns about long-term toxicity have restricted their translation into the clinic. Here, we address this problem by developing a flexible platform for the kinetically-controlled assembly of sub-5 nm ligand-coated gold particles to produce metal/polymer biodegradable nanoclusters smaller than 100 nm with strong NIR absorbance for multimodal application. A key novel feature of the proposed synthesis is the use of weakly adsorbing biodegradable polymers that allows tight control of nanocluster size and, in addition, results in nanoclusters with unprecedented metal loadings, and thus optical functionality. Over time, the biodegradable polymer stabilizer degrades under physiological conditions that leads to disassembly of the nanoclusters into sub-5nm primary gold particles which are favorable for efficient body clearance. This synthesis of polymer/inorganic nanoclusters combines the imaging contrast and therapeutic capabilities afforded by the NIR-active nanoparticle assembly with the biodegradability of a polymer stabilizer.

To date, no specific marker exists for the detection of circulating tumor cell from different types of sarcomas, though tools are available for detection of circulating tumor cell (CTC) in peripheral blood of cancer patients for epithelial cancers. Here we report cell-surface vimentin (CSV) as an exclusive marker on sarcoma CTC regardless of the tissue origin of the sarcoma as detected by a novel monoclonal antibody. Utilizing CSV as a probe we isolated and enumerated sarcoma CTC with high sensitivity and specificity from the blood of patients bearing different types of sarcoma, validating their phenotype by single cell genomic amplification, mutation detection and fluorescence in situ hybridization. Our results establish the first universal and specific CTC marker described for enumerating CTC from different types of sarcoma, thereby providing a key prognosis tool to monitor cancer metastasis and relapse.

Synovial sarcoma (SS) is defined by the hallmark SS18-SSX fusion oncoprotein, which renders BAF complexes aberrant in two manners: gain of SSX to the SS18 subunit and concomitant loss of BAF47 subunit assembly. Here we demonstrate that SS18-SSX globally hijacks BAF complexes on chromatin to activate an SS transcriptional signature that we define using primary tumors and cell lines. Specifically, SS18-SSX retargets BAF complexes from enhancers to broad polycomb domains to oppose PRC2-mediated repression and activate bivalent genes. Upon suppression of SS18-SSX, reassembly of BAF47 restores enhancer activation, but is not required for proliferative arrest. These results establish a global hijacking mechanism for SS18-SSX on chromatin, and define the distinct contributions of two concurrent BAF complex perturbations.

Malignant peripheral nerve sheath tumors (MPNSTs) are aggressive sarcomas that can show overlapping features with benign neurofibromas as well as high-grade sarcomas. Additional diagnostic markers are needed to aid in this often challenging differential diagnosis. Recently mutations in two critical components of the polycomb repressor 2 (PRC2) complex, SUZ12 and EED, were reported to occur specifically in MPNSTs while such mutations are absent in neurofibromas, both in the setting of neurofibromatosis (NF) and sporadic cases. Furthermore, both SUZ12 and EED mutations in MPNSTs were associated with loss of H3K27 tri-methylation, a downstream target of PRC2. Therefore we tested whether H3K27me3 immunohistochemistry is useful as a diagnostic and prognostic marker for MPNSTs. We performed H3K27me3 immunohistochemistry in 162 primary MPNSTs, 97 neurofibromas and 341 other tumors using tissue microarray. We observed loss of H3K27me3 in 34% (55/162) of all MPNSTs while expression was retained in all neurofibromas including atypical (n=8) and plexiform subtypes (n=24). Within other tumors we detected loss of H3K27me3 in only 7% (24/341). Surprisingly, 60% (9/15) of synovial sarcomas and 38% (3/8) of fibrosarcomatous dermatofibrosarcoma protuberans (DFSP) showed loss of H3K27 trimethylation. Only 1 out of 44 schwannomas showed loss of H3K27me3 and all 4 perineuriomas showed intact H3K27me3. Furthermore, MPNSTs with loss of H3K27 tri-methylation showed inferior survival compared to MPNSTs with intact H3K27 tri-methylation, which was validated in two independent cohorts. Our results indicate that H3K27me3 immunohistochemistry is useful as a diagnostic marker in which loss of H3K27me3 favours MPNST above neurofibroma. However H3K27me3 immunohistochemistry is not suitable to distinguish MPNST from its morphological mimicker synovial sarcoma or fibrosarcomatous DFSP. Since loss of H3K27 tri-methylation was related to poorer survival in MPNST, chromatin modification mediated by this specific histone seems to orchestrate more aggressive tumour biology.

Objectives Expression of strong nuclear STAT6 is thought to be a specific marker for solitary fibrous tumors (SFT). Little is known about subtle expression patterns in other mesenchymal lesions. Methods We performed immunohistochemical studies against the C-terminus of STAT6 in tissue microarrays and whole sections, comprising 2366 mesenchymal lesions. Results Strong nuclear STAT6 was expressed in 285/2021 tumors, including 206/240 SFT, 49/408 well/dedifferentiated liposarcomas, 8/65 unclassified sarcomas, and 14/184 desmoids, among others. Expression in SFT was predominately limited to the nucleus. Other positive tumors typically expressed both nuclear and cytoplasmic STAT6. Complete absence of STAT6 was most common in pleomorphic liposarcoma and alveolar soft part sarcoma (60% and 72% cases negative, respectively). Conclusions Strong nuclear STAT6 is largely specific for SFT. Physiologic low-level cytoplasmic/nuclear expression is common in mesenchymal neoplasia, and is of uncertain significance.