Proteomic Comparison Of DSRCT, Normal Mesenteric Tissues and ES. Proteomic Comparison Of DSRCT, Normal Mesenteric Tissues and ES

Desmoplastic small round cell tumor (DSRCT) is an aggressive malignancy that occurs predominantly in young adult males and is characterized by abdominopelvic sarcomatosis exhibiting multi-lineage cellular nests of epithelial, muscular, mesenchymal, and neural differentiation admixed with desmoplastic stroma. Prior to the recognition of the disease as a distinct clinical entity, DSRCT was invariably misclassified as poorly differentiated atypical cancer of the testes, ovary, mesentery, or gastrointestinal tract, and the chemotherapies used for those malignancies elicited poor clinical response. As previously reported, a tectonic shift in the treatment of these patients occurred after researchers made two astute observations: 1) DSRCT microscopically resembles other small round “blue cell” sarcoma subtypes (e.g., ES, rhabdomyosarcoma, synovial sarcoma), and 2) DSRCT and ES have the same N-terminal EWSR1 fusion partner. Proteomic analysis using a reverse-phase protein lysate array (RPPA) was used to elucidate biomarkers that distinguish DSRCT from adjacent normal tissue and Ewing sarcoma. This proteomic analysis revealed novel proteins, such as the androgen receptor and Syk, that may be susceptible to drug targeting, as well as oncogenic pathways like Akt-PI3K that are highly expressed in DSRCT. Overall design: Snap-frozen DSRCT (n=16) or ES (n=6) specimens, and normal-appearing mesenteric tissue (n=8), were collected from core needle biopsies or Complete cytoreductive surgery using clinical protocols approved by MDACC’s Institutional Review Board. Lysates were created, protein concentrations were determined, and individual protein expression was measured using a well-validated reverse-phase protein array (RPPA) as previously described37,38. Raw log2 intensity values were normalized for global protein expression by median centering across 149 antibodies tested. Individual protein expressions in DSRCT specimens, ES specimens, and mesenteric normal tissue specimens were compared using two-sided unpaired t-tests with the GeneSpring GX software program version 12.6.1-GX (Agilent Technologies). For multiple comparison testing, proteins whose expression levels between specimen types were 2-fold and significantly different (p≤0.05), were subjected to unsupervised hierarchical clustering. The protein lysates from DSRCT and ES were subjected to RPPA analysis for 149 proteins and phosphoproteins (red, increased signal; green, decreased signal). Unsupervised double-hierarchical clustering using the Pearson correlation distance metric between proteins (rows) and Centroid linkage (a clustering method) separated the 22 samples into two groups by tumor type (columns). Of the 22 proteins, 8 had expression that differed significantly between ES and DSRCT (p≤0.05; fold-change ≥2). Proteomic profiling between DSRCT and normal mesenteric tissue specimens. The protein lysates from DSRCT and normal mesenteric tissue were subjected to RPPA analysis for 149 proteins and phosphoproteins. Unsupervised double-hierarchical clustering using the Pearson correlation distance metric between proteins (rows) and Centroid linkage (a clustering method) separated the 16 samples into two groups (columns). The heat map of all 16 samples showed 20 proteins significantly distinguishing the proteomic profile of DSRCT from that of normal mesenteric tissue (p ≤0.05; fold-change ≥2).