UNASSIGNED: 432 Background: Novel agents that inhibit kinases in the @entity519 and @entity19213 pathways can achieve response rates of 30-60% in @entity1358 ( @entity1358 ); however, protein biomarkers to diagnose @entity1358 and guide treatment have not been identified. METHODS: We developed the use of two high-throughput, nanoscale immunoassays to profile @entity519 and downstream signaling in clinical specimens from @entity1 with @entity1358 . These automated technologies minimize errors due to manual variability, and make 96 independent measurements per overnight experiment. We used size-based protein separation (Sally Instrument, Protein Simple) to quantify proteins using antibody-specific detection, normalized to loading control. Next, we used a nano-immunoassay with charge-based separation (NIA, Nanopro1000 Instrument, Protein Simple) to distinguish multiple charged modifications of individual proteins, and measure relative ratios of individual unphosphorylated and phosphorylated isoforms. RESULTS: We first optimized assays for proteins in MAPK, @entity10412 and STAT pathways, and loading controls, to analyze frozen surgical specimens and ex-vivo fine needle aspirates (FNA's) performed immediately following nephrectomy. We analyzed more than 200 FNA's from solid @entity5 , comparing @entity1358 with paired adjacent non- @entity5 tissue, and other @entity5 . Basal MAPK signaling of @entity5 varied across 3 logs of expression. Unique to NIA, we analyzed percent phosphorylation and resolved differences in single phosphorylations. Subtle differences in basal MAPK signaling between @entity5 calls and adjacent non- @entity5 tissue can distinguish @entity1358 from other @entity5 . Lastly, we prospectively collected and analyzed blood peripheral mononuclear cells from 20 @entity1358 @entity1 before and during treatment with standard targeted therapies. Kinase inhibitors can preferentially inhibit or activate specific protein phospho-isoforms. CONCLUSIONS: Our studies demonstrate that rapid and quantitative nanoproteomic profiling in very small amounts of clinical specimen may accelerate translational studies for novel diagnostic and predictive biomarkers.

Nanoscale proteomic profiling to define diagnostic signatures and biomarkers of therapeutic activity in @entity1 with XXXX .

@entity1 :: ('9606', 'Species') :: ['patients'] @entity5 :: ('MESH:D009369', 'Disease') :: ['epithelial malignancies', 'tumor', 'malignancies', 'tumors'] @entity1358 :: ('MESH:D002292', 'Disease') :: ['renal cell carcinoma', 'RCC'] @entity519 :: ('MESH:D000860', 'Disease') :: ['hypoxia'] @entity10412 :: ('5293', 'Gene') :: ['PI3K'] @entity19213 :: ('2475', 'Gene') :: ['MTOR']
@entity1358 :: (MESH:D002292,Disease) :: ['RCC']
