Browsing by Author "Robles, Ana I."

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    Frozen tissue coring and layered histological analysis improves cell type-specific proteogenomic characterization of pancreatic adenocarcinoma
    (2024-01-30) Savage, Sara R.; Wang, Yuefan; Chen, Lijun; Jewell, Scott; Newton, Chelsea; Dou, Yongchao; Li, Qing K.; Bathe, Oliver F.; Robles, Ana I.; Omenn, Gilbert S.; Thiagarajan, Mathangi; Zhang, Hui; Hostetter, Galen; Zhang, Bing
    Abstract Background Omics characterization of pancreatic adenocarcinoma tissue is complicated by the highly heterogeneous and mixed populations of cells. We evaluate the feasibility and potential benefit of using a coring method to enrich specific regions from bulk tissue and then perform proteogenomic analyses. Methods We used the Biopsy Trifecta Extraction (BioTExt) technique to isolate cores of epithelial-enriched and stroma-enriched tissue from pancreatic tumor and adjacent tissue blocks. Histology was assessed at multiple depths throughout each core. DNA sequencing, RNA sequencing, and proteomics were performed on the cored and bulk tissue samples. Supervised and unsupervised analyses were performed based on integrated molecular and histology data. Results Tissue cores had mixed cell composition at varying depths throughout. Average cell type percentages assessed by histology throughout the core were better associated with KRAS variant allele frequencies than standard histology assessment of the cut surface. Clustering based on serial histology data separated the cores into three groups with enrichment of neoplastic epithelium, stroma, and acinar cells, respectively. Using this classification, tumor overexpressed proteins identified in bulk tissue analysis were assigned into epithelial- or stroma-specific categories, which revealed novel epithelial-specific tumor overexpressed proteins. Conclusions Our study demonstrates the feasibility of multi-omics data generation from tissue cores, the necessity of interval H&E stains in serial histology sections, and the utility of coring to improve analysis over bulk tissue data.
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    Neoplastic cell enrichment of tumor tissues using coring and laser microdissection for proteomic and genomic analyses of pancreatic ductal adenocarcinoma
    (2022-10-20) Li, Qing K.; Hu, Yingwei; Chen, Lijun; Schnaubelt, Michael; Cui Zhou, Daniel; Li, Yize; Lu, Rita Jui-Hsien; Thiagarajan, Mathangi; Hostetter, Galen; Newton, Chelsea J.; Jewell, Scott D.; Omenn, Gil; Robles, Ana I.; Mesri, Mehdi; Bathe, Oliver F.; Zhang, Bing; Ding, Li; Hruban, Ralph H.; Chan, Daniel W.; Zhang, Hui
    Abstract Background The identification of differentially expressed tumor-associated proteins and genomic alterations driving neoplasia is critical in the development of clinical assays to detect cancers and forms the foundation for understanding cancer biology. One of the challenges in the analysis of pancreatic ductal adenocarcinoma (PDAC) is the low neoplastic cellularity and heterogeneous composition of bulk tumors. To enrich neoplastic cells from bulk tumor tissue, coring, and laser microdissection (LMD) sampling techniques have been employed. In this study, we assessed the protein and KRAS mutation changes associated with samples obtained by these enrichment techniques and evaluated the fraction of neoplastic cells in PDAC for proteomic and genomic analyses. Methods Three fresh frozen PDAC tumors and their tumor-matched normal adjacent tissues (NATs) were obtained from three sampling techniques using bulk, coring, and LMD; and analyzed by TMT-based quantitative proteomics. The protein profiles and characterizations of differentially expressed proteins in three sampling groups were determined. These three PDACs and samples of five additional PDACs obtained by the same three sampling techniques were also subjected to genomic analysis to characterize KRAS mutations. Results The neoplastic cellularity of eight PDACs ranged from less than 10% to over 80% based on morphological review. Distinctive proteomic patterns and abundances of certain tumor-associated proteins were revealed when comparing the tumors and NATs by different sampling techniques. Coring and bulk tissues had comparable proteome profiles, while LMD samples had the most distinct proteome composition compared to bulk tissues. Further genomic analysis of bulk, cored, or LMD samples demonstrated that KRAS mutations were significantly enriched in LMD samples while coring was less effective in enriching for KRAS mutations when bulk tissues contained a relatively low neoplastic cellularity. Conclusions In addition to bulk tissues, samples from LMD and coring techniques can be used for proteogenomic studies. The greatest enrichment of neoplastic cellularity is obtained with the LMD technique.