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1.
Sci Rep ; 7(1): 10060, 2017 08 30.
Article in English | MEDLINE | ID: mdl-28855593

ABSTRACT

Pancreatic ductal adenocarcinoma (PDAC) has single-digit 5-year survival rates at <7%. There is a dire need to improve pre-malignant detection methods and identify new therapeutic targets for abrogating PDAC progression. To this end, we mined our previously published pseudopodium-enriched (PDE) protein/phosphoprotein datasets to identify novel PDAC-specific biomarkers and/or therapeutic targets. We discovered that integrin alpha 1 (ITGA1) is frequently upregulated in pancreatic cancers and associated precursor lesions. Expression of ITGA1-specific collagens within the pancreatic cancer microenvironment significantly correlates with indicators of poor patient prognosis, and depleting ITGA1 from PDAC cells revealed that it is required for collagen-induced tumorigenic potential. Notably, collagen/ITGA1 signaling promotes the survival of ALDH1-positive stem-like cells and cooperates with TGFß to drive gemcitabine resistance. Finally, we report that ITGA1 is required for TGFß/collagen-induced EMT and metastasis. Our data suggest that ITGA1 is a new diagnostic biomarker and target that can be leveraged to improve patient outcomes.


Subject(s)
Adenocarcinoma/genetics , Biomarkers, Tumor/genetics , Drug Resistance, Neoplasm/genetics , Gene Expression Regulation, Neoplastic , Integrin alpha Chains/genetics , Pancreatic Neoplasms/genetics , Adenocarcinoma/diagnosis , Adenocarcinoma/metabolism , Adenocarcinoma/pathology , Aldehyde Dehydrogenase 1 Family , Animals , Antimetabolites, Antineoplastic/pharmacology , Biomarkers, Tumor/antagonists & inhibitors , Biomarkers, Tumor/metabolism , Cell Line, Tumor , Cell Movement/drug effects , Cell Proliferation/drug effects , Chick Embryo , Chorioallantoic Membrane/blood supply , Chorioallantoic Membrane/drug effects , Collagen/genetics , Collagen/metabolism , Deoxycytidine/analogs & derivatives , Deoxycytidine/pharmacology , Epithelial-Mesenchymal Transition , Humans , Integrin alpha Chains/antagonists & inhibitors , Integrin alpha Chains/metabolism , Isoenzymes/genetics , Isoenzymes/metabolism , Pancreatic Neoplasms/diagnosis , Pancreatic Neoplasms/metabolism , Pancreatic Neoplasms/pathology , Prognosis , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Retinal Dehydrogenase/genetics , Retinal Dehydrogenase/metabolism , Signal Transduction , Tissue Array Analysis , Transforming Growth Factor beta/pharmacology , Tumor Microenvironment/genetics , Gemcitabine
2.
Oncotarget ; 8(4): 5885-5894, 2017 Jan 24.
Article in English | MEDLINE | ID: mdl-27602776

ABSTRACT

Next-generation sequencing (NGS) can identify and validate new biomarkers of cancer onset, progression and therapy resistance. Substantial archives of formalin-fixed, paraffin-embedded (FFPE) cancer samples from patients represent a rich resource for linking molecular signatures to clinical data. However, performing NGS on FFPE samples is limited by poor RNA purification methods. To address this hurdle, we developed an improved methodology for extracting high-quality RNA from FFPE samples. By briefly integrating a newly-designed micro-homogenizing (mH) tool with commercially available FFPE RNA extraction protocols, RNA recovery is increased by approximately 3-fold while maintaining standard A260/A280 ratios and RNA quality index (RQI) values. Furthermore, we demonstrate that the mH-purified FFPE RNAs are longer and of higher integrity. Previous studies have suggested that pancreatic ductal adenocarcinoma (PDAC) gene expression signatures vary significantly under in vitro versus in vivo and in vivo subcutaneous versus orthotopic conditions. By using our improved mH-based method, we were able to preserve established expression patterns of KRas-dependency genes within these three unique microenvironments. Finally, expression analysis of novel biomarkers in KRas mutant PDAC samples revealed that PEAK1 decreases and MST1R increases by over 100-fold in orthotopic versus subcutaneous microenvironments. Interestingly, however, only PEAK1 levels remain elevated in orthotopically grown KRas wild-type PDAC cells. These results demonstrate the critical nature of the orthotopic tumor microenvironment when evaluating the clinical relevance of new biomarkers in cells or patient-derived samples. Furthermore, this new mH-based FFPE RNA extraction method has the potential to enhance and expand future FFPE-RNA-NGS cancer biomarker studies.


Subject(s)
Carcinoma, Pancreatic Ductal/genetics , Neoplasm Transplantation/methods , Pancreatic Neoplasms/genetics , RNA/isolation & purification , Animals , Biomarkers, Tumor/genetics , Genetic Heterogeneity , Heterografts , High-Throughput Nucleotide Sequencing , Humans , Mice , Paraffin Embedding , Sequence Analysis, RNA , Tissue Fixation
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