Direct molecular dissection of tumor parenchyma from tumor stroma in tumor xenograft using mass spectrometry-based glycoproteomics

// Xiaoying Ye 1 , Brian T. Luke 2 , Bih-Rong Wei 3 , Jan A. Kaczmarczyk 4 , Jadranka Loncarek 5 , Jennifer E. Dwyer 3 , Donald J. Johann 6 , Richard G. Saul 4 , Dwight V. Nissley 1 , Frank McCormick 7 , Gordon R. Whiteley 1 and Josip Blonder 1 1 National Cancer Institute RAS Initiative, Cancer Research Technology Program, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, MD 21702, USA 2 Advanced Biomedical Computing Center, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, MD 21702, USA 3 Laboratory of Cancer Biology and Genetics, Center for Cancer Research, National Cancer Institute, Bethesda, MD 20892, USA 4 Cancer Research Technology Program, Antibody Characterization Laboratory, Frederick National Laboratory for Cancer Research, Leidos Biomedical Research, Inc., Frederick, MD 21702, USA 5 Laboratory of Protein Dynamics and Signaling, Center for Cancer Research, National Cancer Institute, Frederick, MD 21702, USA 6 Winthrop P. Rockefeller Cancer Institute, University of Arkansas for Medical Sciences, Little Rock, AR 72209, USA 7 UCSF Helen Diller Family Comprehensive Cancer Center, San Francisco, CA 94158, USA Correspondence to: Josip Blonder, email: blonderj@mail.nih.gov Keywords: lung cancer; tumor microenvironment Received: March 22, 2018     Accepted: May 02, 2018     Published: May 29, 2018 ABSTRACT The most widely used cancer animal model is the human-murine tumor xenograft. Unbiased molecular dissection of tumor parenchyma versus stroma in human-murine xenografts is critical for elucidating dysregulated protein networks/pathways and developing therapeutics that may target these two functionally codependent compartments. Although antibody-reliant technologies (e.g., immunohistochemistry, imaging mass cytometry) are capable of distinguishing tumor-proper versus stromal proteins, the breadth or extent of targets is limited. Here, we report an antibody-free targeted cross-species glycoproteomic (TCSG) approach that enables direct dissection of human tumor parenchyma from murine tumor stroma at the molecular/protein level in tumor xenografts at a selectivity rate presently unattainable by other means. This approach was used to segment/dissect and obtain the protein complement phenotype of the tumor stroma and parenchyma of the metastatic human lung adenocarcinoma A549 xenograft, with no need for tissue microdissection prior to mass-spectrometry analysis. An extensive molecular map of the tumor proper and the associated microenvironment was generated along with the top functional N-glycosylated protein networks enriched in each compartment. Importantly, immunohistochemistry-based cross-validation of selected parenchymal and stromal targets applied on human tissue samples of lung adenocarcinoma and normal adjacent tissue is indicative of a noteworthy translational capacity for this unique approach that may facilitate identifications of novel targets for next generation antibody therapies and development of real time preclinical tumor models.