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.
Highlights
Adherent two-dimensionally (2D) cultured human cancer cells are a widely used model system in cancer research and drug screening [1]
In human-murine tumor xenografts, the parenchymal proteome is a product of the human neoplastic tumor cells, while the stromal proteome is principally a product of the host’s tumor microenvironment [25]
Based on the critical role played by protein glycosylation in cancer biology [26] and self/nonself immune recognition [27], we hypothesized that the difference and/or dissimilarity rate between the human and mouse N-glycopeptidome/proteome ought to be substantially larger than the actual dissimilarity rate between the corresponding ordinary human/mouse mass spectrometry (MS)-identifiable unmodified peptidome/proteome
Summary
Adherent two-dimensionally (2D) cultured human cancer cells are a widely used model system in cancer research and drug screening [1]. Speaking, tumor xenografts have a microanatomic configuration analogous to common solid tumors These are characterized by the existence of two morphologically well-defined and functionally interdependent compartments: i) the parenchyma, comprising neoplastic tumor cells of diverging morphology, antigenicity, and metastatic capacity, and ii) the stroma, comprising different cellular elements including tumor fibroblasts, endothelial cells, and immune cells [4]. Parenchymal neoplastic tumor cells secrete cytokines, growth factors, and proteolytic enzymes to sustain their survival and induce and/or modify the tumor stroma formation [6]. In this regard, any solid tumor bigger than two millimeters in diameter must induce its own blood supply, since it cannot survive without a stroma that provides a vascular network for nutrient supply and waste removal [7]
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