The National Cancer Institute Clinical Proteomic Atlas Consortium (CPTAC) herein reports our deep characterization of 228 grade IV IDH1 WT and mutant astrocytomas (including 28 matched primary and recurrent GBMs) using 15 proteogenomic and metabolomic platforms. Major advances over our first CPTAC GBM report (Wang et al., 2021, Cancer Cell), are the inclusion of many more samples including paired primary and recurrent tumors, application of new platforms, including glycoproteomics and targeted mass spectrometry methods, development and application of new computational techniques, and the integration of an atlas of experimentally determined, functional, kinase substrate interactions from Kinase Library. Paired primary-recurrent GBM analyses showed increased clonal diversity in recurrent tumors as a function of time, and treatment-induced mutation signatures. Proteomic and metabolomic analyses showed that different drivers can cause similar downstream effects. Only EGFR altered tumors were associated with dual EGFR glycosylation (N352 and N603) and EGFR phosphorylation (Y316) events. IDH1 mutation was associated with activated RTK signaling and decreased hypoxia pathway activities, concordant with epigenetic and metabolic profiles. Protein-protein interaction and kinase/phosphatase-substrate analyses uncovered detailed signaling events from different upstream drivers (e.g., EGFR, PDGFRA, and IDH1) converged through a PTPN11 hub to downstream effectors, including GAB1, IRS1, MAP3K5, and PTK2B. In summary, this multiscale resource presents new and deeper biological insights regarding treatment impact on tumor evolution, shared downstream consequences of independent drivers, and the potential importance of PTPN11 signaling circuitry across high-grade gliomas. We hope that reporting this new international resource to the SNO community will advance therapeutic development, including targeted therapies that may avoid known mechanisms of resistance.

Both the induction of SPARC expression and the loss of the p53 tumor suppressor gene are changes that occur early in glioma development. Both SPARC and p53 regulate glioma cell survival by inverse effects on apoptotic signaling. Therefore, during glioma formation, the upregulation of SPARC may cooperate with the loss of p53 to enhance cell survival. This study determined whether the loss of Sparc in astrocytes that are null for p53 would result in reduced cell survival and tumor formation and increased tumor immunogenicity in an in vivo xenograft brain tumor model. In vitro, the loss of Sparc in p53-null astrocytes resulted in an increase in cell proliferation, but a loss of tumorigenicity. At 7 days after intracranial implantation, Sparc-null tumors had decreased tumor cell survival, proliferation and reduced tumor size. The loss of Sparc promoted microglia/macrophage activation and phagocytosis of tumor cells. Our results indicate that the loss of p53 by deletion/mutation in the early stages of glioma formation may cooperate with the induction of SPARC to potentiate cancer cell survival and escape from immune surveillance.

Abstract Both the induction of SPARC expression and the loss of the p53 tumor suppressor gene are changes that occur early in glioma development. Therefore, the upregulation of SPARC may cooperate with the loss of p53 to enhance cell survival and inhibit apoptosis during glioma formation. This study determined whether the loss of Sparc in astrocytes that are null for p53 (p53-null/Sparc-null) would result in reduced cell survival and tumor formation and increased tumor immunogenicity in an in vivo xenograft brain tumor model. In vitro, the loss of Sparc in p53-null astrocytes resulted in an increase in cell proliferation (15-33%, p<0.01); however, there was an inhibition of growth in soft agar. Intracranial xenografts of p53-null/Sparc-wt and p53-null/Sparc-null astrocytes were assessed for tumor size, proliferation rate, and SPARC expression. At 7 days post-implantation, Sparc-null astrocytes produced significantly smaller tumors (Wilcoxon rank-sum test p = 0.0091, median = 0.709mm2 for Sparc-wt and 0.240mm2 for Sparc-null) with a significantly lower MIB-1 proliferation index (Wilcoxon rank-sum test p = 0.0345, median = 8.6% for Sparc-wt and 0.4% for Sparc-null). By CD68 and periodic acid Schiff +/- diastase staining of xenograft tumors, it was found that Sparc-null tumors had a massive infiltration of microglia/macrophages with a phagocytic appearance compared to the activated, but non-phagocytic, microglia/macrophages present within the Sparc-wt tumors. The loss of Sparc in astrocytes and the resulting increase in microglia/macrophage activation lead to an alteration in the tumor microenvironment with increased collagen deposition and altered collagen structure at both 7 and 50 days post-implantation as assessed by picrosirius red and polarized light microscopy. Sparc-null tumors had increased collagen deposition with a long fiber structure compared to the small bundles of collagen present in Sparc-wt tumors. Our results indicate that the loss of p53 by deletion/mutation in the early stages of glioma formation may cooperate with the induction of SPARC to potentiate cancer cell survival and escape from immune surveillance. Citation Format: Stacey L. Thomas, Chad R. Schultz, Ezekiell Mouzon, William A. Golembieski, Nancy Lemke, Laila M. Poisson, Jorge A. Gutierrez, Sandra Cottingham, Sandra A. Rempel. Loss of Sparc in p53-null astrocytes alters collagen deposition and promotes macrophage activation and tumor phagocytosis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2363. doi:10.1158/1538-7445.AM2015-2363