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Supplemental Figure 1. CD11b+ myeloid cells in glioma correlate to tumor grade. Supplemental Figure 2. Schematic depicted for bone marrow transplant of RCAS tumor bearing animals. Supplemental Figure 3. Analysis of tumor sections for necrosis and microvascular proliferation. Supplemental Figure 4. AZD1480 impairs phosphorylation of Stat3 but has no effect in vitro and IDH status

Purpose: While the tumor microenvironment has been known to play an integral role in tumor progression, the function of nonresident bone marrow–derived cells (BMDC) remains to be determined in neurologic tumors. Here we identified the contribution of BMDC recruitment in mediating malignant transformation from low- to high-grade gliomas.Experimental Design: We analyzed human blood and tumor samples from patients with low- and high-grade gliomas. A spontaneous platelet-derived growth factor (PDGF) murine glioma model (RCAS) was utilized to recapitulate human disease progression. Levels of CD11b+/GR1+ BMDCs were analyzed at discrete stages of tumor progression. Using bone marrow transplantation, we determined the unique influence of BMDCs in the transition from low- to high-grade glioma. The functional role of these BMDCs was then examined using a JAK 1/2 inhibitor (AZD1480).Results: CD11b+ myeloid cells were significantly increased during tumor progression in peripheral blood and tumors of glioma patients. Increases in CD11b+/GR1+ cells were observed in murine peripheral blood, bone marrow, and tumors during low-grade to high-grade transformation. Transient blockade of CD11b+ cell expansion using a JAK 1/2 Inhibitor (AZD1480) impaired mobilization of these cells and was associated with a reduction in tumor volume, maintenance of a low-grade tumor phenotype, and prolongation in survival.Conclusions: We demonstrate that impaired recruitment of CD11b+ myeloid cells with a JAK1/2 inhibitor inhibits glioma progression in vivo and prolongs survival in a murine glioma model. Clin Cancer Res; 23(12); 3109–19. ©2016 AACR.

Purpose:KRASG12C is the most common KRAS mutation in primary lung adenocarcinoma. Phase I clinical trials have demonstrated encouraging clinical activity of KRASG12C inhibitors in the metastatic setting. We investigated disease-free survival (DFS) and tumor genomic features in patients with surgically resected KRASG12C-mutant lung adenocarcinoma.Experimental Design:Patients who underwent resection of stage I–III lung adenocarcinoma and next-generation sequencing (NGS) were evaluated. Exclusion criteria were receipt of induction therapy, incomplete resection, and low-quality NGS. Mutations were classified as KRAS wild-type (KRASwt), G12C (KRASG12C), or non-G12C (KRASother). DFS was compared between groups using the log-rank test; factors associated with DFS were assessed using Cox regression. Mutual exclusivity and cooccurrence, tumor clonality, and mutational signatures were assessed.Results:In total, 604 patients were included: 374 KRASwt (62%), 95 KRASG12C (16%), and 135 KRASother (22%). Three-year DFS was not different between KRAS-mutant and KRASwt tumors. However, 3-year DFS was worse in patients with KRASG12C than KRASother tumors (log-rank P = 0.029). KRASG12C tumors had more lymphovascular invasion (51% vs. 37%; P = 0.032) and higher tumor mutation burden [median (interquartile range), 7.0 (5.3–10.8) vs. 6.1 (3.5–9.7); P = 0.021], compared with KRASother tumors. KRASG12C mutation was independently associated with worse DFS on multivariable analysis. Our DFS findings were externally validated in an independent The Cancer Genome Atlas cohort.Conclusions:KRASG12C mutations are associated with worse DFS after complete resection of stage I–III lung adenocarcinoma. These tumors harbor more aggressive clinicopathologic and genomic features than other KRAS-mutant tumors. We identified a high-risk group for whom KRASG12C inhibitors may be investigated to improve survival.

PDF - 9162K, Supplementary Methods: Including additional materials and methods Supplementary References: Including reference cited in the supplementary methods Supplementary Figures and Legends: S1. expression of stromal PDGFRalpha in various animal models. S2. Lineage and distribution of stromal PDGFRalpha+ cells. S3. Blood vessel density and pericytes in Gl261 glioma model. S4. Knocking out PDGFRalpha suppresses infiltration of BMDCs without affecting hematopoiesis. S5. Isolation and verification of OPCs from glioma S6. Endothelial tubule formation, endothelial sprouting, and permeability with GA-OPCs and astrocytes. S7. Proliferation of glioma cells with or without activated endothelial cells S8. Microarray analysis on GFP+/F4/80+ BMDCs S9. Hematopoiesis in PDGF-C -/- mice. S10. Astrocytes and endothelial cells express PDGF-C & pro-angiogenic profile of OPCs. S11. Blood vessel staining of grade III Patients S12. Expression of PDGF-C was analyzed in REMBRANDT database. S13. the correlation of PDGF-C with paitents' survial in Mesenchymal, Neural, Pro-neural subsets of GBM. S14. The correlation of PDGF-A and PDGF-B with paitent's survial in classical subset of GBM.

Enhanced platelet-derived growth factor (PDGF) signaling in glioma drives its development and progression. In this study, we define a unique role for stroma-derived PDGF signaling in maintaining tumor homeostasis within the glioma microenvironment. Large numbers of PDGF receptor-α (PDGFRα)–expressing stromal cells derived from oligodendrocytes progenitor cells (OPC) were discovered at the invasive front of high-grade gliomas, in which they exhibited a unique perivascular distribution. In PDGFRα-deficient host mice, in which orthotopic Gl261 tumors displayed reduced outgrowth, we found that tumor-associated blood vessels displayed smaller lumens and normalized vascular morphology, with tumors in host animals injected with the vascular imaging agent gadolinium also being enhanced less avidly by MRI. Notably, glioma-associated OPC promoted endothelial sprouting and tubule formation, in part by abrogating the inhibitory effect that perivascular astrocytes exert on vascular endothelial conjunctions. Stromal-derived PDGF-CC was crucial for the recruitment and activation of OPC, insofar as mice genetically deficient in PDGF-CC phenocopied the glioma/vascular defects observed in PDGFRα-deficient mice. Clinically, we showed that higher levels of PDGF-CC in glioma specimens were associated with more rapid disease recurrence and poorer overall survival. Our findings define a PDGFRα/PDGF-CC signaling axis within the glioma stromal microenvironment that contributes to vascular remodeling and aberrant tumor angiogenesis in the brain. Cancer Res; 74(4); 1011–21. ©2013 AACR.

PDF - 93K, Authentication of Gl261 cells in related to "cell lines and mice lines" section in Materials and Methods.

The PI3K/AKT/mTOR pathway is aberrantly activated in many pediatric solid tumors including gliomas and medulloblastomas. Preclinical data in a pediatric glioma model demonstrated that the combination of perifosine (AKT inhibitor) and temsirolimus (mTOR inhibitor) is more potent at inhibiting the axis than either agent alone. We conducted this study to assess pharmacokinetics and identify the maximum tolerated dose for the combination.We performed a standard 3+3 phase I, open-label, dose-escalation study in patients with recurrent/refractory pediatric solid tumors. Four dose levels of perifosine (25-75 mg/mTwenty-three patients (median age 8.5 years) with brain tumors (diffuse intrinsic pontine glioma [DIPG] n = 8, high-grade glioma n = 6, medulloblastoma n = 2, ependymoma n = 1), neuroblastoma (n = 4), or rhabdomyosarcoma (n = 2) were treated. The combination was generally well tolerated and no dose-limiting toxicity was encountered. The most common grade 3 or 4 toxicities (at least possibly related) were thrombocytopenia (38.1%), neutropenia (23.8%), lymphopenia (23.8%), and hypercholesterolemia (19.0%). Pharmacokinetic findings for temsirolimus were similar to those observed in the temsirolimus single-agent phase II pediatric study and pharmacokinetic findings for perifosine were similar to those in adults. Stable disease was seen in 9 of 11 subjects with DIPG or high-grade glioma; no partial or complete responses were achieved.The combination of these AKT and mTOR inhibitors was safe and feasible in patients with recurrent/refractory pediatric solid tumors.

Exosomes mediate intercellular communication and regulation in many processes, including cancer-immune cell interactions. Exosomes are small (~100 nm) membrane vesicles that arise from the late endosomal pathway and contain bioactive molecules that are transferred to and affect the function of target cells. Tumor-derived exosomes are important regulators of anti-tumor immunity and have been shown to exert stimulatory and inhibitory effects on immune cells, depending on the experimental setting. Using a proteomic approach, we found that exosomes derived from tumors that undergo immune rejection (“regressors”) contain different proteins than exosomes from progressively growing tumors (“progressors”). Specifically, the signaling molecule MAP2K1 (“MEK1”) is highly enriched in regressor compared to progressor exosomes, leading us to hypothesize that exosomal MEK1 stimulates anti-tumor immunity and inhibits tumor progression. We found that forced expression of MEK1 in progressor exosomes delays tumor growth and increases anti-tumor immune cell infiltration, and that MEK1-hi exosomes are sufficient to delay tumor growth, but MEK1-low exosomes are not. Moreover, we found that MEK1 protein can be physically transferred from tumor cells to macrophages via exosomes, and that MEK1-hi exosomes induce pro-inflammatory cytokine production in macrophages, but MEK1-low exosomes do not. We propose that tumor-derived MEK1 is transferred to macrophages via exosomes, which enhances their anti-tumoricidal effector function and delays tumor growth. This cell extrinsic, anti-tumor activity of MEK1 contrasts with the known oncogenic properties of MEK1 and could limit the efficacy of MEK1 inhibitors currently in clinical trials.