For PyMT experiments, syngeneic PyMT breast tumor cells were implanted into the fourth mammary fat pads of female FVB mice at four to five weeks of age
For PyMT experiments, syngeneic PyMT breast tumor cells were implanted into the fourth mammary fat pads of female FVB mice at four to five weeks of age. sufficient to further sensitize angiogenic blockade as TAM would compensate for their lack and vice versa leading to an oscillating pattern of distinct immune cell populations. However, PI3K inhibition in CD11b+ myeloid cells generated an enduring angiostatic and immune-stimulatory environment in which anti-angiogenic therapy remained efficient. Graphical Abstract INTRODUCTION Antiangiogenic therapy represents one of the most widely used anti-cancer strategies today, with most approved therapies targeting the vascular endothelial growth factor (VEGF) signaling pathway. However, the beneficial effects observed across the multitude of cancers that respond are typically short-lived; therefore much effort has focused on uncovering the various mechanisms whereby tumors bypass the tumor-inhibitory effects of therapy (Bergers and Hanahan, 2008; Kerbel, 2008). One such resistance mechanism Regadenoson entails reinstatement of angiogenesis by tumor-infiltrating innate immune cells (Dierickx et al., 1963; Fischer et al., 2007; Shojaei et al., 2007a; Shojaei et al., 2007b). Tumors can contain a significant percentage of different infiltrating myeloid cells with bivalent functions but predominantly are thought to support tumor progression by promoting angiogenesis and suppressing anti-tumor immunity. Tumor-associated macrophages (TAM) are typically characterized as either classically activated tumoricidal macrophages (M1) or alternatively activated protumorigenic macrophages (M2) (Mantovani et al., 2008). Extending upon this nomenclature, neutrophils (TAN) have also been categorized as N1 or N2 based on their anti-or pro-tumor activity in tumors (Fridlender et al., 2009). In addition, immature Gr1+ cells with either a mononuclear or granular morphology have been recognized in tumors that convey immune-suppressive functions and are therefore also termed myeloid-derived suppressor cells (M-MDSC and G-MDSC respectively) (Talmadge and Gabrilovich, 2013). Typically, surface marker profiling based on expression of CD11b, F4/80, Gr1, Ly6C, and Ly6G is used to categorize these subsets of tumor-infiltrating myeloid cells (Fridlender et al., 2009; Talmadge and Gabrilovich, 2013; Wynn et al., 2013). There is mounting evidence that tumors recruit these unique populations where they become an additional source of angiogenic chemokines and cytokines to promote angiogenesis (Coussens Regadenoson et al., 2000; Du et al., 2008; Giraudo et al., 2004; Lin et al., 2006; Shojaei et al., 2007b). As hypoxia is usually a major driver of myeloid cell recruitment (Du et al., 2008; Mazzieri et al., 2011) it is conceivable that therapy-induced hypoxia via an angiogenic blockade can induce factors that mobilize cells from your bone marrow and attract them to the tumor site. Indeed, tumor-associated myeloid cells have been shown to sustain angiogenesis in the face of antiangiogenic therapy, in part by stimulating VEGF-independent pathways. For example, macrophages induced expression of several angiogenic molecules, including and in response to antiangiogenic therapy (Casanovas et al., 2005; Fischer et al., 2007; Rigamonti et al., 2014), while Gr1+ myeloid cells were found to convey resistance to anti-VEGF treatment via secretion of the angiogenic PKR-1/2 ligand Bv8 (Shojaei et al., 2007a; Shojaei Regadenoson et al., 2007b). As much as inhibitors of macrophages or Gr1+ cells enhanced the effects of antiangiogenic therapy, in many of these models tumor growth was still apparent at a slower pace throughout the duration of treatment. Here, we investigated the overall contributions of the different tumor-associated myeloid populations to evasion of antiangiogenic therapy. We analyzed the composition and function of TAM, TAN, and two Gr1+ immature monocyte populations in two distinct tumor models that responded differently to angiogenic inhibition. In the Rip1Tag2 model of pancreatic neuroendocrine tumors (PNET), angiogenic blockade was able to transiently reduce vessel density and block tumor growth (response) followed by reinstatement of neovascularization and robust tumor growth (relapse) thereby enabling us to evaluate true response and relapse phases in a single model. In the PyMT mammary carcinoma model, angiogenic blockade was only able to slow down tumor growth with some reduction in vessel density, a feature that is commonly observed in various tumor models. Analysis of myeloid cell content within tumors revealed that the angiogenic relapse was associated with an increase in tumor-specific subsets of Gr1+ myeloid cells. By investigating the role of these cells during relapse, we were able to Regadenoson uncover a compensatory nature of myeloid cell-mediated resistance to antiangiogenic therapy. In the present study, we inquired about the nature and mechanisms by which distinct innate immune cells compensate for each other to maintain resistance and identify means that modulate inflammation to sustain the Adcy4 effects of antiangiogenic therapy. RESULTS Targeting distinct myeloid subtypes leads to a compensatory oscillation between innate immune cells enabling reneovascularization during antiangiogenic.