Daniela F. Quail

Microenvironmental regulation of tumor progression and metastasis

Cancers develop in complex tissue environments, which they depend on for sustained growth, invasion and metastasis. Unlike tumor cells, stromal cell types within the tumor microenvironment (TME) are genetically stable and thus represent an attractive therapeutic target with reduced risk of resistance and tumor recurrence. However, specifically disrupting the pro-tumorigenic TME is a challenging undertaking, as the TME has diverse capacities to induce both beneficial and adverse consequences for tumorigenesis. Furthermore, many studies have shown that the microenvironment is capable of normalizing tumor cells, suggesting that re-education of stromal cells, rather than targeted ablation per se, may be an effective strategy for treating cancer. Here we discuss the paradoxical roles of the TME during specific stages of cancer progression and metastasis, as well as recent therapeutic attempts to re-educate stromal cells within the TME to have anti-tumorigenic effects.

CSF-1R inhibition alters macrophage polarization and blocks glioma progression

Glioblastoma multiforme (GBM) comprises several molecular subtypes, including proneural GBM. Most therapeutic approaches targeting glioma cells have failed. An alternative strategy is to target cells in the glioma microenvironment, such as tumor-associated macrophages and microglia (TAMs). Macrophages depend on colony stimulating factor-1 (CSF-1) for differentiation and survival. We used an inhibitor of the CSF-1 receptor (CSF-1R) to target TAMs in a mouse proneural GBM model, which significantly increased survival and regressed established tumors. CSF-1R blockade additionally slowed intracranial growth of patient-derived glioma xenografts. Surprisingly, TAMs were not depleted in treated mice. Instead, glioma-secreted factors, including granulocyte-macrophage CSF (GM-CSF) and interferon-γ (IFN-γ), facilitated TAM survival in the context of CSF-1R inhibition. Expression of alternatively activated M2 markers decreased in surviving TAMs, which is consistent with impaired tumor-promoting functions. These gene signatures were associated with enhanced survival in patients with proneural GBM. Our results identify TAMs as a promising therapeutic target for proneural gliomas and establish the translational potential of CSF-1R inhibition for GBM.

The tumor microenvironment underlies acquired resistance to CSF-1R inhibition in gliomas

Another pathway to cancer resistance Therapies targeting the tumor microenvironment show promise for treating cancer. For example, antibodies targeting colony-stimulating factor-1 receptor (CSF-1R) inhibit protumorigenic macrophages and regress tumors in mouse models of glioblastoma multiforme (GBM), a deadly form of brain cancer. Quail et al. found that although CSR-1R blockade prolonged survival in mouse models of GBM, more than 50% of tumors eventually recurred. Recurrence was correlated with elevated PI3-K activity in tumors, driven by macrophage-secreted IGF-1. Blocking PI3-K and IGF-1 signaling in rebounding tumors prolonged survival. Thus, tumors can acquire resistance to therapy through intrinsic changes and through changes in their microenvironment. Science, this issue p. 10.1126/science.aad3018 Brain tumors can acquire resistance to therapy through changes to their microenvironment. INTRODUCTION Therapies targeted against the tumor microenvironment (TME) represent a promising approach for treating cancer. This appeal arises in part from the decreased likelihood of acquired resistance through mutations in target TME cells, as is frequently observed with cancer cell–targeted therapies. Although classical mechanisms of tumor cell–intrinsic resistance to cytotoxic and targeted agents have been well-defined—including aberrant drug metabolism and transport, drug target mutation, and activation of alternative survival pathways—it still remains unclear whether resistance to TME-directed therapies follows similar principles. Given that TME-targeted agents are increasingly being evaluated in the clinic, it is becoming critical to mechanistically define how resistance may evolve in response to these therapies in order to provide long-term disease management for patients. RATIONALE Macrophages and microglia are of the most abundant noncancerous cell types in glioblastoma multiforme (GBM), in some cases accounting for up to 30% of the total tumor composition. Macrophages accumulate with GBM progression and can be acutely targeted via inhibition of colony-stimulating factor–1 receptor (CSF-1R) to regress high-grade gliomas in animal models. However, it is currently unknown whether and how resistance emerges in response to sustained CSF-1R blockade in GBM. Despite this, multiple clinical trials are currently underway testing the efficacy of CSF-1R inhibition in glioma patients. Therefore, determining whether long-term CSF-1R inhibition can stably regress GBM by using animal models is an important and timely question to address. RESULTS Using genetic mouse models of GBM, we show that although overall survival is significantly prolonged in response to CSF-1R inhibition, tumors recur eventually in >50% of mice. Upon isolation and transplantation of recurrent tumor cells into naïve animals, gliomas reestablish sensitivity to CSF-1R inhibition, indicating that resistance is microenvironment-driven. Through RNA-sequencing of glioma cells and macrophages purified from treated tumors and ex vivo cell culture assays, we found elevated phosphatidylinositol 3-kinase (PI3K) pathway activity in recurrent GBM after CSF-1R inhibition, driven by macrophage-derived insulin-like growth factor–1 (IGF-1) and tumor cell IGF-1 receptor (IGF-1R). Consequently, combining IGF-1R or PI3K blockade with continuous CSF-1R inhibition in recurrent tumors significantly prolonged overall survival. In contrast, monotherapy with IGF-1R or PI3K inhibitors in rebound or treatment-naïve tumors was less effective, indicating the necessity of combination therapy to expose PI3K signaling dependency in recurrent disease. Mechanistically, we found that activation of macrophages in recurrent tumors by IL4 led to elevated Stat6 and nuclear factor of activated T cells (NFAT) signaling upstream of Igf1, and inhibition of either of these pathways in vivo was sufficient to significantly extend survival. CONCLUSION We have identified a mechanism of drug resistance that can circumvent therapeutic response to a TME-targeted therapy and promote disease recurrence in the absence of tumor cell–intrinsic alterations. Specifically, we have uncovered a heterotypic paracrine signaling interaction that is initiated by the TME and drives resistance to CSF-1R inhibition through IGF-1R/PI3K signaling. Given that PI3K signaling is aberrantly activated in a substantial proportion of GBM patients, and that recent clinical trial results show limited efficacy in recurrent (albeit very advanced) GBM, it is possible that this pathway could similarly contribute to intrinsic resistance to CSF-1R inhibition. Our findings underscore the importance of bidirectional feedback between cancer cells and their microenvironment and support the notion that although stromal cells are less susceptible to genetic mutation than are cancer cells, a tumor can nonetheless acquire a resistant phenotype by exploiting its extracellular environment. Resistance to CSF-1R inhibition in glioma. (A) Macrophages contribute to GBM progression by creating a protumorigenic niche associated with M2-like gene expression. CSF-1R is a critical receptor for macrophage biology and is under clinical evaluation as a therapeutic target in glioma . (B) Targeting CSF-1R early in gliomagenesis significantly prolongs survival in mouse models. CSF-1R inhibition reprograms macrophages to become antitumorigenic by down-regulating M2-like genes and enhancing phagocytosis. Tumor-derived survival factors sustain macrophage viability despite CSF-1R blockade

The Microenvironmental Landscape of Brain Tumors

The brain tumor microenvironment (TME) is emerging as a critical regulator of cancer progression in primary and metastatic brain malignancies. The unique properties of this organ require a specific framework for designing TME-targeted interventions. Here, we discuss a number of these distinct features, including brain-resident cell types, the blood-brain barrier, and various aspects of the immune-suppressive environment. We also highlight recent advances in therapeutically targeting the brain TME in cancer. By developing a comprehensive understanding of the complex and interconnected microenvironmental landscape of brain malignancies we will greatly expand the range of therapeutic strategies available to target these deadly diseases.

Microenvironmental regulation of cancer stem cell phenotypes.

Cancer is a complex set of diseases, driven by genomic instability overlaid with epigenetic modifications. Two prevailing concepts, the stochastic theory and the hierarchical theory, are traditionally used to understand tumor progression. These seemingly contradictory theories can be reconciled with the concept of cellular plasticity, such that certain genetic mutations enable epigenetic alterations in cell fate. A growing body of evidence suggests that cancer cells co-opt embryonic stem cell-associated regulatory networks in order to sustain tumor cell plasticity concomitant with growth and progression. The expression of these stem cell associated factors is regulated by dynamic niches, characterized by cellderived proteins as well as biophysical features such low oxygen tensions. In this review we describe specific embryo-associated proteins such as NODAL, NOTCH, and canonical WNT, which cooperate to maintain stem cell phenotypes in cancer. We also illustrate how biophysical factors, in particular oxygen, can orchestrate plasticity by modulating the expression of stem cell-associated proteins. As the microenvironment is known to play a key role in cellular regulation, it is essential to understand its role in cancer progression in order to improve and create new therapies.

Embryonic Protein Nodal Promotes Breast Cancer Vascularization

Tumor vascularization is requisite for breast cancer progression, and high microvascular density in tumors is a poor prognostic indicator. Patients bearing breast cancers expressing human embryonic stem cell (hESC)-associated genes similarly exhibit high mortality rates, and the expression of embryonic proteins is associated with tumor progression. Here, we show that Nodal, a hESC-associated protein, promotes breast cancer vascularization. We show that high levels of Nodal are positively correlated with high vascular densities in human breast lesions (P = 0.0078). In vitro, we show that Nodal facilitates breast cancer-induced endothelial cell migration and tube formation, largely by upregulating the expression and secretion of proangiogenic factors by breast cancer cells. Using a directed in vivo angiogenesis assay and a chick chorioallantoic membrane assay, we show that Nodal promotes vascular recruitment in vivo. In a clinically relevant in vivo model, whereby Nodal expression was inhibited following tumor formation, we found a significant reduction in tumor vascularization concomitant with elevated hypoxia and tumor necrosis. These findings establish Nodal as a potential target for the treatment of breast cancer angiogenesis and progression.

Obesity alters the lung myeloid cell landscape to enhance breast cancer metastasis through IL5 and GM-CSF

Obesity is associated with chronic, low-grade inflammation, which can disrupt homeostasis within tissue microenvironments. Given the correlation between obesity and relative risk of death from cancer, we investigated whether obesity-associated inflammation promotes metastatic progression. We demonstrate that obesity causes lung neutrophilia in otherwise normal mice, which is further exacerbated by the presence of a primary tumour. The increase in lung neutrophils translates to increased breast cancer metastasis to this site, in a GM-CSF- and IL5-dependent manner. Importantly, weight loss is sufficient to reverse this effect, and reduce serum levels of GM-CSF and IL5 in both mouse models and humans. Our data indicate that special consideration of the obese patient population is critical for effective management of cancer progression.

Molecular Pathways: Deciphering Mechanisms of Resistance to Macrophage-Targeted Therapies

Tumor-associated macrophages (TAMs) are a major cellular component of numerous tumor types. TAM-targeted therapies include depletion strategies, inhibiting their effector functions or reprogramming toward an antitumorigenic phenotype, with varying degrees of efficacy. Here, we review preclinical and clinical strategies to target macrophages in cancer and discuss potential explanations for why some strategies are effective while other approaches have shown limited success. Clin Cancer Res; 23(4); 876–84. ©2016 AACR.

Obesity and the tumor microenvironment

Obesity-associated inflammation promotes tumor growth and metastatic spread Obesity is a growing global epidemic and rivals smoking as the leading preventable risk factor for cancer incidence and mortality, being responsible for an estimated ∼20% of cancer-related deaths in adults (1). Obesity underlies a number of distinct but interconnected health conditions that have profound consequences for physiology, including hypernutrition, dysbiosis, hypercholesterolemia, metabolic syndrome, and chronic inflammation. Although each of these health conditions may affect cancer pathogenesis, inflammation, in particular, is known to be a potent driver of cancer initiation and progression through its ability to cultivate a microenvironment that is permissive to neoplastic transformation. Thus, as immuno-oncology continues to gain clinical importance, understanding the relationship between cancer and various inflammatory conditions, including obesity, is critical.

Tumor-Associated Macrophages Suppress the Cytotoxic Activity of Antimitotic Agents

SUMMARY Antimitotic agents, including Taxol, disrupt microtubule dynamics and cause a protracted mitotic arrest and subsequent cell death. Despite the broad utility of these drugs in breast cancer and other tumor types, clinical response remains variable. Tumor-associated macrophages (TAMs) suppress the duration of Taxol-induced mitotic arrest in breast cancer cells and promote earlier mitotic slippage. This correlates with a decrease in the phosphorylated form of histone H2AX (γH2AX), decreased p53 activation, and reduced cancer cell death in interphase. TAMs promote cancer cell viability following mitotic slippage in a manner sensitive to MAPK/ERK kinase (MEK) inhibition. Acute depletion of major histocompatibility complex class II low (MHCIIlo) TAMs increased Taxol-induced DNA damage and apoptosis in cancer cells, leading to greater efficacy in intervention trials. MEK inhibition blocked the protective capacity of TAMs and phenocopied the effects of TAM depletion on Taxol treatment. TAMs suppress the cytotoxic effects of Taxol, in part through cell non-autonomous modulation of mitotic arrest in cancer cells, and targeting TAM-cancer cell interactions potentiates Taxol efficacy.