Targeting the Tumor Microenvironment in Radiation Oncology ... · 2/5/2019 · 22 Radiation Oncology (ASTRO) and the American Association of Cancer Research (AACR) organized a scientific

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Targeting the Tumor Microenvironment in Radiation Oncology: Proceedings from the 2018 ASTRO-AACR

Research Workshop

Heather M. McGee, MD, PhD*1, Dadi Jiang, PhD*2, David R. Soto-Pantoja, PhD*3, Avinoam Nevler, MD*4,5, Amato J. Giaccia, PhD6 and Wendy A. Woodward, MD, PhD+2

* These authors contributed equally to this paper 1 Department of Radiation Oncology, Icahn School of Medicine at Mount Sinai, New York, NY 2 Department of Radiation Oncology, The University of Texas MD Anderson Cancer Center, Houston, TX 3 Department of Radiation Oncology, Comprehensive Cancer Center Wake Forest School of Medicine, Winston-Salem, NC

4 Department of Surgery, Thomas Jefferson School of Medicine, Philadelphia, PA 5 Talpoit Medical Leadership Program, Sheba Medical Center, Ramat-Gan, Israel 6 Department of Radiation Oncology, Stanford University School of Medicine, Stanford, CA +Corresponding Author: Wendy A. Woodward, MD, PhD Department of Radiation Oncology The University of Texas MD Anderson Cancer Center 1515 Holcombe Blvd #1202 Houston, TX, 77030 Phone: 713-792-1207 Fax: 713-794-5369 E-mail: [email protected]

Running Title: Proceedings of ASTRO-AACR Workshop on Radiation and the TME

Keywords: radiation oncology, tumor microenvironment, microbiome, immune response, metabolism, stroma

Abbreviations: radiotherapy, RT; tumor microenvironment, TME; American Society for Radiation Oncology, ASTRO; American Association of Cancer Research, AACR, National Cancer Institute, NCI; azoxymethane/dextran sulfate sodium, AOM/DSS; reactive oxygen species, ROS; programmed cell death protein-1, PD-1; programmed cell death-ligand 1, PD-L1; T helper 2 cell; Th2, T helper 1 cell, Th1; undifferentiated pleomorphic sarcoma, UPS; DNA damage response, DDR; immunotherapy, IO; multivariate analysis, MVA; stereotactic body radiation therapy, SBRT; interferon, IFN; cytotoxic T-lymphocyte antigen-4, CTLA-4; dendritic cell, DC; Cyclic GMP-AMP Synthase, cGAS; stimulator of interferon genes, STING; double-stranded-DNA, dsDNA; three prime repair exonuclease 1, TREX1; T cell receptor, TCR; tumor immune microenvironment, TIME; Non-Small Cell Lung Cancer, NSCLC; Small cell lung cancer, SCLC; Body Mass Index, BMI,

phosphorylated AKT, pAKT; Peroxisome proliferator-activated receptor gamma, PPAR-; 2-deoxy-D-glucose, 2-DG; fibroblast growth factor 2, FGF2; fibroblast growth factor receptor, FGFR; extracellular matrix, ECM; inflammatory breast cancer, IBC; triple negative breast cancer, TNBC; mesenchymal stem cells, MSC; inducible nitric oxide synthase, iNOS;

Immunogenic cell death, ICD; DAMPs; Inositol-requiring enzyme 1, IRE1; X- box binding protein 1, XBP1; Unfolded protein response, UPR; Epidermal growth factor receptor, EGFR; bortezomib, BTZ; High mobility group box 1 protein, HMGB1; indoleamine 2,3-dioxygeanse, IDO; The Cancer Genome Atlas, TCGA

Acknowledgements: The authors would like to acknowledge the ASTRO and AACR scientific leadership who organized the ASTRO-AACR workshop (Judy Keen, PhD, Tyler Beck, PhD, and Michael Powell, PhD) along with Dr. Wendy Woodward and Dr. Amato Giaccia for chairing the session. They would also like to acknowledge all speakers who presented their research or moderated sessions or breakout session at the workshop, including: Giorgio Trichinieri, MD, Ann Klopp, MD, PhD, Andrea Facciabene, PhD, Phuoc Tran, MD, PhD, Dorthe Schaue, PhD, Robert Samstein, MD, PhD, Claire Vanpouille-Box, PhD, Michael Gough, PhD, Mohamed Abazeed, MD, PhD, Amato Giaccia, PhD, Julie Schwarz, MD, PhD, Nicolas Denko, MD, PhD, Nicole Simone, MD, Wendy Woodward, MD, PhD, Simon Powell, MD, Catherine Park, MD, Ruth Muschel, Michael Spiotto, MD, PhD, Dadi Jiang, PhD, David R. Soto-Pantoja, PhD and Avinoam Nevler, MD and Heather McGee, MD, PhD. Due to space limitations, we were unable to include a discussion of all of the research

Research. on November 5, 2020. © 2019 American Association for Cancerclincancerres.aacrjournals.org Downloaded from

Author manuscripts have been peer reviewed and accepted for publication but have not yet been edited. Author Manuscript Published OnlineFirst on February 5, 2019; DOI: 10.1158/1078-0432.CCR-18-3781

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presented. Conflict of Interest Disclosure Statement: H. McGee- Advisory Board: AstraZeneca; Research Funding: Adaptive Biotechnologies




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Statement of Translational Relevance: 1

Historically, the field of radiobiology has emphasized the cell autonomous effects of radiation on DNA damage and cell 2

death, but recent advances in immunology, stromal biology metabolism and microbiome research have highlighted the 3

need to understand how radiation is influenced by diverse biologic processes. While radiation serves as a cytotoxic 4

agent that induces DNA damage, it also modifies various components of the tumor microenvironment (TME) and alters 5

the threshold for anti-tumor immune activity. Radiation oncologists must consider the spatial and temporal 6

characteristics of each of these interactions in order to understand the heterogeneous responses seen after radiation 7

therapy. Ultimately, as radiation is paired with therapies that target other components of the TME, obtaining a more 8

comprehensive understanding of interactions between radiation and the TME will help us improve clinical outcomes for 9

our patients. 10




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Abstract: 11

The development of cancers and their response to radiation are intricately linked to the tumor microenvironment in 12

which they reside. Tumor cells, immune cells, and stromal cells interact with each other and are influenced by the 13

microbiome and metabolic state of the host, and these interactions are constantly evolving. Stromal cells not only 14

secrete extracellular matrix and participate in wound contraction, but they also secrete fibroblast growth factors (FGFs), 15

which mediate macrophage differentiation. Tumor associated macrophages (TAMs) migrate to hypoxic areas and 16

secrete VEGF to promote angiogenesis. The microbiome and its byproducts alter the metabolic milieu by shifting the 17

balance between glucose utilization and fatty acid oxidation, and these changes subsequently influence the immune 18

response in the tumor microenvironment (TME). Not only does radiation exert cell autonomous effects on tumor cells, 19

but it influences both the tumor-promoting and tumor-suppressive components of the TME. To gain a deeper 20

understanding of how the tumor microenvironment influences the response to radiation, the American Society for 21

Radiation Oncology (ASTRO) and the American Association of Cancer Research (AACR) organized a scientific workshop 22

on July 26-27, 2018, which focused on how the microbiome, the immune response, the metabolome, and the stroma all 23

can shift the balance between radiosensitivity and radioresistance. The proceedings from this workshop are discussed 24

here and highlight recent discoveries in the field as well as the most important areas for future research. 25

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I. Introduction: 27

We are at a crucial moment in the field of radiobiology – the cell autonomous effects of radiation on DNA 28

damage and cell death are well-established, but it is becoming increasingly clear that we cannot view tumor cells in 29

isolation because they are intricately linked to the tumor microenvironment in which they reside. Decades of work on 30

clonogenic survival assays has demonstrated that radiation causes DNA damage which causes irradiated cells to undergo 31

mitotic catastrophe. Since the induction of reactive oxygen species contributes to DNA damage, hypoxia and 32

reoxygenation have become key determinants of the radiation response. Therefore, radiobiology research on the tumor 33

microenvironment has focused on hypoxia for decades. However, there are also many cell types in the tumor 34

microenvironment which could influence the response to radiation, including stromal cells, endothelial cells, innate 35

immune cells and adaptive immune cells. Each tumor is defined by differences in the composition of tumor cells and 36

other cellular elements; for example, pancreatic cancer contains a significant stromal component, small cell lung 37

carcinoma contains mesenchymal, epithelial, neuroendocrine and neural cells, and lymphoma contains cells derived 38

from the rich lymphatic environment of secondary lymphoid organs. In the clinic, this heterogeneity accounts for varied 39

clinical responses to stereotactic body radiation (SBRT) using the same radiation dose and fractionation (1). 40

Given that radiation activates the immune system, it is becoming clear that many of the factors which influence 41

responsiveness to immune checkpoint inhibitors (ICI) also influence the response to radiation. Outcomes after ICI are 42

influenced by the genetic mutational load in the tumor (2), mutations in DNA damage response and repair pathways (3), 43

and adaptive resistance, a process in which IFN- production by activated CD8+ T cells leads to an upregulation of PD-L1 44

(4). The gut microbiome plays a key role in regulating -PD1-elicited immune responses, since antibiotic treatment 45

reduces bacterial biodiversity and decreases the effectiveness of immunotherapy (5–7). Metabolic pathways such as 46

glycolysis, amino acid metabolism and fatty acid-oxidation (FAO) influence the response to PD-1 inhibitors (8,9). 47

Components of the stroma contribute to an immune-excluded environment and mediate epithelial-mesenchymal 48

transition (EMT) in response to immunotherapy and radiation (10,11). 49

To gain a deeper understanding of how these components of the tumor microenvironment influence the 50

response to radiation, the American Society for Radiation Oncology (ASTRO) and the American Association of Cancer 51

Research (AACR) organized a scientific workshop on July 26-27, 2018, which focused on the following key issues: 1. The 52




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microbiome’s influence on the response to radiation, 2. Immune responses to radiation in the TME compared to the 53

periphery, 3. How metabolism and obesity affect the TME and responses to radiation, and 4. The role of the stroma in 54

shifting the balance between radiosensitivity and radioresistance. 55

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II. Influence of the Microbiome on the Response to Radiation 57

The gut microbiome is rapidly becoming an important clinical biomarker and therapeutic target because 58

microbiome-host crosstalk is important for both metabolism and immune regulation. Dr. Giorgio Trinchieri, Director of 59

the National Cancer Institute’s (NCI) Cancer and Inflammation Program, described the crucial role of the microbiome in 60

setting the threshold for activating the immune response, largely mediated by the IL-23-IL-22 axis, which promotes 61

maintenance of the intestinal barrier and therefore prevents dysbiosis or dysregulated symbiosis between bacteria and 62

host. This is important in the context of cancer because germ-free mice without microbiota develop larger tumors in the 63

azoxymethane/dextran sulfate sodium (AOM/DSS) colon cancer model (12), and show decreased responsiveness to -64

PD-1 therapies (13). Multiple studies have shown that response to immunotherapies are linked to the microbiota in 65

patients as well, but differences in geography and diet lead to microbiome variability which make it challenging to 66

interpret and generalize results from these studies (5–7). 67

In order to elucidate mechanisms of microbiome-associated radioresistance, Dr. Klopp illustrated that broad-68

spectrum antibiotic depletion of the microbiome results in radioresistance of cervical cancer to a single fraction of 6 Gy 69

(14). Dr. Facciabene has shown that vancomycin, but not neomycin or metronidazole, decreased growth of tumors and 70

sensitized the tumors to radiation therapy. Interestingly, vancomycin decreases bacterial diversity, by mainly 71

eliminating many sub-species of the Clostridiales family, leading to a decrease in production of bacteria-derived short 72

chain fatty acids (SCFA). Dr. Facciabene’s research has shown that some of the absorbed SCFAs (e.g, the C4-SCFA, 73

Butyrate) have immunosuppressive effects in the TME and a decrease in SCFA levels can sensitize tumors to radiation 74

therapy by enhancing antigen presentation and T cell priming. The implication from this work is that inhibitors of SCFA 75

could potentially be delivered in combination with radiation to serve as radiosensitizers. In addition, the microbiome is 76

increasingly appreciated for its role in the processing of metabolites of microbial origin such as tryptophan, and many of 77

these byproducts (i.e. indoleamine 2,3-dioxygenase (IDO)) are immunosuppressive molecules in the TME, which have 78




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recently been found to modulate the response to radiation in certain forms of cancer (14)(15). Active areas of 79

investigation in this field focus on developing and implementing methods of microbiome manipulation (spore therapy, 80

probiotics, transplant of specific organisms or fecal transfer. etc.) in combination with radiation therapy. 81

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III. The Immune Response in the Tumor Microenvironment 83

Radiation can act as an in situ tumor vaccine by inducing immunogenic cell death which releases tumor-84

associated antigens and other damage-associated molecular patterns (DAMPS) that activate dendritic cells to migrate to 85

the draining lymph nodes and prime antigen-specific CD8+ T cells. While there is a strong interest in studying how 86

localized radiotherapy induces systemic immune responses, effective anti-tumor immunity is counter-balanced by 87

immune tolerance and immune suppression in the TME (16). In pre-clinical studies using subcutaneous tumor models in 88

the context of CTLA-4 blockade with radiation, hypofractionated radiation regimens (8 Gy x 3 fractions) produce a more 89

robust CD8+ T-cell-mediated abscopal response and improve survival compared to 20 Gy x 1 (17). Hypofractionated 90

radiation can cause double-stranded DNA (dsDNA) to accumulate in the cytoplasm, and this stimulates the cyclic 91

GMP/AMP synthase (cGAS)/ STimulator of INterferon Genes (STING)/interferon- signaling, which recruits Batf3+ 92

dendritic cells to activate CD8+ T cells. Dr. Claire Vonpouille-Box discussed her work showing that radiation doses 93

greater than 12 Gy per fraction induce expression of the DNA exonuclease three prime repair exonuclease 1 (TREX1; 3’ 94

5’), which degrades cytosolic dsDNA, abrogates radiation induction of IFN- and decreases systemic anti-tumor 95

immunity (18). Taken together, this research suggests that we need to incorporate assays for DNA damage, 96

immunogenic cell death, and innate immune sensing into clinical trials in order to better understand how radiation leads 97

to immune activation. 98

Importantly, Dr. Michael Gough’s work has shown how anti-tumor immune responses after combined radiation 99

and ICI depend on pre-existing immunity (19). His murine studies have shown that tumor implantation leads to 100

establishment of resident memory CD39+ CD103+ CD8+ T cells in the tumor milieu. Radiation decreases, but does not 101

eliminate, resident memory T cells in the tumor. In addition, his work has shown that treatment with anti-CD40L, 102

depletion of CD8 T cells, or delivery of FTY-720 (which blocks SIP1-mediated exit of lymphocytes from the lymph nodes) 103

prior to tumor implantation prevents tumor control caused by the combination of radiation and immune checkpoint 104




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inhibition (19). The timing of the radiation treatment in conjunction with immunotherapy is also likely to be a key factor 105

in determining outcomes, as discussed by Dr. Robert Samstein, based on his analysis of retrospective data. 106

Dr. Dorthe Schaue described her translational work studying differences in patients’ immune response to 107

radiation based on dose, fraction size, site of disease and volume irradiated. Together with Dr. Nicholas Nickols and Dr. 108

Anusha Kalbasi, she has observed an increased myeloid cell infiltrate after hypofractionated radiation to the prostate 109

and an increased T helper cell infiltrate after conventional fractionation for undifferentiated pleomorphic sarcoma. This 110

work emphasizes the need to expand our approach to studying the immune response to radiation beyond the classic 111

paradigm of radiation as “in situ vaccine” discussed above, which focuses on dendritic cells and CD8+ T cells. Lastly, the 112

breakout session focused on “hot” (inflamed) vs. “cold” (non-inflamed) tumors, and emphasized that “cold tumors” can 113

be either “immune excluded tumors” characterized by altered vasculature and cancer associated fibroblasts, or 114

“immune desert tumors” characterized by anergy, tolerance, or immunologic ignorance (20). Future studies are needed 115

to investigate how radiation influences each of these unique tumor immune microenvironments (TIMEs) to enhance the 116

local and systemic anti-tumor immune response (20). 117

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IV. How the Metabolome Affects the Response to Radiation 119

Hypoxia and oxidative stress alter metabolism in the TME, and therefore it is not surprising that metabolic 120

changes in the TME affect the response to radiation. Specifically, metabolic adaptations to hypoxia cause decreased 121

mitochondrial function and increased lactate production. This leads to hypoxia-inducible inhibitory phosphorylation of 122

the pyruvate dehydrogenase E1α subunit, which enhances tumor growth (21). Dr. Nicolas Denko discussed work 123

suggesting that hypoxia is not a problem of oxygen delivery, but rather is due to excessive oxygen consumption by 124

tumors that cannot be met by supply demands (22). Thus, interventions that specifically target oxygen consumption may 125

sensitize tumors to conventional cancer therapies such as ionizing radiation. In addition, endocrine imbalances 126

associated with obesity affect responsiveness to radiotherapy. Highly glycolytic cervical tumors are associated with 127

alterations in the PI3K/AKT pathway and are resistant to chemoradiation (23), and the glucose inhibitor 2-DG can 128

sensitize these tumors to ionizing radiation, as described by Dr. Julie Schwartz (24). In addition, targeting glutathione 129

and thioredoxin alone or in combination with 2-DG enhances the efficacy of radiation in these resistant tumors. 130




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Therefore, both redox metabolism and glycolytic flux could be targeted pharmacologically to improve radiation 131

responsiveness. 132

Fatty acids upregulate peroxisome proliferator-activated receptor gamma (PPAR-) and inhibit p53, which both 133

sensitize cancer cells to radiation. Supplementation of fatty acids to culture media upregulates (PPAR-) and inhibits 134

p53, which sensitizes cancer cells to ionizing radiation. In cervical cancer cells, radiation alone seems to increase uptake 135

of unsaturated fatty acids, in part by regulating the receptor fatty acid uptake receptors such as CD36. As mentioned in 136

the previous section, the antibiotic vancomycin reduces bacterial-derived SCFAs highlighting an intersection between 137

microbiota and metabolism regulation that it is likely to influence tumor growth and the response to therapy through 138

the PPAR-pathway. Dr. Nicole Simone presented results from a clinical study showing that patients with diabetes 139

treated with intracranial radiation therapy have reduced overall survival and reduced median intracranial progression-140

free survival compared to non-diabetic individuals, emphasizing a role of host metabolism in radiotherapy response (25). 141

She discussed results where caloric restriction led to radiosensitization of tumors in preclinical models and also 142

suggested the potential of ketogenic diets, which are low in carbohydrates, to sensitize tumors to radiotherapy, thus 143

suggesting that dietary host modification of metabolism may improve outcomes after radiotherapy (26,27). Future 144

studies the metabolome in radiation biology will help us elucidate specific metabolite signatures associated with 145

favorable responses to radiation, and how dietary modifications cause metabolic changes that improve clinical 146

outcomes after radiation. 147

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V. Role of the Tumor Stroma in the Response to Radiation 149

The stroma consists of fibroblasts which can either form fibrovascular areas throughout the tumor or can form a 150

desmoplastic matrix that separates the tumor core from other cells in the microenvironment. Fibroblasts can secrete 151

collagen and fibronectin which serve as components of the extracellular matrix, and can also secrete Matrix 152

metalloproteinases (MMPs) which degrade this scaffold. In addition, cancer-associated fibroblasts (CAFs) secrete 153

growth factors such as epidermal growth factor (EGF), hepatocyte growth factor (HGF), fibroblast growth factor and 154

insulin-like group factor 1 (28). Neutrophils secrete Fibroblast growth factor 2 (FGF2) into the extracellular matrix, where 155

FGF2 is sequestered until it is released upon proteolysis (29). Dr. Ruth Muschel has shown that in experimental models 156




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of colon cancer, radiation stimulates intratumoral infiltration of macrophages, with simultaneous induction of FGF2 and 157

its receptors (FGFRs). Dr. Muschel’s work has revealed that FGF signaling causes macrophages to switch from an M1 158

phenotype to an M2 phenotype, which mediate resistance to radiation (unpublished). Dr. Wendy Woodward’s work on 159

stromal elements of the tumor microenvironment in inflammatory breast cancer has illustrated intricate crosstalk 160

between tumor-associated macrophages and mesenchymal stem cells (MSCs) (30). M2 polarized macrophages educate 161

MSCs to develop an M2 phenotype themselves, which stimulates invasion of inflammatory breast cancer cells in vitro, 162

and leads to an increase in the aggressiveness of tumors in vivo. Similar to Dr. Muschel, Dr. Woodward has shown that 163

M1 macrophages promote radiosensitivity of inflammatory breast cancer cells, but M2 macrophages induce 164

radioresistence via IL-4/IL-13-mediated STAT6 phosphorylation and M2 polarization (31). 165

Another element of cross-talk between cancer cells and the stroma involves the often-overlooked role of 166

stromal cells as antigen-presenting cells. Dr. Michael Spiotto discussed his experimental transplant tumor model, tumor 167

cells that are poorly antigenic which are challenged with stroma can escape host immune surveillance (32). Dr. Spiotto 168

mentioned that negative signaling pathways induced by the stroma counteract an anti-tumor immune response, but 169

then discussed that radiation increases stromal cells’ ability to present antigens and promote anti-tumor immunity. In 170

addition, Dr. Spiotto found that loss of expression of Notch1 increases the expression of ECM proteins and the 171

infiltration of macrophages (33), which echoes Dr. Muschel’s findings that there is cross-talk between fibroblasts and 172

macrophages. Moreover, Dr. Spiotto discussed distinct transcriptomic subtypes identified in lymph node metastases 173

that are absent in primary tumors, and emphasized that this is likely due to the unique lymph node microenvironment. 174

Efforts to understand how the genetic makeups of cancer affects radiation responses were presented by Dr. Mohamed 175

Abazeed, his results showed that Non-small cell lung cancer patients with squamous histological subtype were 176

associated with increased risk for local failure, and patients with adenocarcinoma had significantly higher responses 177

rates to SBRT when compared to patients with squamous subtype (34). The breakout session emphasized that we need 178

to characterize the tumor stroma with a higher definition utilizing both single cell-based sequencing, multiplexed 179

imaging, and deconvolution of bulk cell data to identify (i) stromal characteristics that are specific for individual tumors 180

and/or shared across different tumors, (ii) normal stroma- versus tumor stroma-specific targets, and (iii) stromal 181

characteristics that characterize response to radiation therapy. In addition, leaders in this field emphasized the 182




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importance of developing improved pre-clinical model systems to study cancer cell-stroma interactions, such as 3-D 183

culture systems (organoids and/or bioengineered model systems), syngeneic mouse models and genetically engineered 184

primary tumor models, and patient sample-derived models (PDXs and human tumor organoids). 185

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VI. Conclusion 187

Understanding the contributions from diverse components of the tumor microenvironment will be essential for 188

optimizing radiation therapy (Figure 1). The microbiome is increasingly appreciated for its role in the processing of 189

metabolites of microbial origin such as bacteria-derived short chain fatty acids (SCFA), and many of these byproducts 190

(e.g, the C4-SCFA, Butyrate) have immunosuppressive effects in the TME. The mechanisms by which radiation can 191

induce tissue damage and initiate an immune response are intricately involved with DNA damage response and DNA 192

repair processes. The byproducts of fatty acid metabolism can interact with stromal cells and influence cancer 193

progression (35), but these metabolic processes are only beginning to be understood in the context of the TME. Lastly, 194

the stromal cells, which form the extracellular matrix, interact with immune cells, and often influence the threshold for 195

tumor invasion and metastases; therefore, it is not surprisingly that TGF- and FGF2 are emerging as key mediators of 196

response and toxicity after radiation and therapeutic targets. 197

Moving forward, radiation oncology must merge with scientific other fields (metabolism, immunology, cancer 198

biology, bioengineering, and microbiology) and develop a collaborative, interdisciplinary approach to determine how 199

radiation can most effectively target the tumor microenvironment. There is a critical need to invest in diverse research 200

areas in order to enhance our understanding of the tumor microenvironment. Determining how to apply discoveries in 201

other scientific fields to modulate the response to radiation is essential for our next generation of radiobiology research. 202

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Figure Legends 314

FIGURE 1: The intersection of radiation and the tumor immune microenvironment. 315

A. Microbiome. Broad-spectrum antibiotics reduce microbiome diversity , which decreases gut microbiota-derived short 316

chain fatty acids (SCFA), leading to increased antigen presentation and T cell priming, which sensitizes tumors to 317

radiation therapy. Bacterial processing of SCFA can also influence M1-M2 macrophage polarization. 318

B. Immune Response. Immunogenic cell death leads to release of damage-associated molecular patterns (DAMPs) such 319

as high mobility group box protein 1 (HMGB1), calreticulin, and ATP. Cytosolic DNA activates the cGAS/STING pathway, 320

leading to an increase in IFN-, which activates Baft3+ dendritic cells that play a key role in cross-presentation and cross-321

priming of CD8+ T cells. CD39+ CD103+ tissue resident memory T cells also increase in sensitivity to ionizing radiation. 322

C. Metabolome. Hypoxia causes increases glucose uptake and glycolysis, which contributes to radioresistance. During 323

hypoxic conditions, mitochondria act as O2 sensors and convey signals to Hypoxia-inducible factor (HIF)-1. Impaired 324

glucose and lipid metabolism leads to an increase in lipid and fatty acid stores, and fatty acid uptake by CD36, along with 325

PPAR-inhibition of p53 can increase sensitivity to ionizing radiation. 326

D. Stroma. FGF2 is sequestered in the extracellular matrix and released upon proteolysis. FGF signaling converts myeloid 327

cells into mesenchymal cells via STAT6, leading to M1 to M2 macrophage conversion. M1 macrophages promote 328

radiosensitivity while M2 polarized macrophages promote MSCs to develop an M2 phenotype, which mediates radiation 329

resistance. 330

331




B. Immune response

D. Stroma

A. Microbiome Radiation

C. Metabolome

Reduced microbiome diversity - IR resistance

Bacterial processing of SCFA

cytosolic DNA cGAS/STING

IFN-b

Baft3+ dendritic cell

Cytokines

CD8:CD4 CD39+ CD103+ tissue resident memory cells

Hypoxia

2-DG IR Sensitivity

CD36 FA

PPAR

p53

IR Sensitivity

Glucose uptake Glycolysis

M1M2

polarization

FGFR

FGF-2

pSTAT6

Immunogenic cell death

DAMPS: HMGB1 Calreticulin ATP

IR Sensitivity

Antibiotics

Mesenchymal celll

IR Sensitivity

M1-M2 polarization

O2 sensor Mitochondria

IR Sensitivity

Cytokines

Myeloid cell

Figure 1




Published OnlineFirst February 5, 2019.Clin Cancer Res Heather M. McGee, Dadi Jiang, David R Soto-Pantoja, et al. Proceedings from the 2018 ASTRO-AACR Research WorkshopTargeting the Tumor Microenvironment in Radiation Oncology:

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