As tumors grow, they upregulate glycolytic and oxidative metabolism to support their increased and altered energetic demands. These metabolic changes have major effects on the tumor microenvironment. One of the properties leading to this aberrant metabolism is hypoxia, which occurs when tumors outgrow their often-chaotic vasculature. This scarcity of oxygen is known to induce radioresistance but can also have a disrupting effect on the antitumor immune response. Hypoxia inhibits immune effector cell function, while immune cells with a more suppressing phenotype become more active. Therefore, hypoxia strongly affects the efficacy of both radiotherapy and immunotherapy, as well as this therapy combination. Inhibition of oxidative phosphorylation (OXPHOS) is gaining interest for its ability to combat tumor hypoxia, and there are strong indications that this results in a reactivation of the immune response. This strategy decreases oxygen consumption, leading to better oxygenation of hypoxic tumor areas and eventually an increase in immunogenic cell death induced by radio-immunotherapy combinations. Promising preclinical improvements in radio- and immunotherapy efficacy have been observed by the hypoxia-reducing effect of OXPHOS inhibitors and several compounds are currently in clinical trials for their anticancer properties. Here, we will review the pharmacologic attenuation of tumor hypoxia using OXPHOS inhibitors, with emphasis on their impact on the intrinsic antitumor immune response and how this affects the efficacy of (combined) radio- and immunotherapy.

Tumors quickly outgrow their vasculature, thereby limiting the amount of oxygen available for diffusion into the tissue in areas distant from vessels, while oxygen demand is also increased in proliferative tissues (1). The diffusion-limited deprivation of oxygen is known as chronic hypoxia, which occurs in most solid tumors. Moreover, the chaotic and unreliable nature of the tumor vasculature can also result in a more acute form of hypoxia by the temporal impaired blood flow through these vessels (Fig. 1). Hypoxia results in metabolic changes and hypoxic tumor cells are characterized by an increased glycolytic metabolism (2). Glycolysis leads to accumulation of glycolytic metabolites, such as lactate, and subsequent acidification of the tumor microenvironment (TME; ref. 2). Hypoxic areas in solid malignancies are highly therapy resistant (3, 4). A well-known example is hypoxia-mediated resistance to radiotherapy, primarily caused by the reversion of DNA-free radicals under hypoxia, whereas this DNA damage is fixed under normoxia (oxygen fixation hypothesis; ref. 5). Furthermore, radiation produces reactive oxygen species (ROS) that interact with DNA, inducing mitotic cell death. These ROS cannot be formed when oxygen is deprived, resulting in radioresistance (5). Several studies describe how radiotherapy can be enhanced by the alleviation of hypoxia, both in preclinical models, as well as in clinical studies (6–8). Another possible reason for the failure of radiotherapy is hypoxia-mediated immune escape, as a substantial part of the efficacy of radiotherapy relies on a durable anticancer immune response (9). This is, for instance, mediated by the depletion of immune effector cells from hypoxic areas and acidification of the TME (Fig. 1; refs. 10–12).

Figure 1.

Immunofluorescence image of a syngeneic mouse oral carcinoma model (MOC1) showing that leukocytes (CD45.2, blue) are virtually absent in hypoxic tumor regions (pimonidazole, green). Vessels: 9F1, red; A: acute hypoxia, illustrated by vessels with surrounding hypoxic tumor cells; D: diffusion-limited hypoxia; indicated by increasing hypoxia at increasing distance from vessels. Magnification, 10×; scale bar, 100 μm.

Figure 1.

Immunofluorescence image of a syngeneic mouse oral carcinoma model (MOC1) showing that leukocytes (CD45.2, blue) are virtually absent in hypoxic tumor regions (pimonidazole, green). Vessels: 9F1, red; A: acute hypoxia, illustrated by vessels with surrounding hypoxic tumor cells; D: diffusion-limited hypoxia; indicated by increasing hypoxia at increasing distance from vessels. Magnification, 10×; scale bar, 100 μm.

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In this review, we will describe the mechanisms by which hypoxic stress causes immune suppression, involving different immune components, and how this contributes to treatment resistance. There are strong indications that by alleviating hypoxia and concomitant immune suppression, resistant tumor areas can become sensitive to treatment (13, 14). An effective way to accomplish this goal is the reduction of the oxygen consumption in tumors, hereby normalizing oxygen levels in remote tumor areas. Reduction of the oxygen consumption rate can be achieved by pharmacologic inhibition of oxidative phosphorylation (OXPHOS). As opposed to the longstanding notion that tumors are primarily glycolytic, including under aerobic conditions, as postulated by the Warburg effect, it has recently been described that OXPHOS remains active in many cancers and is a potential therapeutic target (15). Many important oncogenic pathways rely on mitochondrial metabolism and metabolic plasticity is important for tumor progression at every stage of development (16). For instance, in B16 melanoma, the Warburg effect has proven to be dispensable all together because of an upregulated mitochondrial metabolism (17). Current studies indicate that metabolic reprogramming might be an important strategy in reactivating the immune response and describe how this ultimately can also lead to an increased efficacy of the combination of immuno- and radiotherapy (18).

To counter hypoxic stress, several mechanisms are activated, such as the hypoxia-inducible factor 1 (HIF1) pathway (19), the unfolded protein response, and the mTOR pathway (20). The hypoxia response affects tumor metabolism, angiogenesis, pH regulation, and also impacts antitumor immunity (2). Immune effector cells can be influenced directly by hypoxia; for instance, expression of HIF1α (the O2 regulated subunit of HIF1) leads to a transcriptional responses affecting immune cell metabolism and differentiation (21). Immune effector cells can also be affected indirectly via the release of immunosuppressive molecules (e.g., adenosine) by surrounding tumor cells and/or cancer-associated fibroblasts (22, 23).

Metabolic changes can be important drivers of the immune system, often governing the action and development of different immune cell subsets (24). Different populations of immune cells respond differently to hypoxic stress. Many effector cells that are important for tumor eradication, like CD8+ and CD4+ T cells, dendritic cells (DCs), and natural killer (NK) cells are suppressed (25) and less present in hypoxic areas (Figs. 1 and 2). The HIF1 transcriptional hypoxia response directly prevents activation of CD8+ T cells and CD8+ T cells are mostly absent from hypoxic areas in a syngeneic fibrosarcoma model (12, 26). Reduction of hypoxia can also restore the infiltration of CD3+ T cells in a spontaneous prostate cancer model in mice (27). On the other hand, immunosuppressive cell types, like tumor-associated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs), and regulatory T cells (Tregs), are often overrepresented and overly active in hypoxic tumor areas (Fig. 2; ref. 25). In mice, hypoxia induces the accumulation of TAMs in 3LL lung and 4T1 mammary tumors (11). Hypoxia-associated activation of the HIF1 pathway in tumor cells indirectly promotes the accumulation of MDSCs in Hepa1-6 hepatocellular cancer via the extracellular conversion of ATP to 5′-AMP (28), while it directly promotes Treg activation in induced colon cancer (29). Acidosis of the TME, which is the consequence of hypoxia-induced glycolytic metabolism, has an immunosuppressive effect as well (10). Acidification of the TME in B16 melanoma skews macrophage polarization toward a noninflammatory phenotype, promoting tumor growth (30). In contrast, hypoxia can promote antitumor immunity in some cases. In GL261 glioblastomas, for example, although associated with an enhanced animal survival, HIF1α-deficient Tregs showed enhanced CD8+ T-cell–suppressing activity (31). Furthermore, hypoxic cell culture enhances the cytolytic activity of CD8+ T cells, while their capacity to produce cytokines is potentiated in hypoxic CT26 colorectal cancer (32–34). This highlights that the effect of hypoxia on the immune system can be heterogeneous within tumors and, being either direct or indirect, may likely be tumor type dependent.

Figure 2.

Schematic model for the recruitment of immune cells to hypoxic and normoxic tumor areas. In remote hypoxic tumor areas, immune cells with an immunosuppressive phenotype are recruited to the TME. This includes Tregs, MDSCs, and M2 macrophages or TAMs. Furthermore, under hypoxic conditions, immunosuppressive metabolites and cytokines are excreted by both tumor cells and immune cells, while immune checkpoint molecules are upregulated, this dampens an effective immune response. However, in tumor areas with normal oxygen supply, effector immune cells are recruited to the TME. This results in immune cell–mediated attack of tumor cells. Immune cells recruited to these areas include cytotoxic T lymphocytes (CTLs), NK cells, DCs, and M1 or classic macrophages.

Figure 2.

Schematic model for the recruitment of immune cells to hypoxic and normoxic tumor areas. In remote hypoxic tumor areas, immune cells with an immunosuppressive phenotype are recruited to the TME. This includes Tregs, MDSCs, and M2 macrophages or TAMs. Furthermore, under hypoxic conditions, immunosuppressive metabolites and cytokines are excreted by both tumor cells and immune cells, while immune checkpoint molecules are upregulated, this dampens an effective immune response. However, in tumor areas with normal oxygen supply, effector immune cells are recruited to the TME. This results in immune cell–mediated attack of tumor cells. Immune cells recruited to these areas include cytotoxic T lymphocytes (CTLs), NK cells, DCs, and M1 or classic macrophages.

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In addition to the suppressing effects on immune effector cells, hypoxia can also increase the intrinsic resistance of tumor cells themselves to antitumor immune responses. This is, for instance, mediated by increased expression of cell-surface immune checkpoint molecules, like programmed death-ligand 1 (PD-L1) on tumor cells, which have emerged as promising therapeutic targets (35). Programmed death-protein 1 (PD-1) is expressed on activated T cells and functions as a feedback mechanism to protect against tissue pathology–induced autoimmunity upon binding to its ligand PD-L1 (36). Furthermore, HIF1α has been identified as a direct regulator of PD-L1, suggesting that combining hypoxia targeting with PD-L1 blockade may improve treatment outcome (37). Another therapeutically targeted immune checkpoint molecule, cytotoxic T-lymphocyte–associated protein 4 (CTLA-4), is under control of the adenosine-A2A adenosine receptor pathway in Tregs (38). As adenosine accumulates in hypoxic tumors (22), CTLA-4 upregulation has a potential role in hypoxia-mediated immune evasion as well. Expression of other immune checkpoint molecules, including VISTA, TIM-3, and LAG3, targeting of which is currently under clinical investigation, is regulated by hypoxia as well, contributing to immune repression in hypoxic areas (Fig. 2; refs. 34, 39, 40). As opposed to immune checkpoints regulating the adaptive immune response, the don't eat me signal CD47, which mediates resistance against innate immunity through the prevention of phagocytosis by macrophages, is also upregulated under hypoxic conditions as the CD47 gene is under direct control of HIF1α in breast cancer in vitro (41).

Tumor hypoxia and the presence of immunosuppressive cell types can be major obstacles for effective immunotherapy (42, 43). In a murine BrafV600E-induced/Pten-deficient melanoma model, tumor hypoxia caused exhaustion of CD8+ T cells, acting as a barrier to PD-1 blockade immunotherapy (44). Also, the immunosuppressive properties of the metabolite adenosine, which is excessively produced in hypoxic areas, have been described as a potential target to improve immunotherapy (45). This is also relevant for the efficacy of radiotherapy, as potential systemic responses rely on an effective immune system (9). This is especially true for the eradication of metastases outside of the irradiated area, known as the abscopal effect (9). The mechanisms by which hypoxia influences the immune response have recently been reviewed in more detail (Barsoum and colleagues 2014, Noman and colleagues 2015, and Vito and colleagues 2020; refs. 25, 46, 47). A sustained reduction of hypoxia might be an effective strategy to lower immunosuppression and enhance the efficacy of radio- and immunotherapy in therapy-resistant hypoxic areas (13, 18).

Most treatments countering hypoxia-mediated radioresistance, such as accelerated radiotherapy with carbogen and nicotinamide (6), lead to temporary increases in tissue oxygen concentrations, which may be too short to result in a sustained effect on the local immune cell repertoire (48). Prolonging oxygenation can be achieved by reducing the oxygen consumption of tumor cells (49). Specifically, this can be achieved by inhibiting OXPHOS as has been demonstrated using [18F]fluoroazomycin arabinoside ([18F]FAZA)-PET imaging in an A375-R1 melanoma model 24 hours after administration of the inhibitor (50). Furthermore, increased proliferation of reoxygenated cells is not expected as these cells are inhibited in their capability to use the available oxygen. Pharmacologic inhibition of OXPHOS is already proposed as a therapeutic strategy for cancers in which this pathway is upregulated (Table 1), thereby blocking the tumor cells' primary source of ATP production. Inhibition of oxidative metabolism has also been suggested as a strategy to enhance the direct cytotoxic effects of radiotherapy (7, 8). As modulation of the TME can improve the efficacy of immune checkpoint blockade, radiotherapy, and its combination (51), inhibition of oxidative metabolism is a potential strategy to, by alleviating hypoxia, specifically sensitize hypoxic tumors to these treatment modalities. Overall and progression-free survival of patients with melanoma, following PD-1 blockade, more than doubled at 3 years, which was statistically significant, for tumors with a low oxidative metabolism and subsequent lesser hypoxia, posing OXPHOS as a potential target for improving immunotherapeutic response (44). Other studies also associate an increased mitochondrial metabolism with immunotherapy resistance or an immunosuppressive phenotype in melanoma (18, 52, 53). Several OXPHOS inhibitors have already been assessed for their anticancer properties and are making their way into the clinic (Table 1; Fig. 3). To select patients for treatment and to evaluate the antihypoxic effects of these OXPHOS inhibitors, functional hypoxia imaging is a useful tool that has already been used in some preclinical models and clinical trials.

Table 1

. List of selected OXPHOS inhibitors under recent and current clinical study as anticancer therapeutics.

OXPHOS inhibitorSelected clinical trialsPurposeMitochondrial complex targetedFunctional imaging in these trials
Biguanides 
Metformin NCT04114136, NCT04275713, NCT02394652, many more trialsa Many trials in progress, including phase I and II trials combining metformin with anti–PD-1 mAbs, nivolumab or pembrolizumab, in several cancers. Also, several trials aiming to use the antihypoxic properties of metformin in combination with radiotherapy. I (54) [18F]FAZA-PET, DWI- and DCE-MRI 
Phenformin NCT03026517 Phase I, in combination with MEK and B-Raf inhibitors in metastatic melanoma. I (54) — 
IM156 NCT03272256 Phase I, in advanced solid tumors. I (64) — 
Other antidiabetic drugs 
Canagliflozin NCT04073680 Phase II, in combination with PI3K inhibitor in advanced solid tumors. I (62) — 
Rosiglitazone NCT04114136 Phase II, in combination with anti–PD-1 mAb in several advanced malignancies. I (63) — 
Statins 
Simvastatin NCT02104193 Phase II, in combination with radiotherapy in brain metastases. III, V (63) — 
Atorvastatin NCT02029573 Phase II, in combination with chemoradiotherapy in glioblastoma. I, III (66) — 
Atovaquone NCT02628080 Early phase I, as a hypoxia modifier in NSCLC. III (72) [18F]FAZA-PET 
Small-molecule mitochondrial inhibitors 
IACS-010759 NCT03291938, NCT02882321 Phase I, treatment of advanced solid and hematopoietic malignancies. I (75) — 
ME-344 NCT02806817 Early phase I, in combination with VEGF-A inhibitor in breast cancer. I (78) [18F]FDG-PET 
NO NCT01210378, NCT01171170 Phase II, as a radiosensitizer in NSCLC. Phase II, as a hypoxia modifier in NSCLC. IV (79) [18F]FDG-PET, [18F]FAZA-PET, [18F]HX4-PET 
Hormone therapy 
Tamoxifen NCT03280563, NCT00003857 Phase II, anti–PD-L1 mAb in progressed breast cancer, also combined with tamoxifen. Phase III, radiotherapy with or without tamoxifen in ductal carcinoma. I (84), III, IV, V (63) — 
Other FDA-approved drugs 
Arsenic trioxide NCT00045565 Phase II, in combination with radiotherapy for malignant glioma. IV (90) — 
Papaverine NCT03824327 Phase I, as a radiosensitizer in NSCLC and lung metastases. I (93) BOLD MRI 
Chlorpromazine NCT04224441 Phase II, for treatment of glioblastoma. V (63) — 
OXPHOS inhibitorSelected clinical trialsPurposeMitochondrial complex targetedFunctional imaging in these trials
Biguanides 
Metformin NCT04114136, NCT04275713, NCT02394652, many more trialsa Many trials in progress, including phase I and II trials combining metformin with anti–PD-1 mAbs, nivolumab or pembrolizumab, in several cancers. Also, several trials aiming to use the antihypoxic properties of metformin in combination with radiotherapy. I (54) [18F]FAZA-PET, DWI- and DCE-MRI 
Phenformin NCT03026517 Phase I, in combination with MEK and B-Raf inhibitors in metastatic melanoma. I (54) — 
IM156 NCT03272256 Phase I, in advanced solid tumors. I (64) — 
Other antidiabetic drugs 
Canagliflozin NCT04073680 Phase II, in combination with PI3K inhibitor in advanced solid tumors. I (62) — 
Rosiglitazone NCT04114136 Phase II, in combination with anti–PD-1 mAb in several advanced malignancies. I (63) — 
Statins 
Simvastatin NCT02104193 Phase II, in combination with radiotherapy in brain metastases. III, V (63) — 
Atorvastatin NCT02029573 Phase II, in combination with chemoradiotherapy in glioblastoma. I, III (66) — 
Atovaquone NCT02628080 Early phase I, as a hypoxia modifier in NSCLC. III (72) [18F]FAZA-PET 
Small-molecule mitochondrial inhibitors 
IACS-010759 NCT03291938, NCT02882321 Phase I, treatment of advanced solid and hematopoietic malignancies. I (75) — 
ME-344 NCT02806817 Early phase I, in combination with VEGF-A inhibitor in breast cancer. I (78) [18F]FDG-PET 
NO NCT01210378, NCT01171170 Phase II, as a radiosensitizer in NSCLC. Phase II, as a hypoxia modifier in NSCLC. IV (79) [18F]FDG-PET, [18F]FAZA-PET, [18F]HX4-PET 
Hormone therapy 
Tamoxifen NCT03280563, NCT00003857 Phase II, anti–PD-L1 mAb in progressed breast cancer, also combined with tamoxifen. Phase III, radiotherapy with or without tamoxifen in ductal carcinoma. I (84), III, IV, V (63) — 
Other FDA-approved drugs 
Arsenic trioxide NCT00045565 Phase II, in combination with radiotherapy for malignant glioma. IV (90) — 
Papaverine NCT03824327 Phase I, as a radiosensitizer in NSCLC and lung metastases. I (93) BOLD MRI 
Chlorpromazine NCT04224441 Phase II, for treatment of glioblastoma. V (63) — 

Abbreviations: DCE, dynamic contrast enhanced; DWI, diffusion-weighted imaging.

aMany trials are ongoing involving metformin as an anticancer therapeutic, trials listed here are exemplar for metformin as a hypoxia modifier or for the combination with immuno- and radiotherapy.

Figure 3.

Inhibitors of the electron transport chain. To generate ATP through OXPHOS, a proton gradient is maintained across the inner mitochondrial membrane as electrons (dotted line) pass through mitochondrial membrane complexes I–IV, coenzyme Q10 (CoQ), and cytochrome c (Cyt c). Because of this gradient, protons flow back over the membrane through the fifth mitochondrial complex, ATP synthase, converting ADP to ATP in the process. In this process, oxygen is used as a crucial electron acceptor in mitochondrial complex IV. If the electron transport chain comes to a standstill, for instance, by inhibition of one of the transmembrane protein complexes, the oxygen consumption will fall. This inhibition can be accomplished by several compounds that are under clinical investigation and are listed here. *Compounds targeting more than one of the mitochondrial membrane complexes.

Figure 3.

Inhibitors of the electron transport chain. To generate ATP through OXPHOS, a proton gradient is maintained across the inner mitochondrial membrane as electrons (dotted line) pass through mitochondrial membrane complexes I–IV, coenzyme Q10 (CoQ), and cytochrome c (Cyt c). Because of this gradient, protons flow back over the membrane through the fifth mitochondrial complex, ATP synthase, converting ADP to ATP in the process. In this process, oxygen is used as a crucial electron acceptor in mitochondrial complex IV. If the electron transport chain comes to a standstill, for instance, by inhibition of one of the transmembrane protein complexes, the oxygen consumption will fall. This inhibition can be accomplished by several compounds that are under clinical investigation and are listed here. *Compounds targeting more than one of the mitochondrial membrane complexes.

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Biguanides

Biguanides are a group of mitochondrial complex I–inhibiting molecules for treatment of type II diabetes mellitus (54). Metformin, one of the most commonly used biguanides, reduced hypoxia as measured by [18F]FAZA-PET imaging in LNCAP prostate cancer and HCT116 colorectal cancer xenografts or [18F]2-(4-((2-nitro-1H-imidazol-1-yl)methyl)-1H-1,2,3-triazol-1-yl)propan-1-ol ([18F]HX4)-PET imaging in A549 non–small cell lung cancer (NSCLC) and Colo205 colorectal cancer xenografts (8, 14, 55). Moreover, metformin induces a CD8+ T-cell–mediated antitumor effect by increasing the number of tumor-infiltrating lymphocytes and preventing T-cell exhaustion in the TME of several syngeneic tumor models (56). Also, metformin administered every 2 days intraperitoneally was able to potentiate PD-1 blockade in MC38 mouse colon cancer and B16 melanoma models by improving CD8+ T-cell function through upregulation of immunologic checkpoints, thereby reducing tumor growth (13). The closely related compound, phenformin, a more potent inhibitor of OXPHOS, shows a similar effect in BrafV600E-induced/Pten-deficient melanoma by inducing CD8+ T-cell infiltration and inhibiting MDSC function (57). Metformin and phenformin also increase radiotherapy efficacy by alleviating tumor hypoxia in the CT26 colorectal cancer model, presumably by both directly sensitizing tumor cells to irradiation, and the aforementioned immune effects (58). Recruiting trials, investigating a combined use with radiotherapy or immune checkpoint inhibition are listed in Table 1. Although putative overall and disease-free survival benefits of metformin as a radio- and immunotherapy enhancer have been reported, the overall results so far remain inconsistent (59–61). Other antidiabetic drugs, like canagliflozin (a sodium-glucose cotransporter 2 inhibitor) and rosiglitazone (a peroxisome proliferator-activated receptors activator), both affecting OXPHOS (62, 63), are currently under clinical investigation. Canagliflozin has been suggested as a radiosensitizer in a hypoxic A549 NSCLC xenograft model (62), while the rosiglitazone trial is performed in combination with PD-1 blockade (Table 1). IM156 is another newly developed biguanide that inhibits mitochondrial complex I and impairs the growth of MYC-dependent lymphoma in vivo (64). IM156 is currently in clinical trials for advanced solid tumors (Table 1).

Statins

Statins, used for reducing cholesterol in cardiovascular disease through 3-hydroxy-3-methylglutaryl-coA reductase inhibition, are also inhibitors of HIF1α activation (65). However, the precise mechanisms involved remain unclear. Simvastatin has been identified as a potent inhibitor of mitochondrial complexes III and V, thereby potentially reducing hypoxia and explaining the inhibition of HIF1α (63). Moreover, simvastatin-mediated inhibition of tumor growth is attributed to HIF1α reduction and the suppression of TAM-meditated oxidative stress in B16 melanoma (65). The related atorvastatin is also likely an inhibitor of OXPHOS (66), and has been found to attenuate the activation of the HIF1 pathway under hypoxic culture conditions and enhance radiosensitivity in prostate cancer cells in vitro (67). This suggests that statins may alleviate tumor hypoxia via the inhibition of oxidative metabolism. Postdiagnosis statin use, especially atorvastatin and simvastatin, was associated with an increased survival of patients with lung and pancreatic cancer, as well as of patients with prostate cancer treated with radiotherapy (68–70). Clinical trials combining simvastatin and atorvastatin with radiotherapy for brain tumors have been conducted and the atorvastatin trial met criteria for continued accrual (Table 1).

Atovaquone

Atovaquone, which is used to treat and prevent malaria, targets mitochondrial complex III and thereby collapses the malarial parasites mitochondrial membrane potential and energy supply (71). Atovaquone possesses the same inhibitory effect on mitochondrial complex III in MCF7 breast cancer cells (72). In pharynx and colorectal xenografts, administration of atovaquone daily via drinking water, decreased oxygen consumption, reduced tumor hypoxia, and sensitized tumors to radiotherapy (7). In addition, in a renal cell carcinoma xenograft model, the antitumor effect of the immunotherapeutic agent IFNα was potentiated by atovaquone (73). Furthermore, as a consequence of mitochondrial respiration inhibition, atovaquone has been shown to inhibit STAT3, a transcription factor involved in differentiation of immune suppressor cells, which may contribute to the reactivation of an immune response (74). Lately, atovaquone was investigated in clinical trials as a hypoxia modifier (Table 1).

Small-molecule mitochondrial inhibitors

IACS-010759 is a novel compound that potently and specifically inhibits mitochondrial complex I and can inhibit proliferation and induces apoptosis in tumor cells reliant on OXPHOS for their energetic needs (75). IACS-010759 is currently in clinical trials for advanced solid and hematopoietic malignancies (Table 1). Furthermore, daily IACS-010759 treatment showed to be effective in decreasing oxygen consumption and tumor hypoxia considerably in several xenograft models, which was illustrated by [18F]FAZA-PET imaging or pimonidazole staining (50, 53, 76, 77). Together, radiotherapy and IACS-010759 enhanced antitumor immunity and overcame PD-1 blockade resistance in a 344SQ murine lung cancer model, while triple therapy also promoted a systemic response and prolonged survival (18). Recently, another small-molecule inhibitor of mitochondrial complex I, ME-344 (78), has entered clinical trials in combination with bevacizumab, an antiangiogenic compound (Table 1). Results showed on target effects of ME-344, rendering it a promising OXPHOS inhibitor for combination therapies.

Nitric oxide

Nitric oxide (NO) has been suggested as a hypoxia modulator in cancer as it both decreases oxygen consumption (inhibiting OXPHOS) as well as induces vasodilatation (79, 80). NO inhibits mitochondrial complex IV and is administered via prodrugs, like nitroglycerine, via micellar systems, or other nanoscale carriers (80, 81). Daily in situ release of NO has proven to be effective in reducing tumor hypoxia, using [18F]fluoromisonidazole ([18F]FMISO)-PET imaging. It increases the levels of ROS formed by irradiation and improves the radiotherapeutic response in a Lewis lung carcinoma (LLC) model (81). In both B16 melanoma and 4T1 mammary carcinoma, hypoxia-induced PD-L1 expression was reduced by NO treatment, promoting T-cell–mediated lysis and inhibiting immune evasion (82). Moreover, in a prostate cancer xenograft model, NO reduced hypoxia and inhibited the abundance of TAMs (83), while NO increased the presence of tumor-infiltrating CD8+ and CD4+ T cells in murine HCA-1 hepatocellular carcinoma (80). The prodrug nitroglycerin, applied systemically, has been in clinical trials recently as a radiosensitizing agent (Table 1), but no survival benefit has been reported so far. A targeted approach using precision delivery of NO combined with radiotherapy yielded better results in preclinical models (81).

Hormone therapy

The most commonly used drug for estrogen receptor (ER)-positive breast cancer, tamoxifen, has been identified as a potent ER-independent inhibitor of mitochondrial complexes III–V (63), as well as mitochondrial complex I (84). By this mechanism, tamoxifen reduced oxygen consumption in an ER-negative MDA-MB231-LM2-4175 breast cancer xenograft (84). Targeted tamoxifen delivery reduced hypoxia 24 hours after administration, which increased the efficacy of photodynamic therapy in a mouse 4T1 breast cancer model, and suggest the same approach could be used to enhance radiotherapeutic response (85). In MCF7 breast cancer xenografts, however, the antiangiogenic and thrombogenic properties of tamoxifen induced hypoxia, underlining that its antihypoxic effect might be tumor dependent (86). In vitro, MCF7 cells were sensitized by tamoxifen to radiotherapy and tamoxifen is currently in clinical trials to elucidate whether the combination with radiotherapy can be beneficial for patient outcome (Table 1; ref. 87). The possible hypoxia-reducing properties of tamoxifen also render it a potential adjuvant for immune checkpoint blockade or other immune-stimulating therapies (88). Currently, a clinical trial (Table 1) is studying PD-L1 inhibition in patients with progressed breast cancer who are also treated with endocrine therapies, including tamoxifen, possibly enhancing the immunotherapeutic response.

Other FDA-approved drugs

Several other drugs, already approved for clinical use, have been accredited with the disruption of OXPHOS. An example is arsenic trioxide, used for the treatment of acute promyelocytic leukemia, which has also been suggested for the treatment of lung cancer, partly because of its potential radiosensitizing and immunostimulatory effects (89). In syngeneic transplantable liver tumor (TLT) and LLC models, treatment with arsenic trioxide reduced oxygen consumption and increased tissue oxygenation up to 2 hours after administration as measured by electron paramagnetic resonance oximetry, while also augmenting radiotherapy response in TLTs (90). Furthermore, arsenic trioxide treatment has been associated with the depletion of immunosuppressive cell types, like MDSCs and Tregs, while enhancing the antitumor activity of CD8+ T cells in several syngeneic mouse models, posing it as an immune activator (91, 92). Completed trials tested the efficacy of combining arsenic trioxide with radiotherapy, but no conclusive results have been reported yet (Table 1). Papaverine, an antispasmodic drug, was found to be a mitochondrial complex I inhibitor, causing reduced tumor hypoxia and increased radiotherapy response in A549 NSCLC and EO771 mammary cancer xenografts (93). Currently, papaverine is in phase I clinical trials as a radiosensitizing agent (Table 1). Finally, chlorpromazine, an antipsychotic dopamine antagonist with antiserotonergic and antihistaminergic properties, also inhibits OXPHOS (63) and is being investigated in the treatment of glioblastoma multiforme (Table 1).

Several trials have demonstrated that hypoxia-modifying agents can improve radiotherapy outcome, although these effects are overall modest (4). Clinical studies to validate whether hypoxia modifiers can enhance immunotherapy efficacy are needed and have only started recently. In this respect, measuring tumor hypoxia will be of utmost importance as most previous trials have not assessed patients on this basis and possibly overlooked beneficial effects (94). Monitoring hypoxic status can be used to select hypoxic patients, as tumors with pronounced hypoxia are more likely to respond to the proposed combination therapies. Furthermore, monitoring of hypoxia is important to validate whether the hypoxia-modifying treatment has the intended on-target effects. This is essential to better interpret the results of these clinical trials (95). Imaging, as opposed to taking a biopsy, is a preferred approach as it can assess the hypoxic status of the entire tumor, which is known to be very heterogeneous (95). Functional imaging also allows for noninvasive monitoring of hypoxic status over time, which can be helpful to determine follow-up changes in hypoxia during OXPHOS inhibition, both in clinical and preclinical settings. Several methods for imaging hypoxia have made their way into the clinic already (96). The most commonly applied methods to date involve PET using redox-sensitive probes, of which nitroimidazoles are currently the most used class. Of these, [18F]HX4-PET (14, 55), [18F]FAZA-PET (50, 76), and [18F]FMISO-PET (81) have been used to asses hypoxia-alleviating effects of OXPHOS inhibitors in preclinical models, while [18F]FAZA-PET and [18F]HX4-PET are also used in some clinical trials (Table 1). Alternatively, targeting an endogenous hypoxia marker could be advantageous as it visualizes tumor cells that adapted to the harsh hypoxic TME. A loss of signal in this case would represent a durable immune-permissive reprogramming of the TME. In this respect, carbonic anhydrase IX is a promising marker and is currently under preclinical investigation as an imaging target (97).

Several preclinical studies have shown that the inhibition of OXPHOS can structurally alleviate hypoxia by repressing oxygen consumption, thereby increasing the efficacy of both radio- and immunotherapy and its combination. Although these results are very promising in preclinical models, hypoxia-alleviating effects and subsequent survival benefits have yet to be proven in the clinic. OXPHOS inhibition to alleviate hypoxia will most likely only benefit patients with tumor hypoxia because of an enhanced oxygen consumption. Furthermore, the immunosuppressive action caused by acidosis of the TME as a consequence of glycolytic metabolism will not be eliminated by OXPHOS inhibition, and may even be intensified by inhibiting mitochondrial metabolism. Another point of discussion is the influence of tumor mitochondrial metabolism on immunotherapy efficacy, as upregulated OXPHOS has been described to be associated with immunotherapy response (98), as well as resistance (18, 44, 52, 53). A possible explanation could be that oxidative metabolism is not in all cases correlated with tumor hypoxia, once more emphasizing the complexity of cancer biology and the importance of personalized medicine.

Several OXPHOS inhibiting compounds have made their way into clinical testing. Commonly used drugs, such as biguanides and statins, have shown a beneficial effect for patients with cancer in retrospective studies. They may be easy to implement in clinical practice, as are other commonly used drugs, like tamoxifen, NO, and arsenic trioxide. However, as these so-called “dirty drugs” are not very selective, they might have significant off-target effects (99) and it could be hard to pinpoint whether the reason for alleviated immunosuppression is a sustained increase in tissue oxygenation. Furthermore, clinical trials focused on the validation of hypoxia modification by these drugs are still needed. To date, only a few clinical trials have incorporated functional imaging to confirm antihypoxic effects and fewer use imaging for patient selection. More potent and specific mitochondrial inhibitors, like atovaquone and IACS-010759, have shown to effectively alleviate hypoxia and increased the efficacy of immunotherapy in preclinical studies, suggesting this is because of a sustained increase in tissue oxygenation (18, 73). These drugs might be better candidates for a targeted antihypoxia approach and together with appropriate imaging and response monitoring lead the way to increased clinical benefit in selected patients when combined with radiotherapy and immunotherapy.

This review describes how the inhibition of OXPHOS may be a promising strategy to structurally alleviate hypoxia and reactivate an antitumor immune response. Several OXPHOS inhibitors have shown promising preclinical results and have proceeded into clinical trials. Meticulous patient selection and hypoxia monitoring, however, are crucial to validate their hypoxia-modifying effects, these are absent in many trials. Such future clinical studies should show the value of OXPHOS-inhibiting compounds in improving the efficacy of radio- and immunotherapy combinations.

No potential conflicts of interest were disclosed.

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

1.
Span
PN
,
Bussink
J
. 
Biology of hypoxia
.
Semin Nucl Med
2015
;
45
:
101
9
.
2.
Meijer
TWH
,
Kaanders
JHAM
,
Span
PN
,
Bussink
J
. 
Targeting hypoxia, HIF-1, and tumor glucose metabolism to improve radiotherapy efficacy
.
Clin Cancer Res
2012
;
18
:
5585
94
.
3.
Kaanders
JHAM
,
Bussink
J
,
Van Der Kogel
AJ
. 
Clinical studies of hypoxia modification in radiotherapy
.
Semin Radiat Oncol
2004
;
14
:
233
40
.
4.
Overgaard
J
. 
Hypoxic modification of radiotherapy in squamous cell carcinoma of the head and neck - a systematic review and meta-analysis
.
Radiother Oncol
2011
;
100
:
22
32
.
5.
Joiner
MC
,
van der Kogel
AJ
.
Basic clinical radiobiology
. 5th ed.
Boca Raton, FL:
CRC Press
; 
2018
.
6.
Janssens
GO
,
Rademakers
SE
,
Terhaard
CH
,
Doornaert
PA
,
Bijl
HP
,
van den Ende
P
, et al
Accelerated radiotherapy with carbogen and nicotinamide for laryngeal cancer: results of a phase III randomized trial
.
J Clin Oncol
2012
;
30
:
1777
83
.
7.
Ashton
TM
,
Fokas
E
,
Kunz-Schughart
LA
,
Folkes
LK
,
Anbalagan
S
,
Huether
M
, et al
The anti-malarial atovaquone increases radiosensitivity by alleviating tumour hypoxia
.
Nat Commun
2016
;
7
:
1
13
.
8.
Zannella
VE
,
Pra
AD
,
Muaddi
H
,
McKee
TD
,
Stapleton
S
,
Sykes
J
, et al
Reprogramming metabolism with metformin improves tumor oxygenation and radiotherapy response
.
Clin Cancer Res
2013
;
19
:
6741
50
.
9.
Gaipl
US
,
Multhoff
G
,
Scheithauer
H
,
Lauber
K
,
Hehlgans
S
,
Frey
B
, et al
Kill and spread the word: stimulation of antitumor immune responses in the context of radiotherapy
.
Immunotherapy
2014
;
6
:
597
610
.
10.
Damgaci
S
,
Ibrahim-hashim
A
,
Enriquez-navas
PM
,
Pilon-thomas
S
,
Guvenis
A
. 
Hypoxia and acidosis: immune suppressors and therapeutic targets
.
Immunology
2018
;
154
:
354
62
.
11.
Movahedi
K
,
Laoui
D
,
Gysemans
C
,
Baeten
M
,
Stangé
G
,
Van Den Bossche
J
, et al
Different tumor microenvironments contain functionally distinct subsets of macrophages derived from Ly6C(high) monocytes
.
Cancer Res
2010
;
70
:
5728
39
.
12.
Hatfield
SM
,
Kjargaard
J
,
Lukashev
D
,
Schreiber
TH
,
Belikoff
B
,
Abbott
R
, et al
Immunological mechanisms of the antitumor effects of supplemental oxygenation
.
Sci Transl Med
2015
;
7
:
263
9
.
13.
Scharping
NE
,
Menk
AV
,
Whetstone
RD
,
Zeng
X
,
Delgoffe
GM
. 
Efficacy of PD-1 blockade is potentiated by metformin-induced reduction of tumor hypoxia
.
Physiol Behav
2017
;
176
:
139
48
.
14.
De Bruycker
S
,
Vangestel
C
,
Van Den Wyngaert
T
,
Pauwels
P
,
Wyffels
L
,
Staelens
S
, et al
18 F-flortanidazole hypoxia PET holds promise as a prognostic and predictive imaging biomarker in a lung cancer xenograft model treated with metformin and radiotherapy
.
J Nucl Med
2019
;
60
:
34
40
.
15.
Vasan
K
,
Werner
M
,
Chandel
NS
. 
Mitochondrial metabolism as a target for cancer therapy
.
Cell Metab
2020
;
32
:
341
52
.
16.
Faubert
B
,
Solmonson
A
,
DeBerardinis
RJ
. 
Metabolic reprogramming and cancer progression
.
Science
2020
;
368
:
eaaw5473
.
17.
Ždralević
M
,
Brand
A
,
Ianni Di
L
,
Dettmer
K
,
Reinders
J
,
Singer
K
, et al
Double genetic disruption of lactate dehydrogenases A and B is required to ablate the “Warburg effect” restricting tumor growth to oxidative metabolism
.
J Biol Chem
2018
;
293
:
15947
61
.
18.
Chen
D
,
Barsoumian
HB
,
Fischer
G
,
Yang
L
,
Verma
V
,
Younes
AI
, et al
Combination treatment with radiotherapy and a novel oxidative phosphorylation inhibitor overcomes PD-1 resistance and enhances antitumor immunity
.
J Immunother Cancer
2020
;
8
:
e000289
.
19.
Wang
GL
,
Jiang
BH
,
Rue
EA
,
Semenza
GL
. 
Hypoxia-inducible factor 1 is a basic-helix-loop-helix-PAS heterodimer regulated by cellular O2 tension
.
Proc Natl Acad Sci U S A
1995
;
92
:
5510
4
.
20.
Wouters
BG
,
Koritzinsky
M
. 
Hypoxia signalling through mTOR and the unfolded protein response in cancer
.
Nat Rev Cancer
2008
;
8
:
851
64
.
21.
Palazon
A
,
Goldrath
AW
,
Nizet
V
,
Johnson
RS
. 
HIF transcription factors, inflammation, and immunity
.
Immunity
2014
;
41
:
518
28
.
22.
Eltzschig
HK
,
Thompson
LF
,
Karhausen
J
,
Cotta
RJ
,
Ibla
JC
,
Robson
SC
, et al
Endogenous adenosine produced during hypoxia attenuates neutrophil accumulation: Coordination by extracellular nucleotide metabolism
.
Blood
2004
;
104
:
3986
92
.
23.
Ansems
M
,
Span
PN
. 
The tumor microenvironment and radiotherapy response; a central role for cancer-associated fibroblasts
.
Clin Transl Radiat Oncol
2020
;
22
:
90
7
.
24.
Rabold
K
,
Aschenbrenner
A
,
Thiele
C
,
Boahen
CK
,
Schiltmans
A
,
Smit
JWA
, et al
Enhanced lipid biosynthesis in human tumor-induced macrophages contributes to their protumoral characteristics
.
J Immunother cancer
2020
;
8
:
1
13
.
25.
Vito
A
,
El-Sayes
N
,
Mossman
K
. 
Hypoxia-driven immune escape in the tumor microenvironment
.
Cells
2020
;
9
:
992
.
26.
Thiel
M
,
Caldwell
CC
,
Kreth
S
,
Kuboki
S
,
Ghen
P
,
Smith
P
, et al
Targeted deletion of HIF-1α gene in T cells prevents their inhibition in hypoxic inflamed tissues and improves septic mice survival
.
PLoS One
2007
;
2
:
1
7
.
27.
Jayaprakash
P
,
Ai
M
,
Liu
A
,
Budhani
P
,
Bartkowiak
T
,
Sheng
J
, et al
Targeted hypoxia reduction restores T cell infiltration and sensitizes prostate cancer to immunotherapy
.
J Clin Invest
2018
;
128
:
5137
49
.
28.
Chiu
DKC
,
Tse
APW
,
Xu
IMJ
,
Di Cui
J
,
Lai
RKH
,
Li
LL
, et al
Hypoxia inducible factor HIF-1 promotes myeloid-derived suppressor cells accumulation through ENTPD2/CD39L1 in hepatocellular carcinoma
.
Nat Commun;
2017
;
8
:
1
12
.
29.
Westendorf
AM
,
Skibbe
K
,
Adamczyk
A
,
Buer
J
,
Geffers
R
,
Hansen
W
, et al
Hypoxia enhances immunosuppression by inhibiting CD4+ effector T cell function and promoting Treg activity
.
Cell Physiol Biochem
2017
;
41
:
1271
84
.
30.
Bohn
T
,
Rapp
S
,
Luther
N
,
Klein
M
,
Bruehl
TJ
,
Kojima
N
, et al
Tumor immunoevasion via acidosis-dependent induction of regulatory tumor-associated macrophages
.
Nat Immunol
2018
;
19
:
1319
29
.
31.
Miska
J
,
Lee-Chang
C
,
Rashidi
A
,
Muroski
ME
,
Chang
AL
,
Lopez-Rosas
A
, et al
HIF-1α is a metabolic switch between glycolytic-driven migration and oxidative phosphorylation-driven immunosuppression of Tregs in glioblastoma
.
Cell Rep
2019
;
27
:
226
37
.
32.
de Almeida
PE
,
Mak
J
,
Hernandez
G
,
Jesudason
R
,
Herault
A
,
Javinal
V
, et al
Anti-VEGF treatment enhances CD8+ T-cell antitumor activity by amplifying hypoxia
.
Cancer Immunol Res
2020
;
8
:
806
18
.
33.
Gropper
Y
,
Feferman
T
,
Shalit
T
,
Salame
TM
,
Porat
Z
,
Shakhar
G
. 
Culturing CTLs under hypoxic conditions enhances their cytolysis and improves their anti-tumor function
.
Cell Rep
2017
;
20
:
2547
55
.
34.
Doedens
AL
,
Phan
AT
,
Stradner
MH
,
Fujimoto
JK
,
Nguyen
JV
,
Yang
E
, et al
Hypoxia-inducible factors enhance the effector responses of CD8+ T cells to persistent antigen
.
Nat Immunol
2014
;
14
:
1173
82
.
35.
Iwai
Y
,
Ishida
M
,
Tanaka
Y
,
Okazaki
T
,
Honjo
T
,
Minato
N
. 
Involvement of PD-L1 on tumor cells in the escape from host immune system and tumor immunotherapy by PD-L1 blockade
.
Proc Natl Acad Sci U S A
2002
;
99
:
12293
7
.
36.
Nishimura
H
,
Nose
M
,
Hiai
H
,
Minato
N
,
Honjo
T
. 
Development of lupus-like autoimmune diseases by disruption of the PD-1 gene encoding an ITIM motif-carrying immunoreceptor
.
Immunity
1999
;
11
:
141
51
.
37.
Noman
MZ
,
Desantis
G
,
Janji
B
,
Hasmim
M
,
Karray
S
,
Dessen
P
, et al
PD-L1 is a novel direct target of HIF-1α, and its blockade under hypoxia enhanced: MDSC-mediated T cell activation
.
J Exp Med
2014
;
211
:
781
90
.
38.
Ohta
A
,
Kini
R
,
Ohta
A
,
Subramanian
M
,
Madasu
M
,
Sitkovsky
M
. 
The development and immunosuppressive functions of CD4+ CD25+ FoxP3+ regulatory T cells are under influence of the adenosine-A2A adenosine receptor pathway
.
Front Immunol
2012
;
3
:
1
12
.
39.
Deng
J
,
Li
J
,
Sarde
A
,
Lines
JL
,
Lee
YC
,
Qian
DC
, et al
Hypoxia-induced VISTA promotes the suppressive function of myeloid-derived suppressor cells in the tumor microenvironment
.
Cancer Immunol Res
2019
;
7
:
1079
90
.
40.
Koh
HS
,
Chang
CY
,
Jeon
SB
,
Yoon
HJ
,
Ahn
YH
,
Kim
HS
, et al
The HIF-1/glial TIM-3 axis controls inflammation-associated brain damage under hypoxia
.
Nat Commun
2015
;
6
:
1
15
.
41.
Zhang
H
,
Lu
H
,
Xiang
L
,
Bullen
JW
,
Zhang
C
,
Samanta
D
, et al
HIF-1 regulates CD47 expression in breast cancer cells to promote evasion of phagocytosis and maintenance of cancer stem cells
.
Proc Natl Acad Sci U S A
2015
;
112
:
E6215
23
.
42.
Jacobs
JFM
,
Nierkens
S
,
Figdor
CG
,
de Vries
IJM
,
Adema
GJ
. 
Regulatory T cells in melanoma: the final hurdle towards effective immunotherapy?
Lancet Oncol
2012
;
13
:
32
42
.
43.
Lindau
D
,
Gielen
P
,
Kroesen
M
,
Wesseling
P
,
Adema
GJ
. 
The immunosuppressive tumour network: myeloid-derived suppressor cells, regulatory T cells and natural killer T cells
.
Immunology
2013
;
138
:
105
15
.
44.
Najjar
YG
,
Menk
AV
,
Sander
C
,
Rao
U
,
Karunamurthy
A
,
Bhatia
R
, et al
Tumor cell oxidative metabolism as a barrier to PD-1 blockade immunotherapy in melanoma
.
JCI Insight
2019
;
4
:
1
11
.
45.
Kjargaard
J
,
Hatfield
S
,
Jones
G
,
Ohta
A
,
Sitkovsky
M
. 
A2A adenosine receptor gene-deletion or synthetic A2A antagonist liberate tumor-reactive CD8+ T-cells from tumor-induced immunosuppression
.
J Immunol
2018
;
201
:
782
91
.
46.
Barsoum
IB
,
Koti
M
,
Siemens
DR
,
Graham
CH
. 
Mechanisms of hypoxia-mediated immune escape in cancer
.
Cancer Res
2014
;
74
:
7185
91
.
47.
Noman
MZ
,
Hasmim
M
,
Messai
Y
,
Terry
S
,
Kieda
C
,
Janji
B
, et al
Hypoxia: a key player in antitumor immune response. A review in the theme: cellular responses to hypoxia
.
Am J Physiol Cell Physiol
2015
;
309
:
C569
79
.
48.
Bussink
J
,
Kaanders
JHAM
,
Strik
AM
,
van der Kogel
AJ
. 
Effects of nicotinamide and carbogen on oxygenation in human tumor xenografts measured with luminescence based fiber-optic probes
.
Radiother Oncol
2000
;
57
:
21
30
.
49.
Secomb
TW
,
Hsu
R
,
Ong
ET
,
Gross
JF
,
Dewhirst
MW
. 
Analysis of the effects of oxygen supply and demand on hypoxic fraction in tumors
.
Acta Oncol
1995
;
34
:
313
6
.
50.
Gammon
ST
,
Pisaneschi
F
,
Bandi
ML
,
Smith
MG
,
Sun
Y
,
Rao
Y
, et al
Mechanism-specific pharmacodynamics of a novel complex-I inhibitor quantified by imaging reversal of consumptive hypoxia with [18F]FAZA PET in vivo
.
Cells
2019
;
8
:
1487
.
51.
Newton
JM
,
Hanoteau
A
,
Liu
H
,
Gaspero
A
,
Parikh
F
,
Gartrell-Corrado
RD
, et al
Immune microenvironment modulation unmasks therapeutic benefit of radiotherapy and checkpoint inhibition
.
J Immunother Cancer
2019
;
7
:
216
.
52.
Jaiswal
AR
,
Liu
AJ
,
Pudakalakatti
S
,
Dutta
P
,
Jayaprakash
P
,
Bartkowiak
T
, et al
Melanoma evolves complete immunotherapy resistance through the acquisition of a hypermetabolic phenotype
.
Cancer Immunol Res
2020
;
8
:
1365
80
.
53.
Fischer
GM
,
Jalili
A
,
Kircher
DA
,
Lee
W-C
,
McQuade
JL
,
Haydu
LE
, et al
Molecular profiling reveals unique immune and metabolic features of melanoma brain metastases
.
Cancer Discov
2019
;
9
:
628
645
.
54.
Bridges
HR
,
Jones
AJY
,
Pollak
MN
,
Hirst
J
. 
Effects of metformin and other biguanides on oxidative phosphorylation in mitochondria
.
Biochem J
2014
;
462
:
475
87
.
55.
De Bruycker
S
,
Vangestel
C
,
Staelens
S
,
wyffels
L
,
Detrez
J
,
Verschuuren
M
, et al
Effects of metformin on tumor hypoxia and radiotherapy efficacy: a [18F]HX4 PET imaging study in colorectal cancer xenografts
.
EJNMMI Res
2019
;
9
:
74
.
56.
Eikawa
S
,
Nishida
M
,
Mizukami
S
,
Yamazaki
C
,
Nakayama
E
,
Udono
H
. 
Immune-mediated antitumor effect by type 2 diabetes drug, metformin
.
Proc Natl Acad Sci U S A
2015
;
112
:
1809
14
.
57.
Kim
SH
,
Li
M
,
Trousil
S
,
Zhang
Y
,
Pasca di Magliano
M
,
Swanson
KD
, et al
Phenformin inhibits myeloid-derived suppressor cells and enhances the anti-tumor activity of PD-1 blockade in melanoma
.
J Invest Dermatol
2017
;
137
:
1740
8
.
58.
De Mey
S
,
Jiang
H
,
Corbet
C
,
Wang
H
,
Dufait
I
,
Law
K
, et al
Antidiabetic biguanides radiosensitize hypoxic colorectal cancer cells through a decrease in oxygen consumption
.
Front Pharmacol
2018
;
9
:
1073
.
59.
Afzal
MZ
,
Mercado
RR
,
Shirai
K
. 
Efficacy of metformin in combination with immune checkpoint inhibitors (anti-PD-1/anti-CTLA-4) in metastatic malignant melanoma
.
J Immunother Cancer
2018
;
6
:
1
10
.
60.
Afzal
MZ
,
Dragnev
K
,
Sarwar
T
,
Shirai
K
. 
Clinical outcomes in non-small-cell lung cancer patients receiving concurrent metformin and immune checkpoint inhibitors
.
Lung Cancer Manag
2019
;
8
:
LMT11
.
61.
Rao
M
,
Gao
C
,
Guo
M
,
Law
BYK
,
Xu
Y
. 
Effects of metformin treatment on radiotherapy efficacy in patients with cancer and diabetes: a systematic review and meta-analysis
.
Cancer Manag Res
2018
;
10
:
4881
90
.
62.
Angelopoulou
A
,
Kolokithas-Ntoukas
A
,
Papaioannou
L
,
Kakazanis
Z
,
Khoury
N
,
Zoumpourlis
V
, et al
Canagliflozin-loaded magnetic nanoparticles as potential treatment of hypoxic tumors in combination with radiotherapy
.
Nanomedicine
2018
;
13
:
243
54
.
63.
Nadanaciva
S
,
Bernal
A
,
Aggeler
R
,
Capaldi
R
,
Will
Y
. 
Target identification of drug induced mitochondrial toxicity using immunocapture based OXPHOS activity assays
.
Toxicol Vitr
2007
;
21
:
902
11
.
64.
Izreig
S
,
Gariepy
A
,
Kaymak
I
,
Bridges
HR
,
Donayo
AO
,
Bridon
G
, et al
Repression of LKB1 by miR-17∼92 sensitizes MYC-dependent lymphoma to biguanide treatment
.
Cell Reports Med
2020
;
1
:
100014
.
65.
Alupei
MC
,
Licarete
E
,
Patras
L
,
Banciu
M
. 
Liposomal simvastatin inhibits tumor growth via targeting tumor-associated macrophages-mediated oxidative stress
.
Cancer Lett
2015
;
356
:
946
52
.
66.
Broniarek
I
,
Jarmuszkiewicz
W
. 
Atorvastatin affects negatively respiratory function of isolated endothelial mitochondria
.
Arch Biochem Biophys
2018
;
637
:
64
72
.
67.
Chen
B
,
Zhang
M
,
Xing
D
,
Feng
Y
. 
Atorvastatin enhances radiosensitivity in hypoxia-induced prostate cancer cells related with HIF-1α inhibition
.
Biosci Rep
2017
;
37
:
1
7
.
68.
Li
K
,
Si-Tu
J
,
Qiu
J
,
Lu
L
,
Mao
Y
,
Zeng
H
, et al
Statin and metformin therapy in prostate cancer patients with hyperlipidemia who underwent radiotherapy: a population-based cohort study
.
Cancer Manag Res
2019
;
11
:
1189
97
.
69.
Huang
BZ
,
Chang
JI
,
Li
E
,
Xiang
AH
,
Wu
BU
. 
Influence of statins and cholesterol on mortality among patients with pancreatic cancer
.
J Natl Cancer Inst
2017
;
109
:
1
8
.
70.
Ung
MH
,
MacKenzie
TA
,
Onega
TL
,
Amos
CI
,
Cheng
C
. 
Statins associated with improved mortality among patients with certain histological subtypes of lung cancer
.
Lung Cancer
2018
;
126
:
89
96
.
71.
Srivastava
IK
,
Rottenberg
H
,
Vaidya
AB
. 
Atovaquone, a broad spectrum antiparasitic drug, collapses mitochondrial membrane potential in a malarial parasite
.
J Biol Chem
1997
;
272
:
3961
6
.
72.
Fiorillo
M
,
Lamb
R
,
Tanowitz
HB
,
Mutti
L
,
Krstic-Demonacos
M
,
Cappello
AR
, et al
Repurposing atovaquone: targeting mitochondrial complex III and OXPHOS to eradicate cancer stem cells
.
Oncotarget
2016
;
7
:
34084
99
.
73.
Chen
D
,
Sun
X
,
Zhang
X
,
Cao
J
. 
Targeting mitochondria by anthelmintic drug atovaquone sensitizes renal cell carcinoma to chemotherapy and immunotherapy
.
J Biochem Mol Toxicol
2018
;
32
:
1
7
.
74.
Lv
Z
,
Yan
X
,
Lu
L
,
Su
C
,
He
Y
. 
Atovaquone enhances doxorubicin's efficacy via inhibiting mitochondrial respiration and STAT3 in aggressive thyroid cancer
.
J Bioenerg Biomembr
2018
;
50
:
263
70
.
75.
Molina
JR
,
Sun
Y
,
Protopopova
M
,
Gera
S
,
Bandi
M
,
Bristow
C
, et al
An inhibitor of oxidative phosphorylation exploits cancer vulnerability
.
Nat Med
2018
;
24
:
1036
46
.
76.
Vashisht Gopal
YN
,
Gammon
ST
,
Prasad
R
,
Knighton
B
,
Pisaneschi
F
,
Roszik
J
, et al
A novel mitochondrial inhibitor blocks MAPK pathway and overcomes MAPK inhibitor-resistance in melanoma
.
Clin Cancer Res
2019
;
25
:
6429
42
.
77.
Sun
Y
,
Bandi
M
,
Lofton
T
,
Jones
P
,
Heffernan
TP
,
Marszalek
JR
, et al
Functional genomics reveals synthetic lethality between phosphogluconate dehydrogenase and oxidative phosphorylation
.
Cell Rep
2019
;
26
:
469
82
78.
Lim
SC
,
Carey
KT
,
McKenzie
M
. 
Anti-cancer analogues ME-143 and ME-344 exert toxicity by directly inhibiting mitochondrial NADH: ubiquinone oxidoreductase (complex I)
.
Am J Cancer Res
2015
;
5
:
689
701
.
79.
Clementi
E
,
Brown
GC
,
Foxwell
N
,
Moncada
S
. 
On the mechanism by which vascular endothelial cells regulate their oxygen consumption
.
Proc Natl Acad Sci U S A
1999
;
96
:
1559
62
.
80.
Sung
YC
,
Jin
PR
,
Chu
LA
,
Hsu
FF
,
Wang
MR
,
Chang
CC
, et al
Delivery of nitric oxide with a nanocarrier promotes tumour vessel normalization and potentiates anti-cancer therapies
.
Nat Nanotechnol
2019
;
14
:
1160
9
.
81.
Tu
J
,
Tu
K
,
Xu
H
,
Wang
L
,
Yuan
X
,
Qin
X
, et al
Improving tumor hypoxia and radiotherapy resistance via in situ nitric oxide release strategy
.
Eur J Pharm Biopharm
2020
;
150
:
96
107
.
82.
Barsoum
IB
,
Smallwood
CA
,
Siemens
DR
,
Graham
CH
. 
A mechanism of hypoxia-mediated escape from adaptive immunity in cancer cells
.
Cancer Res
2014
;
74
:
665
74
.
83.
Arora
H
,
Panara
K
,
Kuchakulla
M
,
Kulandavelu
S
,
Burnstein
KL
,
Schally
AV
, et al
Alterations of tumor microenvironment by nitric oxide impedes castration-resistant prostate cancer growth
.
Proc Natl Acad Sci U S A
2018
;
115
:
11298
303
.
84.
Daurio
NA
,
Tuttle
SW
,
Worth
AJ
,
Song
EY
,
Davis
JM
,
Snyder
NW
, et al
AMPK activation and metabolic reprogramming by tamoxifen through estrogen receptor-independent mechanisms suggests new uses for this therapeutic modality in cancer treatment
.
Cancer Res
2016
;
76
:
3295
306
.
85.
Yang
Z
,
Chen
Q
,
Chen
J
,
Dong
Z
,
Zhang
R
,
Liu
J
, et al
Tumor-pH-responsive dissociable albumin–tamoxifen nanocomplexes enabling efficient tumor penetration and hypoxia relief for enhanced cancer photodynamic therapy
.
Small
2018
;
14
:
1
10
.
86.
Evans
SM
,
Koch
CJ
,
Laughlin
KM
,
Jenkins
WT
,
Van Winkle
T
,
Wilson
DF
. 
Tamoxifen induces hypoxia in MCF-7 xenografts
.
Cancer Res.
1997
;
57
:
5155
61
.
87.
Zeng
ZJ
,
Li
JH
,
Zhang
YJ
,
Zhao
ST
. 
Optimal combination of radiotherapy and endocrine drugs in breast cancer treatment
.
Cancer Radiother
2013
;
17
:
208
14
.
88.
Recchia
F
,
Sica
G
,
Candeloro
G
,
Necozione
S
,
Bisegna
R
,
Bratta
M
, et al
Beta-interferon, retinoids and tamoxifen in metastatic breast cancer: long-term follow-up of a phase II study
.
Oncol Rep
2009
;
21
:
1011
6
.
89.
Huang
W
,
Zeng
YC
. 
A candidate for lung cancer treatment: arsenic trioxide
.
Clin Transl Oncol
2019
;
21
:
1115
26
.
90.
Diepart
C
,
Karroum
O
,
Magat
J
,
Feron
O
,
Verrax
J
,
Calderon
PB
, et al
Arsenic trioxide treatment decreases the oxygen consumption rate of tumor cells and radiosensitizes solid tumors
.
Cancer Res
2012
;
72
:
482
90
.
91.
Gao
Q
,
Jiang
J
,
Chu
Z
,
Lin
H
,
Zhou
X
,
Liang
X
. 
Arsenic trioxide inhibits tumor-induced myeloid-derived suppressor cells and enhances T-cell activity
.
Oncol Lett
2017
;
13
:
2141
50
.
92.
Thomas-Schoemann
A
,
Batteux
F
,
Mongaret
C
,
Nicco
C
,
Chéreau
C
,
Annereau
M
, et al
Arsenic trioxide exerts antitumor activity through regulatory t cell depletion mediated by oxidative stress in a murine model of colon cancer
.
J Immunol
2012
;
189
:
5171
7
.
93.
Benej
M
,
Hong
X
,
Vibhute
S
,
Scott
S
,
Wu
J
,
Graves
E
, et al
Papaverine and its derivatives radiosensitize solid tumors by inhibiting mitochondrial metabolism
.
Proc Natl Acad Sci U S A
2018
;
115
:
10756
61
.
94.
Walsh
JC
,
Lebedev
A
,
Aten
E
,
Madsen
K
,
Marciano
L
,
Kolb
HC
. 
The clinical importance of assessing tumor hypoxia: relationship of tumor hypoxia to prognosis and therapeutic opportunities
.
Antioxidants Redox Signal
2014
;
21
:
1516
54
.
95.
Bussink
J
,
van Herpen
CML
,
Kaanders
JHAM
,
Oyen
WJG
. 
PET-CT for response assessment and treatment adaptation in head and neck cancer
.
Lancet Oncol
2010
;
11
:
661
9
.
96.
Bonnitcha
P
,
Grieve
S
,
Figtree
G
. 
Clinical imaging of hypoxia: current status and future directions
.
Free Radic Biol Med
2018
;
126
:
296
312
.
97.
Huizing
FJ
,
Hoeben
BAWW
,
Franssen
GM
,
Boerman
OC
,
Heskamp
S
,
Bussink
J
. 
Quantitative imaging of the hypoxia-related marker CAIX in head and neck squamous cell carcinoma xenograft models
.
Mol Pharm
2019
;
16
:
701
8
.
98.
Harel
M
,
Ortenberg
R
,
Varanasi
SK
,
Mangalhara
KC
,
Mardamshina
M
,
Markovits
E
, et al
Proteomics of melanoma response to immunotherapy reveals mitochondrial dependence
.
Cell
2019
;
179
:
236
50
.
99.
Jin
Y
,
Xu
K
,
Chen
Q
,
Wang
B
,
Pan
J
,
Huang
S
, et al
Simvastatin inhibits the development of radioresistant esophageal cancer cells by increasing the radiosensitivity and reversing EMT process via the PTEN-PI3K/AKT pathway
.
Exp Cell Res
2018
;
362
:
362
9
.