Oxidative stress in pituitary neuroendocrine tumors: Affecting the tumor microenvironment and becoming a new target for pituitary neuroendocrine tumor therapy

Abstract Pituitary adenomas (PAs), or pituitary neuroendocrine tumors (PitNETs), are commonly found in the anterior pituitary gland. Although the majority of PitNETs are benign and stable, several tumors have malignant characteristics. The tumor microenvironment (TME) plays an important role in the process of tumorigenesis and is composed of several types of cells. Various cells in the TME are significantly affected by oxidative stress. It has been reported that immunotherapeutic strategies have good effects in several cancers. However, the clinical potential of immunotherapies in PitNETs has not yet been fully discussed. Oxidative stress can regulate PitNET cells and immune cells in the TME, thus affecting the immune status of the TME of PitNETs. Therefore, modulation of oxidative stress‐regulated immune cells using a combination of several agents and the immune system to suppress PitNETs is a promising therapeutic direction. In this review, we systematically analyzed the oxidative stress process within PitNET cells and various immune cells to elucidate the potential value of immunotherapy.


| INTRODUC TI ON
Pituitary adenomas (PAs), recently named pituitary neuroendocrine tumors (PitNETs) according to the new classification and nomenclature established by WHO 2022, are commonly found in the anterior pituitary gland and are harmful to the endocrine system and human health. 1 It has been suggested that the prevalence of PitNETs is approximately 20%; however, most tumors present no clinical manifestation. [2][3][4] Additionally, the incidence of clinically significant PitNETs is 80-100 cases per 100,000 people. 2,5 PitNETs are generally classified as benign (~65%), invasive (~35%) or malignant (only ~0.2%), or as macroadenomas (≥10 mm) and microadenomas (<10 mm) according to their size. 1,6,7 Although most PitNETs are stable benign tumors with a good clinical prognosis after surgical excision, a minority of PitNETs are still difficult to control, even with advanced drugs and radiotherapy, so-called aggressive tumors. 8 Oxidative stress usually occurs when intracellular reactive oxygen species (ROS) levels exceed the cellular antioxidant capacity, resulting in an imbalance in oxidative homeostasis. 16 Numerous studies have confirmed the relationship between oxidative stress and the formation and progression of various human pathologies. 17,18 Intracellular ROS are mainly generated during mitochondrial oxidative phosphorylation and play a crucial role in cellular signaling pathways, but excessive ROS damage genomic and mitochondrial DNA, leading to DNA damage, molecular mutations, and altered signaling pathways. 16,19 Tumor cells often have abnormal alterations in metabolic pathways and mitochondrial function, which lead to the production of ROS and oxidative stress within tumor cells and further affect tumorigenesis. 20 However, the process of oxidative stress and its role in PitNETs have not been fully discussed.
The importance of the TME as a therapeutic target and prognostic factor in tumor treatment has been widely recognized. 21,22 Multiple factors in the TME can have an impact on the progression and persistence of tumors; the effects of immunotherapy (IMT), a central part of current cancer treatment, are strongly associated with the TME. 23 Reactive oxygen species are important signaling mediators for multiple physiological processes in the TME and for the immune cells within it. 24,25 Therefore, the interaction between oxidative stress and the TME is considered of great importance for tumor treatment. This review focuses on the effects of oxidative stress and the TME in PitNETs and discusses novel therapeutic strategies based on the modulatory role of oxidative stress in the TME that can be used to treat PitNETs.

| OVERVIE W OF THE OXIDATIVE S TRE SS RE S P ON S E IN TUMOR S
Oxidative stress occurs when the balance between ROS and the antioxidant system is disrupted, and it is mainly generated by mitochondria. 20 In addition, inflammatory cells, such as neutrophils, macrophages, and eosinophils, generate exogenous ROS through NADPH oxidase reactions. Excessive ROS will destroy and/or change DNA, RNA, lipids, proteins, and other intracellular molecules. 26 Correspondingly, a cytokinesis-block micronucleus cytome (CBMN-cyt) assay showed that the frequency of biomarkers of chromosome breakage and/or loss, biomarkers of DNA misrepair and/or telomere end-fusions, and biomarkers of elimination of amplified DNA and/or DNA repair complexes were significantly increased in PitNET patients. [27][28][29] The researchers believe that these findings indicate that chromosome/oxidative DNA damage is related to the invasion of PitNETs. 29 Furthermore, ionizing radiation and some environmental factors participate in all stages of carcinogenesis induced by ROS. 20 PitNETs show increased ROS levels and signs of oxidative damage, and the secretion of a large amount of ROS/reactive nitrogen species (RNS) recruits more activated immune cells, magnifies the dysregulated processes and eventually leads to a preneoplastic condition. 14,20 When entering a state of oxidative stress, irreversible oxidative damage to DNA, RNA, lipids and proteins may lead to genetic changes, thus leading to the imbalance of oncogenes and tumor suppressor genes. 30 These physiological changes indicate that oxidative stress plays an important role in the development of PitNETs. Notably, although the ROS level has increased to a certain extent, tumor cells can induce a new redox balance so that they can adapt and proliferate, which is a significant feature of cancer cells that is different from normal cells. 30 In fact, pituitary cancer and PitNET are still difficult to distinguish in terms of pathology, immunohistochemistry, genetic analysis, and ultrastructural imaging findings. 31 Therefore, an understanding of oxidative stress-related pathways may aid in the identification of pituitary cancers and the treatment of PitNETs.
The oxidative stress pathway is very complex. In addition to the mainstream pathway, there are some special pathways associated with PitNETs. The Ras pathway is an important pathway related to oxidative stress and cancer. Approximately 30% of human tumors contain activated mutations of Ras family oncogenes, and these mutations cause the proteins to have constitutive activity. 32,33 Overexpression of the Ras family causes increased mitochondrial mass and ROS accumulation, leading to DNA damage and promoting transformation. 34 The Ras pathway relies on the NADPH oxidase Nox4. 34 Ras mutation has been proven to be related to human pituitary tumors, and H-ras point mutation has been found in distant metastatic pituitary cancer, which indicates that H-ras gene mutation may play an important role in the formation of pituitary cancers and the metastasis of PitNETs. 31,35,36 In addition, studies have shown that the epidermal growth factor receptor (EGFR) signaling pathway, including nuclear factor erythroid 2-related factor 2 (Nrf2), mitogen-activated protein kinase ERK1/2, MEK and protein kinase C, is related to cell proliferation and affected by oxidative stress. 37,38 Multiomics studies on PitNETs have revealed significant changes in the Nrf2-mediated oxidative stress response pathway and the corresponding regulatory factors. 39,40 Nrf2 regulates the expression of hundreds of genes, including those related to antioxidant enzymes, tissue remodeling and fibrosis, immune and inflammatory reactions, carcinogenesis and metastasis, which can regulate ROS levels together with Kelch-like ECH-associated protein 1 (Keap1). 41 Nrf2 can also regulate cell protective enzymes such as glutathione peroxidase (GPx) and oxidoreductase to combat oxidative stress. 42,43 In general, Nrf2 plays a central role in the oxidative stress-related pathways of tumors, including PitNETs.

| Nrf2-mediated oxidative stress response pathway
Nrf2 is a key regulator of oxidative stress, and it interacts with Keap1 to induce an antistress response. 44 Previous studies have confirmed that Nrf2 expression is increased in PitNETs, and the level of Nrf2 phosphorylation is also increased. 14 Increased ROS levels activate the Nrf2-Keap1 complex in the cytoplasm through multiple interacting signaling pathways. [45][46][47] In the presence of oxidants, the DGR structural domain of Keap1 releases the DLG motif of Nrf2, thereby preventing ubiquitination and degradation of Nrf2. 44,[48][49][50][51] Upon activation, isolated and phosphorylated Nrf2 rapidly enters the nucleus to interact with antioxidant response elements (AREs). [52][53][54] This reduces oxidative damage by inducing the synthesis of antioxidant proteins 54-56 but also promotes tumorigenesis by regulating cellular metabolism. 54,57 Antioxidant genes such as heme oxygenase 1 (HO-1) contain upstream ARE sequences, which is an important reason why Nrf2 is considered to have a central position in the oxidative stress pathway. 44,58,59 The role of the Nrf2 signaling pathway in PitNETs has been verified. Irradiation-treated C57/BL6 mice exhibited oxidative damage due to Nrf2 activation and HO-1 expression, which could be inhibited by the antioxidant agent pituitary adenylate cyclase-activating polypeptide 38 (PACAP38). 54,60 A study found that patients with PitNETs had significantly more chromosomal and oxidative DNA damage than normal subjects and that oxidative stress was an important cause of this phenomenon. 29 Similar to other tumors, the occurrence and development of PitNETs are closely related to oxidative stress. 44,54 Thus, Nrf2, which is at the core of the response, plays a broad and critical role in PitNET and may be a new target for PitNET treatment.

| Mitochondrial dysfunction pathways
Studies have shown that mitochondrial dysfunction is significantly associated with the pathogenesis of PitNETs. 39,61 Mitochondria, as a source of ROS, are the center of oxidative stress and the main location of energy metabolism. 62 Altered energy metabolism has been recognized as a marker of tumors, and such alterations may lead to increased ROS production. 63 Accordingly, enlargement of the mitochondrial area can be detected in the early stages of PitNET, and significantly increased expression of major factors that regulate mitochondrial gene expression, such as Nrf1, Nrf2, and mitochondrial transcription factor A (TFAM), was found and maintained. 14,64 Mitochondrial fusion and fission processes regulate mitochondrial morphology, and the expression of the fusion-related kinetic proteins mitofusin 2 (MFN2) and optic atrophy type 1 (OPA1) was found to be enhanced with the development of PitNET, which was associated with a decrease in mitochondrial elongation and a tendency toward roundness. 14,65 In contrast, increased levels of the fission-related protein dynamin-related protein 1 (DRP1) indicated that mitochondrial fission was inhibited, which explains the mitochondrial morphology in PitNETs. 14,65, 66 An elevated rate of glycolysis is a common feature of tumor cells, and it is logical that there is also a consistent increase in lactate in PitNETs. 14,67,68 All of the above findings imply the presence of significant mitochondrial dysfunction in PitNETs.
In addition, the mitochondrial OXPHOS system, which is the main generator of endogenous ROS, is usually defective in tumor cells, but OXPHOS activity is not disrupted in PitNETs. 14,69 In addition to ROS production, mitochondrial nitric oxide synthase (mtNOS) induces the production of much of nitric oxide (NO), which reacts with superoxide radicals to generate peroxynitrite anion (ONOO-) or hydroxyl radical ·OH. 62,70 The increased rate of glycolysis and the maintenance of OXPHOS system activity may provide PitNETs with adequate ATP and raw materials to help them have a higher metabolic rate. 71 In conclusion, mitochondrial dysfunction leads to an increase in ROS in tumor cells, which mediates relevant signaling pathways and activates procancer signaling pathways that regulate tumor progression, angiogenesis, metastasis, and survival. 62 However, this also activates the antioxidant system of tumor cells, allowing them to survive at higher ROS levels. 72 This endows PitNET cells with a redox system and associated signaling pathways that are different from those of normal cells.

| Overview of the TME
The TME is the environment in which tumors exist, and the significance of TME angiogenesis for tumors has been extensively studied; disorganized, leaky vessels cause an increase in interstitial pressure and a reduction in the efficiency of oxygen and nutrient delivery. 23,73 Vascular endothelial growth factor (VEGF) and its receptor (VEGFR) are central to tumor angiogenesis. [73][74][75] Researchers have found that PitNETs exhibit the senescence-associated secretory phenotype (SASP), which is regulated by NF-κB. VEGF and matrix metalloproteinase 9, as paracrine components of SASP, may contribute to the metastasis of PitNETs. [76][77][78][79][80] Upregulation of VEGF usually implies tumor development and malignant transformation, and this is indeed the case in PitNETs. 15,81 Compared with adjacent normal pituitary tissue, PitNET tissue shows decreased VEGF expression, while pituitary carcinoma tissues show increased expression. 82 Furthermore, angiogenesis directly affects the oxygen and nutrient supply to the tumor, and the effects on oxygen tension and immune infiltration are also important. 23,83 This has led to the use of antiangiogenic therapy and treatment modalities in combination with immunotherapy in clinical trials. 73,84 NF-κB-activated proinflammatory molecules are an important method through which NF-κB affects PitNETs. 76 In the development of PitNETs, TNFα and IL-1β are specific targets for NF-κB, and they increase with the level of p-p38 MAPK protein, which accompanies strong activation of NF-κB. 76,85 Studies conducted in invasive PitNETs have shown that both TNFα levels and IL-6 levels were upregulated in invasive PitNETs. 86 They are believed to enhance the invasiveness of tumor cells and protect them from immune cell attacks. 87,88 Interestingly, some researchers have found that IL-6 has a dual role, inhibiting PitNET growth and malignant transformation by triggering senescence. 89,90 The release of IL-6 and IL-8 was detected in cultured adenoma cells, which was influenced by IL-1β dose-dependent stimulation. Correspondingly, IL-8 also has a similar effect to IL-6. 91 Another downstream factor of NF-κB that plays an important role in the senescence of tumors is CXCR2. Knocking down CXCR2 alleviates both replicative and oncogene-induced senescence and reduces DNA damage responses. 92 In contrast, ectopic expression of CXCR2 leads to premature senescence via a p53-dependent mechanism. 92 CXCR2 is upregulated in preneoplastic lesions in vivo, and its role in promoting the invasiveness of PitNETs has also been confirmed. 92,93 In addition, TGFβ is also believed to have a regulatory effect on the development of PitNETs. 94 As the regulatory protein of TGFβ, P27 gradually decreases during the development of pituitary tissue tumorigenesis. 95 97 Studies have found that the K ATP channel is expressed in FS cells, and its activation inhibits VEGF secretion at the transcriptional level, which plays a regulatory role in the development of PitNETs. 98 Another study based on TtT/ GF cells showed that IL-6 promotes cell proliferation in a dose-and time-dependent manner under low-density seeding conditions. For high-density seeding, researchers believe that the high concentration of IL-6 secreted by cultured cells themselves playes a promoting role. 99 This indicates that FS cells also have a mechanism of autocrine growth stimulatory loop. 99 In addition, TNFα was also found to be involved in the diffusion mechanism of inflammatory signals related to FS cells. 96

| Modulation of various immune cells by oxidative stress
Oxygen has a dual role in the TME and is not only involved in hypoxia.
There is evidence that oxidative stress also has a profound effect on immune cells in the TME (Figure 1). 109 Appropriate levels of ROS are necessary for antigen-presenting cells to maintain normal function. 110 Macrophages and dendritic cells (DCs) have been found to regulate antigen crosspresentation by producing phagosomal ROS in a process mediated by NOX2. 111 The antigen presentation function of DCs is also believed to exist in PitNETs. 112 Researchers stained macrophages with CD68 to study PitNETs. CD68 + cells were detected in all cases, and their number was positively correlated with tumor size and Knosp grade. 15,113 In addition, endoplasmic reticulum (ER) stress has an immunosuppressive effect and can lead to an increase in DCs and immunosuppressive signaling molecules, including arginase, which is related to antigen presentation disorders. 23,114,115 In addition, ER oxidative stress induces immunogenic death of cancer cells through additional pathways. 110,[116][117][118] Immunogenic cell death (ICD) leads to the release of damage-associated molecular patterns (DAMPs) that bind to receptors on DCs and activate antigen crosspresentation by DCs, ultimately generating an antitumor CD8 + T-cell response. 110,119 Such effects provide a variety of viable options for regulating antigen presentation via ROS.
As one of the multiple immune cells in the TME, regulatory T cells (Tregs) suppress immune responses via oxidative stress. [120][121][122] Increased Treg numbers and function are characteristics of PitNET patients and can be reversed by surgical resection of the tumor. 123 On the one hand, Tregs are susceptible to oxidative stress-induced apoptosis due to their weak Nrf2-associated antioxidant system and susceptibility to free oxygen in the TME. 124 On the other hand, apoptotic Tregs can inhibit the expression of interferonγ (IFNγ) and tumor necrosis factor (TNF) in CD4 + and CD8 + T cells, and this inhibitory effect is even stronger than that of live Tregs. Related studies have confirmed that apoptotic Tregs promote tumor growth more than live Tregs and are even able to nullify the antitumor effect of PD-L1 blockade. The inhibitory effect of IFNγ on the development of PitNETs has been confirmed by single-cell sequencing of PIT1-positive PitNETs. 125 In addition, immunosuppression by Tregs is somewhat dependent on macrophage function. 124 Tumor Tregs were found to have higher mitochondrial activity and produce more endogenous ROS than conventional T cells, and these features make Tregs more sensitive to oxidative stress in the TME and consequently undergo apoptosis. 124 Activation of T cells and NK cells leads to an increase in ROS, and conversely, ROS are important for T-cell activation, expansion and effector functions. [126][127][128][129][130] In addition, ROS, while contributing to T-cell activation-induced cell death, are also secondary messengers for nuclear factor of activated T cells (NFAT) activation and inhibition of negative regulatory phosphatases. 110,127 Increased ROS levels also trigger anti-glutathione (GSH) responses that affect the metabolic integration and reprogramming of inflammatory T cells. 131 Thus, it appears that oxidative stress leads to impaired T-cell antitumor function, while ROS neutralization slows T-cell terminal differentiation. 132,133 A study of somatotroph PitNETs revealed that the number of CD8 + lymphocytes was decreased in the cavernous sinus invasion group, suggesting that this factor may be able to predict the F I G U R E 1 Effect of ROS on immune cells in tumor microenvironment. prognosis of somatotroph PitNETs. 134 However, while high levels of ROS damage T cells, ROS are also necessary for T cells. Reactive oxygen species were found to maintain the metabolic and functional status of CD8 + T cells through SEPN family members in some tumors, thus enhancing the antitumor function of CD8 + T cells. 129,[135][136][137][138] The ability of NK cells to distinguish between malignant and healthy cells and their antigen-independent tumor cell killing mechanism makes them an interesting tool for tumor therapy. 139,140 NK cells, when activated, tend to kill tumor cells and secrete a series of cytokines and chemokines that activate and recruit adaptive immune cells to the TME to induce antitumor effects. [141][142][143] In addition to their cytotoxic function, NK cells also have a dual regulatory effect on antigen-presenting cells (APCs). [144][145][146] Overall, the TME promotes tumor growth and inhibits NK cells. NK cells use glucose as an energy source to generate the antitumor response. 147,148 A study found that NK cells in the TME had significantly increased expression of lipid peroxidation-and oxidative damage-related proteins, which was related to NK cell dysfunction. 148 In contrast, expanded NK (exNK) cells with metabolic adaptations similar to those of tumor cells had a greater capacity for nutrient uptake and glutamine export and were also able to survive better in the TME. [148][149][150] In addition, excess ROS in NK cells blocks NF-kB activation, leading to insufficient production of TNFα, IFNγ, and IL-2. 151 In addition, the PD pathway also negatively affects DCs, TAMs, and NK cells, but the mechanism of PD-1-mediated inhibition is very complex, and there can be dual effects. [156][157][158][159][160] Many factors are involved in the regulation of ROS and PD-L1, among which NF-kB, HIF-1α and Yes-associated protein 1 (YAP1) are some of the more critical. 161,162 HIF-1α is a cellular oxygen sensor that is upregulated together with ROS under hypoxic conditions. Typically, activation of HIFα leads to upregulation of PD-L1, which in turn leads to elevation of the expression of PD-L1 on the cancer cell surface. 154 The regulation of PD-L1 and HIF-1α is bidirectional. PD-L1 induces upregulation of HIF-1α through ROS production, thereby upregulating YAP1 expression in cancer cells. 161 In addition, NF-kB also contributes to the regulatory relationship between ROS and PD-L1. Subsequently, MDSCs, Tregs, and TAMs infiltrate tumors, which in turn stimulates PD-L1 expression in tumor cells. 154 The increase in ROS is often due to mitochondrial dysfunction, which has been demonstrated in PitNET cells. 66 This implies that the regulatory relationship between ROS and PD-L1 may be present in metastatic PitNETs, representing a new therapeutic target.

| Induction of oxidative stress in PitNETs
Mitochondria have been mentioned previously as the source of endogenous ROS production. Therefore, there have been several studies targeting mitochondrial oxidative stress for the treatment of PitNETs ( Table 1), some of which have identified feasible strategies. 163,164 18β-Glycyrrhetinic acid is a natural compound extracted from liquorice that has been shown to have anticancer and antiinflammatory effects. 165

Inhibited proliferation and activated the apoptosis of MMQ cells
Mice after high-dose CsA treatment. Similarly, the Mn-SOD level also decreased in a dose-dependent manner, potentially revealing one of the pathways by which CsA leads to GH3 cell apoptosis. 182 In addition, similar to 18β-glycyrrhetinic acid, CsA was able to decrease mitochondrial membrane potential and alter mitochondrial membrane permeability, which in turn caused the release of cytochrome C and ultimately inhibited Bcl2 expression and induced apoptosis. 183 Interestingly, in studies on CsA nephrotoxicity, ERK, JNK, p38 and NF-kB were found to be targets of CsA, and CsA was also found to strongly activate Nrf2 in proximal tubular epithelial cells, which, as previously mentioned, are key factors in the induction of oxidative stress. 181,[184][185][186] In addition, CsA has been found to directly trigger a decrease in antioxidant capacity in a variety of tissue cells. 187,188 This mechanism renders cellular redox homeostasis unstable and makes cells more susceptible to oxidative stress. These findings suggest that CsA may have unprecedented potential in inducing oxidative stress in PitNET cells. 182 Numerous other drugs have also been studied. Dopamine agonists, mainly bromocriptine (BRC) and cabergoline (CAB), are effective in reducing the size of prolactinomas. BRC induces apoptosis of prolactinoma cells primarily through the ERK/EGR1 signaling pathway, while CAB induces autophagy by inhibiting the AKT/mTOR signaling pathway. [189][190][191] Gossypol acetate (GAA) has the ability to inhibit enzymatic activity, causing changes in mitochondrial morphology and inhibiting Bcl2 expression, thereby inducing apoptosis. 192 Melatonin is able to selectively block the production of VEGF and ROS and even reverse angiogenesis, which can result in reduced ROS and a lack of tumor nutrition in the TME. 193 Paeoniflorin increased the expression of cleaved caspase-9, caspase-3, and Bax and inhibited the expression of Bcl-2 in MMQ cells and GH3 cells, thereby inducing mitochondria-mediated apoptosis, which ultimately manifested as a decrease in cell viability and inhibited proliferation. 194,195

| Immunotherapies for PitNET targeting tumor redox
The human immune system achieves tumor surveillance under normal conditions through innate and adaptive immune responses against tumor antigens. However, tumor cells can evade monitoring by the immune system and further proliferate, infiltrate, and metastasize. 155 Therefore, immunotherapies have been developed to activate immune cells in the TME to enable them to generate an immune response and ultimately inhibit tumor growth. TNF and IL-2 were explored in early studies, but they are no longer used for routine treatment due to unacceptable systemic side effects. 196 In the quest for additional immunotherapies, strategies targeting immune cells in the TME and modulating oxidative stress pathways are emerging due to the abnormal ROS levels in tumor cells.

| Melatonin
Melatonin is a natural hormone secreted mainly by the pineal gland. Melatonin has antioxidant effects because it reacts directly with free radicals and induces the production of antioxidant enzymes. 197,198 Correspondingly, increased melatonin release at night reduces oxidative DNA damage, and conversely, oxidative DNA damage is increased during the day. [199][200][201] The inhibition of indoleamine 2,3-dioxygenase 1 (IDO1) induced by melatonin is associated with the activation and maturation of DCs and activates cytotoxic CD8 + T lymphocytes. [201][202][203][204] In addition, investigators found a dose-dependent decrease in ROS levels in melatonin-treated CD4 + T cells. 205 This suggests that melatonin has the potential to rescue CD4 + T cells that die in the TME due to excessive ROS levels. Studies on prolactinoma showed that melatonin inhibited the activity of the mitochondrial respiratory complex and ATP production in prolactinoma cells. 206 Another study found that melatonin can decrease Bcl-2 mRNA expression and mitochondrial membrane potential. 207 These effects enable melatonin to regulate the mitochondrial function and energy metabolism of prolactinoma cells. 206,207 In the previous section, we mentioned the ability of melatonin to inhibit angiogenesis, and here, we have reported the additional ability of melatonin to activate immune cells in the TME. Such dual effects suggest that melatonin has potential in the treatment of PitNETs, and in vitro tests have demonstrated that melatonin inhibits the growth of prolactin tumor cells in rats. 197
Although these compounds have the potential to be used in the treatment of PitNETs, few compounds have been investigated in PitNETs. [208][209][210][211][212][213][214] Phenformin is a biguanide that is used as an antidiabetic agent. In addition, phenformin has been found to have antitumor effects on several tumors. 215 Phenformin is thought to exert antitumor effects through oxidative stress pathways, as evidenced by decreased GSH expression and a reduction in the GSH reduction ratio. 216,217 Furthermore, phenformin induces ROS production in MDSCs. 215 In addition, phenformin also inhibits mTOR phosphorylation and HIF-1α expression, which attenuates PD-L1 expres- Toll-like receptor 4 (TLR4) is thought to mediate the proliferation of primary PitNET cells. 226 Coincidentally, TLR4 is inhibited by baicalein, and the expression of its downstream signaling factor HIF-1α is also reduced. 227 As we mentioned, the reduction in HIF-1α expression causes PD-L1 expression to be reduced. 154 In addition, baicalein significantly inhibited IFNγ-induced PD-L1 upregulation, 228 and baicalein inhibited the expression of VEGF. 227 These results suggest that baicalein may be a target for PD-L1-related PitNET immunotherapy. treatment of many tumors. Therefore, immunotherapy has likewise been attempted for the treatment of PitNETs. To identify additional therapy targets and achieve better results, oxidative stress has been considered. Oxidative stress is of great importance in both tumors and the TME, mediating numerous biochemical reactions. The potential of strategies related to oxidative stress in the treatment of PitNETs has revealed many new research directions.

| CON CLUS I ON S AND PER S PEC TIVE S
On one hand, oxidative stress can act directly on tumor cells to inhibit tumor cell proliferation and metastasis. However, oxidative stress can act on immune cells in the TME and mediate the immunotherapy response.
Popular targets of immunotherapy include PD-L1 and cytotoxic T lymphocyte-associated antigen 4 (CTLA4), but CTLA4 has been little studied in PitNETs, so we focused on PD-L1. 229 Tumor- In addition to the compounds mentioned in this review, some emerging ROS-modulating agents have been suggested to modulate the response of tumors to immunotherapy. 110 High-dose ascorbate showed a synergistic effect with anti-PD-1 treatment in mouse tumor models and was able to enhance CD8 + T-cell infiltration, and this effect seemed to be independent of its antioxidant capacity. 230,231 Nonsteroidal anti-inflammatory drugs (NSAIDs), including celecoxib and aspirin, have also been reported to stimulate antitumor immune responses, and enhance the efficacy of anti-PD-1 treatment in mouse tumor models. [232][233][234] In addition, xCT inhibitors and ROS-response prodrugs have been suggested to affect the immunotherapy response of tumors. 110 In summary, oxidative stress mediates antitumor therapy by acting on PitNETs themselves and immune cells in the TME. This emerging therapeutic pathway has surprising potential and remains to be investigated further.

CO N FLI C T O F I NTE R E S T S TATE M E NT
The authors declare no conflict of interest.

DATA AVA I L A B I L I T Y S TAT E M E N T
The data that support the findings of this study are available from the corresponding author upon reasonable request.