1 Introduction

The intricate relationship between altered metabolism and disease has become central to biomedical research, particularly cancer biology. Among the hallmark traits of cancer is metabolic reprogramming, which allows tumor cells to thrive in nutrient-deprived environments. One of the pivotal elements of this reprogramming is the dependency on external resources provided by the tumor microenvironment (TME) [1]. Cancer cells adapt to the harsh conditions within the TME by modifying their metabolic pathways to support rapid proliferation, evade immune responses, and resist treatment. These adaptations primarily involve changes in the metabolism of carbohydrates, lipids, and amino acids-key drivers of cellular growth and survival [2]. Glucose, glutamine, and leucine are vital nutrients in nutrient-poor microenvironments that provide energy and generate building blocks such as nucleotides, proteins, and lipids. Glucose and glutamine, two critical nutrients, are indispensable substrates in tumor cell metabolism, supporting bioenergetics and biosynthesis [3-5]. Glutamine, the most abundant amino acid in the body, plays a crucial role in these processes. Cancer cells exhibit a marked dependence on glutamine to fuel anaplerosis, generate metabolic intermediates, and maintain redox balance. This “glutamine addiction” is not limited to providing energy but extends to supporting the biosynthesis of nucleotides, proteins, and lipids, all essential for the uncontrolled proliferation of cancer cells [6, 7]. Besides serving as a fuel source, glutamine is involved in cytoprotective programs that protect cancer cells against harmful agents in the TME [8, 9]. Cancer cells metabolize glutamine through glutaminolysis within the mitochondria, converting it to glutamate and tricarboxylic acid cycle (TCA) intermediary α-ketoglutarate (αKG) [10].

The diversion of pyruvate from the TCA cycle leads to increased dependence on glutamine as a carbon source for anaplerosis [11, 12]. Glutamine is transported into the cell by solute carrier (SLC) type transporters and is then catabolized by the enzyme glutaminase (GLS1), which converts glutamine into glutamate and ammonia. αKG is then produced by the metabolism of glutamate by transaminases or glutamate dehydrogenase (GLUD) [13]. αKG then undergoes carboxylation to produce isocitrate catalyzed by aconitase to produce citrate. Isocitrate is converted to citrate through aconitase reverse reaction, allowing carbon for ATP citrate lyase to produce acetyl CoA. This process enables ATP production and provides necessary biosynthetic precursors for cancer cell growth and proliferation [14-16]. This heightened demand for glutamine underscores its significance in cancer and other diseases characterized by metabolic dysregulation. Over the years, research into glutamine metabolism has uncovered its broad influence on various biological processes, including cell signaling, autophagy, and apoptosis [17-20]. These findings have positioned glutamine metabolism as a potential therapeutic target across multiple diseases.

The central role of glutamine metabolism in diseases has prompted research exploring its molecular regulation and biological functions. Recent studies have shed light on how glutamine metabolism is controlled at the molecular level, revealing key regulators such as oncogenes and tumor suppressors that orchestrate its uptake and utilization in cancer cells. Additionally, the discovery of glutaminase inhibitors and other metabolic modulators has opened new avenues for therapeutic intervention, particularly in targeting the vulnerabilities of cancer cells’ reliance on glutamine. However, despite these advances, significant gaps remain in our understanding of how glutamine metabolism interacts with other metabolic pathways and contributes to disease progression.

This review aims to provide a comprehensive overview of glutamine metabolism, focusing on its regulation, functions, and implications in various diseases. We first explored the historical background and current state of research on glutamine metabolism. Then, we searched the molecular mechanisms governing glutamine uptake, transport, and utilization, particularly in cancer. Finally, we highlighted emerging therapeutic strategies targeting glutamine metabolism, emphasizing the potential for future clinical applications. By synthesizing these findings, this review provides insights into the critical role of glutamine metabolism and its therapeutic potential in disease management.

2 Role of Glutaminolysis in Cancer

Cancer metabolism has gained interest for nearly a century due to its ability to uncover fundamental aspects of malignancy and its potential to improve cancer diagnosis, monitoring, and treatment. Glutamine metabolism, the most abundant amino acid in plasma, is crucial in cancer due to its ability to donate nitrogen and carbon into growth-promoting pathways. During periods of rapid growth or stress, glutamine becomes conditionally essential, especially in cancer cells that display oncogene-dependent addictions. Many cancer cells rely heavily on glutamine for survival, a phenomenon known as glutamine dependence. This underscores glutamine's essential role in their ability to thrive. Cancer cells adapt to support the citric acid cycle by increasing glutamine metabolism, which is crucial for their growth. Glutamine provides carbon for the cycle and supplies nitrogen needed to synthesize hexosamines, nucleotides, and various nonessential amino acids [21, 22].

In cancers like non-small cell lung cancer (NSCLC), brain tumors, and breast cancer (BC), glutamine metabolism is a vital process. Additionally, it influences the TME by regulating oxidative stress through glutathione (GSH) production [23]. Glutamine metabolism regulators, including amino acid transporters SLC1A5, SLC7A5, SLC7A11, SLC3A2, and Myc, are critical for maintaining the balance between glutamine metabolism and cell viability [24, 25]. Increased expression of glutamine metabolism regulators has been linked to a high survival rate in multiple cancers. Tumor cells upregulate most glycolytic enzymes due to increased c-Myc and hypoxia-inducible factor 1 alpha (HIF-1α) transcriptional activity, insufficient control by p53 and other tumor suppressors, and oncogenes such as mutant Kirsten rat sarcoma virus (KRAS). These factors induce glutaminolysis by directly or indirectly activating glutamine transporters and glycolytic genes in cancer cells [3]. Glutamine and its metabolites play crucial roles in various cellular mechanisms, including mTOR activation and the biosynthesis of sugars, nucleic acids, amino acids, and fatty acids [9]. Dysregulation of the mTORC1 signaling pathway is associated with pathological conditions, including cancer, obesity, diabetes, and neurodegeneration [26]. In certain types of cancer, overexpression of mTORC1 occurs due to Tuberous sclerosis protein 1/2 (TSC) phosphorylation and inactivation by Cyclin D1–CDK4/6. Glutamine flux has been reported to modulate mTOR to coordinate cell proliferation and growth [27]. Recent studies have indicated that mTORC1, a signaling pathway involved in cellular metabolism, regulates aerobic glycolysis through HIF-1α. Recent studies have found new functions for glutamine in regulating cell proliferative events. For example, cancer cells under hypoxia or with defective mitochondria can use glutamine-derived α-KG to produce citrate, which is crucial for lipid synthesis, highlighting the importance of glutamine in cell proliferation. Because of the energy requirements of rapidly proliferating cells, tumors produce a hypoxic environment as they develop. According to current research, mTORC1 regulates aerobic glycolysis using HIF-1α  lowers hypoxia by inducing angiogenesis and regulates the metabolism of cells [28]. Furthermore, dysregulation of oxidative phosphorylation (OXPHOS) and autophagy are key metabolic characteristics of metastatic tumor cells, which frequently experience metabolic stress [29]. NcRNAs, including miRNAs [30], long noncoding RNAs (lncRNAs) [31], and circular noncoding RNAs (cncRNAs) [32], have been discovered as glutaminolysis regulators, and they may interact with oncogenes and tumor suppressors genes and influence the metabolism of cancer. Research has confirmed the contribution of ncRNAs to cancer progression, impacting crucial cancer signatures such as glutaminolysis. LncRNAs, newly discovered functional noncoding RNAs, exert significant regulatory effects through various mechanisms. Recent studies have demonstrated the extensive function of lncRNAs in controlling many biological functions, such as metabolic processes. Gaining insight into the intricate nature of lncRNAs will help us comprehend tumor metabolic machinery and make it easier to build lncRNA-based cancer metabolism-targeting therapeutics. Tumor suppressor genes like p53 or oncogenes like c-Myc can control lncRNA functions. In contrast, lncRNAs and circular RNAs (circRNAs) can impact the production of HIF-1α [33] and c-Myc [34]. The atypical metabolic rewiring observed in tumors is an important cancer characteristic. It has been explored for diagnostic, monitoring, and targeted therapeutic techniques, making it a promising target for cutting-edge therapies. In addition to cancer, glutaminolysis is an important player in the pathophysiology of several other diseases, including neurological, kidney, autoimmune, and cardiovascular diseases [35-37]. Recent research on immune metabolism has highlighted the role of glutamine in immune system regulation [38]. It has been observed that glutamine is necessary for immune cells to survive, proliferate, differentiate biologically, and defend against various diseases [39, 40]. Recent research suggests that glutamine is also crucial in cardiovascular physiology and pathology, as it aids in synthesizing DNA, ATP, proteins, and lipids, driving vital vascular cell processes [41]. The dysregulation of glutamate metabolism is responsible for glutamate excitotoxicity [42, 43]. The glutamate excitotoxicity idea holds that excessive glutamate promotes neuronal dysfunction and degeneration [44]. It has important implications for both acute CNS insults, such as ischemia and traumatic brain damage, as well as chronic neurodegenerative illnesses, including amyotrophic lateral sclerosis (ALS), multiple sclerosis, and Parkinson's disease (PD) [45]. Despite continuous research, no pharmaceutical therapies are available to provide considerable neuroprotection in brain ischemia or damage cases. Also, metabolic reprogramming influences the progression and prognosis of kidney diseases. At the same time, glutaminolysis generates ammonia, essential for maintaining renal pH and cellular and systemic homeostasis, and is released from glutamine breakdown [46]. Several different regulators have a distinct role in how this process functions, as seen by the metabolic pathway that breaks down glutamine. Different strategies employed by these regulators can impact the efficiency and results of glutaminolysis. The significance of these activities is especially evident in different diseases, as the disruption of glutamine metabolism can potentially exacerbate the disease's genesis or progression. For example, certain regulators may block the route, which might have therapeutic effects, while others may strengthen it, promoting the fast proliferation of cancer cells. As a result, identifying the major regulators of glutaminolysis is critical for successful targeting. This could be useful in developing strategies to prevent or mitigate the effects of these diseases.

3 Altered Metabolic Genes in Glutaminolysis

Mutations in metabolic genes related to glutaminolysis can significantly influence cancer metabolism, affecting both cell growth and survival. These genetic alterations often enhance the conversion of glutamine to αKG, which fuels the TCA cycle, promoting energy production and biosynthesis. Tumor suppressors and oncogenes play essential roles in regulating glutamine metabolism, and their activity can profoundly impact the function of glutamine and its metabolites in cancer cells [47] (Figure 1). It has been determined that certain cancer forms contain mutations in metabolic enzymes, such as TP53, retinoblastoma protein (Rb), and HIF-1α, as well as enzymes like hexokinase (HK), pyruvate kinase isozymes M2 (PKM2), isocitrate dehydrogenase (IDH), succinate dehydrogenase (SDH), and nicotinamide phosphoribosyl transferase (Nampt), CDKN2A, or activating mutations of NFE2L2, NOTCH1/2, MLL2, and EP300 [48] driving tumor progression and metastasis [49, 50]. Also, many oncogenic agents and tumor suppressors directly control the metabolic reprogramming of cancer cells. Myc, MycL, and MycN are among the Myc family members of oncoproteins, the master regulators of metabolic reprogramming in a wide range of human malignancies. In this context, the c-Myc oncogene plays a pivotal role, particularly in glutamine metabolism. The c-Myc oncogene acts as a master regulator of cellular growth and metabolism, orchestrating a wide range of metabolic changes in transformed cells to facilitate rapid proliferation. When c-Myc is overexpressed, it leads to coordinated changes in the expression of various gene families, which collectively enhance cellular growth and division. One significant effect of c-Myc overexpression is the upregulation of GLS1, an enzyme responsible for converting glutamine into glutamate. In various types of cancers, it enhances the glutamate-ammonia ligase (GLUL) expression level, which is involved in the fresh synthesis of glutamine. Hyperactivation of c-Myc has been associated with glutamine addiction [51, 52]. mTORC1 modulates GLS levels via S6K1-dependent c-Myc regulation, improving translation efficiency by modifying eIF4B phosphorylation, which is required to unravel the 5′UTR structure [53]. Dysregulation of the mTORC1 signaling pathway is linked to several pathological conditions, including cancer, obesity, diabetes, and neurodegeneration [26]. Phosphorylation and inactivation of TSC1/2 by Cyclin D1–CDK4/6 lead to overexpression of mTORC1 in certain types of cancer [54, 55]. The study of p53's role in regulating the mTOR pathway has gained interest due to its crucial role in tumorigenesis. The coordinated regulation of p53 and the mTOR pathway is essential for maintaining homeostasis in response to stimuli. p53 controls the mTOR pathway at multiple levels, including direct regulating signaling mechanisms, posttranscriptional regulation by miRNAs, and inhibiting autophagy through protein-protein interaction [56]. p53 plays an essential role in glutamine metabolism by regulating the gene expression of glucose transporters GLUT1 and GLUT4. Elevations in glycolysis and energy availability are caused by polymorphisms in the DNA-binding domain of p53, which eliminates its suppression of GLUT1 and GLUT4 genes being expressed. p53 upregulated the glutaminase 2 (GLS2) enzyme, leading to increased GSH levels and decreased reactive oxygen species (ROS) levels, safeguarding cells against damage to DNA [57]. In response to oxidative stress or DNA damage, p53 promotes GLS2 synthesis in a p53-dependent manner, and p53 interacts with the GLS2 promoter. The tumor suppressor p53 regulates the expression of the genes that control mitochondrial oxidative respiration, namely GLS2 and cytochrome c oxidase deficient homolog 2 (SCO2). The balance of glucose consumption is shifted from mitochondrial respiration to the anaerobic glycolytic route by altered SCO2. At the same time, overexpression of GLS2 by p53 increases the level of GSH and decreases ROS, finally defending cells against DNA damage [57, 58]. Aberrant expression of GLS1 has been found in hepatocellular carcinoma (HCC), contributing to malignancy and poor prognosis. GLS1 knockdown inhibits the proliferation of cancerous cells in the liver and prevents tumor formation [59-61]. Overexpression of GLS1 has also been observed in human colorectal cancer (CRC) tissues and NSCLC. Data from the TCGA database reveal overexpression of GLS1 in various solid tumors, including stomach adenocarcinoma, head and neck squamous cell carcinoma, thymoma, testicular germ cell tumors, HCC, colon adenocarcinoma, and others. Overexpression of glutamine transporters (ASCT2) has been observed in various cancers, contributing to increased glutamine uptake and offering it a possible therapeutic target for the control of cancer. c-Myc and HIF-1α regulate multiple glycolytic enzymes and proteins involved in glutaminolysis and fatty acid synthesis [62]. HIF-1α is involved in regulating glutamate transporters and glutamate receptors [63]. A study showed that HIF-1α stimulates glutamine metabolism in CRC by increasing GLS1 expression and activity [64]. The activation and stabilization of HIF-1α play a crucial role in cellular metabolic adaptations to hypoxia. Prolyl hydroxylase domain (PHD) proteins are oxygen-sensing enzymes that hydroxylate HIF-1α at a proline residue at normal oxygen levels. The ubiquitin ligase von Hippel Lindau (VHL) then degrades the protein. Then, ubiquitin ligase VHL degrades this hydroxylated enzyme [65]. Cancer cells with elevated HIF-1α levels tend to exhibit higher malignancy and poorer response to radiotherapy, leading to a worse prognosis.

IDH1 and IDH2 isoforms are frequently mutated in various cancers, including glioma, acute myeloid leukemia, thyroid carcinomas, cartilaginous tumors, and intrahepatic cholangiocarcinoma (ICC) [66-69]. Mutations in IDH result in the accumulation of R-2-hydroxyglutarate (R-2-HG) [70-72], which activates PHD, leading to increased prolyl hydroxylation of HIF-1α and subsequent degradation. Inactivation of SDH, also known as succinate-coenzyme Q reductase or Complex II, leads to the accumulation of succinate, which inhibits PHDs and causes an increase in HIF-1α protein levels [73]. Accordingly, the build-up of R-2-HG caused by IDH1/2 mutations decreases HIF-1α levels and encourages tumor development, which includes astrocyte cancer [74, 75]. Cancer cell metabolic plasticity relies on activating and inhibiting various genes, oncogenes, growth factors, and tumor suppressors. A critical factor in this process is the interaction between HIF-1α and AMP-activated protein kinase (AMPK), which serves as an energy sensor and essential regulator of cellular metabolism. Glutaminolysis is critical in ATP production to turn off AMPK and mTORC1 [76]. AMPK activation, triggered by the binding of AMP or ADP, redirects metabolism toward increased catabolism and decreased anabolism by phosphorylating downstream critical protein networks that, in the end, cause mTORC1 suppression [77]. When there are dietary imbalances, glutaminolysis drives mTORC1 activation, which results in abnormal suppression of autophagy and glutaminolysis [78, 79]. Two parallel pathways of glutamine metabolism promote mTORC1 activation: one that relies on glutaminolysis and is mediated by RagB, and the other that is dependent on ATP and not on GLS or glutamine dehydrogenase (GDH) [80].

Metabolic reprogramming in idiopathic pulmonary fibrosis (IPF) lung fibroblasts involves glutaminolysis, contributing to their resistance to apoptosis. Reducing glutamine metabolism induces apoptosis in IPF fibroblasts, leading to changes in antiapoptotic gene expression and various epigenetic processes [81, 82]. Survivin and X-linked inhibitors of apoptosis protein (XIAP), which belong to the regulator of apoptosis protein (IAP) family, have less expression when glutaminolysis is inhibited [83]. Glutamate plays a role in invasion through the involvement of GLS1 and the metabotropic glutamate receptor GRM3. GRM3 is associated with the growth of malignant brain tumors, including glioma and breast and melanoma cancers [84-89]. Mutations in GRM3 result in constitutive receptor activation, provide signals for cell proliferation and survival in melanoma [89], and are closely associated with invasive behavior [90]. Mutations in PIK3CA are also related to the glutamine metabolism imbalance, as seen in intestinal cancer, which makes cancer cells rely on glutamine through overactivation of glutamate-pyruvate transaminase 2 (GPT2) [91]. Dysregulated RAS signaling has been shown to promote the rewiring of glutamine in pancreatic ductal adenocarcinoma (PDAC) and CRC [49, 92, 93]. Recent evidence indicates that metabolic enzymes can be modulated using posttranslational changes, such as butyrylation, crotonylation, propionylation, glutarylation, methylation, acetylation, succinylation, and malonylation [94, 95]. Sirtuin5 (SIRT5), a member of the sirtuin family and a regulator of posttranslational modifications (PTMs), globally regulates lysine succinylation [96-98], malonylation [96, 99], and glutarylation [100]. However, the reduction of biological ROS is brought about by the desuccinylation of IDH2 and the deglutarylation of glucose-6-phosphate dehydrogenase by SIRT5, which shields cells from oxidative damage [101]. In CRC tissues and cells, SIRT5 is highly elevated [102]. It sustains the TCA cycle and enhances glutaminolysis by activating GLUD1 through deglutarylation, making SIRT5 a potential target for anti-CRC therapy. To assist cancer cells overcome oxidative stress barriers during carcinogenesis, mutations in KEAP1 trigger the nuclear factor erythroid-related factor 2 (NRF2) antioxidant program and work in tandem with mutant KRAS to drive the development of lung adenocarcinoma (LUAD) [103-108]. Other malignancies with genetic mutations may be open to therapy due to the metabolic need for glutaminolysis in KEAP1–NRF2-mutant LUAD tumors [109-112] like epigenetic [113, 114], or posttranscriptional changes [115] in the KEAP1–NRF2 axis.

Furthermore, ammonia release from tumor cells involved in glutaminolysis acts as an autocrine and paracrine signal, promoting autophagy and protecting cells in different tumor regions from internal or environmental stress [20]. Recent findings in BC patients indicate an inverse correlation between GLS2 levels and epithelial–mesenchymal transition (EMT). Decreased GLS2 expression is associated with reduced mitochondrial activity. FOXC2, a critical regulator of EMT, plays a role in this process. Inhibiting FOXC2 expression restores GLS2 expression and enables glutamine utilization. These findings suggest that epithelial cancer cells undergoing EMT become independent of glutamine. Inhibiting EMT induces a metabolic shift directed by GLS2 in mesenchymal cancer cells, making them more susceptible to chemotherapy. Further validation is required to explore the predictive value of the inverse relationship between GLS2 and FOXC2 in BC outcomes [116].

4 Signaling Pathways Regulating Glutamine Metabolism

Glutamine metabolism is regulated by various signaling pathways crucial for disease and cellular homeostasis. Several critical signaling pathways, including c-Myc, mTOR, and KRAS, are tightly linked to the regulation of glutamine metabolism. The mTOR pathway, a key regulator of cell growth and metabolism, enhances glutamate influx and metabolism in cancer cells, making it a potential therapeutic target. Other signaling pathways, such as AMPK, AKT, and Wnt/β-catenin, have been implicated in regulating glutamine metabolism and offer potential avenues for therapeutic intervention. Understanding the intricate signaling networks that control glutamine metabolism opens up opportunities for developing novel therapies for diseases associated with dysregulated glutamine metabolism, including cancer, neurodegenerative disorders, and metabolic syndromes (MetS). Further research into these signaling pathways and their specific roles in glutamine metabolism will be crucial for developing effective targeted therapies and identifying diagnostic biomarkers for various diseases.

5 Role of MicroRNAs in the Regulation of Glutaminolysis

MicroRNAs, small noncoding RNAs, regulate biological processes like gene expression and RNA silencing. Around 2200 conserved miRNAs have been identified, affecting cell growth, differentiation, metabolism, viral infection, and tumorigenesis [159]. MiRNAs are also involved in pathological settings, with cancer being a leading area of investigation. MiRNAs regulate energy metabolism in tumors, affecting genes and enzymes involved in metabolic pathways [160] (Table 1). Understanding miRNAs' role in these pathways is crucial for developing new therapeutics and identifying biomarkers for cancer diagnosis.

6 Role of lncRNAs in the Regulation of Glutaminolysis

Eukaryotes possess a diverse array of RNA molecules that are crucial for transmitting genetic information and often exhibit specific subcellular localization. RNA synthesis, processing, and transport play crucial and interconnected roles in controlling various cellular activities and functions. Depending on their size, ncRNAs can be categorized into many categories, such as transfer RNA-derived short RNAs, PIWI-interacting RNAs, miRNAs, small nucleolar RNAs, and newly identified category of lncRNAs. Over 68% of the genes expressed in the human transcriptome are transcribed into lncRNAs. LncRNAs are comparatively large RNA transcripts that span more than 200 nucleotides, possess minimal or no capacity to code for proteins and exhibit restricted conservation among different species [179]. Various forms of cancer frequently exhibit changes in lncRNAs, which impact metabolic reprogramming, a characteristic of cancer (Table 2). It is unclear exactly how lncRNAs control these biological processes. Still, they control glutamine uptake-a vital fuel source for cancer cells, supporting their survival and growth [180].

Interestingly, lncRNAs and other factors can regulate and activate different isoforms of GLS in distinct ways, and their upregulation is linked to a higher chance of developing certain malignancies [181, 182]. Therefore, the internalization and metabolism of glutamine are critical for multiple biological processes in cancer cells, including energy production, synthesis of macromolecules, and the maintenance of redox equilibrium and levels of ROS [11]. Consequently, glutamine is involved in many biological functions, and lncRNAs target various enzymes involved in glutamine metabolism (Figure 2).

7 Circular RNAs-Mediated Glutaminolysis in Cancers

CircRNAs are endogenous biomolecules with closed-loop structures specific to cells and tissue and are resistant to exonuclease digestion [202, 203]. They can function as transcription regulators and microRNA sponges and have been linked to the progression of many human diseases, including cancer [204]. circRNAs have emerged as novel noncoding RNAs that play essential roles in various tumors [205], particularly in metabolic reprogramming [206, 207]. Several functional circRNAs associated with cancer have been identified [206, 207], including circ-002013 [208], circ-ABCB10 [209], circ-0032821 [210], and has-circ-0006168, which are implicated in LC, BC, Gastric cancer (GC), and esophageal cancer, respectively. The circRNA circ-HECTD1 is highly expressed in GC and promotes glutaminolysis by modulating the miR-1256/USP5 axis. Increased expression of circ-HECTD1 in GC is associated with overall survival. miR-137, a tumor suppressor in several malignancies, including stomach cancer, is another possible target of circ-HECTD1 [211, 212]. Depletion of circHECTD1 enhances sensitivity to drug treatment via the miR-137/PBX3 axis [213]. CircRNA circ-0000517 interacts with miR-330-5p in NSCLC to enhance Yin yang-1 (YY1) expression and boost cell proliferation and glutamine catabolism [214]. Previous research demonstrated that circRNA circ_0000003 facilitated GLS expression in tongue squamous cell carcinoma cells, indicating that circRNAs can regulate GLS expression and affect glutamine metabolism and cancer progression. According to several studies, miR-330-3p suppresses tumor growth in various tumor types and is inhibited by circ-0016068, which causes PCa cells to proliferate, invade, and migrate more freely [215]. GLS was shown to be a putative target gene of miR-579-3p by target prediction and screening; however, more research is required to determine its precise function and mode of action in ESCC [216]. Furthermore, it was shown that circ-0001093 was upregulated in ESCC tissues and cell lines and that circ_0001093 expression was positively connected with ESCC malignant phenotype and poor survival. It was discovered that circ-0001093 may bind to the tumor repressor miR, miR-579-3p, and negatively regulate it [217]. According to a study, circ-0001093 functions as a molecular sponge for miR-579-3p to promote GLS expression, glutamine metabolism, and the malignant phenotype of ESCC [218]. Similarly, the host gene of circ-OGDH regulates the interaction between glutamine metabolism and the TCA cycle. Research findings showed that circ-OGDH silencing reduced ATP content, α-KG synthesis, glutamine consumption, and GLS1 protein level in ESCC cells, suggesting that circ-OGDH promoted glutamine metabolism in these cells. Mechanical sponging inhibited miR-615-5p, allowing circ-OGDH to release PDX1, enhancing glutamine metabolism, and supporting tumor growth in ESCC, suggesting circ-OGDH as a promising therapeutic target [219].

Moreover, the control of autophagy has been linked to cirRNAs. Deletion of circRNAs enhances autophagy, leading to apoptosis and reduced proliferation in cervical cancer cells. For instance, circ-cTICRR binds with HuR protein, which stabilizes GLUD1 mRNA and increases the level of GLUD1 protein, indicating its oncogenic role in cervical cancer. There may be hope for cervical cancer treatments if the circ-TICRR relationship with the HuR protein is addressed [220]. Additionally, circ-SLC25A16 induces a rise in the rate of extracellular acidification, ATP synthesis, intake of glucose, and lactate formation, all of which support A549 cell growth. It operates as a ceRNA by binding to miR-488-3p and enhancing the synthesis of HIF-1α [221]. These studies highlight the multiple roles that circRNAs play in tumor development, including their involvement in metabolic reprogramming, modulation of miRNA activity, and regulation of vital cancer-related genes. Targeting circRNAs and their associated pathways could have potential therapeutic implications for cancer treatment.

8 Diseases Associated with Glutaminolysis Deregulation

Glutaminolysis, the metabolic pathway involving the conversion of glutamine to various intermediates, plays a crucial role in cellular function and energy production. This process has garnered significant attention due to its association with a range of diseases, particularly neurodegenerative diseases, Autoimmune diseases, kidney diseases, and cardiovascular diseases. Understanding the intricacies of glutaminolysis sheds light on the underlying mechanisms of these diseases and opens avenues for potential therapeutic interventions to target this metabolic pathway. Here, we discuss several diseases associated with glutaminolysis (Figure 3).

9 Glutaminolysis: Target for Therapeutic Intervention in Cancers

Various drugs are being tested in clinical trials to specifically target glutamine metabolism and inhibit glutaminolysis in different types of cancers (Figure 4 and Table 2). In addition, emerging therapeutic targets focus on mutated or overexpressed KRAS, which are present in multiple cancers. Clinical trials evaluate different inhibitors and targets of KRAS, with some inhibitors having already passed the initial clinical trial phase, while others are currently in phase 2 or 3 trials. One promising approach is targeting the downstream pathways of KRAS protein using MEK inhibitors such as selumetinib and trametinib. Selumetinib and docetaxel combination treatment significantly increased response rates for KRAS-mutant NSCLC as compared with when docetaxel (NCT01933932) is used alone (NCT01933932) [314]. Phase 3 studies are still underway to assess the efficacy of docetaxel with a placebo or selumetinib in treating NSCLC with KRAS mutations [315, 316]. To successfully sequester KRAS in an inactive state, these efforts require finding allele-specific drugs that disrupt the nucleotide exchange process and using a unique allosteric location in the binding pocket of the mutant cysteine residue [315, 316]. Clinical trials are also underway for EGLN1 inhibitors of KRAS, which are being tested for treating anemia [317]. These inhibitors have shown good tolerability and safety in early clinical trials.

Furthermore, the activation of AKT by the KRAS protein leads to the EphA2 phosphorylation at Ser897, resulting in increased cancer proliferation [318-320]. To target this pathway, there are ongoing phase 1 clinical trials for BT5528, a bicyclic peptide that binds to EphA2, and the phase 1 clinical trials for DS-8895a, an EphA2 monoclonal antibody (Phase 1, NCT02252211), are complete. In many malignancies, KRAS and its downstream signaling cascades are the focus of several more recent clinical trials, such as  (NCT01274624, NCT03808558, NCT03785249, NCT04006301, NCT03600883, and NCT03948763) [321]. These trials aim to explore novel approaches and potential therapeutic options. Even though cancer cell-targeted medicines have progressed, long-term use can frequently result in relapse and resistance to drugs, highlighting the necessity for more study and development of more potent treatments.

The importance of autophagy in the development and progression of tumors with KRAS mutation may be one of the reasons for investigating its inhibition [322]. Targeting autophagy and other pathways, such as the ERK/MAPK pathway, is a strategy to improve outcomes in cancers with KRAS mutations. Clinical studies evaluate a range of medications targeting HIF-1α, a protein involved in cancer growth. Some of these drugs include 2-methoxyestradiol, which downregulates HIF-1α at the posttranscriptional level [323], and tanespimycin, a heat-shock protein 90 inhibitor that destabilizes HIF-1α protein [324]. HIF-1α protein expression is inhibited at the translational level by vorinostat [325]. Other drugs like EZN-2968 and EZN-2208 are being tested to inhibit the expression of HIF-1α mRNA, leading to downregulating HIF-1α protein in cancer cells [326, 327]. Autophagy, a cellular process, has gained attention in cancer research, particularly in Burkitt's lymphoma. The autophagy inhibitors chloroquine and hydroxychloroquine have been approved by the US FDA [328]. Through the neddylation path, the autophagy activator pevonedistat has shown potential as a novel inhibitor. Combination therapies targeting multiple therapeutic locations using autophagy inhibitors like chloroquine and pevonedistat have shown intriguing results in clinical trials [329]. Several additional autophagy blockers, including the Lys05 family [330, 331], ROC-305 [332], and GNS561 [333], are currently being investigated in clinical trials. These inhibitors target the regulatory functions of lysosomes that are involved in autophagy. In addition to autophagy, lysosomotropic drugs like hydroxychloroquine have been found to inhibit micropinocytosis, which could potentially hinder tumor growth [334].

Moreover, these medications may have antitumor solid effects via lysosomal permeability independent of autophagy inhibition. Most evidence suggests that autophagy inhibition is beneficial in PDAC, and ongoing clinical trials explore the synergistic suppression of the ERK/MAPK pathway and autophagy. These trials focus on various ERK/MAPK pathway inhibitors combined with hydroxychloroquine. This combination therapy has shown promise in preclinical studies and is evaluated for its efficacy in clinical trials (NCT04145297, NCT03825289, NCT04132505) [322]. Inhibition of glutamine transporters to inhibit glutaminolysis in different cancers is also being investigated in NCT02771626 and NCT02071862 [335]. Clinical trials are underway for GLS inhibitors such as CB-839 (NCT02071862, NCT02071888, and NCT02071927), as well as OXPHOS inhibiting molecules like atovaquone, metformin, IACS-010759, and phenformin NCT03291938, NCT03026517, NCT01101438 and NCT03568994) [336-338]. MS0 (l-methionine sulfoximine), a GS inhibitor, is additionally being explored as a possible treatment for glutamine-addicted cancers [339, 340].

Many prospective paths are being investigated for cancer treatment, and targeting glutamine metabolism has emerged as a practical approach. Strategies include blocking receptors responsible for glutamine uptake, depleting glutamine levels in the plasma, and inhibiting glutamine synthesis and breakdown enzymes [341] (Table 3). In the case of acute lymphoblastic leukemia, targeting glutaminolysis has been demonstrated through the administration of l-asparaginase, which reduces plasma glutamine concentration by converting glutamine to glutamate [342]. However, a limitation of using l-asparaginase for targeting glutamine is the potential development of immunosuppression due to low glutamine levels required for the proliferation of immune cells [343]. Glutamine mimetics have been developed to inhibit glutaminolysis in various cancers. As an illustration, the glutamine analog DON acts as an antimetabolite and a GLS1 inhibitor. Another inhibitor, telaglenastat (CB-839), explicitly targets GLS1 and has shown promising results in combination with paclitaxel, demonstrating tumor inhibition [344]. Telaglenastat has been proven to be effective in suppressing GLS1 in tumors in patients with multiple myeloma and lymphoma, either alone or in combination with other treatments [345]. MLN4924, a small molecule inhibitor, inhibits neddylation, a form of posttranslational modification responsible for controlling glutamine metabolism. This inhibits the degradation of the glutamine transporter ASCT2, which increases glutamine influx and utilization and enhances anticancer efficacy [346]. Blocking has been targeted at several plasma membrane glutamine transporters, including SLC7A11, SLC6A14, and SLC38A1. Agents like erastin, α-Me-Trp, and MeAIB have been found to inhibit these transporters, leading to altered glutamine metabolism [347]. Pharmacological targeting of the ASCT2 receptor using -l-glutamyl-p-nitroanilide (GPNA) resulted in glutamine deprivation, specifically in ASCT2 overexpressing cells [348]. Another competitive antagonist of the transmembrane glutamine transporter, V-9302, has shown promising results by attenuating cell growth, promoting cell death, and inducing oxidative stress, leading to antitumor responses [349]. Glutamine metabolism in cancer cells is a growing area of interest due to its role in various cellular functions beyond its metabolic role. Understanding how cancer cells coordinate these functions and reprogram their metabolic pathways in response to environmental stress could provide therapeutic intervention opportunities. However, challenges remain, and more specific pharmacological interventions targeting cancer cells' vulnerabilities will be the future focus of the field.

10 Therapeutic Opportunities and Clinical Trials of lncRNAs in Controlling Cancer

The therapeutic use of RNA offers several common obstacles to developing lncRNA-based medication. These challenges render it more difficult to design treatments that target lncRNAs, including the absence of effective delivery systems, insufficient dose guidelines, and unknown adverse effects. While lncRNAs have been shown to regulate various human malignancies, the exact mechanisms through which they modulate metabolism are still largely unknown. There have been attempts to investigate lncRNA treatment in animal models using multiple techniques. Targeting lncRNAs with antisense oligonucleotides (ASOs) shows promise as a cancer treatment strategy. However, ASOs have poor membrane permeability, mainly confined to the cytoplasm, making it challenging to manipulate sub-nuclear lncRNAs. Linking nanotechnology with ASOs may hold the potential for addressing this limitation. CRISPR/Cas9 technology has received considerable attention in cancer treatment for specific DNA alteration of targeted genes. Recent research has shown successful silencing of lncRNA-expressing loci using CRISPR/Cas9 [350-352].

Nonetheless, there is still considerable uncertainty within the therapeutic use of CRISPR/Cas9 for lncRNA targeting in cancer therapy, as off-target cleavage events can occur despite the system's target specificity [392, 393]. Developing more specific gene-editing tools and techniques is crucial in this regard [394]. Viral vectors, including recombinant adenovirus, lentivirus, and retrovirus vectors, are superior RNAi transfection techniques. By neutralizing targeted RNA with exogenous double-stranded RNA molecules like siRNAs and shRNAs, RNAi is a technique for precise gene inhibition [395, 396]. Extensive research has been conducted on applying shRNAs to target lncRNAs in cancer therapy. However, the complexity of clinical trials using viral transfection is significant, and accurate viral infection and dose control are important considerations for future applications due to species-specific off-target effects [396, 397]. CircRNAs and lncRNAs are being studied in the context of LC metabolism, but their specific roles are still limited. Propofol, a drug used in metabolism-oriented treatment for LC that relies on lncRNAs or circRNAs, requires further analysis due to the inadequate clinical data on effectiveness and safety [398, 399]. Phase 2 or 3 clinical development is currently underway for miRNA-based treatments, including anti-miRNAs and miRNA mimics. Nevertheless, clinical practice does not now utilize circRNA- or lncRNA-based therapies [221]. It is important to note that these drugs and therapies are still in the clinical trial phase, and further research is needed to determine their effectiveness and safety in treating cancer.

Further study is required to understand better the regulatory framework involved in cancer metabolism and to discover possible targets for developing cancer treatment approaches. It is anticipated that future research on the most effective treatment options for tumor patients with metabolic disorders will be influenced by the advancements achieved in mechanical analysis of lncRNA activity and metabolic signaling in recent years. But, expecting lncRNA-targeted treatment to restore normal metabolism at this point would be unrealistic.

11 Conclusion and Future Perspectives

Glutamine, the most prevalent amino acid in blood and muscle, is essential to many biological functions and plays a significant part in maintaining metabolic homeostasis. While glutamine is considered a nonessential amino acid for normal cells, it becomes necessary for cancer cells to generate ATP and synthesize crucial cellular building blocks in a challenging TME. Numerous diseases, such as cancer, neurological problems, and metabolic abnormalities, are associated with elevated levels of glutamine. Nevertheless, there are insufficient data to determine a direct causative link between these diseases and aberrant glutamine levels. Although the precise processes are yet unknown, some research indicates that glutamine variations may impact the extent or progress of the disease.

Glutamate transporter and other modulator expression and function changes are linked to several diseases. Understanding whether these alterations contribute to pathogenesis or result from preexisting disease is critical. In many situations, the involvement of glutamate transporters and other modulators in pathogenesis remains unclear due to inconsistent data, potentially due to different methodologies. The metabolic process of glutaminolysis has been established as essential for multiple tumors. Neoplastic cells must absorb and metabolize external nutrients to provide energy and building blocks for unchecked tumor growth. There is growing interest in understanding how cancer cells sustain their metabolic functions by utilizing available resources. This glutamine reliance is often referred to as an Achilles’ heel of cancer, as it represents a metabolic vulnerability that can be targeted therapeutically. The complicated characteristics of cancer metabolism are highlighted by the variations in the dependency of cancer cells on metabolic sources of fuel caused by mutational heterogeneity. The regulation of glutamine metabolism largely depends on oncogenes and tumor suppressor genes. Glutamine addiction in cancer is driven by oncogenes such as c-Myc, KRAS, and EGFR, as well as the loss of tumor suppressor genes like p53, INK4, and PTEN. Understanding the metabolic dynamics within tumors has driven the development of biologically targeted therapies that exploit these metabolic vulnerabilities. Although research suggests that both oncogene and tumor suppressor expression levels may influence glutamine metabolism in cancer cells, little is known about glutamine metabolism in cancer, especially concerning glucose metabolism.

Additionally, deregulated enzymes in glutamine metabolism contribute to cancer metabolic variations, acting as signaling molecules and activators of cancer progression factors like mTOR and autophagy. KRAS mutations may have specific effects on metabolism in tumor tissues due to their intrinsic metabolic wiring. Studying these effects requires considering the tumor suppressor background and investigating how oncogenic KRAS-dependent metabolic changes are altered in the TME, including hypoxia, limited nutrients, and crosstalk between tumor cells and stromal cells. Myc, a common deregulated oncoprotein in cancer, is a challenging target due to its lack of enzymatic activity or structural features. Inhibiting Myc can lead to cell cycle arrest, ATP production collapse, and apoptosis. Efforts have included altering Myc DNA-binding sites, Myc-Max heterodimers, and transcriptional activation machinery. However, early Myc inhibitors have shown disappointing therapeutic efficacy in vivo. Non-neoplastic settings allow for considering downstream Myc effectors, such as PDH, which could be therapeutic. Other targetable enzymes in the Myc axis have the potential for cotargeting Myc downstream genes/pathways. It found enrichment of mTOR signaling, antioxidant, and redox balance pathways in Myc target genes, providing the basis for a clinical trial using GLS and mTOR cotargeting strategy. KRAS mutations may have specific effects on metabolism in tumor tissues due to their intrinsic metabolic wiring. Studying these effects requires considering the tumor suppressor background and investigating how oncogenic KRAS-dependent metabolic changes are altered in the TME, including hypoxia, limited nutrients, and crosstalk between tumor cells and stromal cells.

Certain cancers, like those driven by Myc, rely on glutamine, making targeting glutamine metabolism pharmacologically beneficial. Oncogenic drivers may result in tumor cells bypassing glutamine's need. Targeted inhibition of some oncogenic drivers can rewire cells to become dependent on glutamine, making targeted inhibitors potentially lethal. The field of cancer metabolism has made progress in understanding alternative fuel sources for cancers, including glutamine, which can be exploited for therapeutic purposes under specific circumstances. Targeting glutaminolysis with inhibitors of oncogenic proteins and dysregulated proteins in the glutamine pathway has shown promising therapeutic results for aggressive cancers. Clinical trials investigating small molecular inhibitors will enhance understanding of their efficacy in humans and correlate with preclinical models. As mutational heterogeneity in cancer metabolism deepens, new targets are emerging to effectively target each aspect of metabolic reprogramming. Combining dual therapy strategies may help overcome chemoresistance in glutamine-dependent metabolic cancers.

Furthermore, ncRNAs, such as miRNAs, lncRNAs, and circRNAs, regulate glutamine addiction and metabolism. These ncRNAs modify essential hallmarks of cancer, including metabolic rewiring, allowing cancer cells to adapt to the challenging microenvironment and sustain deregulated proliferation. However, it is still unclear how exactly lncRNAs affect the hallmarks of cancer metabolic reprogramming, work, and what species they belong to. In vitro studies provided the majority of information about the metabolic activities and regulatory mechanisms of lncRNAs. Further in vivo studies using lncRNA knockdown or overexpression are required to examine their involvement in metabolic regulation. LncRNA-based diagnostics and treatments for cancer metabolism are expected to benefit cancer patients, while clinical trials are still in the early phases. The development of lncRNA inhibitors and analytical techniques will make it possible to quickly screen for lncRNAs differently expressed in cancer and elucidate the mechanisms underlying cancer metabolism, opening the door to potential clinical applications and future approaches to cancer treatment. Altered glutamine levels are linked to various human pathologies, but limited evidence supports direct causality. Although glutamine-targeting interventions have shown potential, they have generally lacked efficacy in vivo. The extent to which glutamine dysregulation contributes to disease development in specific mouse models is unknown due to the absence of specialized pharmacological techniques or genetically altered mouse models.

In conclusion, glutamine is a versatile molecule that plays a crucial role in various biological processes, including antioxidant defense, neurotransmission, and cellular metabolism. Alterations in glutamine levels or its metabolic pathways have been associated with multiple health disorders, ranging from neurodegenerative diseases to cancer. However, the exact causal relationships remain often unclear, complicating our understanding of how changes in glutamine metabolism contribute to these conditions. Additionally, targeting glutamine metabolism for therapeutic purposes has proven to be challenging. This difficulty arises from the complexity of glutamine's functions in the body and the need for precise interventions that do not disrupt its essential roles in normal physiological processes.

Author Contributions

M.A.K., S.K.B., I.R.K., A.A.B., and M.A.M. wrote the manuscript and generated figures. M.A.M. and A.A.B. contributed to the concept and design and critically edited the manuscript. M.S.K., F.M.H., S.A., M.H., M.S., A.S.A.A., A.A.B., and M.A.M. performed critical revision and editing of the scientific content. All authors read and approved the final manuscript.

Ethics Statement

Not appliable.

Conflicts of Interest

The authors declare no conflicts of interest.