Given the expanding global population and increasing life expectancy, the alarming incidence cancer in 2030 was predicted to reach 20.3 million worldwide, and is poised to become the main cause of morbidity and mortality across all regions of the world in the next decade [
29]. Specifically, the highest lifetime risk of developing of cancers is lung cancer, the most common type of cancer death in 94 countries, in both high and low Human Development Index(HDI) regions [
29]. What’s worse, despite many years of efforts, the metabolic reprogramming of cancer cells is far beyond fully understanding and there has been limited clinical progress in addressing this mechanism [
30] due to the complex features of aberrant cancer metabolism.
Recent studies have reported a significant upregulation of SFXN1 mRNA level in various malignancies compared with normal tissues [
15]. Despite some studies investigating the differential expression and function of the SFXN family genes in cancer, no evidence regarding the association of SFXN1 expression with immunotherapy response has been identified. Therefore, this study employed data mining techniques alongside clinical validation to identify promising prognostic biomarkers within the SFXN family for LUAD. By conducting a comprehensive and comparative analysis of the compound genes, we aim to unveil their interrelations and pinpoint novel biomarkers. Targeting a specific gene in the SFXN family aligns with “Precision Medicine” and has the potential to induce maximally synergistic effects in clinical practice. Our results showed a significant increase in the expression of SFXN1 and SFXN4 in LUAD. However, only SFXN1 had a significant impact on the overall survival of LUAD patients. Previous research has shown that all SFXN family members are present in pancreatic islet cells while SFXN3, in particular, is a crucial carrier molecule for the differentiation and regeneration of pancreatic β-cells [
31]. Likewise, while many SFXN family members are highly expressed in LUAD, SFXN1 exerts a significant influence on the development and progression of the disease. Our investigation was focused on SFXN1 as a potential prognostic biomarker for LUAD. We found that SFXN1 expression was correlated with clinical stage, tumor size, lymph node invasion, and distant metastasis. Our study also demonstrated that patients with high SFXN1 expression had a significantly higher TMB, which is consistent with previous findings linking high TMB to a poor prognosis for non-small cell lung cancer [
32]. Furthermore, both univariate and multivariate Cox regression analyses confirmed that SFXN1 remained a prognostic indicator associated with overall OS, indicating that this gene acts as an independent predictor for LUAD. After extending the follow-up period, the accuracy of nomogram based on the SFXN1 protein expression and clinical stage has significantly improved, indicating a more reliable prediction of the outcome. This extension allowed for a more thorough analysis of the data and additional insights into the long-term effects of the intervention.
The KEGG pathway and GO analysis of SFXN1 interactive genes in this study evidenced that SFXN1 participates in the cell cycle, including DNA replication, meiosis, and signaling pathways such as p53 and HIF-1 pathways. As mitochondria carry their own DNA (mtDNA) in eukaryotic cells, they also regulate the cell cycle and mitosis through their metabolic activities [
10]. Mutations in mtDNA can generate a more favorable metabolic profile in rapidly proliferating tumor cells, accelerating cancer progression and metastasis [
33]. Thus, SFXN1 may exacerbate LUAD progression by promoting cancer cell proliferation, inhibiting apoptosis, and activating invasion and metastasis pathways. Therefore, altered expression levels of SFXN1 in LUAD might be a critical factor, and further investigation is necessary to determine whether it acts as a driver or passenger oncogene. Functionally, the GO molecular function (MF) analysis revealed that SFXN1 has a broad function in cytoskeletal movement and microtubule binding, both of which require mitochondrial energy in process.
Mitochondria serve as the primary energy-producing organelles in cells and have key roles in generating reactive oxygen and regulating iron metabolism. Mutation of the SFXN1 gene, which is responsible for facilitating the transportation of iron into the mitochondria, can result in pathologic accumulation of iron within erythrocytes, as observed in flexed-tail (f/f) mice [
34]. In addition, researchers discovered that SFXN1 is a mitochondrial iron transporting protein that carries excess free iron into the mitochondria, resulting in mitochondrial damage and subsequent cardiomyocyte hypertrophy in the long term [
35]. Sousa and colleagues also demonstrated that erythrocytes in patients with iron overload exhibit elevated ATPase activity [
36]. These findings highlight the critical role of proper regulation of iron metabolism in mitochondria for maintaining cellular function. Therefore, SFXN1 might facilitate iron transport by boosting the ATPase activity, which needs additional experimental verification. Additionally, during the last decade, metabolic reprogramming has been the most significant factor observed among the pathways of central carbon metabolism in cancer cells [
37]. Our study’s KEGG enrichment analysis revealed that SFXN1 participated in the folate-dependent central carbon metabolism in cancer, which is consistent with the remarkable work that identified SFXN1 as the primary SFXN carrier protein for transporting serine into mitochondria during the one-carbon metabolism process [
15]. Our study also found that tumor purity was higher in LUAD patients with SFXN1 overexpression, indicating that there were fewer infiltrating immune cells in the tumor microenvironment. This may be one of the reasons for the poorer prognosis of patients with SFXN1 upregulation. A growing body of evidence has shown that mitochondria not only serve as the “energy factories” of immune cells, but also modulate immune responses by regulating metabolic and physiological states in different types of immune cells [
38,
39]. Serine is a primary source of one-carbon units and abnormal metabolism of serine is closely related to cancer progression [
40]. Studies have confirmed that restricting serine can inhibit tumor growth in mice [
41]. Thus, inhibiting the expression of SFXN1, which is the key transporter of serine into mitochondria [
15], may reduce tumor development. Importantly, our study found that a large number of MCs, eosinophils, macrophages, MDSCs, and Treg cells infiltrated the SFXN1 high-expression group, all of which can shape an immunosuppressive microenvironment and promote tumor growth under certain conditions. For example, MCs release FGF-2, NGF, PDGF, VEGF, IL-8, and IL-10, which promote the expansion of tumor cells [
42]. Li et al. demonstrated that eosinophils promote tumor cell migration and bone metastasis by secreting C-C motif chemokine ligand 6 (CCL6) in mice [
43]. MDSCs have been shown to suppress immune responses and protect tumor cells from attack, making them valuable prognostic biomarkers and potential targets for anti-cancer therapies [
44]. Macrophages are classified into pro-inflammatory (M1) and immunosuppressive (M2) macrophages, with M2-polarized macrophages associated with poorer prognosis in cancer patients. Therefore, SFXN1 may modulate the tumor immune microenvironment by directly or indirectly influencing the infiltration of immune cells. Importantly, we found that SFXN1 overexpression was associated with higher expression of immune checkpoints CD274 (PD-L1) and IDO1. CD274 is predominantly expressed by tumor cells, binding with programmed cell death-1(PD-1) on the surface of T cells and triggering immune escape [
44]. IDO1, indoleamine 2, 3-dioxygenase 1, is a widely expressed enzyme in human cancers that metabolizes tryptophan to kynurenine, which mainly interacts with effector T cells to impair their antitumor effects and facilitate immune escape [
45,
46]. Additional studies indicated that IDO1 enhances the proliferation of regulatory T cells (Tregs) and activates MDSCs [
47,
48]. Therefore, these findings suggest that the high expression of SFXN1 in the tumor immune microenvironment may lead to immunosuppression and possibly compromise the efficacy of immunotherapeutic strategies.
While our research has confirmed the role of SFXN1 in LUAD using publicly available cohorts and our own clinical samples, further investigations are required to fully understand the biological processes and oncogenic mechanisms that involve SFXN1, both in vitro and in vivo.