2.1 m6A writers

m6A writers primarily comprise the m6A methyltransferase complex, which includes methyltransferase-like 3 (METTL3), methyltransferase-like 14 (METTL14), and Wilms tumor 1-associating protein (WTAP).²³ METTL3, the first discovered writer and a key member of this complex, is a 70-kDa protein that contains a methyltransferase domain that methylates RNA. Similarly, METTL14 can methylate RNA. In addition, when METTL3 and METTL14 form a complex with a 1:1 stoichiometry,²⁴ METTL3 and METTL14 have much better m6A methyltransferase activity than either alone.²⁵ Although WTAP does not directly methylate RNA, it significantly influences the structure and substrate specificity of the METTL3/METTL14 complex. In nuclear speckles, WTAP is associated with the accumulation of METTL3 and METTL14.²⁶ Moreover, other important methyltransferases have been reported, including methyltransferase-like 5 (METTL5),²⁷ methyltransferase-like 7A (METTL7A),^{28, 29} methyltransferase-like 7B (METTL7B),^{30, 31} methyltransferase-like 16 (METTL16),^{32, 33} KIAA1429,^{34, 35} RNA binding motif protein 15 (RBM15),³⁶ zinc finger CCHC domain-containing protein 4 (ZCCHC4),³⁷ and zinc finger CCCH domain containing protein 13 (ZC3H13).³⁸ Emerging evidence has revealed the important role of writers in tumorigenesis,³⁹ tumor metastasis,⁴⁰ and immunotherapy.⁴¹ m6A writers can upregulate the m6A methylation of cancer-related genes. For example, Wei et al.⁴² demonstrated that METTL3 accelerates gastric cancer (GC) progression through the ADAMTS9-mediated phosphatidylinositol-3 kinase (PI3K)/V-akt murine thymoma viral oncogene homolog (AKT) pathway. Zhou et al.⁴³ found that the METTL3/YTHDF2 m6A axis accelerated colorectal carcinogenesis. Collectively, m6A writers promote RNA methylation and induce multiple biological functions.

2.2 m6A erasers

m6A erasers, also known as m6A demethylases, remove m6A from RNA and decrease m6A levels. Fat mass and obesity-associated protein (FTO) has been considered as the first m6A demethylase since He et al.⁴⁴ discovered that m6A was the main substrate of FTO in nuclear RNA in 2011. FTO not only plays an essential role in obesity by regulating adipogenic pathways and inducing preadipocyte differentiation to facilitate adipogenesis but also participates in tumor processes.^{45, 46} Another key demethylase, human AlkB homolog 5 (ALKBH5), also plays a role in m6A modifications associated with various diseases, especially tumors. ALKBH5 belongs to the alkB family of dioxygenases, which regulate oxidative demethylation to modulate the repair of N-alkylated nucleobases.^{47, 48} ALKBH5 participates in multiple cancer or noncancer processes.⁴⁹ Protein arginine methyltransferase 5 (PRMT5) is a new demethylase that inhibits RNA m6A modification by enhancing the nuclear translocation of ALKBH5.⁵⁰ In summary, m6A erasers play important roles in demethylating RNA and inducing subsequent functions.

2.3 m6A readers

m6A readers, also called m6A-binding proteins, can recognize and bind to m6A-modified transcripts to regulate the expression and function of several genes during various processes. Several m6A readers have been identified, including the YTHDF family,^51-54 YTHDC family,^{55, 56} IGF2BP family,^57-59 RNA-binding protein heterogeneous nuclear ribonucleoprotein (HNRNP) family,^{60, 61} proline-rich coiled-coil 2A (PRRC2A),⁶² ELAV-like RNA-binding protein 1 (ELAVL1),⁶³ and fragile X mental retardation protein (FMRP).⁶⁴ The YTHDF family includes three proteins: YTHDF1, YTHDF2, and YTHDF3. YTHDFs have different functions. YTHDF1 recognizes m6A-modified RNA and promotes its translation into the cytoplasm. YTHDF2 participates in the degradation of m6A-modified RNA. YTHDF3 enhances mRNA translation and degradation. YTHDFs regulate cancer progression in an m6A-dependent manner. For instance, Chen et al.⁶⁵ found that YTHDF1 could facilitate the translation of FOXM1 via m6A modification, which subsequently promotes breast cancer (BC) progression. However, a new model for YTHDFs provided by Zaccara and Jaffrey⁶⁶ illustrates that YTHDFs function together to mediate the degradation of m6A-modified RNA. Liquid‒liquid phase separation (LLPS) is the formation of several membraneless condensates. Interestingly, YTHDFs have the potential to form condensates. Specifically, the N-terminal of YTHDFs is mainly an intrinsically disordered region (IDR), whereas the C-terminal consists of the m6A-binding YTH domain. Both play vital roles in the formation of the condensates.⁶⁷ Zou et al.⁶⁸ found that YTHDF1 and YTHDF2 can form different granules because of their diverse low-complexity regions. In addition, Fu et al.⁶⁷ found that under oxidative stress, YTHDF1 and YTHDF3 are abundant in stress granules rather than in processing bodies (P-bodies). However, YTHDF2 was abundant in both stress granules and P-bodies.⁶⁷ Stress granules and P-bodies have several functions, including metabolic reprogramming, targeting and silencing of specific mRNAs via the RNA-induced silencing complex, and restoring and permitting the translation of key specific mRNAs.⁶⁹ YTHDCs are YTH domain-containing proteins, including YTHDC1 and YTHDC2. YTHDC1 mediates nuclear export of m6A-modified mRNA and facilitates mRNA splicing.^{55, 70} Recent studies have also demonstrated its ability to form membraneless organisms such as nuclear condensates via LLPS, which may be associated with gene expression, transcript splicing, and nucleocytoplasmic export.⁷¹ YTHDC2 enhances translation efficacy and destabilizes cytoplasm.⁵⁶ IGF2BP family refers to Insulin-like growth factor 2 mRNA-binding proteins, which mainly consist of IGF2BP1, IGF2BP2, and IGF2BP3. IGF2BPs are key readers of m6A modifications that stabilize mRNA and promote its translation in the cytoplasm.⁵⁹ HNRNPA2B1 has multiple functions in m6A modification, including processing of miRNA,⁷² promotion of nucleocytoplasmic trafficking,⁷³ and stabilization of m6A-modified mRNA.^{60, 61} HNRNPC is an abundant nuclear RNA-binding protein that recognizes and splices m6A-modified RNA.⁷⁴ HNRNPG recognizes RNA via RNA recognition motif (RRM) and Arg-Gly-Gly (RGG) motifs and splices m6A-modified RNA.⁷⁵ PRRC2A plays a role in target RNA stabilization. For example, PRRC2A stabilizes Olig2 mRNA via the GGACU motif.⁶² ELAVL1 (also known as HuR) interacts with other m6A regulators and promotes m6A-modified RNA stabilization.^{40, 76} FMRP sustains m6A-modified stability and maintains its expression via m6A modification.⁷⁷ Collectively, m6A-binding proteins recognize m6A-modified mRNA and influence their stability, localization, and translation. Notably, condensate formation of reader proteins induced by LLPS is an important direction for future research.

3.1 Methylation

Arginine methylation of m6A regulatory proteins, induced by protein arginine methyltransferases (PRMTs), plays a vital role in cancer initiation and progression. Methylated METTL14 participates in cancer development. For instance, PRMT1-mediated methylation of METTL14 at the C-terminus is important for its function in catalyzing m6A modification, which promotes cancer cell proliferation.⁷⁸ PRMT3 methylates METTL14, which downregulates its expression and promotes EC progression by influencing glutathione peroxidase 4 (GPX4) expression.⁷⁹ WTAP is methylated by PRMT1, which targets the oxidative phosphorylation of multiple myeloma (MM) cells and promotes carcinogenesis via m6A modification of NDUFS6.⁸⁰ PRMT6 can methylate the m6A eraser ALKBH5 at R283, which upregulates LDHA expression and promotes aerobic glycolysis and tumorigenesis in BC.⁸¹ In conclusion, arginine methylation of the m6A regulator is a potent target for cancer modulation.

3.2 Acetylation

Acetylation is a PTM.⁸² METTL3 is acetylated by P300 at K177. Acetylated METTL3 dampens its subcellular localization and function in BC cells, impeding cancer metastasis.⁸³ Similarly, METTL3 is acetylated by acetyl-CoA acetyltransferase 1 (ACAT1) at residues K263 and K388, which stabilizes METTL3 and suppresses triple-negative BC migration and invasion via the NR2F6/ACAT/METTL3 axis.⁸⁴ Acetylation of the m6A writer complex METTL3–METTL14–WTAP can be induced by sulfatide, thus regulating MTF1 expression to promote the growth of hepatocellular carcinoma (HCC) cells.⁸⁵ KAT2B, a lysine acetyltransferase, catalyzes METTL14 acetylation at K398 and increases METTL14 protein stability to upregulate m6A methylation of Spi2a mRNA, which inactivates the NF-κB pathway.⁸⁶ ALKBH5 is acetylated by lysine acetyltransferase 8 (AT8) at K235, which enhances its demethylase activity.⁸⁷ IGF2BP2 is acetylated at K530. Loss of its deacetylase SIRT1 recruits nuclease XRN2 to degrade the ATP6V1A transcript.⁸⁸ All the abovementioned studies show that acetylation can regulate the function of m6A regulatory proteins to influence disease processes.

3.3 Lactylation

The Warburg effect involves an increase in glycolytic metabolism even in the presence of O₂, which is crucial for carcinogenesis, metastasis and drug resistance.^{89, 90} Lactate, a product of glycolytic metabolism, has recently gained increasing attention owing to its potential biological functions in cancer. Lactylation is a novel epigenetic modification.⁹¹ In 2019, Zhang et al.⁹² reported the metabolic regulation of histone lactylation gene expression. Current evidence indicates that lactylation is involved in m6A modifications. Histone lactylation is abundant in the promoters of m6A regulators and significantly influences their expression to a great extent. For instance, YTHDF1 and YTHDF2 expression are facilitated by H3K18la.^{93, 94} In contrast, lactylation directly modifies m6A regulators. Xiong et al.⁹⁵ identified two lactylation modification sites, K281 and K345, in the zinc-finger domain of METTL3. The lactylation of METTL3 enhances its binding to Jak1 mRNA and therefore promotes the immunosuppression of colorectal cancer (CRC). Sun et al.⁹⁶ found that high copper stress could induce METTL16 lactylation at K229, which upregulates the activity of METTL16 and, therefore, promotes FDX1 expression to facilitate carcinogenesis in GC. In summary, lactylation is important and may provide a new strategy for m6A modification in cancer cells.

3.4 Ubiquitination

Ubiquitin is a small protein consisting of 76 amino acids that assists in protein degradation by the S26 proteasome.⁹⁷ Protein ubiquitination is a ubiquitin-dependent PTM that affects all cellular processes.⁹⁸ Ubiquitination plays a key role in translation by modifying ribosomal and regulatory proteins.⁹⁹ In this study, we focused on its role as an m6A regulator. METTL14 is ubiquitinated and degraded by HRD1 to suppress endoplasmic reticulum-related liver diseases.¹⁰⁰ Serine/threonine kinase receptor-associated protein (STRAP) ubiquitinates FTO at K216, promotes its degradation, stabilizes MTA1 mRNA, and promotes CRC metastasis.¹⁰¹ KRT17 promotes YTHDF2 degradation via ubiquitination, which stabilizes CXCL10 mRNA and induces T-cell infiltration in CRC.¹⁰² IGF2BP protein ubiquitination can be upregulated by circNDUFB2 and TRIM25 (a kind of E3 ubiquitin ligase), leading to antitumor immunity in non-small cell lung cancer (NSCLC).¹⁰³ IGF2BP1 is ubiquitinated by the E3 ubiquitin ligase FBXO45 at K190 and K450, which subsequently induces PLK1 upregulation and HCC carcinogenesis.¹⁰⁴ Additionally, blocking the ubiquitination of IGF2BP proteins can suppress their degradation and promote their binding to RNA.^105-107 These studies demonstrate that ubiquitination mainly affects m6A regulator proteins by promoting degradation.

3.5 SUMOylation

SUMOylation is a novel PTM associated with small ubiquitin-like modifiers that participate in cancer development of cancer.¹⁰⁸ SUMOylation of m6A regulators influences their functions. For example, extracellular signal-regulated kinase (ERK)/c-Jun N-terminal kinases (JNK) signaling-induced SUMOylation of ALKBH5 represses its demethylase activity.¹⁰⁹ YTHDF2 can be SUMOylated by PIAS1 at K281, K571, and K572, thereby facilitating the degradation of viral RNA and suppresses the replication of EBV.¹¹⁰ Additionally, YTHDF2 SUMOylation at K571 promotes mRNA decay and tumorigenesis.¹¹¹ FTO is modified by SIRT1 at K216, which promotes FTO SUMOylation and degradation, decreases GNAO1 expression, and promotes HCC.¹¹² METTL3 is reported to be SUMOylated at K177, K211, K212, and K215, which represses METTL3 m6A methyltransferase activity.¹¹³

3.6 Phosphorylation

Phosphorylation is an extensively studied PTM of proteins.¹¹⁴ Over the past 25 years, protein phosphorylation has been observed at the serine, threonine, and tyrosine residues.¹¹⁵ Protein phosphorylation regulates important cellular functions.¹¹⁶ ERK phosphorylates and stabilizes the METTL3/14-WTAP methyltransferase complex, which increases m6A methylation and promotes tumorigenesis.¹¹⁷ Specifically, METTL3 is typically modified at S43, S50, and S525,^{117, 118} whereas WTAP is modified at S306 and S341.^{117, 119} METTL3 is also phosphorylated at residue S67, which promotes its activation and helps m6A modification to stabilize IRF3 mRNA, leading to antiviral immunity.¹²⁰ YTHDF2 can be phosphorylated and stabilized by EGFR/SRC/ERK signaling at S39 and T381, which facilitates the degradation of LXRA and HIVEP2 mRNA, leading to glioblastoma (GBM) tumorigenesis.¹²¹ Therefore, phosphorylation mainly stabilizes m6A regulators and activates their function in target mRNAs to modulate downstream gene expression and cellular processes. Current studies on SUMOylation of m6A regulators are limited and require further research.

3.7 O-GlcNAcylation

O-linked N-acetylglucosaminylation (O-GlcNAcylation) is a type of glycosylation related to O-GlcNAc at serine or threonine residues.¹²² Yang et al.¹²³ found that YTHDF2 is modified by O-GlcNAc transferase 8(OGT8) at S263, which stabilizes YTHDF2 and enhances its activity by inhibiting its ubiquitination. Subsequently, O-GlcNAc-modified YTHDF2 stabilizes the minichromosome maintenance protein 2/5 transcripts to facilitate the onset of HBV-related HCC.¹²³ YTHDF1 O-GlcNAcylation at S196, S197, and S198 promotes YTHDF1 cytosolic localization and upregulates the expression of downstream target genes to promote CRC tumorigenesis.¹²⁴ The O-GlcNAcylation of m6A regulators has a significant impact on diseases, and related research has great potential.

These studies show that diverse PTMs in m6A regulators significantly influence their functions in cancer.

5.1 Autophagy

Autophagy is highly conserved in eukaryotes. It maintains cellular homeostasis and metabolism.¹³⁵ When stressed by internal or external stimuli, cells combine autophagosomes with lysosomes to form autolysosomes, membrane structures that engulf and degrade aged or injured organelles, misfolded proteins, and pathogens to regulate cell homeostasis of cells.¹³⁶ Additionally, this process plays a vital role in tumors by promoting and inhibiting tumorigenesis and progression.¹³⁷

m6A modifications are closely associated with autophagy. The upregulation of m6A RNA modification is helpful for autophagosome formation when nutrients are deficient.¹³⁸ The m6A modification can regulate autophagy-related genes (ATGs), ultimately influencing their function and promoting the onset of various diseases, including tumors.¹³⁹ For instance, Hao et al.¹³⁸ reported that YTHDF3 responds to METTL3-associated m6A hypermethylation and recruits eIFs to promote FOXO3 translation, consequently activating a subset of ATGs and leading to autophagy. In liver tissues, m6A RNA modification plays a complex role in tumor progression. If not addressed, liver fibrosis can progress to cirrhosis and cancer. Hepatic stellate cells (HSCs) play an important role in myofibroblast matrix production during this physiological and pathological process. Shen et al.¹⁴⁰ showed that YTHDF1 stabilizes BECN1 mRNA and promotes autophagy activation via m6A modification in HSCs. The hypoxia-inducible factor-1α (HIF-1α) drove YTHDF1 to bind to the m6A-modified mRNA of ATG2A and ATG14, which can contribute to the translation of ATG2A and ATG14, thereby promoting the survival of HCC under hypoxic conditions and its progression.¹⁴¹ One study identified an oncogenic circRNA, circMDK, as a potential biomarker for HCC, because its upregulation with m6A modification upregulates ATG16L1, resulting in the activation of the PI3K/AKT/mTOR signaling pathway to promote cell proliferation, migration, and invasion.¹⁴² Interestingly, circRNAs have gained considerable attention in recent years. CircRAB11FIP1 mediates the expression of ATG5 and ATG7 via m6A, thus promoting epithelial ovarian cancer.¹⁴³ LKB1 is regulated by WTAP via m6A modification and then phosphorylated AMPK. At the same time, researchers found that knockdown of WTAP could upregulate the level of autophagy and inhibit hepatoblastoma (HB) cell proliferation.¹⁴⁴ Similarly, the progression of clear cell renal cell carcinoma (ccRCC) can be regulated by FTO because it inhibits autophagy and promotes tumorigenesis through an m6A-IGF2BP2-dependent mechanism, indicating that FTO can be a prognostic biomarker and a promising target in ccRCC.¹⁴⁵

In parallel, m6A-dependent autophagy plays an important role in antitumor drug resistance. From a pharmacokinetic perspective, m6A modifications influence drug transport and metabolism. This may be related to several membrane transporter proteins such as ATP-binding cassette proteins. In addition, m6A can alter drug targets to regulate drug response and resistance.¹⁴⁶ Autophagy promotes anticancer drug resistance to protect tumor cells from survival.¹⁴⁷ Furthermore, m6A modification can modulate ATGs (ATG5 and ATG7) to regulate the formation and progression of autophagosomes, thus influencing autophagy and promoting the survival and anticancer resistance of tumor cells.¹⁴⁸ However, the underlying mechanisms remain unclear. FTO-mediated cisplatin resistance in GC is attributed to the inhibition of Unc-51-like kinase (ULK1)-mediated autophagy.¹⁴⁹ Sun et al.¹⁵⁰ found that METTL3 is a marker for poor prognosis of small-cell lung cancer (SCLC) and is highly expressed in chemoresistant SCLC cells through Pink1–Parkin pathway-mediated mitophagy. Translation machinery-associated 7 homolog (TMA7) plays a key role in the carcinogenesis of laryngeal squamous cell carcinoma (LSCC) and cisplatin resistance via the IGF2BP3/TMA7/UBA2 axis.¹⁵¹ In liver cancer, sorafenib resistance is induced by m6A-dependent, FOXO3-mediated autophagy.¹⁵² m6A plays a crucial role in the onset, progression, and drug resistance of multiple tumors by regulating autophagy, suggesting a promising breakthrough in future antitumor treatments.

5.2 Ferroptosis

Ferroptosis, first described by Dixon et al. in 2012,¹⁵³ is a new nonapoptotic form of PCD. Biochemically, ferroptosis is a ROS-dependent PCD characterized by iron accumulation and lipid peroxidation.¹⁵⁴ Emerging evidence has demonstrated its mechanisms, including the suppression of system Xc−, GPX4, mitochondrial voltage-dependent anion channels, and P53.¹⁵⁵ System Xc is an amino acid antitransporter composed of two subunits, SLC7A11 and SLC3A2.¹⁵³ GPX4 plays a pivotal role in the induction and regulation of ferroptosis by inhibiting lipid peroxide formation.

Studies have indicated that m6A modification can regulate cancer cell ferroptosis via m6A-modifying ferroptosis-associated mRNA to modulate these mechanisms. For instance, SLC7A11 has been widely demonstrated to be m6A-modified by several m6A regulators. SLC7A11 mRNA stability can be promoted, and its expression can be upregulated in a METTL3 manner, resulting in tumor growth and ferroptosis resistance in HB¹⁵⁶ and lung adenocarcinoma.¹⁵⁷ SLC7A11 can be downregulated by FTO to inhibit thyroid cancer progression via ferroptosis.¹⁵⁸ ALKBH5 can decrease the expression of SLC7A11 by repressing the m6A modification, which promotes ferroptosis in CRC¹⁵⁹ and thyroid cancer.¹⁶⁰ NF-κB activating protein (NKAP) serves as a novel suppressor of ferroptosis. NKAP binds to m6A and promotes SLC7A11 mRNA splicing to protect GBM cells from ferroptosis.¹⁶¹ METTL16 increases GPX4 expression by modifying m6A to inhibit ferroptosis and promote BC.¹⁶² The mature GPX4 mRNA contains three m6A binding motifs. RUNX1 intronic transcript 1 (RUNX1-IT1) directly binds to IGF2BP1 and promotes LLPS to increase GPX4 mRNA stability, thereby blocking ferroptosis and promoting BC carcinogenesis.¹⁶³ Erianin, a low-molecular-weight bibenzyl natural product extracted from Dendrobium chrysotoxum, induces ferroptosis in renal cancer stem cells by promoting the m6A methylation of ALOX12 and P53 mRNA.¹⁶⁴ m6A-modified FGFR4 reduces ferroptosis in recalcitrant HER2-positive BC via the β-catenin/TCF4-SLC7A11/FPN1 axis.¹⁶⁵

Furthermore, m6A-associated ferroptosis plays an important role in anticancer immunity and immunotherapy. Immunotherapy-activated CD8+ T cells can regulate tumor ferroptosis to enhance the antitumor effects.¹⁶⁶ Several studies have aimed to bioinformatically analyze the relationship between m6A modification, ferroptosis, and immunity in cancers. SLC17A9 is associated with tumor immune infiltration, m6A modification, and ferroptosis in HCC.¹⁶⁷ Li et al.¹⁶⁸ found that the expression of BTBD10 (an activator of the Akt family) was correlated with some m6A-associatedgenes, ferroptosis-related genes, and immune checkpoints in HCC. Wang et al.¹⁶⁹ showed that YTHDF1 suppresses CD8+ T cell-related anticancer effects and ferroptosis by stabilizing programmed cell death ligand 1 (PD-L1) transcripts, which are important for prostate cancer cells to evade effector T cell cytotoxicity and CD8+ T cell-mediated ferroptosis. In summary, the m6A modification regulates ferroptosis in cancer cells by modulating the expression of ferroptosis-associated mRNAs. m6A-modulated ferroptosis also participates in cancer immunity and immunotherapy. However, the underlying mechanisms remain largely unknown. Therefore, further research is required to develop novel cancer treatment strategies.

5.3 Pyroptosis

Pyroptosis is a gasdermin-mediated PCD process mainly activated by caspases. Pyroptotic cells are swollen and their plasma membrane ruptures. METTL3 suppresses pyroptosis in retinal pigment epithelium cells by targeting the miR-25-3p/PTEN/Akt signaling cascade.¹⁷⁰ YTHDF1 inhibits caspase-1-dependent pyroptosis by upregulating the WW domain-containing E3 ubiquitin protein ligase 1.¹⁷¹ LINC00969 promotes acquired gefitinib resistance by decreasing NLRP3 levels via m6A modification to inhibit pyroptosis in lung cancer.¹⁷² Additionally, bladder cancer, GC, BC, and melanoma are associated with pyroptosis-related lncRNAs and m6A modification.^173-176 However, the underlying mechanisms remain to be elucidated.

5.4 Cuproptosis

Cuproptosis is a newly identified form of PCD that is dependent on copper (Cu). This process is characterized by Cu-targeting and binding to lipoylated components within the tricarboxylic acid cycle, which ultimately induces proteotoxic stress and leads to cell death. Current research indicates that cuproptosis may be correlated with various cancer signaling pathways, including EGFR, PDK1, PI3K, MAPK, MYC, and Notch.¹⁷⁷ Sun et al.⁹⁶ found that tumor tissues had higher Cu and lactate contents than normal tissues in GC. In addition, they demonstrated that under high Cu stress, lactylation of METTL16 at K229 upregulated the m6A methylation levels of FDX1 mRNA and FDX1 protein expression, which triggered cuproptosis.⁹⁶ Nucleophosmin 1 (NPM1) is a biomarker for gastrointestinal cancer. Researchers found that NPM1 is associated with anticancer immunity, m6A modification, and cuproptosis. Cuproptosis-related genes can be used to predict the prognosis of various cancers, including lung adenocarcinoma,^{178, 179} BC,¹⁸⁰ and HCC^{181, 182} in an m6A-associated manner. However, little is known about the specific role of m6A and further studies are needed to uncover this mystery.

5.5 Disulfidptosis

In contrast to other types of PCDs, cells with high SLC7A11 expression can accumulate cellular disulfides such as cystine. This accumulation induces disulfide stress, which leads to an increasing number of disulfide bonds within the actin cytoskeleton, damaging the cytoskeletal structure, and consequently resulting in cell death.^183-185 Disulfidopathy is a novel approach for metabolic cancer therapy. Recent studies have suggested that metabolic therapy using glucose transporter inhibitors can facilitate disulfidptosis and dampen cancer development.¹⁸⁶ Disulfidopathy is a potential target for cancer treatment. However, current studies require bioinformatics analyses. Further research is required to explore this mechanism, and m6A modifications may provide an important perspective.

m6A modification plays a pivotal role in regulating the survival and death of cancer cells. PCD serves as a critical modulator of cancer immunity, influencing the function of immune cells and leading to diverse outcomes. Consequently, targeting m6A could be considered for cancer immunotherapy.

6.1 Macrophages

Macrophages, derived from the mononuclear phagocyte system, play essential roles in various biological and pathological processes.¹⁹² Macrophages are commonly divided into M1 and M2 types. M1 macrophages are usually considered to have antitumor functions, whereas M2 macrophages are thought to have protumor functions. Additionally, tumor-associated macrophages (TAMs) significantly influence various stages of cancer progression, including angiogenesis, tumorigenesis, metastasis, invasion, and hypoxia induction, by modulating the tumor tissues.^193-195 m6A RNA modification is a regulator of macrophage activation.¹⁹⁶ METTL3-deficient TAMs eventually lead to tumorigenesis by increasing the infiltration of regulatory T cells (Tregs) and reducing the number of Th1 and CD8+ cells through the NF-κB and STAT3 signaling pathways, resulting in the reduced therapeutic efficacy of anti-PD-1 therapy.¹⁹⁷ A previous study showed that silencing ALKBH5 significantly reduces the infiltration of M2 macrophages in gliomas, suggesting that ALKBH5 may be a potent predictor of sensitivity to immunotherapy in some cancers, especially gliomas.¹⁹⁸ Liu et al.¹⁹⁹ found that lncRNA–PACERR increases the number of M2 TAMs in pancreatic cancer (PC) cells in an IGF2BP2/m6A manner. IGF2BP3 upregulates and activates the STAT3 pathway to promote M2-TAM polarization and immunosuppression in gallbladder cancer.²⁰⁰ CircASPH interacts with IGF2BP2, stabilizes it to activate STING signaling, and promotes M2-TAM infiltration in CRC.²⁰¹ In hepatoma tissues, M2 macrophages exhibit high PD-L1 expression. ALKBH5 recruits PD-L1+ TAMs by regulating MAP3K8 expression and activating the ERK/JNK and IL-8 pathways to promote HCC progression.²⁰² RBM15, an oncogene in a number of tumors, enhances the stability of CXCL11 mRNA via m6A RNA modification to facilitate M2 macrophage polarization and infiltration, boosting the progression of ccRCC.²⁰³ LINC00657 activates the transforming growth factor-β (TGF-β) signaling pathway and induces M2-TAM infiltration by m6A modification in BC.²⁰⁴ In conclusion, m6A plays a crucial role in the activation, polarization, migration, and infiltration. Targeting m6A in TAMs may alter the suppressive immune microenvironment to promote anticancer immunity.

6.2 Neutrophils

Neutrophils, which account for the majority of granular leukocytes, play essential roles in tumorigenesis, progression, and invasion. Tumor-associated neutrophils (TANs) influence the activation and function of other immune cells.²⁰⁵ The m6A modification plays a crucial role in neutrophil activation. METTL3 enhances m6A methylation, thereby increasing the translation of toll-like receptor 4 (TLR4) mRNA, a key factor in neutrophil activation.²⁰⁶ The m6A modification also influences neutrophil infiltration and migration. In papillary thyroid cancer (PTC), METTL3 deficiency leads to increased IL-8 expression, which recruits TANs and promotes PTC progression through c-Rel and Rel A inactivation of the NF-κB pathway, highlighting the anticancer function of METTL3 in PTC.²⁰⁷ piRNA-17560 increases FTO levels in BC cells by enhancing its stability. Senescent neutrophils are abundant in therapy-treated tissues and can produce exosomal piRNA-17560, contributing to chemoresistance and epithelial–mesenchymal transition (EMT) in BC cells in an m6A-dependent manner.²⁰⁸ In addition, Ou et al.¹¹⁹ revealed a correlation between m6A and TANs in BC, identifying a novel subset of C5aR1-positive neutrophils implicated in promoting BC progression. Mechanistically, C5aR1-positive neutrophil-derived IL1β and TNFα can activate ERK1/2 signaling, phosphorylating and stabilizing WTAP to promote BC cells by facilitating glycolysis in BC cells.¹¹⁹ Solute carrier family 2 member 1 (SLC2A1) plays an essential role in cellular glycometabolism. A pancancer analysis identified SLC2A1 as a m6A-related potential biomarker for prognosis and immunotherapy. SLC2A1 positively correlates with neutrophils, providing a new strategy for cancer immunotherapy.²⁰⁹ Neutrophil extracellular traps (NETs) are extracellular fibrous structures produced by neutrophils that regulate NETosis.^210-213 NETs can impair autophagic flux, resulting in abnormal autophagy, which induces sepsis-associated acute lung injury via METTL3.²¹⁴ Researchers have found that METTL5 is strongly and positively associated with immune cell infiltration, including neutrophil infiltration, in HCC.²¹⁵ Recent studies have demonstrated that NETs are associated with m6A modifications in sepsis-associated acute lung injury. NETs mediate m6A modification through METTL3, subsequently inducing ferroptosis inacute lung injury.²¹⁶ NETs also participate in cancer invasion, evasion, angiogenesis, and metastasis by regulating the TME.^217-220 However, the association between m6A and NETs remains unclear. These studies indicate that m6A participates in TAN activation and infiltration of TANs. Understanding the link between m6A and NETs is a potential direction for future research.

6.3 Dendritic cells

DCs, known for dendritic pseudopodia, were first discovered by Steinman in 1973.²²¹ DCs are regarded as the most professional antigen-presenting cells (APCs); they play a key role in anticancer immunity and therapy.²²² For instance, DCs are necessary for T cell-mediated antitumor immunity by activating T cells and presenting tumor antigens.²²³ Recently, m6A modification-mediated DCs and cancer cells have been subjected to new trials. YTHDF1 can increase the expression of lysosomal cathepsins in DCs. Downregulation of cathepsins enhances the cross-presenting ability of wild-type DCs. Moreover, the efficacy of PD-L1 therapy is promoted in Ythdf1^−/− mice, indicating that m6A modification and YTHDF1 can modulate anticancer immunity in DCs.²²⁴ Similarly, in gastric tumors, the loss of YTHDF1 can recruit mature DCs, which subsequently promote sensitivity to antitumor effect.²²⁵ In HCC, Wang et al.²¹⁵ showed that METTL5 expression was positively correlated with the infiltration of immune cells, including DCs. Gong et al.²²⁶ demonstrated that the expression levels of METTL14, ZC3H13, and APC (an antagonist of the Wnt signaling pathway) positively correlated with DCs in BC. Downregulation of METTL14 and ZC3H13 correlated with a poor prognosis. Endothelin-converting enzyme 2 is a prognostic biomarker associated with m6A modifications in lung adenocarcinoma. Endothelial-converting enzyme 2 expression was significantly negatively correlated with DC infiltration.²²⁷ Glycolipid transfer protein expression is associated with m6A-related genes and DCs in cervical cancer (CC).²²⁸ YTHDC2 is correlated with immune infiltration levels of DCs in head and neck squamous cell carcinoma (HNSCC).²²⁹ Briefly, m6A modification influences DC activation and infiltration. More importantly, as the strongest APC, targeting the m6A modification of DCs may shed new light on DC-based immunotherapy.

6.4 T cells

T lymphocytes (T cells) play a major role in adaptive cellular immunity and participate in humoral immunity induced by thymus-dependent antigens.²³⁰ Functionally, T cells are divided into helper T cells (Ths), cytotoxic T lymphocytes (CTLs), Tregs, and exhausted T cells (Texs).^{230, 231} T cells are important for anticancer immunity. Recent studies have shown that the growth and activation of T cells are modulated by m6A RNA modification.^{232, 233} For instance, m6A-modified circIGF2BP3 can inhibit the CD8+ T cell response and induce cancer escape via PD-L1 deubiquitination in NSCLC.²³⁴ In BC, the upregulation of the m6A modification of PD-L1 mRNA via the METTL3/IGF2BP3 axis can downregulate T cell antitumor immune activation to inhibit tumor immune surveillance.²³⁵ Dong et al.²³⁶ demonstrated a negative relationship between m6A levels and dysfunctional CD8+ T cells in CRC. METTL3 methylates BHLHE41 mRNA and upregulates its expression, leading to increased CXCL1 expression, which, in turn, recruits immunosuppressive myeloid-derived suppressor cells (MDSCs) to dampen T cells. Additionally, silencing METTL3 in CRC sustains the activation and proliferation of both CD4+ and CD8+ T cells, thereby suppressing tumor progression.²³⁷ YTHDF1 in CRC also recruits MDSCs by activating the m6A–p65–CXCL1 axis to inhibit T-cells, subsequently promoting CRC. This implies that targeting FTHDF1 is a good strategy for boosting anti-PD1 therapy.²³⁸ Keratin 17 (KRT17) plays a protective role in CRC by promoting T cell infiltration in a YTHDF2 dependent manner.¹⁰² In HCC, CD8+ T cell dysfunction can lead to immune evasion. HCC cell-derived exosomal circCCAR1 is stabilized by WTAP and IGF2BP3, which can be taken up by CD8+ T-cells. It stabilizes PD-1 by promoting its deubiquitination. At the same time, it facilitates the dysfunction of CD8+ T cells, resulting in immunosuppression.²³⁹ Intriguingly, Li et al. found that methionine restriction reduces tumor growth and enhances antitumor immunity by increasing the number and cytotoxicity of tumor-infiltrating CD8+ T cells in a YTHDF1-dependent manner, suggesting that targeting methionine metabolism or YTHDF1 is a potential target for tumor immunotherapy.²⁴⁰ Zhang et al.²⁴¹ showed that YTHDF2 inhibits its downstream target RIG-I, thereby facilitating immune evasion in bladder carcinoma (BLCA). YTHDF2-deficient BLCA cells implanted in recipient mice activated innate immune responses and recruited CD8+ T cells.²⁴¹ HNRNPC interacts with m6A modifications during the immune processes. Cheng et al.²⁴² suggested that this gene could enhance the activation of Tregs, leading to immune escape and poor prognosis in prostate cancer. Additionally, some studies have revealed a crosstalk between m6A modification and T-cell exhaustion in anticancer immunity and immunotherapy. The inhibition of METTL3 or IGF2BP3 can enhance antitumor immunity through PD-L1-mediated T-cell exhaustion in BC as well.²³⁵ A previous study reported that lncRNA–AC026356.1, a downstream target of METTL14/IGF2BP2, is positively correlated with T cell exhaustion in lung adenocarcinoma.²⁴³ CCL8 andIL-1b can make hypoxia zones to recruit TAMs and cytotoxic T cells. The recruited immune cells can then be reprogrammed for immunosuppression in GBM.²⁴⁴ This study illustrates a new mechanism of hypoxia in tumors. Previous studies have also reported a correlation between hypoxia and m6A modification.^{187, 245} In conclusion, m6A participates in T cell-mediated antitumor immunity mostly by regulating T cell activation and PD-1/PD-L1. Therefore, Tregs and Texs are potential targets for future immunotherapy.

6.5 B cells

B lymphocytes (B cells), a type of lymphocyte differentiated from mammalian bone marrow lymphoid stem cells, participate in B-cell-mediated humoral immunity.^{246, 247} m6A is a key factor in the development, differentiation, and function of B cells. METTL3–METTL14 complex-mediated m6A modification is crucial for IL-7-induced pro-B cell proliferation via YTHDF2.²⁴⁸ Loss of METTL14 eventually represses the transition of pre-B cells from large to small.^{248, 249} Interestingly, another study showed that the loss of METTL3 in the pro-B stage slightly influences B cells in liver fibrosis.²⁵⁰ These results indicate that the link between m6A modification and B cell development may be associated with different stages. B-cell differentiation is important for antibody-mediated immunity and is determined by transcription factors. YTHDF2 promotes the formation of germinal centers by suppressing the plasmablast genetic program.²⁵¹ Several B cell lymphomas are associated with m6A modifications. piRNA-30473 upregulates WTAP expression. Increased WTAP stabilizes its target mRNA, hexokinase 2 (HK2), which promotes tumorigenesis in diffuse large B-cell lymphoma (DLBCL).²⁵² METTL3 is also functionally upregulated in DLBCL tissues.²⁵³ Furthermore, Meng et al.²⁵⁴ verified that NCBP1 enhances METTL3 by maintaining METTL3 mRNA stabilization and mediating c-MYC to promote DLBCL proliferation. Chen et al.²⁵⁵ found that KIAA1429/YTHDF2 suppression of carbohydrate sulfotransferase 11 (CHST11) inactivates the Hippo– yes associated protein (YAP) pathway via m6A RNA modification in DLBCL. Overall, m6A plays key roles in the development, differentiation, and function of B cells. In addition, m6A modification occurs in B cell lymphomas, especially DLBCL, thus providing an avenue to better understand and treat these malignancies.

6.6 NK cells

NK cells are cytotoxic lymphocytes derived from bone marrow lymphoid progenitor cells that play a vital role in nonspecific and specific immunity.^{256, 257} Unlike T and B cells, NK cells can kill pathogens or cancer cells without prior sensitization.²⁵⁸ NK cells have gained attention owing to their heterogeneous characteristics and functions in antitumor immunity and immunotherapy.²⁵⁹ Studies have reported alterations in NK cell levels in various tumors, including lung adenocarcinoma,²⁶⁰ HCC,^{261, 262} bladder cancer,²⁶³ GC,²⁶⁴ and HNSCC.²⁶⁵ IL-15 is an important factor in the proliferation, development, and function of NK cells. Interactions between IL-15 and m6A regulate the anticancer immunity of NK cells. A previous study reported that the expression level of YTHDF2 is regulated in NK cells. Upregulated YTHDF2 forms a STAT5–YTHDF2 loop that promotes the proliferation and antitumor immunity of NK cells.²⁶⁶ METTL3 enhances the NK cell response to IL-15, which is dependent on the activation of AKT–mTOR and MAPK–ERK. Src homology 2-containing protein tyrosine phosphatase 2 (SHP-2), encoded by Ptpn11, is an essential factor in IL-15-induced ERK activation. METTL3 deficiency reduces SHP-2 expression.^{267, 268} The relationship between m6A and NK cells is unclear, and further studies are needed to reveal the role of m6A in tumor-infiltrating NK cells.

TAM heterogeneity is closely associated with tumorigenesis and immune evasion. m6A modification plays an important role in the regulation of immune cell infiltration in the TMEs. Therefore, targeting m6A-regulated immune cells may be an attractive therapeutic strategy for restoring antitumor immunity.

7.1 Helicobacter pylori

Hp infection is one of the main causes of GC. Typically, Hp infection first induces nonatrophic gastritis, which can develop into atrophic gastritis, gastric polyps, and ultimately, GC.²⁷² Mechanistically, a recent review summarized that Hp may impair gastric epithelial cells through oxidative stress, DNA damage, impairment of DNA repair pathways, and endoplasmic reticulum stress.²⁷³ Recently, Li et al.²⁷⁴ performed a comprehensive analysis of differences in m6A modifications during Hp infection. They found an increasing level of m6A in Hp infection and a significantly different expression of m6A regulators, indicating that m6A modification might correlate with Hp infection.²⁷⁴ FTO promotes tumorigenesis in chronic CagA + Hp infection by regulating CD44 mRNA m6A methylations.²⁷⁵ Additionally, Hp infection has an undesirable effect on cancer immunotherapy by decreasing the efficacy of anti-PD1 immunotherapy.²⁷⁶ Some researchers have suggested that increasing the expression of PD-L1 could be an early response to Hp infection.²⁷⁷ The relationship between Hp infection and host cell DNA impairment has been studied to some extent. However, the relationship between Hp and m6A modifications in gastric carcinogenesis and disease progression remains largely unknown. Further studies are required to provide additional insights into Hp-related GC treatment.

7.2 Fusobacterium nucleatum

Fn is an opportunistic pathogen in human body.²⁷⁸ Emerging research has reported that Fn is associated with CRC.^{279, 280} Herein, from the perspective of m6A modification, we present illustrative research on Fn and CRC. Fn decreases METTL3 expression and m6A levels in KIF26B by activating YAP signaling and inhibiting FOXD3 (forkhead Box D3, a transcription factor for METTL3) expression to promote CRC progression.²⁸¹ Xu et al.²⁸² indicated that Fn infection-induced microRNA-4717-3p excessive maturation via METTL3-dependent m6A modification suppressed the expression of mitogen-activated protein kinase 4 and its anticancer function to promote CRC proliferation. Fn not only facilitates CRC, but also correlates with ESCC. Researchers have found that the upregulated expression of METTL3, which is induced by Fn, could promote ESCC proliferation and metastasis by promoting c-Myc mRNA methylation in a YTHDF1-dependent manner.²⁸³ Additionally, Fn-related m6A modifications are involved in CRC immunotherapy. Fn promote the expression of PD-L1 and mediate immune escape in CRC via m6A-modified IFIT1.²⁸⁴ In conclusion, Fn is involved in cancer progression and immunotherapy. Further studies are needed to explore strategies for treating cancers, particularly CRC.

7.3 Hepatitis B virus

HBV is a member of the Hepadnaviridae family of enveloped viruses, with a double-stranded DNA genome of 3200 bp in length.²⁸⁵ HBV infection is also associated with HCC. In the present study, we clarified the function of m6A modification during this process. First, HBV RNA is predominantly modified by m6A in the coding region of HBx.^{286, 287} Kim discovered an m6A site at nt 1616 of the HBV genome, indicating that m6A modification may regulate HBx protein expression to modulate the HBV life cycle.²⁸⁸ Yang et al. found that YTHDF2 stabilizes the transcripts of minichromosome maintenance protein 2 (PPM2) and minichromosome maintenance protein 5 (PPM5) via m6A modification, which promotes the tumorigenesis and progression of HBV-associated HCC.¹²³ HBV-pgRNA (pregenomic RNA) upregulated IGF2BP3 expression to promote HCC. Furthermore, interferon (IFN)-α−2a can increase pgRNA m6A modification and degrade its stability.²⁸⁹ Kim et al. demonstrated that HBV could increase the m6A modification of phosphatase and tensin homolog (PTEN) RNA, leading to decreased RNA stability and PTEN protein expression. PTEN downregulation can affect nonspecific immunity by inhibiting interferon regulatory factor 3 (IRF-3) nuclear import. Simultaneously, it activates the PI3K/AKT pathway, which influence HCC development.²⁹⁰ The m6A modification can also affect HBV RNA localization. YTHDC1 and FMRP promote nuclear export of HBV transcripts. The loss of YTHDC1 and FMRP can inhibit reverse transcription in HBV, affecting the HBV life cycle.²⁹¹ Additionally, the m6A modification modulates immune cell infiltration in HBV-HCC. In a recent study, patients were divided into two clusters: cluster A and B. Relatively, the overall survival rate of cluster A was higher than that of addition, and immune cell infiltration of the two clusters was significantly different.²⁹² Another study pointed out that ALKBH5 interacting with macrophages and WTAP interacting with NK T cells may be key factors in the progression of liver fibrosis during HBV infection.²⁹³ In summary, the m6A modification regulates HBx RNA and protein levels to modulate the HBV life cycle. HBV infection influences both oncogene expression and immune cell infiltration in an m6A-dependent manner to regulate HCC progression.

7.4 Epstein‒Barr virus

EBV is an oncogenic herpes virus linked to various cancers, including Burkitt's lymphoma, nasopharyngeal carcinoma (NPC), and EBV-associated GC.²⁹⁴ Recent studies have indicated that the m6A modification may be involved in this process. First, m6A modification mediates mRNA decay during EBV lytic reactivation to regulate its life cycle.²⁹⁵ The m6A modification can promote EBV lytic reactivation by attenuating IFN signaling.²⁹⁶ Additionally, WTAP deposits m6A marks on EBV transcripts and recruits YTHDF reader proteins to activate the CNOT1 RNA decay pathway.²⁹⁷ The tumorigenic function of EBV may be related to virus-encoded latent oncoproteins such as EBNA2.²⁹⁸ One study examined the role of EBNA3C, which binds METTL14 to promote tumorigenesis.²⁹⁹ In addition, EBER1 can activate the NF-κB signaling pathway, which downregulates WTPA expression to promote the migration of EBV-associated GC.³⁰⁰ Liu et al.³⁰¹ divided patients with NPC into two subgroups: an m6A high-score group and an m6A low-score group. They found that the m6A high-score group was related to immune suppression and poorer survival, while the m6A low-score group was related to a better response to immunotherapy; therefore, m6A modification is likely related to NPC progression and immunotherapy effects.³⁰¹ Zhang et al.³⁰² demonstrated that EBV-circRPMS1 promotes EBV-associated GC progression by recruiting Sam68 to METTL3 and upregulating METTL3 expression. Overall, m6A participates in the lytic reactivation of EBV and its life cycle. The m6A modification may regulate oncoproteins to influence EBV-associated cancers; however, it remains unclear whether this modification can be applied in EBV-associated cancer treatment.

7.5 Human papillomavirus

HPV is a small double-stranded DNA virus that infects the squamous epithelia and promotes tumorigenesis.³⁰³ Globally, 4.5% of cancers are caused by HPV, of which squamous epithelium-associated cancers account for a large proportion, including CC.^{303, 304} HPV is associated with several cancer processes, including initiation, progression, invasion, and metastasis, with unclear mechanisms.³⁰⁵ In the present study, we focused on m6A modifications. Sustained expression of HPV E6/E7 oncogenes alters cancer cell growth.³⁰⁶ The m6A modification can modulate E6/E7 expression to influence cancer cells. E6/E7 proteins regulate the m6A methylation levels of MYC mRNA in an IGF2BP2-dependent manner, which promotes several biological and pathological processes in CC, including aerobic glycolysis, proliferation, and metastasis.³⁰⁷ Wang et al.³⁰⁸ showed that E7 transcripts are stabilized by the m6A reader IGF2BP1. Interestingly, upon heat stress, the m6A-modified E7 reversed the fate of IGF2BP1. Based on this, they provided a treatment strategy for HPV-associated cancers that depend on heat.³⁰⁸ Huo et al.³⁰⁹ found that E7 increases ALKBH5 expression and enhances PAK5 expression by downregulating m6A modification levels, which promotes the tumorigenesis and metastasis of CC. Additionally, the m6A modification influences the antiviral treatment of HPV-associated cancers. IFN-ε is a key cytokine that helps the human body defend against viral infection, especially in epithelial cells. HPV can influence the m6A RNA modification of IFNE via WTAP to regulate IFN-ε, subsequently influencing the nonspecific immune responses to HPV in condyloma acuminata.³¹⁰ Importantly, METTL3 has been identified as a mediator of the immunosuppressive TME in HPV-associated cancers. In vivo cell-derived xenograft models showed that METTL3 inhibitors combined with anti-PD1 therapy enhanced the efficacy of immunotherapy in CC.³¹¹ Collectively, m6A modifications interact with HPV infections and influence immunotherapy.

Microorganisms regulate the expression of host tumor-related genes through m6A modifications. However, even after the microorganisms are eradicated, these epigenetic changes can persist and continue to drive tumorigenesis, which is called “the hit-and-run model.” Understanding this interaction provides crucial insights into the mechanisms underlying microorganism-related cancers and highlights potential targets for therapeutic intervention in m6A modification pathways.

8.1 RNA therapy

As a carrier of genetic information, RNA plays an essential role in diverse biological processes, as well as in disease processes such as cancer. Specific chemical modifications of RNA directly influence its molecular function.³¹⁵ m6A, the most abundant RNA modification, regulates both mRNA and ncRNA stability, translocation, and translation efficacy to modulate anticancer immunity.^{316, 317} Moreover, targeting m6A modifications of both mRNAs and ncRNAs may have potential value in cancer immunotherapy. Thus, we have clarified several recent advances and prospects.

8.2 Immune checkpoint blockade

ICB has been widely applied in diverse diseases, especially cancer, wherein ICIs block the interaction between receptors on the surfaces of immune and tumor cells, thereby inhibiting the dysfunction of immune cells and enhancing their anticancer immunity.³⁴¹ The use of ICIs has significantly prolonged the survival of many cancer patients. Current evidence suggests that m6A modifications closely influence immune checkpoint-blocking therapy by directly regulating immune checkpoint expression and modulating immune cell suppression. This reduces drug resistance and enhances treatment efficacy.

8.3 m6A modification and cytokine therapy

Cytokines are secreted by both immune and nonimmune cells. Cytokines play a key role in anticancer immunity.^{362, 363} Over the past 30 years, cytokines and cytokine receptors have gained increasing attention because of their vital roles in several physiological and pathological processes. Cytokine therapy is a novel cancer immunotherapy. Diverse cytokines have been considered immunotherapeutic targets, such as IL-1,³⁶⁴ IL-2,³⁶⁵ IL-6,³⁶⁶ IL-8,³⁶⁷ and IL-15.³⁶⁸ Here, we reviewed the role of m6A in cancer cytokine immunotherapy. The m6A modification modulates cytokine expression to recruit immune cells. For instance, METTL3 deficiency improves IL-8 production by PTC cells, which recruit TANs and promotes PTC progression.²⁰⁷ Targeting the upregulation of METTL3 and mRNA m6A modifications may reduce IL-8 secretion and decrease TAN infiltration, thereby enhancing the efficacy of immunotherapy in PTC. YTHDF1 promotes HCC progression by increasing IL-6 secretion and recruiting MDSCs. Reducing YTHDF1, for instance with lipid nanoparticle-encapsulated siRNA against YTHDF1 (LNP-siYTHDF1), can alleviate MDSC-induced CD8+ T cell exhaustion and improve anticancer immunity.³⁶⁹ YTHDF1 loss in GC increases IL-12 expression and DC recruitment, which restores sensitivity to anticancer immunity.²²⁵ Cytokines are multifunctional and important biomolecules that participate in anticancer immunity. Cytokine therapy alters the TME and improves interactions between immune and cancer cells. Based on previous studies, the m6A modification may provide a new strategy for cytokine therapy. m6A plays a vital role in cytokine secretion by recruiting tumor-associated immune cells to influence cancer progression. Future research should target cytokine-related mRNA m6A modifications in cancer and immune cells to change specific cytokine expression, thus enhancing the efficacy of immunotherapy. In addition, we suggest that constructing and delivering m6A-modified cytokine-related mRNAs into the TME to persistently increase or decrease cytokine expression may be helpful in cancer immunotherapy. However, further studies are needed to confirm this hypothesis.

8.4 ACT therapy

ACT therapy refers to the modification of immune cells to express a chimeric antigen receptor (CAR) for the treatment of different types of cancers, especially leukemia and lymphoma.^370-372 The CAR consists of an antigen-binding region, a transmembrane region, and a signal transmembrane region. Typically, CAR immune cells are derived from a patient's peripheral blood, engineered to express CARs in vitro, and reinjected into the patient after expansion. Therefore, CAR immune cells can specifically recognize and interact with cancer cells without antigen processing or presentation.³⁷² CAR-T, CAR-NK, and CAR-macrophage therapies have been studied, with CAR-T therapy being the most abundant.³⁷³ Recently, scientists reported that super CAR-T cells target multiple tumor-associated antigens and exhibit improved antitumor ability.³⁷⁴ However, challenges such as primary resistance, relapse, and adverse effects are unsolved.³⁷⁵ Although CAR-T therapy has been successful in treating leukemia and lymphoma, a number of solid tumors cannot obtain satisfactory results, which is probably related to a lack of tumor-specific targets, immunosuppressive TME, problems with homing and access to the tumor site, and lack of CAR-T cell expansion.³⁷⁶ Interestingly, non-m6A-related neoantigen-coding lncRNAs are regarded as vital factors in glioma progression and may provide an important direction for CAR-T therapy.³⁷⁷ The m6A modification is still worth considering for applications in CAR-T therapy. First, m6A modification regulates immunosuppressive TME.³⁷⁸ Targeting m6A may relieve the immunosuppressive TME and enhance the efficacy of CAR immune cell therapy. METTL3 plays a role in T cell dysfunction via MDSC recruitment.²³⁷ METTL14 is also associated with T-cell dysfunction.²³⁶ The expansion of CAR immune cells is essential. Targeting m6A modifications promotes immune cell proliferation. For example, YTHDF2 regulates NK cell function and proliferation.²⁶⁶ These results may aid future CAR-NK cell therapy. In addition, it is worth exploring whether targeting the m6A modification of immune cells derived from patients can promote CAR expression, which may promote CAR immune cell therapy efficacy.

8.5 Direct-targeted treatment of m6A regulators

m6A modifications play various roles in the occurrence and development of tumors, and therapies targeting m6A-related molecules are also diverse.^{379, 380} Multiple studies have shown that m6A modifications affect the sensitivity of chemotherapy drugs as a direct target and thus affect the treatment of cancer.^381-384 Therefore, an increasing number of therapies that directly target m6A regulators have been reported. The two most widely regulated proteins are METTL3 and FTO, which coordinate during the dynamic reversible m6A modification process. At present, a small-molecule inhibitor targeting METTL3 has entered clinical research, and the other four are in preclinical research, all of which are used to treat cancer or leukemia. The same is true for small-molecule inhibitors that target FTO.