The HNSCC microenvironment exhibits highly heterogeneous and complex immune cell infiltration patterns (12) ( Figure 1 ). Key infiltrating immune populations include CD8+ T cells, CD4+ T cell subsets (Th1/Th2/Th17), natural killer (NK) cells, macrophages, and dendritic cells, collectively forming a dynamic immune network that governs tumor progression and therapeutic responses (13). CD8+ T cells, as central antitumor effectors, demonstrate prognostic significance linked to their infiltration density and spatial organization (14). However, tumor-derived immunosuppressive factors such as IL-10 and TGF-β drive CD8+ T cell dysfunction and exhaustion, facilitating immune escape (15). TAMs further contribute to this immunosuppressive milieu. While M1-polarized TAMs exhibit antitumor activity, M2-polarized TAMs dominate the HNSCC microenvironment, promoting tumor progression via ARG1-mediated L-arginine depletion (impairing TCR signaling) and IL-10/VEGF-driven immunosuppressive angiogenesis (16, 17).

Bidirectional interactions between tumor and immune cells critically shape HNSCC progression and therapeutic resistance. Programmed death-ligand 1 (PD-L1), overexpressed in ~50% of HNSCC cases, mediates immune evasion through dual mechanisms: membrane-bound PD-L1 engages PD-1 on T cells to induce exhaustion, while extracellular vesicle (EV)-encapsulated PD-L1 systemically suppresses T cell activity (18). Spatial proteomic analyses reveal PD-L1 enrichment at invasive fronts, particularly on cancer stem-like cells (CSCs), where PD-1/PD-L1 interactions impair immune synapse formation (19). Spatial proteomic analyses reveal PD-L1 enrichment at invasive fronts, particularly on CSCs, where PD-1/PD-L1 interactions impair immune synapse formation (20). Tumor-derived IL-6 and TGF-β synergistically reinforce immunosuppression: IL-6 activates STAT3 to upregulate B7-H3 expression, while TGF-β drives Treg differentiation and confers CD8+ T cells with stem-like exhausted epigenetic states (21). Myeloid-derived suppressor cells (MDSCs) exacerbate immune evasion through peroxynitrite-mediated antigen modification, impairing TCR recognition (22).

Immune infiltration patterns are modulated by tumor-intrinsic mutations, cytokine networks, and physicochemical stressors. High tumor mutational burden (TMB) inversely correlates with immune infiltration, suggesting genomic instability shapes immune evasion (23). NOTCH1 mutations disrupt CD8+ T cell/Treg balance, while PIK3CA-activating mutations recruit MDSCs via CXCL12/CXCR4 signaling (24). In addition, the hypoxic state in the tumor microenvironment also suppresses the function of immune cells, leading to immune escape (25). The study also found that tumor-associated fibroblasts (CAFs) play an important role in the HNSCC microenvironment (26–28). CAFs alter the infiltration pattern of immune cells and promote tumor growth and metastasis by secreting a variety of cytokines and growth factors (29). Hypoxia-driven HIF-1α activation reprograms Treg metabolism to enhance adenosine production via CD39/CD73 upregulation, while extracellular acidosis (pH 6.5–6.8) inhibits NFAT nuclear translocation, blunting T cell activation (30). CAFs emerge as central stromal orchestrators, secreting TGF-β superfamily ligands to polarize M2 macrophages and LOXL2-mediated collagen crosslinking to form T cell-excluding physical barriers (31).

In recent years, the breakthrough of single-cell spatial analysis technology has enabled the detailed analysis of the spatial structure of HNSCC immune microenvironment. Spatial transcriptomic studies have shown that HNSCC has a characteristic “Immune Desert” and “Immune Excluded” phenotype (32). The immune desert region showed substantial loss of effector T cells and dendritic cells, forming a “cold tumor” characteristic without immune monitoring. CD8+ T cells and TAMs were abundant in the immune rejection region, but these cells showed a state of functional inactivation, and their spatial distribution was highly coexisting with the remodeled ECM region (33). This spatial heterogeneity suggests that tumor cells establish hierarchical immune escape mechanisms through physical barrier construction and immunosuppressive signal diffusion.

Pseudotemporal analysis reveals spatial dynamics of T cell exhaustion, with CD8+ T cells transitioning from TCF1+ progenitor states at tumor margins to TIM-3+ terminally exhausted populations in cores—a process governed by TOX/OX40-driven epigenetic reprogramming (34, 35).

Current research paradigms are shifting from targeting oncogenic drivers toward remodeling the immunosuppressive niche. Deciphering the stromal barriers in immune deserts and metabolic suppression networks in excluded zones will enable spatiotemporally precise combination strategies (e.g., ECM degradation coupled with checkpoint blockade). This therapeutic evolution—from tumor eradication to immune ecosystem reconstruction—holds promise for overcoming the spatial limitations of current immunotherapies and achieving durable clinical responses.

The PD-1/PD-L1 signaling axis, a central hub of immune evasion in HNSCC, exhibits marked spatiotemporal heterogeneity in its expression dynamics ( Figure 2 ). Single-cell spatial analyses reveal that PD-L1 is specifically overexpressed in CD44+EpCAM+ cancer stem-like subpopulations at invasive tumor fronts, where it induces T cell inactivation through dual mechanisms: membrane-bound signaling and exosome-mediated delivery (36). This expression pattern correlates with HPV status (37), HPV+ tumors upregulate PD-L1 transcription via NF-κB/p65 signaling driven by E6/E7 oncoproteins, whereas HPV− tumors primarily rely on EGFR/MAPK pathways (15). PD-L1 is expressed in approximately 50% of HNSCC tumors, with higher prevalence in HPV+ cases (18). IFN-γ-induced PD-L1 regulation exhibits biphasic dynamics: acute stimulation rapidly upregulates expression via JAK/STAT signaling, while chronic exposure leads to persistent hypermethylation-associated expression (38). Despite the approval of PD-1/PD-L1 inhibitors for recurrent/metastatic HNSCC, clinical response rates remain modest (15–20%), attributed to spatial heterogeneity, the absence of tertiary lymphoid structures, CXCL13+CD8+ T cell exhaustion, and tumor mutational burden (39). These findings underscore the limitations of monotherapy and emphasize the need for combinatorial strategies targeting complementary immunosuppressive pathways.

CTLA-4, a critical early immune checkpoint, sculpts immunosuppressive niches via two mechanisms: (1) competitive B7 ligand binding in lymph nodes to inhibit naïve T cell priming, and (2) IDO+ dendritic cell-mediated Treg expansion within tumors (40). Preclinical evidence suggests that dual CTLA-4/PD-1 blockade enhances antitumor immunity. However, TIM-3 overexpression exacerbates PD-1-driven T cell exhaustion by suppressing IL-2 secretion in CD4+ T cells, while LAG-3 amplifies CTLA-4 suppression via MHC class II-dependent pathways (41). TIM-3 and LAG-3 further promote immune escape by impairing T cell function through distinct mechanisms (42). Despite these advances, clinical efficacy remains constrained by the redundancy of immune evasion mechanisms, necessitating rational combinations of checkpoint inhibitors and microenvironmental modulators.

Immune checkpoint inhibitors (ICIs), including anti-PD-1 antibodies such as pembrolizumab and nivolumab, are approved therapies for HNSCC, but anti-CTLA-4 antibodies are still under investigation and not yet considered standard of care (40, 43–47). Additionally, the metabolic reprogramming observed in HNSCC, including upregulation of glycolytic pathways and lactate production, contributes to an acidic microenvironment that suppresses T cell function and reduces the efficacy of ICIs (48). Macrophage reprogramming, such as depleting M2-polarized TAMs or blocking CSF-1/IL-10 signaling, reverses immunosuppression and synergizes with PD-1 blockade (49). Furthermore, epigenetic changes in tumor cells and immune cells can lead to a loss of PD-L1 expression or promote immune exhaustion, rendering T cells unresponsive to checkpoint blocked. Emerging therapeutic strategies targeting these resistance mechanisms aim to restore T cell function, such as by reprogramming TAMs or targeting key metabolic enzymes like lactate dehydrogenase (LDH), which has shown promise in enhancing the antitumor efficacy of ICIs. Moreover, combination therapies that incorporate epigenetic modulators to reverse immune evasion mechanisms are being actively explored as a way to overcome resistance to current therapies. These combined approaches hold potential for improving the overall success rates of immunotherapies in HNSCC and other solid tumors.

The PD-1/PD-L1 axis and synergistic checkpoints play pivotal roles in HNSCC immune evasion. Understanding their spatiotemporal regulation, compensatory networks, and microenvironmental interactions is key to developing next-generation immunotherapies. Rational combinations of checkpoint inhibitors with metabolic modulators, stromal disruptors, or epigenetic therapies hold promise for transforming immune “cold” HNSCC into responsive ecosystems, ultimately improving clinical outcomes.

HNSCC exhibits spatially heterogeneous metabolic reprogramming that supports tumor survival and immune evasion. The classical “Warburg effect” (aerobic glycolysis) not only fuels ATP production but also generates an acidic microenvironment via lactate accumulation, especially in hypoxia in multiple cancers (50–52), directly suppressing CD8+ T cell cytotoxicity (53). Single-cell metabolomics reveals compartmentalized metabolic in HNSCC: glucose-dependent metabolism dominates at invasive margins, while core regions favor glutamine metabolism, creating dynamic gradients of metabolites (54). Lipid metabolism also facilitates immune escape, tumor cells utilize FABP4-mediated lipid droplet storage for energy reserves and secrete lipocalin-2 (LCN2) to polarize macrophages toward an M2 immunosuppressive phenotype (55). These reprogrammed pathways reshape the TME, depleting nutrients and altering metabolite profiles to suppress immune cell activity (10).

Tumor-derived metabolites reinforce immune evasion through signaling and epigenetic remodeling. Lactate acts beyond metabolism—via GPR81 signaling, it induces dendritic cell tolerization and inhibits CD8⁺ T cell effector function by hyperacetylating the IFN-γ promoter through HDAC inhibition (38). Acidosis (pH 6.5–6.8) further impairs T cell proliferation and cytotoxicity (15). Additionally, lipid-derived mediators such as prostaglandin E2 (PGE2) promote M2 macrophage polarization and Treg expansion, consolidating the immunosuppressive niche (56).

Targeting tumor metabolism has emerged as a promising approach to enhance immunotherapy. Inhibiting glycolysis or lipid metabolism can restore T-cell function; for instance, targeting HK2 can reverse impaired glucose uptake in PD-1⁺ CD8⁺ T cells (53). Combination strategies are also under investigation: the adenosine receptor antagonist AB928, when paired with anti-PD-1 therapy, significantly improved responses in HPV-negative HNSCC by promoting TLS formation and increasing CXCR3⁺ T cell infiltration (57). Thus, metabolic interventions may overcome immune resistance and broaden treatment options. Continued research into how metabolic rewiring facilitates immune escape could yield novel therapeutic targets and optimize existing immunotherapies (58).

PD-1/PD-L1 inhibitors have made significant progress in the treatment of HNSCC. In recent years, anti-PD-1 monoclonal antibodies such as pembrolizumab and nivolumab have been approved by the FDA and EMA for the treatment of patients with recurrent or metastatic HNSCC. By blocking the interaction between PD-1 and its ligand PD-L1, these drugs restore the anti-tumor activity of T cells, thereby enhancing the immune system ‘s attack on tumor cells. However, their overall clinical efficacy remains limited—approximately 60% of patients do not respond, and only 20–30% achieve durable progression-free survival (47).

Emerging evidence suggests that this limited efficacy is largely attributable to TME factors, including immune exclusion, upregulation of immunosuppressive cytokines, and downregulation of human leukocyte antigen (HLA) expression (40). For example, increased infiltration of MDSCs and regulatory T cells has been associated with resistance to anti-PD-1 therapy. To address these challenges, combination therapies are being actively explored. Clinical trials have demonstrated that combining PD-1/PD-L1 inhibitors with chemotherapy or radiotherapy can synergistically enhance antigen release and immune priming. In KEYNOTE-048, for instance, pembrolizumab plus chemotherapy significantly improved overall survival compared to chemotherapy alone in PD-L1–positive HNSCC. Preclinical models also support combinations with anti-angiogenic agents, epigenetic modulators (e.g., DNMT or HDAC inhibitors), or metabolic modulators targeting lactate dehydrogenase (LDH) and adenosine pathways.

Future directions include rational selection of patients based on biomarkers (TMB, PD-L1 expression, and TLS formation) and spatiotemporal profiling of immune infiltration to guide individualized therapy.

CAR-T cell therapy is an emerging immunotherapeutic approach that has achieved significant success in hematological tumors, but its use in solid tumors such as HNSCC remains challenging. CAR-T cells are genetically engineered to express specific antigen receptors, thereby enhancing the recognition and killing ability of tumor cells. However, the tumor microenvironment of HNSCC is often highly immunosuppressive, which can limit the effectiveness of CAR-T cell therapy (59–61).

In HNSCC, CAFs secrete IL-6, TGF-β, and ECM-modifying proteins such as LOXL2, which restrict T cell trafficking and promote Treg polarization. Additionally, tumor antigen heterogeneity and antigen loss variants further compromise CAR-T targeting. CAFs showed inhibitory effects on CD8 + T cells, further limiting the antitumor activity of CAR-T cells (Qin, 9).

To overcome these obstacles, researchers are exploring multiple strategies, including optimizing the manufacturing process of CAR-T cells, combining other immunotherapies or targeted therapies, and developing novel CAR-T cell designs to improve their efficacy in solid tumors (62). These improvements are expected to overcome the limitations of existing treatments and enhance the clinical application of CAR-T therapy in HNSCC.

Beyond PD-1/PD-L1 inhibitors and CAR-T cell therapy, several alternative and adjunctive immune-therapeutic strategies are under rapid development in HNSCC. For example, interventions directed at the tumor microenvironment, such as by modulating the function of TAMs or CAFs, may effectively enhance antitumor immune responses (63, 64). In addition, cancer vaccines and immunomodulators are being developed to improve the immune response to tumors by activating the patient ‘s immune system (65). Combination regimens involving radiotherapy have shown synergistic effects, where ionizing radiation increases tumor antigen release and upregulates immune checkpoint molecules, thereby sensitizing tumors to PD-1 blockade (Chen, 66).

However, these strategies face several challenges: Lack of validated biomarkers to guide patient selection; Heterogeneity in immune phenotypes among HPV+ vs. HPV− tumors; Incomplete understanding of immune escape mechanisms in immune-desert tumors; Need for robust clinical data to confirm long-term efficacy and safety (67).

In the treatment of HNSCC, monotherapies often yield suboptimal outcomes, underscoring the need for rational combination strategies. Studies demonstrate that multimodal approaches can overcome immune evasion mechanisms in the TME, thereby enhancing therapeutic efficacy (47). Combining immune checkpoint inhibitors with chemotherapy, radiotherapy, or targeted therapies can amplify antitumor immune responses. Additionally, cancer stem cells significantly contribute to metastasis and immune evasion, making CSC-targeting approaches a critical component of combination regimens (19).

Specific combinations, such as PD-1/PD-L1 inhibitors co-administered with other immunomodulators (TGF-β blockers or CSF-1R inhibitors), show promise in reshaping the immunosuppressive TME. For instance, dual PD-1/CTLA-4 blockade has yielded improved response rates and survival outcomes in HNSCC patients (15).

Future clinical trials could explore that sequential or adaptive regimens that tailor dosing intervals and drug sequencing to dynamic TME changes, and combinations with epigenetic therapies or metabolic modulators to restore tumor immunogenicity and T cell function. This comprehensive approach may bolster the durability of immunotherapy responses and overcome primary or acquired resistance (68, 69). These strategies aim to comprehensively reshape the immunosuppressive TME, offering new avenues for improving immunotherapy outcomes.

Personalized therapy has become essential for optimizing HNSCC treatment outcomes. With the advent of omics technologies, clinicians can now profile tumor genetics, proteomics, and immunophenotypes to tailor individualized regimens. For instance, TMB and TME characteristics have been identified as predictive biomarkers for immunotherapy response (70). Integrating genomic profiling of tumor samples allows for precision immunotherapy selection, targeting specific mutations and expression patterns (71).

In addition to genetic and molecular data, clinical features such as HPV status and comorbidities also inform personalized treatment. PD-L1 expression levels can guide therapeutic decisions regarding PD-1/PD-L1 inhibitors (72) (18). Moving forward, prospective clinical trials should incorporate longitudinal biomarker analysis—such as monitoring circulating tumor DNA (ctDNA) or single-cell transcriptomic changes—to refine therapy choices in real time.

Biomarker development is central to early diagnosis, prognostication, and treatment monitoring in HNSCC. Recent high-throughput methods, including single-cell RNA sequencing and bioinformatics-driven integrative analyses, have revealed numerous biomarkers correlated with tumor progression and immune evasion (73). For instance, dysregulated long non-coding RNAs (lncRNAs) may predict immunotherapy responses and disease trajectory (74).

Future research should focus on multiparametric biomarker panels, integrating immune cell composition, cytokine profiles, and metabolic signatures to stratify patients more accurately. In addition to that, TAM and CAF phenotypes, as their polarization states strongly impact patient prognosis and therapy responsiveness (75, 76). By combining multi-biomarker analyses with functional TME characterization, we can drive the evolution of precision medicine. Large-scale clinical trials, employing adaptive designs and real-time biomarker assessment, will accelerate the translation of these findings into standard-of-care practices.