SCC and its precursor lesions are complex systems in which neoplastic cells coexist with the other cell types and tissue components that comprise the tumor microenvironment (TME). The TME contains multiple different cell types, including cancer-associated fibroblasts (CAFs), neutrophils, macrophages, and regulatory T cells. Tumor cells and TME cell populations interact with each other via complex communication networks through the various secreted cytokines, chemokines, growth factors, and proteins of the extracellular matrix (ECM). The TME is known to be implicated in cancer cell survival, tumor progression, and the tumor response to therapy ^[1][50].

The extracellular matrix (ECM) is a non-cellular network of macromolecules (collagen, fibronectin, and laminin, etc.) that offers structural and biochemical support for cellular components, enabling it to influence cell communication, adhesion, and proliferation ^[2][51]. In cancer, the ECM is frequently deregulated and disorganized, which directly stimulates malignant cell transformation. Matrix metalloproteinases (MMPs) are critical molecules for the EMT process because they not only degrade cell adhesion molecules, favoring migration and metastasis, but also promote the initiation and proliferation of primary tumors ^[3][52]. MMPS are produced by tumor and stromal cells, such as fibroblast and inflammatory cells ^[2][3][4][48,51,52]. Alterations in the proteins of the ECM may impact the tumor progression in SCCs. Collagen is the major protein component of the ECM, which provides the cells with tensile strength and support for migration ^[2][51]. The loss of type IV collagen correlates with poorly differentiated OSCC ^[3][52] and CSCC ^[5][53]. Fibronectin is produced by fibroblasts and endothelial cells and mediates the cellular interaction with the ECM. In the development of cancer, increased levels of fibronectin have been associated with increased tumor progression, migration, and invasion, as well as an impaired response to treatment ^[6][7][54,55]. Additionally, the expression of the laminin receptor plays an important role in SCC progression ^[7][8][55,56], as reduced laminin expression has been correlated with an invasive phenotype of OSCC tumors ^[5][53].

Fibroblasts are one of the main cell components of the connective tissue subjacent to the epithelia. The main functions of these cells are the synthesis of the ECM (collagen, laminin, and fibronectin, including those needed to form basal membranes), the regulation of epithelial differentiation, and the promotion of wound closure ^[9][10][57,58]. CAFs are activated fibroblasts with mesenchymal characteristics associated with cancer cells, which contribute to tumor-promoting inflammation and fibrosis. CAFs acquire specific characteristics, such as a distinct morphology (an elongated spindle-like shape), and express differential markers (α-sma—Alpha-Smooth Muscle Actin; FAP-1—Fibroblast Activation Protein-1; vimentin and S100A4) and a lack of lineage markers for epithelial cells, endothelial cells, and hematopoietic cells ^[11][12][59,60]. However, the precise origins and roles of the fibroblast populations within the tumor microenvironment remain poorly understood.

In the case of HNSCC, several populations of CAFs have been described that, according to specific markers, can be classified into three subgroups: classical CAFs, normal activated fibroblasts, and elastic fibroblasts. Classical CAFs are enriched for genes, encoding proteins such as FAP, PDGF (platelet-derived growth factor receptor), lysyl oxidase, and MMPs. Normal activated fibroblasts show a low expression of CAF markers and elastic fibroblasts are enriched for tropoelastin, fibrillin 1, and microfibril-associated protein 4. It seems that the presence of different CAFs can be related to overall patient prognoses ^[13][61].

CAFs are key in different stages of the development of OSSC and CSCC, stimulating the growth and progression of tumors and participating in the maintenance of a state of poor differentiation of the surrounding cells. They act synergistically with epithelial cells to promote carcinogenesis and influence the patterns of invasiveness and metastasis ^[13][14][61,62]. In this sense, CAFs could act in the initiation process of cancer, favoring the mutagenicity of epithelial cells, for example, by secreting ROS, which favor decreases in the pH and hypoxia in the TEM ^[15][63]. In tumor progression, they promote the migration and invasion of tumor cells by chronically maintaining the proinflammatory stimuli via the promotion of oxidative stress. They also contribute to this end by secreting a broad amount of cytokines and chemokines, such as transforming growth factor beta (TGFβ) and interleukins (IL-1 and IL-6), and a broad range of growth factors, such as EGF (epidermal growth factor), bFGF (basic fibroblast growth factor), VEGF (vascular endothelial growth factor), HGF (hepatocyte growth factor), tumor necrosis factor (TNF), interferon-(IFN), CXCL12, IL-6, galectin-1, sonic hedgehog protein (SHH), and bone morphogenetic protein (BMP), among others, which are tumor-promoting. In particular, HGF has been described to promote glycolysis in HNSCC cells ^[15][63]. All these molecules can influence tumor cell growth, angiogenesis, and the recruitment of immunosuppressive immune cells ^[16][17][18][64,65,66]. CAFs are also crucial producers of MMPs, playing an important role in modulating the TME through the remodeling and degradation of the ECM, which ultimately results in the promotion of the invasive phenotype of cancer cells ^[18][19][20][66,67,68].

In addition, they promote the EMT by secreting a variety of soluble activators that initiate the TGFβ cascade in epithelial tumor cells, leading to a change in morphology and response, acquiring mesenchymal characteristics ^[21][22][69,70]. As a major secreted factor of CAFs, TGFβ predominantly mediates the crosstalk between CAFs and cancer cells. Several in vitro studies have demonstrated that, in OSCC and CSCC, the TGFβ secreted by CAFs induces EMT and resistance to different therapies ^{[23][24][25][26][27]}[71,72,73,74,75].

The immune cell component of the TME is formed by tumor-infiltrating lymphocytes (TILs, including CD4+ and CD8+ T cells, B cells, natural killer T cells, and myeloid lineage cells (macrophages, neutrophils, monocytes, eosinophils, myeloid-derived suppressor cells or MDSCs, and mast cells or MCs)). In general, OSCC and CSCC tumors are highly infiltrated by immune cells, although the extent and composition of the immune cell infiltrate vary according to the anatomical subsite and etiological agent ^[28][29][76,77].

Tumor-infiltrating lymphocytes (TILs) are the major cell type in adaptive immunity that recognize specific antigens and produce specific immune responses. High levels of TILs generally correspond to better outcomes in OSCC, but this is dependent on the balance of cells with anti-tumor activity (effector T or Teff cells) versus those with immunosuppressive activity (regulatory T or Treg cells) in the TIL population ^[28][30][76,78].Teffs are related to high levels of cytotoxic CD8+, which produces IFN-ϒ ^[28][76]. On the contrary, high levels of CD4+Foxp3+ or Treg cells through IL-4 and IL-10 production have been correlated with immunosuppression and pro-tumor activity, triggering poor outcomes ^[31][32][79,80]. Likewise, UV radiation also causes an increase in Treg and a decrease in Teff cells in the skin, leading to a change in the T-cell balance and promoting the development of CSCC ^[33][81]. On the other hand, the presence of B lymphocytes is related to higher levels of CD8+ cell infiltration and, therefore, to a better prognosis in OSCC and CSCC ^[34][35][82,83].

Tertiary lymphoid structures (TLS) are crucial elements of the tumor immune microenvironment, corresponding to sites of lymphoid neogenesis with the potential of orchestrating anti-tumor responses. They correspond to ectopic lymphoid organs, emerging in the context of chronic inflammation such as the TME and even allowing for germinal center formation ^[36][37][38][84,85,86]. In OSCC patients, a high density of TLS has been associated with a better overall survival and identified as an independent positive prognostic factor ^[39][40][87,88]. In the case of CSCC, though not much work has been performed, clinically, the presence of TLS has been prominently associated with a better degree of histopathological grades and a higher level of sun exposure. Furthermore, the presence of intratumoral TLS has been associated with lower lymphovascular invasion. Therefore, TLSs are considered to be a positive prognostic factor for CSCC and will provide a theoretical basis for the future diagnostic and therapeutic value in this type of cancer ^[41][89]. The features and clinical significance of TLSs in SCC still remain unknown.

Tumor-associated macrophages (TAMs) interact, modulate, and influence tumor progression, invasion, and metastasis. Macrophages display a great plasticity, oscillating between M1 (antitumoral) and M2 (protumoral) phenotypes. M1 macrophages produce pro-inflammatory cytokines (IL-12 and IL-23), tumor necrosis factor-α (TNF-α), and chemokines (CCL-5, CXCL9, CXCL10, and CXCL5), which promote adaptive immunity. They also express high levels of major histocompatibility complex 2 (MHC-2) molecules, allowing for the presentation of tumor antigens ^[42][43][90,91]. In contrast, M2 macrophages play an immunoregulatory role and are involved in tissue remodeling, angiogenesis, and tumor progression. M2 macrophages act by releasing anti-inflammatory cytokines (IL-4, IL-13, IL-10, and TGFβ, etc.), overexpressing PD-L1 (Programmed death-ligand 1) and expressing comparatively lower levels of MHC-2 molecules ^[42][43][44][90,91,92]. Several studies have suggested a correlation between the level of TAM infiltration and a poor outcome in OSCC, which could be used as a potential prognostic marker ^[45][46][93,94].

Myeloid-derived suppressor cells (MDCSs) comprise a heterogeneous population of cells that play a crucial role in the negative regulation of the immune response in cancer by inhibiting both adaptive and innate immunity, establishing the premetastatic niche in different types of cancer ^[47][48][95,96]. In addition, MDSCs have also been linked to angiogenesis and the degradation of the ECM ^[49][50][97,98]. A high abundance of circulating MDSCs correlates with advanced stages of OSCC and is also known to promote CSCC development ^[50][51][52][98,99,100].

Mast cells (MCs) represent another important myeloid component of the immune system. MCs in the TME may have pro-tumoral functions, such as the promotion of angiogenesis (through VEGF production), ECM degradation (via MMPs production), and the induction of tumor cell proliferation (through tryptase and histamine) ^[53][54][101,102]. In OSCC and CSCC, the protective and pro-tumoral role of MCs has been described in several studies ^[55][56][57][103,104,105].

OSCC and CSCC are generally treated with surgical resection and, depending on the disease state, this is accompanied by radiation or chemotherapy ^[58][59][60][6,12,20]. Traditional tumor treatment methods cannot solve the problems of tumor recurrence and metastasis, which are often associated with the TME, as mentioned in the previous section. Therefore, new single or combined strategies are being developed to address the TME, as described below.

The importance of CAFs in tumor development and their role in therapy resistance have been demonstrated in several types of cancer, including OSCC and CSCC ^[16][64]. CAF-mediated resistance to cetuximab has been reported in OSCC ^[61][62][106,107]. Additionally, in CSCC, the presence of CAFS has been found to increase resistance to photodynamic therapy ^[23][71], suggesting that therapeutic CAF targeting could increase the response rates for a diverse range of treatments.

One of the main mediators related to these resistance effects is TGFβ. This cytokine modulates the tumor progression and therapy response through the CAF activation status, shape, and invasiveness ^[63][108]. Quan et al. ^[64][109] suggested that TGF-β1 induces EMT to increase the capacity of OSCC for invasion, and Gallego et al. ^[23][71] described that an increased secretion of CAF-derived TFGβ mediates resistance in CSCC. However, targeting TGF-β is potentially problematic for its dual role: in the early stages of tumorigenesis it can act as a tumor suppressor, while acting as a tumor promoter in later stages ^[65][110]. Even so, it has been described that a novel TGFβ inhibitor promoted anti-tumor immune responses in OSCC, alone and in combination with anti PD-L1 antibodies ^[66][111]. Although the signaling cascades involving TGFβ are the primary signaling pathways regulating CAF activation, there are other growth factors and signaling molecules also implicated in the differentiation process, including NOX4 (NADPH oxidase 4), FGF, IL-6, or TNF ^[67][112]. Hanley et al. ^[68][113] identified NOX4 as a critical regulator of CAF activation in OSCC, and its inhibitor Setanaxib suppressed CAF activation. In OSCCs and CSCCs, there have not been many more studies targeting CAFS in order to prevent resistance. However, in other cancers, more trials have been described, such as the depletion of FAP-expressing cells as an adjuvant to immunotherapy ^[69][114]. Additionally, a combination of paclitaxel (which suppresses the expression of α-SMA) with gemcitabine improved the overall survival in pancreatic cancer patients ^[70][115], suggesting a possible new therapeutic window for OSCC and CSCC.

Immunotherapy has outstanding application value in the field of tumor therapy, including antibody-based therapy, cytokine therapy, and gene therapy.

One of the main strategies for eliminating SCCs is based on the use of monoclonal antibodies (mAbs), which include immune checkpoint inhibitors or anti-angiogenesis mAbs.

There are several clinical trials focused on mAbs for OSCC and CSCC treatment.

PD-1 is a checkpoint protein belonging to a group of T cell receptors involved in T cell suppression. PD-1 is also expressed by B cells, monocytes, and natural killer and dendritic cells ^[71][116]. This transmembrane protein binds to PD-L1, which is present on the surface of tumor cells, and this interaction triggers a signal that inhibits the activated T cells and induces immunological exhaustion and T cell apoptosis ^[71][72][116,117]. Then, the PD-L1/PD-1 axis is a primary mechanism of cancer immune evasion and has thus become the main target for the development new drugs that have emerged in recent years. Targeting the immune checkpoint proteins with mAbs has yielded a net clinical benefit in cancer ^[73][74][118,119]. So far, several mAbs have been approved for PD-1/PD-L1 blockade in clinical studies for oral cancer treatment, including Cemiplimab, Nivolumab, Sintilimab, Toripalimab, Pembrolizumab, Aezolizumab, Avelumab, Camreluzimab, and Durvalamab. Likewise, mAbs blocking other immune checkpoint receptors such as CTLA-4 (Tremelimumab) are being studied ^[75][120]. Finally, in OSCC, the effect of inhibiting OX40, a costimulatory molecule that can enhance T cell immunity, is also being tested. Anti-human OX40 was used in a phase I clinical trial (NCT02274155) prior to surgery. The results demonstrated that anti-OX40 mAb could induce the activation and proliferation of T cells in hosts, suggesting its successful potential as a clinical strategy ^[76][121].

On the other hand, as angiogenesis plays an important role in tumor development and metastasis, mAbs are also being tested to inhibit these processes in both OSCC and CSCC. Cetuximab, an anti-EGFR mAb, and bevacizumab, an anti-VEGFR mAb, are being administered in clinical trials, either alone or in combination with other treatments.

Macrophages and fibroblasts, among other cell types belonging to the TME, produce different cytokines that can be pro- or anti-tumorigenic ^[25][42][43][73,90,91]. Several cytokines clinical trials have been completed in OSCC, although the results have not been published yet. Some of these trials are related to the administration of IL-2 (NCT00899821 and NCT00019331) or INFα (NCT00276523, NCT00054561, NCT00002506, and NCT00014261), in order to see if they promote anti-tumor responses in combination with other treatments.