CD4⁺ T cells, as major organizers of cellular immunity, participate in all stages of the immune response. They differentiate into different Th subsets, including Th1 and Th2 cells, and present tumor antigen peptides on target cells (e.g. DCs and tumor cells) through the interaction between the T-cell receptor (TCR) and the major histocompatibility complex (MHC) class II molecules (15). It is well known that the surface markers of Th1 cells include interleukin (IL)-12 receptor β2 (β2R) (16), IL-18 receptor (IL-18R) (17), chemokine receptor (CXCR)3(18), CXCR5(19) and selectin ligand (20). Moreover, Th1 cells mainly mediate cellular immunity and function in tumor differentiation, immune regulation and development (21). The antitumor immune mechanisms are summarized as follows: i) Production of interferon-γ (IFN-γ) and IL-2 to activate CD8⁺ T cells and NK cells, as well as promotion of cellular immunity; and ii) activation of antigen-presenting cells (APCs) and induction of antibody production to further enhance tumor cell uptake of APCs (22). Evidence has shown that the differentiation and function of Th1 cells are closely related to those of other immune cells in the TME. For instance, DCs produce a large amount of IL-12, promoting CD4⁺ T cells to differentiate into Th1 cells and triggering a robust Th1 immune response (23). Similar to DCs, B cells and TAMs have been confirmed to enhance the differentiation of Th1 cells (24,25), whereas Tregs mediate immunosuppression by inhibiting the differentiation of Th1 cells (26).

TAMs differentiate under different conditions into two different subtypes, namely M1 and M2 macrophages. When macrophages are exposed to inflammatory cytokines produced by Th1 cells in the TME, such as tumor necrosis factor α (TNF-α) and IFN-γ, these differentiate into proinflammatory M1 macrophages (27) presenting several surface markers, such as CD80(28), CD86(29), vascular endothelial growth factor (VEGF) (30), suppressor of cytokine signaling 3 (SOCS3) (31) and CXCR7(32), useful to distinguish between M1 and M2 macrophages. Regarding their function in the TME, proinflammatory M1 macrophages stimulate the activation of mature T cells and amplify the Th1 response, thus exerting efficient antitumor activity (33). In addition, M1 macrophages are regulated by immune cells. Th1 cells and CD8⁺ T cells can induce M1 polarization of macrophages by secreting IFN-γ (34). However, due to the lack of in-depth research on M1 macrophages, the mechanism by which immune cells regulate M1 macrophages in the TME remains unclear.

After tumor invasion, immature CD8⁺ T cells differentiate into effector CD8⁺ T cells under the influence of antigen peptide-MHC, cytokines and costimulatory signals produced by APCs (35), as well as the secretion, metabolism, epigenetic modification and transcription factors of extracellular cytokines (36). Furthermore, effector CD8⁺ T cells differentiate into cytotoxic and memory CD8⁺ T cells (37). The specific surface markers of CD8⁺ T cells include CD3⁺ and CD8⁺. Notably, CTLs in the TME can produce IL-2, IL-12 and IFN-γ, demonstrating the cytotoxic function of CD8⁺ T cells and mediating antitumor immunity by producing TNF-related apoptosis-inducing ligands, reactive oxygen species (ROS) and perforin (38). In return, CD8⁺ T cells can also be modulated by various immune cells in the TME. DCs can cross-present exogenous tumor-associated antigens to MHC-I molecules to activate CD8⁺ T cells, and participate in the regulation of CD8⁺ T-cell immunity and tumor antigen tolerance (39). In addition to the stimulatory signals derived from DCs, CD4⁺ T cells are also a necessary condition for the initiation of CTLs. CD4⁺ T cells directly promote the activation of CD8⁺ T cells and develop CD8⁺ T cells to memory CD8⁺ T cells by the interaction with CD40-CD40 ligand (40) and costimulatory molecule combinations (41).

DCs, known as the most powerful professional APC, activate primitive antigen-specific CD4⁺ and CD8⁺ T cells by ingesting, processing and presenting antigens, thus initiating adaptive immune responses. DCs originate from macrophages/DC progenitor cells in the bone marrow and then differentiate into DC subsets, including classical DCs (cDCs), plasma cell-like DCs (pDCs) and monocyte-derived inflammatory DCs (MoDCs) (42). cDCs can be further divided into cDC1s and cDC2s. The development of cDC1s is dependent on IFN regulatory factor 8 and basic leucine zipper transcription factor ATF-like 3, and they specifically present internalized exogenous antigens to MHC-I to activate CD8⁺ T cells (43). cDC2s depend on IFN regulatory factor 4 development, presenting internalized antigens on MHC-II to activate CD4⁺ T cells (44). pDCs are a multifunctional population that produces large amounts of type I IFN (45).

There are also differences in DC surface markers among the different subsets: cDC1s are characterized by TLR7, TLR9 and RIG-I-like receptors; cDC2s are characterized by TLR1, TLR3 and TLR6; pDCs are characterized by CD11c, human leukocyte DR antigen (HLA-DR), CD304 and CD303A; whereas MoDCs are characterized by CD11c⁺, MHC-II⁺ and CD11b⁺ (45-47). Studies have confirmed that DCs strengthen immune function in the TME through a series of mechanisms: i) Receptors identify DCs to activate T cells and regulate CD4⁺ T-cell differentiation (48); ii) DCs induce the transformation of effector T cells into memory T cells (49); and iii) DCs activate CD8⁺ T cells, Tregs and NK cells by secreting an array of cytokines and metabolism-related enzymes, such as IL-2, IL-12(50) and indoleamine-2 dioxygenase (IDO1) (51). However, since tumors of different tissues and stages have unique features, DC function will also change in different TMEs, which may suppress tumor-specific immunity under certain conditions (52).

Although DCs promote the antitumor immune effect of the TME, DCs in the TME are often functionally impaired or defective, and various cytokines and cells can inhibit their antitumor effects in the TME As follows: i) IL-6, IL-10, VEGF and transforming growth factor-β (TGF-β) can negatively regulate the function of DCs and induce a tolerant phenotype by inhibiting their maturation and migration (53-57); and ii) Tregs exhibit inhibitory effects on DCs, either by downregulating the expression of the costimulatory molecules CD80, CD86 and CTL-associated protein 4 (CTLA-4) on DCs (58), or by releasing cytokines, such as IL-10 and TGF-β (59).

NK cells are innate immune effector cells. Currently, two different NK cell subsets have been identified: i) CD56^bright NK; and ii) CD56^dim NK cells. Under the action of IL-12, IL-15 and IL-18, they can secrete cytokines and chemokines, and participate in the regulation of acquired immunity (60). Regarding surface markers, NK cells lack the phenotypic markers of B (CD19^-) and T (CD3^-) lymphocytes, but they can be defined by their unique CD56⁺ state (61). Notably, unlike T cells that recognize target cells in an MHC-restricted manner through TCRs, NK cells recognize target cells through the activation and inhibition signal combinations of cell surface receptors, thus playing an antitumor immune role (62). Briefly, the NK cells operate through the following mechanisms: i) NK-activated receptors, as well as the natural cytotoxic receptors NKp30, NKp44 and NKp46, mediate the release of intracellular perforin and granzyme from NK cells after activation, which directly kill tumor cells (63); ii) NK cells regulate the dynamic balance of the immune system by secreting cytokines and chemokines such as IFN-γ to promote the activation and effector function of CTLs and restrain Treg functions by increasing their activity (63,64); and iii) NK cells have memory, self-renewal, long-term proliferation and persistence abilities in vivo and memory NK cells show stronger tumor-specific cytotoxicity, compared with naive NK cells (65). It is worth noting that, similar to DCs, NK cells may play different tumor immune roles in different situations (66). Compared with NK cells in healthy non-tumor tissues, tumor-infiltrating NK cells exhibit weaker cytotoxic activity, accompanied by the downregulation of activated receptors and upregulation of inhibitory receptors. These observations suggest that the function of NK cells is impaired by the following immune cells in the TME (67): i) Tregs and MDSCs, which can secrete TGF-β to directly undermine the killing ability of NK cells, or indirectly inhibit the killing effect mediated by NK cells by reducing the expression of the activated receptors NKG2D and NKp30 through membrane binding to TGF-β (68-70); and ii) similar to Tregs and MDSCs, tumor-associated fibroblasts and tumor cells can secrete IDO, prostaglandin E2 and TGF-β to reduce the expression of NKG2D in NK cells to restrain the cytotoxicity of NK cells (71,72).

CD4⁺ Th2 cells are a type of CD4⁺ T cell that induce B cells to differentiate into antibody-secreting cells to participate in the humoral immune response, secreting a variety of cytokines, such as IL-4, IL-5, IL-9, IL-10, IL-13, IL-25 and dimodulin, to participate in the antitumor immune response in vivo. Different from Th1 cell surface markers, Th2 cell surface markers include CXCR4(73), C-C motif chemokine receptor (CCR)3(74), CCR8(75) and chemokine receptor homologous molecule 2(76). The role of Th2 cells in the TME is also opposite to that of Th1 cells. The number of Th2 cells infiltrated by the tumor stroma is significantly higher than that of Th1 cells, resulting in Th1/Th2 drift. The resulting immunosuppressive state seriously affects antitumor immunity, eventually leading to the occurrence and development of tumors (77). In detail, the secretion of the cytokines IL-4, IL-5 and IL-25, as well as the transcription factor GATA-3 by Th2 cells contribute to the increase in cytokine cascades, therefore inhibiting the proliferation of Th1 cells, DCs and eosinophils, as well as regulating the inflammatory response of mast cells and lymphocytes (78-80). However, it has been speculated that consistent with Th1 cells, immune cells and factors in the TME can affect the differentiation and function of Th2 cells based on the results of the current research, although the specific regulatory mechanism remains unclear (81).

Tregs, also known as suppressor T cells, are a subgroup of CD4⁺ T cells, characterized by the expression of CD4, CD25 and forked/winged helix transcription factors (Foxp3) (82). Human Tregs can be classified according to the expression levels of Foxp3 and CD45RA: i) FoxP3^loCD45RA⁺CD25^lo-immature Tregs; ii) FoxP3^hiCD45RA^-CD25^hi-effect Tregs; and iii) FoxP3^loCD45RA^-CD25^lo-non-Tregs. Tregs mainly express a variety of activated cell surface markers, including Foxp3(83), CD127(84), CTLA-4(85), glucocorticoid-induced tumor necrosis factor receptor (86) and lymphocyte-activation gene 3 (LAG-3) (87).

Extensive research has been performed on Tregs, demonstrating the protumor effect of Tregs on the TME (88,89). The effects of Tregs are summarized as follows: i) Significant expression of CTLA-4 and LAG-3 on Tregs inhibits the proliferation of T cells and the release of cytokines, thus suppressing the maturation of T cells (90,91); ii) Treg-derived perforin and granzyme inhibit CTLs in the TME and prevent the death of tumor cells (92); iii) secretion of IL-10, IL-35 and TGF-β restrains the proliferation and optimal activation of T cells (93-95); and iv) Tregs directly inhibit the proliferation of B cells and increase cell death through cell-cell contact (96).

Tregs, as the main cells mediating immunosuppression in the tumor microenvironment, are also regulated by other immune cells; i) DCs promote the development of Tregs by presenting polypeptide-MHC to TCRs, and secreting IL-2 and IDO1, which play important immunosuppressive functions (50,51); and ii) MDSCs can transform antigen-specific CD4⁺CD25^- cells into CD4⁺CD25^-Foxp3 Tregs and mediate immunosuppression in the presence of IFN-γ and IL-10(97).

As mentioned previously, M2 macrophages are one of the subtypes of TAMs. Macrophages exposed to Th2 mediators (IL-4, IL-13, IL-10 and TGF-β) in the TME have an immunosuppressive phenotype, namely M2 macrophages (27). The M2 phenotype is triggered by a series of different transcription factor cascades, including interferon regulatory factor (IRF)/STAT family members (IRF4, STAT3 and STAT6), inhibitory NF-κB homodimer and hypoxia inducible factor 2, the surface markers of which are composed of CD163, IL-10, SOCS1/2, CD206, CCL-18, PDGF-BB and MMP (98,99). According to the current understanding, M2 macrophages can regulate the TME and mediate immune escape through the following mechanisms: i) By secreting factors, such as IL-10, IL-12 and CCL18(100), promoting the expression of STAT3, and inducing the release of Th2 cytokines, thus regulating immunosuppressive activity (101,102); and ii) by producing metabolic enzymes, such as human arginase 1 and nitric oxide synthase 2, undermining the T-cell immune response (103,104).

In recent years, it has become clear that TAMs differentiate and change under the regulation of the TME, such as through the secretion of tumor-related molecules by tumor cells, metabolic changes in the TME and the presence of other immune cells. The TME promotes the polarization of M2 macrophages via: i) IL-4 secreted from Th2 cells, IL-10 secreted from Tregs and immunoglobulin secreted from B cells; and ii) the regulation of Tregs and MDSCs (105,106). Tregs inhibit the secretion of IFN-γ by CD8⁺ T cells, thus preventing the metabolism of M2 macrophage fatty acids, and indirectly but selectively maintain the metabolic adaptability and survival rate of M2 macrophages (107), whereas MDSCs stimulate M2 macrophage differentiation and promote tumor proliferation by downregulating STAT3(108).

MDSCs are a specialized population of immature heterogeneous cells, consisting of myeloid cell precursors, immature granulocytes, monocytes and DCs. MDSCs can be divided into two types: Polymorphonuclear cells and monocytes, which have relatively common surface markers and the same immunosuppressive function. Generally, they aggregate in the TME to secrete a variety of cytokines and mediate immunosuppression (109). Human MDSCs express the myeloid common surface markers CD11b and CD33, but do not express the MHC class II molecule HLA-DR or mature myeloid or lymphocyte cell surface markers (110). However, MDSCs exhibit different phenotypes and functions in different patients with cancer (111). In the TME, MDSCs suppress the immune microenvironment by regulating the release of cytokines and inhibiting immune responses through several mechanisms: i) The MDSC surface receptor TGF-β mediates the downregulation of IFN-γ and the NK-cell receptor NKG2D, to reduce the cytotoxicity of NK cells, thus restraining the function of NK cells (112,113); ii) MDSCs produce IL-10 to promote the polarization of macrophages to the M2 phenotype and can differentiate into TAMs (114); iii) metabolic factors, such as a decrease in arginine (115) and cysteine levels (116), and increased NO (117), ROS and peroxynitrite (118,119), which are derived from MDSCs, have a negative impact on immune activity inside the TME, thus regulating T-cell function (120); iv) MDSCs can also prevent T cells from homing to the lymph, thus affecting their antigen stimulation and inhibiting activation, resulting in tumor-specific CD4⁺/8⁺ T cells that are unable to respond effectively to tumor antigens (121); and v) can induce the production of Tregs by transforming antigen-specific CD4⁺CD25^- cells into CD4⁺CD25^-Foxp3 Tregs and mediating immunosuppression in the presence of IFN-γ and IL-10(97). Furthermore, MDSCs can be modulated by other immune cells in the TME. It has been found that mast cells can enhance the immunosuppressive function of MDSCs by secreting histamine (122), but the specific regulatory mechanisms of other immune cells remain unclear and further research is needed.

In summary, TILs can exert an immune response in several ways, which can not only promote antitumor immune effects but also enhance immunosuppression to aid tumor cell immune escape. In retrospect, the research on TILs in solid tumors, such as melanoma, colorectal cancer and breast cancer, has been more extensive, but there are few related studies on gynecological malignant tumors, such as ovarian cancer, endometrial cancer and cervical cancer. Moreover, immunotherapy for gynecological malignant tumors is facing new dilemmas regarding personal specificity, effectiveness and, adverse effects. Therefore, the present review focused on the association between TILs and the clinical prognosis of gynecological tumors, as well as the mechanism of the immune response. Based on the aforementioned summary, immunotherapy will hopefully be developed further in the future to improve the survival prognosis of patients.

TIL-ACT takes advantage of the natural ability of TILs to recognize and eliminate tumor cells. The main steps include isolation, induction of differentiation, modification, amplification and infusion back into the body (154). Rather than simply amplifying T cells, it identifies T cells that effectively target specific mutations in different patients, and then screens and amplifies them. This strategy not only activates antitumor immunity in the host but also induces a more targeted and specific immune response.

To improve efficacy, in previous studies, patients have received lymphodepleting chemotherapy prior to infusion, which is designed to consume endogenous T cells and Tregs, and to provide the optimal environment for injected TILs over competing cell populations. This type of steady-state expansion ensures that TILs have persistent stability (155,156). In addition, to further activate TILs, patients are given a high dose of IL-2 intravenously until maximum tolerance is achieved (157).

The presence of TILs in tumor tissues has been associated with clinical outcomes in several tumor types, particularly in melanoma, in which TIL-ACT was revealed to be highly successful in phase I/II clinical trials (158). In addition, different degrees of studies have been performed in nonmelanoma types of cancer, including cervical (159), lung (160), ovarian (161), triple-negative breast (162) and gastrointestinal cancer (163). In all of these studies, TILs have been shown to affect prognosis (155) and studies have also proposed to use TILs as potential prognostic markers (164). Nevertheless, due to the presence of a high mutation load and high neoantigen rates, the efficacy of TIL-ACT therapy is highly variable (165). Therefore, further research and reliable clinical trials on TIL-ACT are warranted.

As early as 1991, Aoki et al (166) used TIL-ACT to treat seven patients with advanced or recurrent epithelial ovarian cancer and found a high response rate. Notably, it was considered that TIL-ACT combined with cisplatin holds promise for improving cure rates and long-term survival. Another clinical study in 1995 verified this conclusion, demonstrating that TIL-ACT may be a promising method for achieving a complete cure for advanced epithelial ovarian cancer (167). A 2015 review on TILs in ovarian cancer concluded that combining other therapies, such as immune checkpoint inhibitors, can better improve the clinical efficacy of TIL-ACT (168). In 2017, a large trial evaluating TILs in ovarian epithelial neoplasms demonstrated that higher levels of intratumor TILs were associated with improved prognosis and may serve as a clinically useful immunological prognostic indicator (161).

In a study of cervical cancer, researchers administered TIL-ACT to patients with HPV E6 and E7 reactivity, and found permanent regression of metastatic cervical cancer (154). More importantly, in a phase II clinical trial, 27 patients with advanced cervical cancer were treated with a novel TIL-ACT, LN145, and achieved 85% disease control and 44% objective response rates, without any serious side effects. The efficacy of LN145 in the treatment of advanced cervical cancer is promising compared with the response rate of chemotherapy or immunotherapy, which ranges between 4 and 14% for second-line treatment. LN145 has been approved by the FDA as a breakthrough therapy for advanced cervical cancer (169). As for ovarian cancer, a clinical trial demonstrated promising clinical benefit in patients treated with a maintenance TIL-ACT therapy after primary cytoreduction and platinum-based chemotherapy, showing significantly improved 3-year OS and 3-year disease-free interval (170). In addition, a TIL-ACT clinical trial is currently being performed on patients with endometrial cancer (NCT01174121), in which patients receive TILs and IL-2 after lymphocyte-depleting chemotherapy with cyclophosphamide and fludarabine; however, there is no published research data yet. Completed trials on ovarian, cervical and endometrial cancer are summarized in Table II. Regarding the selection of treatment strategies, some scholars tend to use combination therapy, e.g., combining radiation therapy with TIL-ACT (154). The sensitivity of immune effectors after tumor cells have received a lethal dose of radiation has been shown to be enhanced. In this case, the combination of TIL-ACT may result in the promotion of antitumor effects. However, this combination therapy still needs further study to balance the immune response in the host.

Overall, the aim of TIL-ACT is to create an individualized treatment that targets only certain tumors in one patient. Over the past few decades, TIL-ACT has made great progress on a technical level as an anticancer therapy. There is also an increasing number of studies focusing on using surface antigens, such as CD137(171) and PD-1(172), to screen TILs with high tumor responsiveness. Among them, PD-1 monoclonal antibody blocks the binding between PD-L1 expressed by tumor cells and PD-1 on the surface of T cells, and promotes a large-scale release of TILs in the TME, contributing to tumor regression (173). Although immunotherapy for gynecological malignancies is receiving significant attention, only a few studies and clinical trials have focused on TIL-ACT; therefore, further studies need to be conducted.

TIL-ACT is becoming popular because of its high specificity and efficiency in killing tumor cells and minimal side effects. However, the cytotoxicity associated with TIL-ACT is the major hurdle in its therapeutic applications (174). Adverse events (AEs) can be classified into two types: Mild and severe. Mild AEs may be due to contamination during the expansion and preparation of TILs in vitro, resulting in subsequent infusion allergy and infection (175). Patients have been reported to exhibit mild fever, chills and other mild inflammatory reactions. Severe AEs include two subcategories: Autoimmune toxicity and cytokine release syndrome (CRS) (176).

Since some tumor-associated antigens are also expressed in normal tissues, once TILs recognize them, an enhanced immune response from the host is triggered, resulting in graft vs. host disease. Adoptive T cells have been shown to target not only antigen-specific tumor cells but also normal skin cells and uveal cells, leading to vitiligo and uveitis (177). It is important to note that with the development of T-cell sorting and modification techniques, autoimmune toxicity-related AEs have become very rare (154).

The extensive nonantigen-related inflammation resulting from TIL infusion is known as CRS or cytokine storm, which is a severe overreaction of the immune system induced by a positive feedback loop between cytokines and immune cells (178). When CRS occurs, T cells in patients are activated and proliferate rapidly, causing an excessive cascading release of cytokines. These cytokines mediate a variety of immune responses, leading to clinical symptoms such as fever, hypotension, heart problems, dyspnea, fatigue, nausea and clotting disorders. In severe cases, they may introduce great damage to body tissues and organs, and even death (179). CRS is not specific to TIL-ACT and may be induced by the use of certain monoclonal antibody drugs and other types of ACT cell therapies, such as chimeric antigen receptor T-cell immunotherapy (180). In addition to the two types of severe AEs described above, Rohaan et al (154) classified whole-blood cytopenia and febrile neutropenia caused by lymphatic depletion as toxicity due to the lymphodepletion preparative regimen.

Despite the risk of AEs, the combination of lymphoid depletion before TIL infusion (181) and the high dose of IL-2 after infusion makes the current TIL-ACT quite safe and serious adverse reactions have rarely been reported.