Tumor-infiltrating lymphocytes (TIL) play a crucial role in TME, which is associated with cancer progression, response to therapy, and clinical outcomes [
98]. In addition, studies of TIL mainly focus on T cells. T cell infiltration formed in human cancer is a regulator of natural disease progression and also determines the probability of clinical response to cancer immunotherapy, which may provide potential prognostic value [
99]. There are three main types of T cells: helper T cells (T
H cells/CD4
+ T cells), cytotoxic T cells (T
C cells/CD8
+ T cells), as well as regulatory T cells (T
reg cells). CD4
+ T cells generally express CD3
+CD4
+CD8
−, while CD8
+ T cells generally express CD3
+ CD8
+CD4
− [
53]. CD4
+CD25
+Foxp3
+ are used to describe T
reg cells [
51,
52]. In addition, different subsets of CD4
+ and CD8
+ T cells have their specific markers, such as Th1 cells (CXCR3), Th2 cells (CCR4), Th17 cells (CCR6), Th22 cells (CCR10), Tc1 cells (CRCX3 and IRF4), Tc2 cells (CCR4, CRTH2, and GATA3), Tc9 cells (CRCX3, IRF4, IL9, and IL10), Tc17 cells (CCR6, IL23R, IRF4, and IL17) [
53,
54]. CD8
+ T cells encounter dysfunction and exhaustion due to immunosuppression within the TME during tumor development and progression [
100]. CD4
+ T cells play a key role in the adapted immune system, which can variously target tumors either directly by eliminating tumor cells through cytolytic mechanisms or indirectly by modulating TME [
101‐
103]. Hiraoka et al. have indicated that deeper infiltration by both CD8
+ and CD4
+ T cells presents a better prognosis for patients with NSCLC [
104]. Another study has mapped the heterogeneity of TILs in NSCLC, which may attribute to cancer immunotherapy [
105]. Yang et al. have shown the association between CD8
+ and CD4
+ T cell-related genes and colon cancer prognosis [
106]. In addition, high infiltration of lymphocytes has been observed in one subpopulation characterized by low peroxisome and high TIM3 of colorectal cancer [
107].
SPOTlight applied to PDAC samples annotated 12 T cells and predicted the proportion within each capture spot. Recently activated CD4
+, pre-exhausted CD8
+, and proliferative CD8
+ T cells significantly increased in tumor regions, while most transitional memory CD4
+ T cells were in normal tissue. Intriguingly, recently activated CD4
+ T cells co-localized with pre-exhausted CD8
+ T cells in tumor areas and could not be detected through their presence alone, indicating a possible target for precise pathology assessments [
87]. In breast tumor samples, 18 T-cell and innate lymphoid clusters were identified. One subset of exhausted CD8
+ T cells named
LAG3/c8 in triple-negative breast cancer (TNBC) had higher expression of PD-1, LAG3, and the ligand-receptor pair of CD27 and CD70, known to enhance T cell cytotoxicity [
49,
108]. In human squamous cell carcinoma, CD8
+ T cells were observed to co-localize with T
reg cells in the compartmentalized tumor stroma, which showed a feature of potential immunosuppression [
35]. Such visualization underlined interactions between T cells that mediate the tumor immune environment and can shed new light on the peculiarities of tumor microenvironments [
10,
87].