CD4⁺ T cells are commonly divided into regulatory T (Treg) cells and conventional T helper (Th) cells. Th cells control adaptive immunity against pathogens and cancer by activating other effector immune cells. Treg cells are defined as CD4⁺ T cells in charge of suppressing potentially deleterious activities of Th cells. This review briefly summarizes the current knowledge in the Treg field and defines some key questions that remain to be answered. Suggested functions for Treg cells include: prevention of autoimmune diseases by maintaining self‐tolerance; suppression of allergy, asthma and pathogen‐induced immunopathology; feto‐maternal tolerance; and oral tolerance. Identification of Treg cells remains problematic, because accumulating evidence suggests that all the presently‐used Treg markers (CD25, CTLA‐4, GITR, LAG‐3, CD127 and Foxp3) represent general T‐cell activation markers, rather than being truly Treg‐specific. Treg‐cell activation is antigen‐specific, which implies that suppressive activities of Treg cells are antigen‐dependent. It has been proposed that Treg cells would be self‐reactive, but extensive TCR repertoire analysis suggests that self‐reactivity may be the exception rather than the rule. The classification of Treg cells as a separate lineage remains controversial because the ability to suppress is not an exclusive Treg property. Suppressive activities attributed to Treg cells may in reality, at least in some experimental settings, be exerted by conventional Th cell subsets, such as Th1, Th2, Th17 and T follicular (Tfh) cells. Recent reports have also demonstrated that Foxp3⁺ Treg cells may differentiate in vivo into conventional effector Th cells, with or without concomitant downregulation of Foxp3.

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death in the world. Immunological analysis of the tumor microenvironment (immunoscore) shows great promise for improved prognosis and prediction of response to immunotherapy. However, the exact immune cell composition in NSCLC remains unclear. Here, we used flow cytometry to characterize the immune infiltrate in NSCLC tumors, non-cancerous lung tissue, regional lymph node, and blood. The cellular identity of >95% of all CD45⁺ immune cells was determined. Thirteen distinct immune cell types were identified in NSCLC tumors. T cells dominated the lung cancer landscape (on average 47% of all CD45⁺ immune cells). CD4⁺ T cells were the most abundant T cell population (26%), closely followed by CD8⁺ T cells (22%). Double negative CD4⁻CD8⁻ T cells represented a small fraction (1.4%). CD19⁺ B cells were the second most common immune cell type in NSCLC tumors (16%), and four different B cell sub-populations were identified. Macrophages and natural killer (NK) cells composed 4.7 and 4.5% of the immune cell infiltrate, respectively. Three types of dendritic cells (DCs) were identified (plasmacytoid DCs, CD1c⁺ DCs, and CD141⁺ DCs) which together represented 2.1% of all immune cells. Among granulocytes, neutrophils were frequent (8.6%) with a high patient-to-patient variability, while mast cells (1.4%), basophils (0.4%), and eosinophils (0.3%) were less common. Across the cohort of patients, only B cells showed a significantly higher representation in NSCLC tumors compared to the distal lung. In contrast, the percentages of macrophages and NK cells were lower in tumors than in non-cancerous lung tissue. Furthermore, the fraction of macrophages with high HLA-DR expression levels was higher in NSCLC tumors relative to distal lung tissue. To make the method readily accessible, antibody panels and flow cytometry gating strategy used to identify the various immune cells are described in detail. This work should represent a useful resource for the immunomonitoring of patients with NSCLC.

The immune system can both promote and suppress cancer. Chronic inflammation and proinflammatory cytokines such as interleukin (IL)-1 and IL-6 are considered to be tumour promoting. In contrast, the exact nature of protective antitumour immunity remains obscure. Here, we quantify locally secreted cytokines during primary immune responses against myeloma and B-cell lymphoma in mice. Strikingly, successful cancer immunosurveillance mediated by tumour-specific CD4⁺ T cells is consistently associated with elevated local levels of both proinflammatory (IL-1α, IL-1β and IL-6) and T helper 1 (Th1)-associated cytokines (interferon-γ (IFN-γ), IL-2 and IL-12). Cancer eradication is achieved by a collaboration between tumour-specific Th1 cells and tumour-infiltrating, antigen-presenting macrophages. Th1 cells induce secretion of IL-1β and IL-6 by macrophages. Th1-derived IFN-γ is shown to render macrophages directly cytotoxic to cancer cells, and to induce macrophages to secrete the angiostatic chemokines CXCL9/MIG and CXCL10/IP-10. Thus, inflammation, when driven by tumour-specific Th1 cells, may prevent rather than promote cancer.

Tumor-associated macrophages may either promote or suppress tumor growth depending on their activation status. Interferon-gamma (IFN-gamma) has been identified as a key factor for inducing tumoricidal M1 phenotype in macrophages. However, it remains unclear whether IFN-gamma is sufficient or if additional stimuli are required. Here, we tested IFN-gamma and a panel of toll-like receptors (TLR) agonists for the ability to activate murine macrophages towards a tumoricidal M1 phenotype. The following TLR ligands were used: TLR1/TLR2 agonist Pam3CSK4, TLR2/TLR6 agonist lipotechoic acid (LTA), TLR3 agonist poly(I:C), TLR4 agonist lipopolysaccharide (LPS), TLR5 agonist flagellin (FLA), TLR7 agonist CL264, and TLR9 agonist CpG. We used an in vitro growth inhibition assay to measure both cytotoxic and cytostatic activity of mouse macrophages against Lewis lung carcinoma (LLC) and MOPC315 plasmacytoma tumor cells. Production of nitric oxide (NO) and cytokines by activated macrophages was quantified. We found that IFN-gamma alone was not able to render macrophages tumoricidal. Similarly, macrophage activation with single TLR agonists was inefficient. In sharp contrast, IFN-gamma was shown to synergize with TLR agonists for induction of macrophage tumoricidal activity and production of both NO and pro-inflammatory cytokines (TNF-alpha, IL-12p40 and IL-12p70). Furthermore, IFN-gamma was shown to suppress macrophage IL-10 secretion induced by TLR agonists. NO production was necessary for macrophage tumoricidal activity. We conclude that two signals from the microenvironment are required for optimal induction of antitumor M1 macrophage phenotype. Combination treatment with IFN-γ and TLR agonists may offer new avenues for macrophage-based cancer immunotherapy.

The importance of the immune system in conferring protection against pathogens like viruses, bacteria, and parasitic worms is well established. In contrast, there is a long-lasting debate on whether cancer prevention is a primary function of the immune system. The concept of immunological surveillance of cancer was developed by Lewis Thomas and Frank Macfarlane Burnet more than fifty years ago. We are still lacking convincing data illustrating immunological eradication of precancerous lesions in vivo. Here, I present eight types of evidence in support of the cancer immunosurveillance hypothesis. First, primary immunodeficiency in mice and humans is associated with increased cancer risk. Second, organ transplant recipients, who are treated with immunosuppressive drugs, are more prone to cancer development. Third, acquired immunodeficiency due to infection by human immunodeficiency virus (HIV-1) leads to elevated risk of cancer. Fourth, the quantity and quality of the immune cell infiltrate found in human primary tumors represent an independent prognostic factor for patient survival. Fifth, cancer cells harbor mutations in protein-coding genes that are specifically recognized by the adaptive immune system. Sixth, cancer cells selectively accumulate mutations to evade immune destruction (“immunoediting”). Seventh, lymphocytes bearing the NKG2D receptor are able to recognize and eliminate stressed premalignant cells. Eighth, a promising strategy to treat cancer consists in potentiating the naturally occurring immune response of the patient, through blockade of the immune checkpoint molecules CTLA-4, PD-1, or PD-L1. Thus, there are compelling pieces of evidence that a primary function of the immune system is to confer protection against cancer.