The implementation of immune checkpoint inhibitors to the oncology clinic signified a new era in cancer treatment. After the first indication of melanoma, an increasing list of additional cancer ...types are now treated with immune system targeting antibodies to PD-1, PD-L1 and CTLA-4, alleviating inhibition signals on T cells. Recently, we published proof-of-concept results on a novel checkpoint inhibitor, NKG2A. This receptor is expressed on cytotoxic lymphocytes, including NK cells and subsets of activated CD8+ T cells. Blocking antibodies to NKG2A unleashed the reactivity of these effector cells resulting in tumor control in multiple mouse models and an early clinical trial. Monalizumab is inhibiting this checkpoint in human beings and future clinical trials will have to reveal its potency in combination with other cancer treatment options.
The surface inhibitory receptor NKG2A forms heterodimers with the invariant CD94 chain and is expressed on a subset of activated CD8 T cells. As antibodies to block NKG2A are currently tested in ...several efficacy trials for different tumor indications, it is important to characterize the NKG2A+ CD8 T cell population in the context of other inhibitory receptors. Here we used a well‐controlled culture system to study the kinetics of inhibitory receptor expression. Naïve mouse CD8 T cells were synchronously and repeatedly activated by artificial antigen presenting cells in the presence of the homeostatic cytokine IL‐7. The results revealed NKG2A as a late inhibitory receptor, expressed after repeated cognate antigen stimulations. In contrast, the expression of PD‐1, TIGIT and LAG‐3 was rapidly induced, hours after first contact and subsequently down regulated during each resting phase. This late, but stable expression kinetics of NKG2A was most similar to that of TIM‐3 and CD39. Importantly, single‐cell transcriptomics of human tumor‐infiltrating lymphocytes (TILs) showed indeed that these receptors were often coexpressed by the same CD8 T cell cluster. Furthermore, NKG2A expression was associated with cell division and was promoted by TGF‐β in vitro, although TGF‐β signaling was not necessary in a mouse tumor model in vivo. In summary, our data show that PD‐1 reflects recent TCR triggering, but that NKG2A is induced after repeated antigen stimulations and represents a late inhibitory receptor. Together with TIM‐3 and CD39, NKG2A might thus mark actively dividing tumor‐specific TILs.
What's new?
The immune inhibitory receptor NKG2A is of emerging interest in cancer immunotherapy, with antibodies that target and interrupt NKG2A, in combination with PD‐L1 blockade, yielding promising antitumor responses in ongoing clinical trials. NKG2A expression on CD8 T cells, however, has not been fully charted and thus little is known about its regulation. Here, in vitro analyses of the kinetics of inhibitory receptor expression identify NKG2A as a late immune checkpoint. Its expression, induced upon repeated antigen encounter, is frequently associated with TIM3 and CD39. The findings suggest that NKG2A is a potential marker for the identification of tumor‐specific T cells.
The success of checkpoint blockade therapy revolutionized cancer treatment. However, we need to increase the fraction of responding patients and overcome acquired resistance to these therapies. ...Recently, the inhibitory receptor NKG2A received attention as a new kid on the block of immune checkpoints. This receptor is selectively expressed on cytotoxic lymphocytes, including natural killer cells and CD8 T cells, and NKG2A
T cells are preferentially residing in tissues, like the tumor microenvironment. Its ligand, histocompatibility leucocyte antigen E (HLA-E), is a conserved nonclassical HLA class I molecule that binds a limited peptide repertoire and its expression is commonly detected in human cancer. NKG2A blockade as a standalone therapy appears poorly effective in mouse tumor models, however, in the presence of activated T cells, for example, induced by PD-1/PD-L1 blockade or cancer vaccines, exerts strongly enhanced efficacy. Clinical trials demonstrated safety of the humanized NKG2A-blocking antibody, monalizumab, and first results of phase II trials demonstrate encouraging durable response rates. Further development of this axis is clearly warranted.
Highlights • Tumor immune escape compromises the efficacy of cancer immunotherapy. • Loss of MHC class I expression is a frequent event in cancer cells. • Three tumor phenotypes determine cancer ...fate: escape, rejection and dormancy. • Recovery of MHC class I expression is required to improve cancer immunotherapy.
High serum levels of interleukin‐6 (IL‐6) correlate with poor prognosis and chemotherapy resistance in several cancers. The underlying mechanisms and its effects on immunotherapy are largely unknown. ...To address this, we developed a human papillomavirus type 16 (HPV16)‐associated tumor model expressing IL‐6 to investigate the impact of tumor‐expressed IL‐6 during cisplatin chemotherapy and HPV16 synthetic long peptide vaccination as immunotherapy. The effects of tumor‐produced IL‐6 on tumor growth, survival and the tumor microenvironment were analyzed. Our data demonstrated that tumor‐produced IL‐6 conferred resistance to cisplatin and therapeutic vaccination. This was not caused by a changed in vitro or in vivo growth rate of tumor cells, or a changed sensitivity of tumor cells to chemotherapy or T‐cell‐mediated killing. Furthermore, no overt differences in the frequencies of tumor‐infiltrating subsets of T cells or CD11b+ myeloid cells were observed. IL‐6, however, affected the systemic and local function of myeloid cells, reflected by a strong reduction of major histocompatibility complex (MHC) class II expression on all major myeloid cell subtypes. Resistance to both therapies was associated with a changed intratumoral influx of MHC class II+ myeloid cells toward myeloid cells with no or lower MHC class II expression. Importantly, while these IL‐6‐mediated effects provided resistance to the immunotherapy and chemotherapy as single therapies, their combination still successfully mediated tumor control. In conclusion, IL‐6‐mediated therapy resistance is caused by an extrinsic mechanism involving an impaired function of intratumoral myeloid cells. The fact that resistance can be overcome by combination therapies provides direction to more effective therapies for cancer.
What's new?
Interleukin‐6 (IL‐6) cytokine has multiple effects on hematopoiesis and immune function and typically circulates at low levels. In cancer, however, IL‐6 serum levels are significantly elevated, with suspected impacts on tumor behavior. In this study, using a mouse model of human papillomavirus‐induced cancer with IL‐6 expression, the authors show that tumor‐produced IL‐6 confers resistance to both chemotherapy and immunotherapy. Resistance was associated with impaired myeloid cell maturation, with no evidence of involvement of mechanisms intrinsic to tumor cells. Resistance was overcome by combining chemotherapy and immunotherapy, providing insight into a potentially effective therapeutic approach for cancers with IL‐6‐mediated resistance.
Therapeutic vaccines preferentially stimulate T cells against tumour-specific epitopes that are created by DNA mutations or oncogenic viruses. In the setting of premalignant disease, carcinoma in ...situ or minimal residual disease, therapeutic vaccination can be clinically successful as monotherapy; however, in established cancers, therapeutic vaccines will require co-treatments to overcome immune evasion and to become fully effective. In this Review, we discuss the progress that has been made in overcoming immune evasion controlled by tumour cell-intrinsic factors and the tumour microenvironment. We summarize how therapeutic benefit can be maximized in patients with established cancers by improving vaccine design and by using vaccines to increase the effects of standard chemotherapies, to establish and/or maintain tumour-specific T cells that are re-energized by checkpoint blockade and other therapies, and to sustain the antitumour response of adoptively transferred T cells.
Uveal melanoma (UM) with an inflammatory phenotype, characterized by infiltrating leukocytes and increased human leukocyte antigen (HLA) expression, carry an increased risk of death due to ...metastases. These tumors should be ideal for T-cell based therapies, yet it is not clear why prognostically-infaust tumors have a high HLA expression. We set out to determine whether the level of HLA molecules in UM is associated with other genetic factors, HLA transcriptional regulators, or microenvironmental factors.
28 enucleated UM were used to study HLA class I and II expression, and several regulators of HLA by immunohistochemistry, PCR microarray, qPCR and chromosome SNP-array. Fresh tumor samples of eight primary UM and four metastases were compared to their corresponding xenograft in SCID mice, using a PCR microarray and SNP array.
Increased expression levels of HLA class I and II showed no dosage effect of chromosome 6p, but, as expected, were associated with monosomy of chromosome 3. Increased HLA class I and II protein levels were positively associated with their gene expression and with raised levels of the peptide-loading gene TAP1, and HLA transcriptional regulators IRF1, IRF8, CIITA, and NLRC5, revealing a higher transcriptional activity in prognostically-bad tumors. Implantation of fresh human tumor samples into SCID mice led to a loss of infiltrating leukocytes, and to a decreased expression of HLA class I and II genes, and their regulators.
Our data provides evidence for a proper functioning HLA regulatory system in UM, offering a target for T-cell based therapies.
In cancer immunotherapy, a patient’s own immune system is harnessed against cancer. Immune checkpoint inhibitors release the brakes on tumor-reactive T cells and, therefore, are particularly ...effective in treating certain immune-infiltrated solid tumors. By contrast, solid tumors with immune-silent profiles show limited efficacy of checkpoint blockers due to several barriers. Recent discoveries highlight transforming growth factor-β (TGF-β)-induced immune exclusion and a lack of immunogenicity as examples of these barriers. In this review, we summarize preclinical and clinical evidence that illustrates how the inhibition of TGF-β signaling and the use of oncolytic viruses (OVs) can increase the efficacy of immunotherapy, and discuss the promise and challenges of combining these approaches with immune checkpoint blockade.
Immune checkpoint blockade is not effective in immune-excluded and -desert tumors due to an immunosuppressive tumor microenvironment and the absence of activated T cells.TGF-β is a pleiotropic cytokine that contributes to immune exclusion and evasion in various cancer types.The therapeutic efficacy of oncolytic viruses is built on the recruitment of T cells and the induction of tumor-reactive immunity.Oncolytic virotherapy and inhibition of TGF-β signaling, either alone or in combination, are two emerging approaches to increase the susceptibility of immune-silent tumors to immune checkpoint therapy.
There is growing interest in HLA‐E‐restricted T‐cell responses as a possible novel, highly conserved, vaccination targets in the context of infectious and malignant diseases. The developing field of ...HLA multimers for the detection and study of peptide‐specific T cells has allowed the in‐depth study of TCR repertoires and molecular requirements for efficient antigen presentation and T‐cell activation. In this study, we developed a method for efficient peptide thermal exchange on HLA‐E monomers and multimers allowing the high‐throughput production of HLA‐E multimers. We optimized the thermal‐mediated peptide exchange, and flow cytometry staining conditions for the detection of TCR and NKG2A/CD94 receptors, showing that this novel approach can be used for high‐throughput identification and analysis of HLA‐E‐binding peptides which could be involved in T‐cell and NK cell‐mediated immune responses. Importantly, our analysis of NKG2A/CD94 interaction in the presence of modified peptides led to new molecular insights governing the interaction of HLA‐E with this receptor. In particular, our results reveal that interactions of HLA‐E with NKG2A/CD94 and the TCR involve different residues. Altogether, we present a novel HLA‐E multimer technology based on thermal‐mediated peptide exchange allowing us to investigate the molecular requirements for HLA‐E/peptide interaction with its receptors.
In this study, we developed a method for efficient peptide thermal exchange on HLA‐E monomers and multimers allowing the high‐throughput production of HLA‐E multimers. We optimized the thermal‐mediated peptide exchange, and flow cytometry staining conditions for the detection of TCR and NKG2A/CD94 receptors, showing that this novel approach can be used for high throughput identification and analysis of HLA‐E‐binding peptides which could be involved in T‐cell and NK cell‐mediated immune responses. Importantly, our analysis of NKG2A/CD94 interaction in the presence of modified peptides led to new molecular insights governing the interaction of HLA‐E with this receptor.
The efficiency of antigen (Ag) processing by dendritic cells (DCs) is vital for the strength of the ensuing T‐cell responses. Previously, we and others have shown that in comparison to protein ...vaccines, vaccination with synthetic long peptides (SLPs) has shown more promising (pre‐)clinical results. Here, we studied the unknown mechanisms underlying the observed vaccine efficacy of SLPs. We report an in vitro processing analysis of SLPs for MHC class I and class II presentation by murine DCs and human monocyte‐derived DCs. Compared to protein, SLPs were rapidly and much more efficiently processed by DCs, resulting in an increased presentation to CD4+ and CD8+ T cells. The mechanism of access to MHC class I loading appeared to differ between the two forms of Ag. Whereas whole soluble protein Ag ended up largely in endolysosomes, SLPs were detected very rapidly outside the endolysosomes after internalization by DCs, followed by proteasome‐ and transporter associated with Ag processing‐dependent MHC class I presentation. Compared to the slower processing route taken by whole protein Ags, our results indicate that the efficient internalization of SLPs, accomplished by DCs but not by B or T cells and characterized by a different and faster intracellular routing, leads to enhanced CD8+ T‐cell activation.
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