Immune checkpoint blockade: timing is everything.

Neoadjuvant immunotherapy effectively uses the in situ tumor as a reservoir of tumor antigens to promote systemic antitumor immunity. Studies indicate that intratumoral responses to immune checkpoint inhibitors (ICIs) are mediated by resident memory T cells cells that are sequestered in tumors and have specificity for a wide range of tumor antigens ICI treatment produces de novo priming of CD8+ T cells in tumor and in tumor-draining lymph nodes, and can boost the antitumor immune response by blocking inhibitory checkpoint proteins that can turn off T cells within the tumor. Neoadjuvant ICI treatment can enhance both intratumoral and systemic antitumor immunity, including expansion of intratumoral T-cell clones which is strongly associated with pathological treatment response. Recent data have shown high rates of pathological response to neoadjuvant immunotherapy with prolongation of survival compared with adjuvant ICI therapy alone in patients with unresectable or advanced melanoma. These data suggest that removal of the reservoir of tumor-specific T cells in the tumor and draining nodes by surgical resection may remove a significant proportion of the patient's antitumor immunity with the potential to compromise ICI outcomes.

CD39 Is Expressed on Functional Effector and Tissue-resident Memory CD8+ T Cells.

The ecto-ATPase CD39 is expressed on exhausted CD8+ T cells in chronic viral infection and has been proposed as a marker of tumor-specific CD8+ T cells in cancer, but the role of CD39 in an effector and memory T cell response has not been clearly defined. We report that CD39 is expressed on Ag-specific CD8+ short-lived effector cells, while it's co-ectoenzyme, CD73, is found on memory precursor effector cells (MPECs) in vivo. Inhibition of CD39 enzymatic activity during in vitro T cell priming enhances MPEC differentiation in vivo after transfer and infection. The enriched MPEC phenotype is associated with enhanced tissue resident memory T cell (TRM cell) establishment in the brain and salivary gland following an acute intranasal viral infection, suggesting that CD39 ATPase activity plays a role in memory CD8+ T cell differentiation. We also show that CD39 is expressed on human and murine TRM cells across several nonlymphoid tissues and melanoma, whereas CD73 is expressed on both circulating and resident memory subsets in mice. In contrast to exhausted CD39+ T cells in chronic infection, CD39+ TRM cells are fully functional when stimulated ex vivo with cognate Ag, further expanding the identity of CD39 beyond a T cell exhaustion marker.

CD39 is expressed on functional effector and tissue resident memory CD8+ T cells.

The ecto-ATPase CD39 is expressed on exhausted CD8+ T cells in chronic viral infection and has been proposed as a marker of tumor-specific CD8+ T cells in cancer, but the role of CD39 in an effector and memory T cell response has not been clearly defined. We report that CD39 is expressed on antigen-specific CD8+ short-lived effector cells (SLECs), while it's co-ecto-enzyme, CD73, is found on memory precursor effector cells (MPEC) in vivo . Inhibition of CD39 enzymatic activity during in vitro T cell priming enhances MPEC differentiation in vivo after transfer and infection. The enriched MPEC phenotype is associated with enhanced tissue resident memory (T RM ) establishment in the brain and salivary gland following an acute intranasal viral infection, suggesting that CD39 ATPase activity plays a role in memory CD8+ T cell differentiation. We also show that CD39 is expressed on human and murine T RM across several non-lymphoid tissues and melanoma, while CD73 is expressed on both circulating and resident memory subsets in mice. In contrast to exhausted CD39+ T cells in chronic infection, CD39+ T RM are fully functional when stimulated ex vivo with cognate antigen. This work further expands the identity of CD39 beyond a T cell exhaustion marker.

CDDO-Methyl Ester Inhibits BRAF Inhibitor Resistance and Remodels the Myeloid Compartment in BRAF-mutant Melanoma.

Approximately 50% of advanced melanomas harbor activating BRAF V600E mutations that are sensitive to BRAF inhibition. However, the duration of the response to BRAF inhibitors (BRAFi) has been limited due to the development of acquired resistance, which is preceded by recruitment of immunosuppressive myeloid cells and regulatory T cells (T regs ). While the addition of MAPK/ERK kinase 1 inhibitors (MEKi) prolongs therapeutic response to BRAF inhibition, most patients still develop resistance. Using a Braf V600E/+ /Pten -/- graft mouse model of melanoma, we now show that the addition of the methyl ester of the synthetic triterpenoid 2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oic acid (C-Me) to the BRAFi vemurafenib analog PLX4720 at resistance significantly reduces tumor burden. Dual treatment remodels the BRAFi resistant-tumor microenvironment (TME), reducing infiltration of T regs and tumor associated macrophages (TAMs), and attenuates immunosuppressive cytokine production. For the first time, we characterize myeloid populations using scRNA-seq in BRAFi-resistant tumors and demonstrate that restoration of therapeutic response is associated with significant changes in immune-activated myeloid subset representation. Collectively, these studies suggest that C-Me inhibits acquired resistance to BRAFi. Use of C-Me in combination with other therapies may both inhibit melanoma growth and enhance therapeutic responsiveness more broadly.

TimiGP: Inferring cell-cell interactions and prognostic associations in the tumor immune microenvironment through gene pairs.

Determining the prognostic association of different immune cell types in the tumor microenvironment is critical for understanding cancer biology and developing new therapeutic strategies. However, this is challenging in certain cancer types, where the abundance of different immune subsets is highly correlated. In this study, we develop a computational method named TimiGP to overcome this challenge. Based on bulk gene expression and survival data, TimiGP infers cell-cell interactions that reveal the association between immune cell relative abundance and prognosis. As demonstrated in metastatic melanoma, TimiGP prioritizes immune cells critical in prognosis based on the identified cell-cell interactions. Highly consistent results are obtained by TimiGP when applied to seven independent melanoma datasets and when different cell-type marker sets are used as inputs. Additionally, TimiGP can leverage single-cell RNA sequencing data to delineate the tumor immune microenvironment at high resolutions across a wide range of cancer types.

Th1-like Treg cells are dressed to suppress anti-tumor immunity.

Mechanisms of Th1-like Treg suppression are unknown in cancer. Two studies in Immunity by Ayala et al. and Zagorulya et al. demonstrate that Th1-like Treg cells interact with type 1 dendritic cells in tumors and draining lymph nodes to potently suppress anti-tumor immunity.

In the right place at the right time: tissue-resident memory T cells in immunity to cancer.

Tissue-resident memory (Trm) cells have recently emerged as essential components of the immune response to cancer. Here, we highlight new studies that demonstrate how CD8+ Trm cells are ideally suited to accumulate in tumors and associated tissues, to recognize a wide range of tumor antigens (Ags), and to persist as durable memory. We discuss compelling evidence that Trm cells maintain potent recall function and serve as principal mediators of immune checkpoint blockade (ICB) therapeutic efficacy in patients. Finally, we propose that Trm and circulating memory T-cell compartments together form a formidable barrier against metastatic cancer. These studies affirm Trm cells as potent, durable, and necessary mediators of cancer immunity.

The Effect of Lung Resection for NSCLC on Circulating Immune Cells: A Pilot Study.

This pilot study sought to evaluate the circulating levels of immune cells, particularly regulatory T-cell (Treg) subsets, before and after lung resection for non-small cell lung cancer. Twenty-five patients consented and had specimens collected. Initially, peripheral blood of 21 patients was collected for circulating immune cell studies. Two of these patients were excluded due to technical issues, leaving 19 patients for the analyses of circulating immune cells. Standard gating and high-dimensional unsupervised clustering flow cytometry analyses were performed. The blood, tumors and lymph nodes were analyzed via single-cell RNA and TCR sequencing for Treg analyses in a total of five patients (including four additional patients from the initial 21 patients). Standard gating flow cytometry revealed a transient increase in neutrophils immediately following surgery, with a variable neutrophil-lymphocyte ratio and a stable CD4-CD8 ratio. Unexpectedly, the total Treg and Treg subsets did not change with surgery with standard gating in short- or long-term follow-up. Similarly, unsupervised clustering of Tregs revealed a dominant cluster that was stable perioperatively and long-term. Two small FoxP3hi clusters slightly increased following surgery. In the longer-term follow-up, these small FoxP3hi Treg clusters were not identified, indicating that they were likely a response to surgery. Single-cell sequencing demonstrated six CD4+FoxP3+ clusters among the blood, tumors and lymph nodes. These clusters had a variable expression of FoxP3, and several were mainly, or only, present in tumor and lymph node tissue. As such, serial monitoring of circulating Tregs may be informative, but not completely reflective of the Tregs present in the tumor microenvironment.

Toward a better understanding of T cells in cancer.

T cells mediate anti-tumor immune responses and are the key target of immune checkpoint therapy, but they can also promote immune tolerance. A clear understanding of the specific contributions and biology of different T cell subsets is required to fully harness the curative potential of immunotherapies. Experts discuss the state of the field and key challenges for moving forward.

Resident memory CD8+ T cells in regional lymph nodes mediate immunity to metastatic melanoma.

The nature of the anti-tumor immune response changes as primary tumors progress and metastasize. We investigated the role of resident memory (Trm) and circulating memory (Tcirm) cells in anti-tumor responses at metastatic locations using a mouse model of melanoma-associated vitiligo. We found that the transcriptional characteristics of tumor-specific CD8+ T cells were defined by the tissue of occupancy. Parabiosis revealed that tumor-specific Trm and Tcirm compartments persisted throughout visceral organs, but Trm cells dominated lymph nodes (LNs). Single-cell RNA-sequencing profiles of Trm cells in LN and skin were distinct, and T cell clonotypes that occupied both tissues were overwhelmingly maintained as Trm in LNs. Whereas Tcirm cells prevented melanoma growth in the lungs, Trm afforded long-lived protection against melanoma seeding in LNs. Expanded Trm populations were also present in melanoma-involved LNs from patients, and their transcriptional signature predicted better survival. Thus, tumor-specific Trm cells persist in LNs, restricting metastatic cancer.