Skin dendritic cells in melanoma are key for successful checkpoint blockade therapy

Abstract

BackgroundImmunotherapy with checkpoint inhibitors has shown impressive results in patients with melanoma, but still many do not benefit from this line of treatment. A lack of tumor-infiltrating T cells is a common reason for therapy failure but also a loss of intratumoral dendritic cells (DCs) has been described.MethodsWe used the transgenic tg(Grm1)EPv melanoma mouse strain that develops spontaneous, slow-growing tumors to perform immunological analysis during tumor progression. With flow cytometry, the frequencies of DCs and T cells at different tumor stages and the expression of the inhibitory molecules programmed cell death protein-1 (PD-1) and T-cell immunoglobulin and mucin-domain containing-3 (TIM-3) on T cells were analyzed. This was complemented with RNA-sequencing (RNA-seq) and real-time quantitative PCR (RT-qPCR) analysis to investigate the immune status of the tumors. To boost DC numbers and function, we administered Fms-related tyrosine 3 ligand (Flt3L) plus an adjuvant mix of polyI:C and anti-CD40. To enhance T cell function, we tested several checkpoint blockade antibodies. Immunological alterations were characterized in tumor and tumor-draining lymph nodes (LNs) by flow cytometry, CyTOF, microarray and RT-qPCR to understand how immune cells can control tumor growth. The specific role of migratory skin DCs was investigated by coculture of sorted DC subsets with melanoma-specific CD8+ T cells.ResultsOur study revealed that tumor progression is characterized by upregulation of checkpoint molecules and a gradual loss of the dermal conventional DC (cDC) 2 subset. Monotherapy with checkpoint blockade could not restore antitumor immunity, whereas boosting DC numbers and activation increased tumor immunogenicity. This was reflected by higher numbers of activated cDC1 and cDC2 as well as CD4+ and CD8+ T cells in treated tumors. At the same time, the DC boost approach reinforced migratory dermal DC subsets to prime gp100-specific CD8+ T cells in tumor-draining LNs that expressed PD-1/TIM-3 and produced interferon γ (IFNγ)/tumor necrosis factor α (TNFα). As a consequence, the combination of the DC boost with antibodies against PD-1 and TIM-3 released the brake from T cells, leading to improved function within the tumors and delayed tumor growth.ConclusionsOur results set forth the importance of skin DC in cancer immunotherapy, and demonstrates that restoring DC function is key to enhancing tumor immunogenicity and subsequently responsiveness to checkpoint blockade therapy.

Local delivery of low-dose anti–CTLA-4 to the melanoma lymphatic basin leads to systemic T reg reduction and effector T cell activation

“…The increased expression of CD83 and CD40 on migratory cDC subsets in tremelimumab-treated SLN in our study was likely an indirect result of enhanced pro-inflammatory cytokine signaling and/or decreased aT reg rates at the dermal injection site. The finding that CTLA-4 blockade preferentially interferes with migratory cDC is in keeping with their higher expression levels of CD80 and CD86 ( 40 ) and their reported cross-talk with T regs during or right after their migration to skin-draining lymph nodes, which is involved in the maintenance of self-tolerance ( 53 – 57 ). We thus provide clinical evidence that CTLA-4–mediated interactions between migratory cDC and aT reg in TDLN (with elevated surface levels of CTLA-4) are pivotal in setting the levels of both local and systemic aT regs .…”

“…Others show no decrease in absolute T reg numbers in the TME by quantitative immunohistochemistry ( 48 ). Differences may in part be explained by assessment of T reg levels at different time points after CTLA-4 blockade ( 24 , 53 , 54 , 58 ). Previous studies, limited to patients with advanced or metastatic melanoma, have not reported a consistent pattern of change in peripheral blood T reg frequency after systemic anti–CTLA-4 treatment ( 59 – 62 ).…”

Glutamatergic Signaling a Therapeutic Vulnerability in Melanoma

“…These mice develop spontaneous melanoma without any chemical or UV induction; therefore we propose that the tumor-stromal interactions as well as the tumor-immune system interactions are physiologically relevant to human melanomas. This notion was supported by the report that these mice have similar immune system dysfunctions as human melanoma patients and is one of the contributors to melanoma development [111][112][113][114]. Further discussion of the immune dysfunction occurring in our Grm1driven melanoma mice have been reviewed by Eddy et al, in "Overcoming Immune Evasion in Melanoma" [1].…”

The Expression Pattern of Pyroptosis-Related Genes Predicts the Prognosis and Drug Response of Skin Cutaneous Melanoma

“…Dendritic cells are specialized antigen-presenting cells that can effectively activate T cell immunity against SKCM [33][34][35], and are signi cantly associated with an improved survival time in SKCM patients [36]. As the key trigger of pyroptosis, the NLRP3 in ammasome activated by dying tumor cells can stimulate DCs, thus effectively priming CD8 + T cells by secreting IL-1β, and thereby inducing a strong and durable tumor immune response [37].…”