Cancer immunoediting is the process whereby immune cells protect against cancer formation by sculpting the immunogenicity of developing tumors. Although the full process depends on innate and adaptive immunity, it remains unclear whether innate immunity alone is capable of immunoediting. To determine whether the innate immune system can edit tumor cells in the absence of adaptive immunity, we compared the incidence and immunogenicity of 3′methylcholanthrene-induced sarcomas in syngeneic wild-type, RAG2−/−, and RAG2−/−x γc−/− mice. We found that innate immune cells could manifest cancer immunoediting activity in the absence of adaptive immunity. This activity required natural killer (NK) cells and interferon γ (IFN-γ), which mediated the induction of M1 macrophages. M1 macrophages could be elicited by administration of CD40 agonists, thereby restoring editing activity in RAG2−/−x γc−/− mice. Our results suggest that in the absence of adaptive immunity, NK cell production of IFN-γ induces M1 macrophages, which act as important effectors during cancer immunoediting.
Despite the proven efficacy of immune checkpoint inhibitor (ICI) therapy in the recurrent/metastatic setting for head and neck squamous cell carcinoma (HNSCC), clinical trials of ICI combined with curative-intent therapies have yielded equivocal results [1–4]. Collectively, this highlights gaps in our understanding of rational immune oncology (IO) treatment sequencing and suggests that the efficacy ICI may be disrupted by standard therapies, which necessarily compromise regional lymphatics.
Methods
We employ a preclinical model of tobacco-signature HNSCC to identify sequences of therapy that maximize durable response. By mapping the cervical lymphatic basins in the mouse, we define patterns of active antitumor immunosurveillance. Additionally, we establish tumors with distinct patterns of regional lymphatic drainage and develop a murine neck dissection (ND) model.
Results
We find that cervical lymphatic ablation, with ND or stereotactic body radiation therapy, in tumor bearing animals abolishes the response to ICI therapy, significantly impacting overall survival. Examination of the tumor immune microenvironment following ND reveals dramatic changes with a ten-fold increase in CD45 cells and exclusion of cytotoxic and antigen-specific lymphocytes. By examining the lymphatics removed at the time of ND, we find that conventional type I dendritic cells (cDC1s) and type I interferon (IFN-I) signaling are significantly increased, suggesting that these effectors are lost after curative-intent therapy. Depleting IFN-I or cDC1s blocks the response to ICI similar to lymphatic ablation. We find that successful primary response to ICI leads to durable immunity, conferred by systemically distributed memory T cells, not impaired by delayed ND. Lastly, we discover a rational IO treatment sequence by delivering neoadjuvant ICI followed by ND. Neoadjuvant ICI leads to complete tumor response, accumulation of nodal cDC1, and durable immunity. Surprisingly, the incidence of nodal metastasis at early timepoints reveals a similar burden of nodal disease between control and ICI-treated animals that decreases at late timepoints only with ICI treatment (44% vs 15%, n=25, p=0.033). This suggests that ICI also drives active immunosurveillance in regional, tumor-draining lymphatics, challenging the landmark findings from the definitive clinical trial demonstrating the benefit of elective versus therapeutic neck dissection for oral SCC patients with clinically negative necks.
Conclusions
This work demonstrates the necessity of preserving tumor-draining lymphatics during the tumor response to ICI therapy in HNSCC. Overall, we define rational IO treatment sequences to achieve optimal primary tumor response, durable antitumor immunity and immunosurveillance of regional metastatic disease. These findings can inform future clinical trials investigating combination IO therapy and treatment sequencing.
References
Harrington, K. J. et al. Nivolumab versus standard, single-agent therapy of investigator's choice in recurrent or metastatic squamous cell carcinoma of the head and neck (CheckMate 141): health-related quality-of-life results from a randomised, phase 3 trial. Lancet Oncology 18, 1104–1115 (2017). Burtness, B. et al. Pembrolizumab alone or with chemotherapy versus cetuximab with chemotherapy for recurrent or metastatic squamous cell carcinoma of the head and neck (KEYNOTE-048): a randomised, open-label, phase 3 study. Lancet (London, England) 394, 1915–1928 (2019). Lee, N. Y. et al. Avelumab plus standard-of-care chemoradiotherapy versus chemoradiotherapy alone in patients with locally advanced squamous cell carcinoma of the head and neck: a randomised, double-blind, placebo-controlled, multicentre, phase 3 trial. Lancet Oncol 22, 450–462 (2021). D'Cruz, A. K. et al. Elective versus Therapeutic Neck Dissection in Node-Negative Oral Cancer. New England Journal of Medicine 373, 521–529 (2015).
PD-1 inhibition (PD1i) has demonstrated no benefit for locally advanced HNSCC, and emerging neoadjuvant PD1i window of opportunity trial data yield promising but limited responses. Our previous work demonstrates that ablating tumor draining lymphatics compromises the response to immune-oncology therapy.1 In concert, a recently completed phase I trial investigating neoadjuvant immunoradiotherapy (IRT) for treatment-naïve HNSCC demonstrated a 60% major pathologic response and 100% clinical-to-pathologic downstaging (NCT03247712). Accordingly, we hypothesize that lymphatic-preserving immune oncology therapy can potentiate PD1i and promote antitumor immunity by enhancing surveillance along the tumor-immune-lymphatic axis.
Methods
To explore this, we employed our recently characterized tobacco-signature, orthotopic murine oral squamous cell carcinoma models, one of which matches the immune infiltrate and PD1i response of human disease and the other of which is immune-cold with limited response to PD1i. In both models we define an IRT scheme, using low-dose tumor-directed radiotherapy (td-RT), that achieves complete response and confers durable immunity.
Results
Mechanistically, we observe that successful td-IRT potentiates the response to PD1i by coordinating antitumor immunity across the tumor and regional lymphatics – specifically, the sentinel lymph node (SLN) – suggesting an active process of locoregional antitumor immunosurveillance. To study surveillance across the tumor-SLN axis, we map the locoregional lymphatics joining the primary tumor to its SLN and develop models to selectively ablate these lymphatic channels. Interestingly, ablation of the lymphatic channels that link the tumor and SLN is sufficient to block the tumor IRT response, leading to a restriction of cytotoxic, tumor-antigen specific CD8 T cells from the TIME and an overall reduced antitumor T cell repertoire (figure 1). To identify the requisite immune effectors that transit across tumor-SLN axis to mediate the td-IRT response, we employ tamoxifen-inducible reporter animal models in which we spatiotemporally label immune effectors during td-IRT with or without lymphatic channel ablation. Using CITE-sequencing, we comprehensively profile the dynamics of locoregional antitumor immunosurveillance at single cell resolution, finding that activated, migratory myeloid effector cells transiting between the SLN and tumor are critical for T cell priming and clonotypic expansion; and, ultimately, the successful tumor response to IRT.
Conclusions
We demonstrate that targeting tumors with stereotactic radiation and PD1i while sparing draining lymphatics enhances anticancer immunity by promoting locoregional immunosurveillance between the tumor and sentinel lymph node, resulting in significantly improved responses. Overall, this work elucidates the mechanistic underpinnings of successful tumor responses to immune oncology therapy, which can immediately inform the design of next-generation therapies for HNSCC.
Reference
Saddawi-Konefka R, et al. Lymphatic-preserving treatment sequencing with immune checkpoint inhibition unleashes cDC1-dependent antitumor immunity in HNSCC. Nat Commun 2022;13:4298.
Pediatric sinus surgery is indicated for a wide range of sinonasal and skull base pathologies, but it is most commonly performed for recalcitrant chronic rhinosinusitis or complicated acute sinusitis. The authors aim to report medical risk factors of morbidity and mortality following inpatient sinus surgery in the pediatric population.Using data from the Kids' Inpatient Database from 2003 to 2012, patients with International Classification of Diseases, Ninth Revision, procedure codes for primary sinus surgery were identified. Mixed-effect multivariable logistic regression was used to identify risk factors of inpatient postoperative morbidity and mortality.The final sample included a weighted estimate of 4965 pediatric patients. The rates of inpatient morbidity and mortality were 6% and 1%, respectively. Respiratory complications (2.5%) were the most prevalent postoperative adverse events. The most prevalent comorbidities were chronic sinusitis (59.8%), acute sinusitis (27.8%), and cystic fibrosis (26.4%). Compared with patients who did not experience any morbidity, patients with inpatient morbidity had higher rates of pneumonia, mycoses, and nasal or paranasal benign neoplasm ( P < .05). The odds of inpatient morbidity and mortality were highest for patients with leukemia (odds ratio, 2.74; 95% confidence interval, 1.59-4.72; P < .001) and mycoses (odds ratio, 15.84; 95% confidence interval, 6.45-38.89; P < .001), respectively.This study is the first to report the national comorbidity burden and risk factors for postoperative adverse events following inpatient sinus surgery. Knowledge of the comorbidities and independent factors associated with morbidity and mortality will help in directing preoperative optimization and counseling.2c.
Head and neck squamous cell carcinomas (HNSCCs) represent the sixth most common cancer worldwide with an estimated 65,630 cases and 14,500 deaths in the United States last year. Fortuitously, by virtue of their high mutational burden and robust neoantigenome, HNSCCs harbor an abundance of tumor-specific antigen (TSA)-T cells; and, thus, represent an ideal target for autologous adoptive therapy. However, the lack of a reliable biomarker to accurately identify bona fide TSA-T cells among a heterogenous population of tumor-infiltrating lymphocytes (TILs) has precluded the complete translation of this otherwise promising therapeutic strategy.
Methods
We developed an interaction-based chemoenzymatic labeling method to rapidly and efficiently identify TSA-T cells: the α-(1,3)-fucosyltransferase–FucoID–strategy. Using the FucoID method, we profiled TSA-T cells identified by proximity-based chemoenzymatic labeling (figure 1).
Results
We find that FucoID labeled T cells in both the tumor and tumor-draining lymph node compartments feature a population of TSA-T cells with exhausted-stem cell like phenotypes and robust antitumoral cytotoxic activity. Additionally, when subjected to conventional ex vivo expansion protocols, these FucoID-labeled T cells are resistant to differentiation and anergy following adoptive transfer in vivo. Through tandem T cell receptor (TCR) and transcriptomic sequencing at the single-cell level, FucoID-labeled T cells from the tumor constitute a defined population with potent effector function (figure 2). An analysis of the FucoID-labeled T cells from the draining lymph node that share a TCR repertoire with those from the tumor reveals a unique population with high TCF-7 expression and stem-like features (figure 3).
Conclusions
The FucoID proximity-based labeling strategy represents a translatable, antigen-agnostic method to exclusively and expediently identify TSA-reactive T cells with phenotypes optimal for ex vivo expansion, in vivo persistence, and antitumor cytotoxicity. This work represents a paradigm shift in the approach to adoptive T cell therapies, which can immediately inform the design of next-generation immune oncology trials for HNSCC.
Abstract Our laboratory has recently identified a novel role for the cytokine IL-17D as an important mediator of the antitumor immune response; and, the loss of IL-17D as a key component of tumor progression. Our data show for the first time that overexpression of the cytokine IL-17D is sufficient to induce tumor rejection or growth delay. Tumors expressing IL-17D displayed an increased infiltration of natural killer (NK) cells, which gave way to a commensurate increase of M1-type macrophages. While examining the regulation of IL-17D, we came across the surprising finding that IL-17D can be induced by oxidative stress signals via the transcription factor nrf2. Nrf2, responsible for regulating the expression of the anti-oxidant response element genes, is known as the primary cellular responder to oxidative stress. Although the role of nrf2 in cancer biology has been well studied, the role of nrf2 in the regulation of immune responses (particularly via IL-17D) has yet to be defined. We find that nrf2 is necessary to induce IL-17D expression in a variety of tumor and non-tumor cells, and that this regulation is active in tumor progression. These studies constitute an intriguing and novel connection between oxidative cellular stress and immune activation. Citation Format: Robert Saddawi-Konefka, Jack Bui. The novel cytokine IL-17D at the intersection of cellular stress and tumor immunity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4888. doi:10.1158/1538-7445.AM2014-4888
The process of cancer immunoediting generates a repertoire of cancer cells that can persist in immune-competent hosts. In its most complex form, this process begins with the elimination of highly immunogenic unedited tumor cells followed by the escape of less immunogenic edited cells. Although edited tumors can release immunosuppressive factors, it is unknown whether unedited tumors produce cytokines that enhance antitumor function. Utilizing gene microarray analysis, we found the cytokine interleukin 17D (IL-17D) was highly expressed in certain unedited tumors but not in edited mouse tumor cell lines. Moreover, forced expression of IL-17D in edited tumor cells induced rejection by stimulating MCP-1 production from tumor endothelial cells, leading to the recruitment of natural killer (NK) cells. NK cells promoted M1 macrophage development and adaptive immune responses. IL-17D expression was also decreased in certain high-grade and metastatic human tumors, suggesting that it can be targeted for tumor immune therapy.
