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T cells engineered to express chimeric-antigen receptors (CAR-T cells) can effectively control relapsed and refractory haematological malignancies in the clinic. However, the successes of CAR-T cell therapy have not been recapitulated in solid tumours due to a range of barriers such as immunosuppression, poor infiltration, and tumour heterogeneity. Numerous strategies are being developed to overcome these barriers, which include improving culture conditions and manufacturing protocols, implementing novel CAR designs, and novel approaches to engineering the T cell phenotype. In this review, we describe the various emerging strategies to improve CAR T cell therapy for solid tumours. We specifically focus on new strategies to modulate cell function and fate that have precipitated from the growing knowledge of transcriptional circuits driving T cell differentiation, with the ultimate goal of driving more productive anti-tumour T cell immunity. Evidence shows that enrichment of particular phenotypic subsets of T cells in the initial cell product correlates to improved therapeutic responses and clinical outcomes. Furthermore, T cell exhaustion and poor persistence are major factors limiting therapeutic efficacy. The latest preclinical work shows that targeting specific master regulators and transcription factors can overcome these key barriers, resulting in superior T cell therapeutic products. This can be achieved by targeting key transcriptional circuits promoting memory-like phenotypes or sustaining key effector functions within the hostile tumour microenvironment. Additional discussion points include emerging considerations for the field such as (i) targeting permutations of transcription factors, (ii) transient expression systems, (iii) tissue specificity, and (iv) expanding this strategy beyond CAR-T cell therapy and cancer., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Srinivasan, Armitage, Nilsson and Waithman.)

Background: Human perinatal life is characterized by a period of extraordinary change during which newborns encounter abundant environmental stimuli and exposure to potential pathogens. To meet such challenges, the neonatal immune system is equipped with unique functional characteristics that adapt to changing conditions as development progresses across the early years of life, but the molecular characteristics of such adaptations remain poorly understood. The application of single cell genomics to birth cohorts provides an opportunity to investigate changes in gene expression programs elicited downstream of innate immune activation across early life at unprecedented resolution., Methods: In this study, we performed single cell RNA-sequencing of mononuclear cells collected from matched birth cord blood and 5-year peripheral blood samples following stimulation (18hrs) with two well-characterized innate stimuli; lipopolysaccharide (LPS) and Polyinosinic:polycytidylic acid (Poly(I:C))., Results: We found that the transcriptional response to LPS was constrained at birth and predominantly partitioned into classical proinflammatory gene upregulation primarily by monocytes and Interferon (IFN)-signaling gene upregulation by lymphocytes. Moreover, these responses featured substantial cell-to-cell communication which appeared markedly strengthened between birth and 5 years. In contrast, stimulation with Poly(I:C) induced a robust IFN-signalling response across all cell types identified at birth and 5 years. Analysis of gene regulatory networks revealed IRF1 and STAT1 were key drivers of the LPS-induced IFN-signaling response in lymphocytes with a potential developmental role for IRF7 regulation., Conclusion: Additionally, we observed distinct activation trajectory endpoints for monocytes derived from LPS-treated cord and 5-year blood, which was not apparent among Poly(I:C)-induced monocytes. Taken together, our findings provide new insight into the gene regulatory landscape of immune cell function between birth and 5 years and point to regulatory mechanisms relevant to future investigation of infection susceptibility in early life., Competing Interests: JFR and AB are co-inventors on a patent application that is related to this work. JFR and AB are co-founders, equity holders, and directors of the startup company Respiradigm Pty Ltd and subsidiary First Breath Health Pty Ltd that are related to this work. AB is the founder of the startup company INSiGENe Pty Ltd that is unrelated to this work. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Read, Serralha, Armitage, Iqbal, Cruickshank, Saxena, Strickland, Waithman, Holt and Bosco.)

Natural killer (NK) cells have an intrinsic ability to detect and eliminate leukaemic cells. Cellular therapies using cytokine-activated NK cells have emerged as promising treatments for patients with advanced leukaemia. However, not all patients respond to current NK cell therapies, and thus improvements in efficacy are required. Type I interferons (IFN-I) are a family of potent immunomodulatory cytokines with a known ability to modulate NK cell responses against cancer. Although the human IFN-I family comprises 16 distinct subtypes, only IFNα2 has been widely explored as an anti-cancer agent. Here, we investigated the individual immunomodulatory effects each IFNα subtype and IFNβ had on NK cell functionality to determine whether a particular subtype confers enhanced effector activity against leukaemia. Importantly, IFNα14 and IFNβ were identified as superior activators of NK cell effector function in vitro . To test the ability of these subtypes to enhance NK cell activity in vivo , IFN-I stimulation was overlaid onto a standard ex vivo expansion protocol to generate NK cells for adoptive cell therapy. Interestingly, infusion of NK cells pre-activated with IFNα14, but not IFNβ, significantly prolonged survival in a preclinical model of leukaemia compared to NK cells expanded without IFN-I. Collectively, these results highlight the diverse immunomodulatory potencies of individual IFN-I subtypes and support further investigation into the use of IFNα14 to favourably modulate NK cells against leukaemia., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Barnes, Audsley, Newnes, Fernandez, de Jong, Waithman and Foley.)

Tissue-resident memory T (T RM ) cells have emerged as key players in the immune control of melanoma. These specialized cells are identified by expression of tissue retention markers such as CD69, CD103 and CD49a with downregulation of egress molecules such as Sphingosine-1-Phosphate Receptor-1 (S1PR1) and the lymphoid homing receptor, CD62L. T RM have been shown to be integral in controlling infections such as herpes simplex virus (HSV), lymphocytic choriomeningitis virus (LCMV) and influenza. More recently, robust pre-clinical models have also demonstrated T RM are able to maintain melanoma in a dormant state without progression to macroscopic disease reminiscent of their ability to control viral infections. The discovery of the role these cells play in anti-melanoma immunity has coincided with the advent of immune checkpoint inhibitor (ICI) therapy which has revolutionized the treatment of cancers. ICIs that target programmed death protein-1 (PD-1) and cytotoxic T lymphocyte antigen-4 (CTLA-4) have led to substantial improvements in outcomes for patients with metastatic melanoma and have been rapidly employed to reduce recurrences in the resected stage III setting. While ICIs mediate anti-tumor activity via CD8 + T cells, the specific subsets that facilitate this response is unclear. T RM invariably exhibit high expression of immune checkpoints such as PD-1, CTLA-4 and lymphocyte activating gene-3 (LAG-3) which strongly implicates this CD8 + T cell subset as a crucial mediator of ICI activity. In this review, we present pre-clinical and translational studies that highlight the critical role of T RM in both immune control of primary melanoma and as a key CD8 + T cell subset that mediates anti-tumor activity of ICIs for the treatment of melanoma., Competing Interests: PL has received honoraria from Bristol Myers Squibb and Pfizer. PL received institutional funding for the purposes of clinical trials from Ambrx, AstraZeneca, Beigene, Bristol Myers Squibb, Pimera, MSD and Roche Genentech. These companies did not provide funding for the publication of this article. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Plunkett, Armitage, Inderjeeth, McDonnell, Waithman and Lau.)

Cancer vaccination drives the generation of anti-tumor T cell immunity and can be enhanced by the inclusion of effective immune adjuvants such as type I interferons (IFNs). Whilst type I IFNs have been shown to promote cross-priming of T cells, the role of individual subtypes remains unclear. Here we systematically compared the capacity of distinct type I IFN subtypes to enhance T cell responses to a whole-cell vaccination strategy in a pre-clinical murine model. We show that vaccination in combination with IFNβ induces significantly greater expansion of tumor-specific CD8 + T cells than the other type I IFN subtypes tested. Optimal expansion was dependent on the presence of XCR1 + dendritic cells, CD4 + T cells, and CD40/CD40L signaling. Therapeutically, vaccination with IFNβ delayed tumor progression when compared to vaccination without IFN. When vaccinated in combination with anti-PD-L1 checkpoint blockade therapy (CPB), the inclusion of IFNβ associated with more mice experiencing complete regression and a trend in increased overall survival. This work demonstrates the potent adjuvant activity of IFNβ, highlighting its potential to enhance cancer vaccination strategies alone and in combination with CPB., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Audsley, Wagner, Ta, Newnes, Buzzai, Barnes, Wylie, Armitage, Kaisho, Bosco, McDonnell, Cruickshank, Fear, Foley and Waithman.)

Immunotherapies harnessing T cell immunity have shown remarkable clinical success for the management of cancer. However, only a proportion of patients benefit from these treatments. The presence of type I interferon (IFN) within the tumor microenvironment is critical for driving effective tumor-specific T cell immunity. Individuals can produce 12 distinct subtypes of IFNα, which all signal through a common receptor. Despite reported differences in anti-viral potencies, the concept that distinct IFNα subtypes can improve anti-cancer treatments remains unclear. We tested whether expression of unique IFNα subtypes confined to the tumor microenvironment enhances tumor control. This was systematically evaluated by transplantation of B16 murine melanoma cells secreting five unique IFNα subtypes (B16_IFNα2; B16_IFNα4; B16_IFNα5; B16_IFNα6; B16_IFNα9) into a pre-clinical murine model. We show that IFNα2 and IFNα9 are the only subtypes capable of completely controlling tumor outgrowth, with this protection dependent on the presence of an adaptive immune response. We next determined whether these differences extended to other model systems and found that the adoptive transfer of tumor-specific CD8 + T cells engineered to secrete IFNα9 delays tumor growth significantly and improves survival, whereas no enhanced survival was observed using T cells secreting IFNα4. Overall, our data shows that the expression of distinct IFNα subtypes within the tumor microenvironment results in different anti-tumor activities, and differentially affects the efficacy of a cancer therapy targeting established disease., (Copyright © 2020 Buzzai, Wagner, Audsley, Newnes, Barrett, Barnes, Wylie, Stone, McDonnell, Fear, Foley and Waithman.)

Conventional dendritic cells (cDC) resident in the lymphoid organs of mice have been classically divided into CD8 + and CD8 neg subsets. It is well-established that CD8 + dendritic cells (DCs) and their migratory counterparts in the periphery comprise the cross-presenting cDC1 subset. In contrast, CD8 neg DCs are grouped together in the heterogeneous cDC2 subset. CD8 neg DCs are relatively poor cross-presenters and drive more prominent CD4 + T cell responses against exogenous antigens. The discovery of the X-C motif chemokine receptor 1 (XCR1) as a specific marker of cross-presenting DCs, has led to the identification of a divergent subset of CD8 + DCs that lacks the ability to cross-present. Here, we report that these poorly characterized CD8 + XCR1 neg DCs have a gene expression profile that is consistent with both plasmacytoid DCs (pDCs) and cDC2. Our data demonstrate that CD8 + XCR1 neg DCs possess a unique pattern of endocytic receptors and a restricted toll-like receptor (TLR) profile that is particularly enriched for TLR5, giving them a unique position within the DC immunosurveillance network.