Unraveling the immunobiology of tumor-specific T follicular helper CD4 T cells

While immune checkpoint blockade therapies have revolutionized the treatment of cancer for patients, many remain unresponsive. There has been increased focus on factors outside of tumor-infiltrating CD8 T cells that contribute to treatment efficacy, particularly the role of helper CD4 T cells. In investigating this question, current studies have identified the CD4 subset, T follicular helper cells and their canonical cytokine Interleukin (IL)-21, to be an important driver of response, particularly to the CTLA-4 axis of immunotherapy. These studies, however, fail to define the precise location of IL-21 delivery for optimal CD8 T cell help or additional required factors, including the generation tumor-specific B cells. Based on preliminary data that tumor draining lymph nodes (tdLNs) contain a reservoir of IL-21 producing CD4 T cells outside of B cell follicles and that CTLA-4-mediated IL-21 production depends on a B cell antigen, I hypothesize that in order to optimally prime CD8 T cells to generate antitumor effectors and respond to CTLA-4, IL-21 production must occur outside of the B cell follicle in tdLNs and that this depends on the presence of tumor-specific B cells. To directly test this, I will pursue the following aims. For my first aim, I will define the factors that give rise to the subset of IL-21+ CD4 T cells in the extrafollicular zone of tdLNs. To accomplish this, I will spatiotemporally track the location, phenotype, and path of differentiation of IL-21-producing cells in tdLNs by cyclic immunofluorescence, flow cytometry, and single cell RNA sequencing. I will target genes predicted to be involved in this differentiation path using Cas9 ribonucleoprotein editing of tumor-specific CD4 T cells to modify their development and function. For my second aim, I will determine the role of IL-21 production on CD8 T cell effector function and anti-CTLA-4 therapeutic efficacy. To do this, I will eliminate the ability to produce IL-21 in tumor-specific CD4 T cells or remove B cells and determine the effects on CD8 T cell effector function and CTLA-4 blockade efficacy. I will then use adoptive transfer studies with a model of lung adenocarcinoma that is capable of generating IL-21+ TFH, HELLO, and one that is not, NINJA, to assess the requirement of CD4 interactions with tumor-specific B cells for CTLA-4 immunotherapy responses. Lastly, I will directly deliver tumor-specific CD4 T cells with ectopic IL-21 expression to NINJA tumor-bearing mice to overcome the requirement of B cells. I expect that my findings will uncover the necessary factors for productive IL-21 delivery within tdLNs and identify new strategies to generate IL-21 producing cells and reinvigorate CD8 T cells in ICB unresponsive tumors. Project Number: 1F30CA310070-01 | Fiscal Year: 2026 | NIH Institute/Center: National Cancer Institute (NCI) | Principal Investigator: Emily Kessler | Institution: YALE UNIVERSITY, NEW HAVEN, CT | Award Amount: $33,384 | Activity Code: F30 | Study Section: Special Emphasis Panel[ZRG1 F09C-H (22)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11315201