Interleukin 9 (IL-9) in anticancer immune responses against melanoma in mice and human

Interleukin 9 (IL-9) is a cytokine produced by T cells that has recently been shown to provide potent and long-lasting protection against B16 melanoma growth in mice. In humans, IL-9 producing T cells are decreased in melanoma metastases. However, the mechanisms of IL-9 anticancer effects remain uncharacterized and effects of IL-9 have never been studied in a human cancer. In healthy individuals, it was shown that human IL-9 production by T cells was transient, preceded the up-regulation of other inflammatory cytokines and that IL-9 enhanced cytokine production by TH1, TH2, TH9 and TH17 cells, suggesting this cytokine may function is as an amplifier of inflammation.Because of its remarkable ability to stop B16 melanoma growth, this project aims to study the role and mechanism of IL-9 in suppressing tumor growth using a novel physiologically relevant mouse model. BRAF(V600E) melanoma cells will be injected into wild type mice infused with transgenic T cells specific for mouse homologues of the known human melanoma antigens gp100 and TRP. Using melanoma tumor cells similar to those seen in patients will lead to a better understanding of the ability of T cells making IL-9 to attack and destroy these tumors.To study IL-9 in human melanoma responses, the applicant will measure IL-9 levels in a rare and valuable set of primary melanoma tumors of intermediate risk that are matched in thickness from 62 patients, 31 who died and 31 who survived, and determine if IL-9 is associated with better antitumor responses and improved survival. Lastly, the applicant will study IL-9 production by T cells in tumors and blood in patients before and after immunomodulatory therapies for advanced (metastasized) melanoma, determining if IL-9 production is increased after reversal of immune evasion and if IL-9 production predicts a better responsiveness to immunotherapy.In summary, this project will be a highly synergistic combination of mouse and human studies to evaluate a novel role for IL-9 as an initiator and sustainer of inflammation and as a novel therapy for melanoma. If successful, these studies will enhance our basic understanding of IL-9 biology and also have the potential to lead to novel IL-9-based therapies for cancer.

This research fellowship enabled me to study the T cell receptor repertoire and lipid metabolism of human melanoma infiltrating T cells after the originally proposed IL-9 based project focus changed due to severe mouse breeding problems. Recent advances in immunotherapy and targeted drug therapy have dramatically improved the prognosis of metastatic melanoma. Primary melanomas > 1mm thickness are potentially curable by resection but may later recur metastatically. To date, the risk of recurrence of primary melanomas is solely based on histopathological predictors: tumor thickness and ulceration. We aimed to identify a clinically translatable molecular marker to identify patients at high risk for disease progression more accurately. We established that T cell fraction (TCFr, proportion of nucleated cells that are T cells), as measured by high-throughput sequencing of the T cell receptor beta-chain, robustly predicted progression-free survival (PFS). 20% TCFr was the best cut-off to identify patients at risk for progression. Primary melanoma patients with low TCFr had a markedly worse prognosis. To our surprise, TCFr was completely independent of any other predictive variable and the second most powerful predictor after tumor thickness. A TCFr > 20% was protective regardless of ulceration status or mitotic rate and even protective for patients with resected regional nodal disease who had a PFS rate comparable to patients without nodal disease but low TCFr. TCFr was more accurate to predict 5-year PFS than conventional histopathological grading. Therefore, TCFr is the first quantitative molecular test that predicts metastatic recurrence in primary melanoma patients whose disease has been completely resected surgically. This tool could be easily applied in the clinic for more accurate prognostic staging and rational stratification of Stage II and III melanoma patients for therapeutic clinical trials involving adjuvant therapies. Furthermore, we characterized the lipid metabolism of tumor infiltrating T cells in human metastatic melanoma as well as squamous cell carcinoma samples. Fatty acid binding protein 5 (FABP5) was highly upregulated on gene and protein level while other lipid mediators (FABP4, LPL, CD36) were not detectable. FABP5 serves as a lipid receptor and intracellular lipid transporter and might enable tumor infiltrating T cells to survive long-term within the tumor microenvironment. Future functional studies will be conducted by members of Dr. Kupper’s laboratory to understand the effect of FABP5 up-regulation in tumor infiltrating T cells.