Expression of lymphoid structure-associated cytokine/chemokine gene transcripts in tumor and protein in serum are prognostic of melanoma patient outcomes

BackgroundProinflammatory chemokines/cytokines support development and maturation of tertiary lymphoid structures (TLS) within the tumor microenvironment (TME). In the current study, we sought to investigate the prognostic value of TLS-associated chemokines/cytokines (TLS-kines) expression levels in melanoma patients by performing serum protein and tissue transcriptomic analyses, and to then correlate these data with patients clinicopathological and TME characteristics.MethodsLevels of TLS-kines in patients’ sera were quantitated using a custom Luminex Multiplex Assay. The Cancer Genomic Atlas melanoma cohort (TCGA-SKCM) and a Moffitt Melanoma cohort were used for tissue transcriptomic analyses. Associations between target analytes and survival outcomes, clinicopathological variables, and correlations between TLS-kines were statistically analyzed.ResultsSerum of 95 patients with melanoma were evaluated; 48 (50%) female, median age of 63, IQR 51-70 years. Serum levels of APRIL/TNFSF13 were positively correlated with levels of both CXCL10 and CXCL13. In multivariate analyses, high levels of serum APRIL/TNFSF13 were associated with improved event-free survival after adjusting for age and stage (HR = 0.64, 95% CI 0.43-0.95; p = 0.03). High expression of APRIL/TNFSF13 tumor transcripts was significantly associated with improved OS in TCGA-SKCM (HR = 0.69, 95% CI 0.52-0.93; p = 0.01) and in Moffitt Melanoma patients (HR = 0.51, 95% CI: 0.32-0.82; p = 0.006). Further incorporation of CXCL13 and CXCL10 tumor transcript levels in a 3-gene index revealed that high APRIL/CXCL10/CXCL13 expression was associated with improved OS in the TCGA SKCM cohort (HR = 0.42, 95% CI 0.19-0.94; p = 0.035). Melanoma differentially expressed genes positively associated with high APRIL/CXCL10/CXCL13 tumor expression were linked to tumor infiltration by a diverse array of proinflammatory immune cell types.ConclusionSerum protein and tumor transcript levels of APRIL/TNFSF13 are associated with improved survival outcomes. Patients exhibiting high coordinate expression of APRIL/CXCL10/CXCL13 transcripts in their tumors displayed superior OS. Further investigation of TLS-kine expression profiles related to clinical outcomes in larger cohort studies is warranted

Checks and Balances of T cells in Inflammatory Environments

The immune system is composed of many checks and balances that are critical to limiting disease and maintaining homeostasis. T cells are critical regulators of this balance. For example, CD8+ T cells can limit diseases such as cancer and infection through induction of apoptosis in malignant and infected cells, respectively. Conversely, Regulatory T cells (Tregs), a subset of CD4+ T cells, are a suppressive population that can limit overt T cell activation and prevent autoimmunity. However, these checks and balances can be potentially damaging if they are off-balance. CD8+ can inappropriately induce cell death of various cells which can induce disease. For example, cytotoxicity towards pancreatic beta cells, can induce autoimmune diabetes. Likewise, Tregs can detrimentally suppress T cell activation and function in the tumor microenvironment (TME) to limit clearance of the tumor. Therefore, identifying these mechanisms and regulation of these mechanisms of inappropriate function in inflammatory environments will be critical to limit disease. Therefore, I examined how this balance may be maintained through epigenetic regulation in the TME and production of cytotoxic molecules in the TME and diabetic islet. Specifically, in Chapter 3, I demonstrate that Tregs alter their epigenome to aid in their suppression of the anti-tumor response. In Chapter 4, I show that Tregs do not require TNF-related apoptosis-inducing ligand (TRAIL) as a means of suppression in the TME nor diabetic islet but rather are capable of using other suppressive molecules in its absence. In Chapter 5, I demonstrate CD8+ T cell-restricted deletion of Tnfsf10 leads to almost complete protection from autoimmune diabetes. Understanding this regulation of checks and balances may aid in future therapeutic approaches to cancer and autoimmune disease. In addition to my thesis study, Appendix A shows data in which I examined various models of tumor growth in a mouse model with genetic deletion of Neuropilin-1 (Nrp1) on Tregs and Appendix B shows data in which I examined the role of Nrp1 on Tregs in fetal maternal tolerance. Finally Appendix C identifies publications that I have contributed to and awards that I have received during my graduate training