Impact of human immunosenescence on the development of solid and hematopoietic malignancies
Final Report Abstract
A considerable amount of data suggests an important role of the immune system in cancer development, prognosis, and therapeutic responses. We hypothesized an association of the development of late onset cancers with proceeding phenotypic and functional changes to the human immune system in the peripheral blood to be detectable with modern technologies. We used cytometry by time-of-flight, phosphoryolation-specific flow cytometry, and next generation sequencing to identify immune phenotypes in multiple myeloma, a plasma cell malignancy, and colorectal cancer as well as their corresponding pre-cancers. In peripheral blood from multiple myeloma patients, we identified a novel B cell population that was up to 14-fold expanded in patients with active multiple myeloma but not in pre-cancer and healthy individuals. This population expressed markers previously associated with both memory (CD27+) and naive (CD24'°CD38+) phenotypes. Single cell immunoglobulin gene sequencing showed polyclonality, indicating that these cells are not direct precursors to the multiple myeloma, and somatic mutations, a characteristic of memory cells. SYK, ERK, and p38 phosphorylation responses confirmed the memory character of this B cell population, calling into question the use of CD24 and CD38 for the identification of naive and memory B cells. Future experiments will determine the cues driving CD24'°CD38+CD27+ B cell expansion and analyze if this expansion occurs in cancers other than multiple myeloma or autoimmune diseases as well. For the analysis of phenotypes and antigen specificity of tumor-infiltrating lymphocytes (TILs) in colorectal cancer, we developed a novel DNA-barcoding technique for single cell sequencing. T celt receptor, cytokine, and transcription factor genes of hundreds of single TILs were amplified, DNA-barcoded, and sequenced in parallel using next generation sequencing. We showed that clonal expansion occurs selectively in CD4*+TILs but not in CD4+ T cells infiltrating macroscopically unaffected colon tissue from the same patient. Further, TILs showed unique immune phenotypes distinct from T cells isolated from unaffected colon tissue, suggesting the tumor being infiltrated with most likely tumor-reactive and potentially functionally incompetent T cell clones. In future experiments we will identify the ligands for expanded TIL clones and analyze the impact of inhibitory receptor-ligand, such as PD1-PDL1, interactions on local anti tumor immunity. Major innovations of the presented research include (1) a focus on human malignancies studied entirely in human subjects, (2) the extensive phenotypic and functional examination of peripheral blood and tumor-infiltrating T cells at a single-cell level, and (3) the complete and simultaneous sequencing of TCR α/β genes in combination with T cell lineage defining transcription factors and cytokines from single TILs that will deeply improve our understanding of colorectal cancer pathophysiology and tumor immunology, leading to (4) the possible identification of novel immune-modulatory strategies for the treatment of multiple myeloma and colorectal cancer.
Publications
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Effects of serum and plasma matrices on multiplex immunoassays. Immunologic Research, May 2014, Volume 58, Issue 2–3, pp 224–233
Rosenberg-Hasson, Y., Hansmann, L., Liedtke, M., Herschmann, I. & Maecker, N.T.
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Linking T-cell receptor sequence to functional phenotype at the single-cell level. Nature biotechnology 32, 684-692 (2014)
Han, A., Glanville, J., Hansmann, L. & Davis, M.M.
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Mass cytometry analysis shows that a novel memory phenotype B cell is expanded in multiple myeloma. Cancer immunology research. 2015 Feb 20 [Epub ahead of print]
Hansmann, L., Blum, L., Ju, C.H., Liedtke, M., Robinson, W.H. & Davis, M.M.
