Type: Oral
Session: 622. Lymphoma Biology—Non-Genetic Studies: Microenvironment and Immune Response in Hodgkin Lymphoma
Hematology Disease Topics & Pathways:
Adult, Diseases, Lymphoma (any), Non-Hodgkin Lymphoma, Biological Processes, T-Cell Lymphoma, Technology and Procedures, immune cells, Cell Lineage, Lymphoid Malignancies, Study Population, immune mechanism, mass cytometry, microenvironment, RNA sequencing, serologic tests
METHODS: We designed two novel CyTOF antibody (Ab) panels to identify and characterize cells of T, B, NK, monocyte and fDC lineages. Samples analyzed included a cohort of 25 biopsy specimens from 8 histologically confirmed AITL patients (5 lymph node (LN), 3 spleen (SP)) and 17 normal controls across key comparator immune tissue types (7 LN, 6 SP, 4 tonsil (TL)). Extensive high-dimensional analysis of CyTOF data was then performed to provide novel insights into key phenotypes and trends of malignant and non-malignant populations in AITL. We then performed CITE-Seq on control and AITL samples to gain further insight into the RNA transcriptome of key T cell populations at the cellular level. Finally, peripheral serum analysis of cytokines, soluble immune receptors, and ligands were then measured by multiplex ELISA from a separate cohort of 22 samples (5 AITL, 17 control) distinct from the individuals analyzed in the original high-dimensional study cohort.
RESULTS: While the presence of “reactive” CD8+ populations is a known histologic hallmark of AITL, we describe the gross expansion of novel CD8+ populations with distinctive immunophenotypic features which have not previously been detailed in this malignancy. Using single-cell protein expression data from CyTOF, these expanded CD8+ populations can be broadly categorized as “effector memory” (CCR7-, CD45RO+, CD45RA-) and further characterized phenotypically by markers of progressive exhaustion, checkpoint inhibition, and terminal differentiation (PD1++, TIGIT++, ICOS+, TIM3+). Further analysis of the single-cell transcriptome from these expanded CD8+ populations via CITE-Seq revealed an expression signature consistent with dysfunction and limited cytotoxic activity (including significant down-regulation of granzyme, perforin, and IFN-g) when compared to benign and malignant controls. Interestingly, when compared to CD8+ populations of identical phenotype found in control tissues, these cells also featured marked upregulation of XCL2 and XCL1 in AITL.
Additionally, global shifts in infiltrating CD19+ B cell phenotypes were seen in AITL, marked specifically by diminished expression of both CXCR5 and CD73. Finally, soluble PD-1 and other key immune molecules implicated in the expanded tumor microenvironment were found to be significantly increased in the peripheral serum of AITL patients compared to controls (1567.9 pg/mL (1109.3 S.E.) in AITL vs 29.79 (8.84 S.E.) in controls; P<0.0001).
CONCLUSIONS: High-dimensional and single-cell transcriptome analysis of the AITL microenvironment yielded several novel insights which have not been previously described in this malignancy. Highlights include the gross expansion of distinct CD8+ populations – the majority of which are of an exhausted, dysfunctional phenotype featuring marked upregulation of XCL2 and XCL1 – and the global loss of CXCR5 and CD73 expression among AITL CD19+ B cell populations. Taken together, this suggests the presence of aberrant non-malignant immune subsets within the AITL microenvironment which may contribute to novel mechanisms of immune escape.
Disclosures: Cerhan: NanoString: Research Funding; BMS/Celgene: Research Funding. Ansell: Bristol Myers Squibb: Research Funding; ADC Therapeutics: Research Funding; Seattle Genetics: Research Funding; Regeneron: Research Funding; Affimed: Research Funding; AI Therapeutics: Research Funding; Trillium: Research Funding; Takeda: Research Funding.
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