Digital PCR-Based T-cell Quantification–Assisted Deconvolution of the Microenvironment Reveals that Activated Macrophages Drive Tumor Inflammation in Uveal Melanoma

Abstract

: Uveal melanoma progression can be predicted by gene expression profiles enabling a clear subdivision between tumors with a good (class I) and a poor (class II) prognosis. Poor prognosis uveal melanoma can be subdivided by expression of immune-related genes; however, it is unclear whether this subclassification is justified; therefore, T cells in uveal melanoma specimens were quantified using a digital PCR approach. Absolute T-cell quantification revealed that T-cell influx is present in all uveal melanomas associated with a poor prognosis. However, this infiltrate is only accompanied by differential immune-related gene expression profiles in uveal melanoma with the highest T-cell infiltrate. Molecular deconvolution of the immune profile revealed that a large proportion of the T-cell-related gene expression signature does not originate from lymphocytes but is derived from other immune cells, especially macrophages. Expression of the lymphocyte-homing chemokine CXCL10 by activated macrophages correlated with T-cell infiltration and thereby explains the correlation of T-cell numbers and macrophages. This was validated by analysis of CXCL10 in uveal melanoma tissue with high T-cell counts. Surprisingly, CXCL10 or any of the other genes in the activated macrophage-cluster was correlated with reduced survival due to uveal melanoma metastasis. This effect was independent of the T-cell infiltrate, which reveals a role for activated macrophages in metastasis formation independent of their role in tumor inflammation. IMPLICATIONS: The current report uses an innovative digital PCR method to study the immune environment and demonstrates that absolute T-cell quantification and expression profiles can dissect disparate immune components.

Combined immune checkpoint blockade for metastatic uveal melanoma: a retrospective, multi-center study

“…Further evidence for a better efficacy of the combined regimen is supported by the observation of complete responders, albeit to a small extent. This is notable as UM is considered a "cold" tumor due to a low mutational burden and a unique immunosuppressive tumor microenvironment [27][28][29]. Further research is urgently needed to identify the radiologic, immunologic, and molecular determinants for treatment response in this small subset of patients.…”

Immunological analyses reveal an immune subtype of uveal melanoma with a poor prognosis

“…Although higher cytotoxic expression patterns are associated with better anti-tumor response and better patient survival in many solid tumors [34,41,42], the prognostic effects of immune infiltration depend on the type of tumor, the location of the cells and the state of activation. In UM, high levels of immune cells are associated with poor prognostic factors, such as M3 and BAP1 mutation [15,17,43]. Immune cell infiltration occurs more frequently in epithelioid-cell-type UM, which also has a poor prognosis [44].…”

“…Moreover, one of the most significant UM studies revealed that lymphocyte infiltration and tumorassociated M2 macrophage levels were associated with a poor prognosis in primary UM after adjustment for other risk factors [16]. In another study, the authors used a digital PCR-based T cell quantification method to characterize the prognostic value of the T cell count and activated macrophage level in the microenvironment of UM [17]. Thus, characterizing the immunological features of UM may provide novel immune biomarkers for prognostic assessment and immunotherapy.…”

Development and validation of an immune and stromal prognostic signature in uveal melanoma to guide clinical therapy

“…Previous studies mainly focused on the intrinsic genes of tumors [ 19 ], and some have provided elegant analyses on expression of immune-related genes or immune-infiltration in UM [ 11 , 20 ]. However, a comprehensive analysis of the UM microenvironment consisting of larger cohorts is needed in studies of the UM microenvironment.…”