The role of SLAMF6- and TIM-3- splicing isoforms in THP-1-derived macrophage and dendritic cell subsets

Treatment of metastatic melanoma with immune checkpoint blockades show promising clinical data, but there is still a large non-responsive cohort. Therefore, there is an unmet need to study the suppressive tumor-microenvironment (TME) and potential immune checkpoints. T cells and myeloid regulatory cells in the TME express the SLAMF6 and TIM-3 immune receptors. We have shown that the splicing isoform V3 (Δ17-65) of the inhibitory receptor SLAMF6 enhances T cell function. However, its function in myeloid regulatory cells remains unknown. We aim toidentify the role of SLAMF6 and TIM-3 splicing patterns in THP-1 myeloid derived subsets.

The THP-1 Wt, SLAMF6-/-, and TIM-3-/- cell lines were differentiated into M0, M1, M2, or mDC subsets. Their phenotypes were investigated by studying CD83, CD86, CD14, CD11c, CD80, CD206, CD163, HLA-DR, and PD-L1 cell surface expression, and the secretion of pro- and anti-inflammatory cytokines TNF-α, IL-6, and IL-10. mRNA expression levels of SLAMF6 and TIM-3 isoforms were analyzed by qRT-PCR.

Preliminary data show increased TIM-3 and SLAMF6 mRNA levels in THP-1 Wt M1 and M2 subsets with a more prominent fold change of SLAMF6 V1 isoform than SLAMF6 V3 when normalized to ActB gene. Remarkably THP-1 Wt mDCs show decreased surface expression of SLAMF6.

Results of THP-1 SLAMF6-/- and TIM-3-/- are expected soon.

SLAMF6 and TIM-3 might play a role in macrophage polarization. We will investigate our findings with U973 and PBMC-derived macrophage subsets. After, we want to unravel the intracellular pathways and eventually find novel interacting partners of these splicing isoforms.