Co-cultures between hgp100-pulsed splenocytes with Pmel mouse CD8 T cells treated with human recombinant AOAH protein or vehicle (10% glycerol PBS)

Secreted proteins are central mediators of intercellular communications and can serve as therapeutic targets in diverse diseases. The ~1903 human genes encoding secreted proteins are difficult to study through common genetic approaches. To address this hurdle and, more generally, to discover cancer therapeutics, we developed the Cancer Immunology Data Engine (CIDE, https://cide.ccr.cancer.gov), which incorporates 90 omics datasets spanning 8575 tumor profiles with immunotherapy outcomes from 17 solid tumor types. CIDE systematically identifies all genes associated with immunotherapy outcomes. Then, we focused on secreted proteins prioritized by CIDE without known cancer roles and validated regulatory effects on immune checkpoint blockade for AOAH, CR1L, COLQ, and ADAMTS7 in mouse models. The top hit, AOAH (Acyloxyacyl Hydrolase), potentiates immunotherapies in multiple tumor models by sensitizing T-cell receptors to weak antigens and protecting dendritic cells through depleting immunosuppressive arachidonoyl phosphatidylcholines and oxidized derivatives. The CD8 T cells were first isolated from the splenocytes of pmel mice and pre-treated in the T-cell medium (Miltenyi) with either rhAOAH-His (Wuxi Biologics, 10 µg/mL or 5 µg/mL) or Buffer (PBS + 10% glycerol) for 22 hours. Next, the splenocytes from C57BL/6 mice were pulsed with 100 nM hgp100 antigens (GeneScript) in RPMI-1640 at 37 °C for 2 hours and mixed with the pre-treated pmel CD8 T cells at 3:1 or 2:1 ratios in T-cell medium (Miltenyi) + 100 IU/mL recombinant human IL-2 (R&D Systems) with either 10 µg/mL rhAOAH-His (Wuxi Biologics) or buffer control (PBS + 10% glycerol) for 24 hours.