Mouse CD8 T cells treated with human recombinant AOAH protein or phosphatidylcholine with C16:0 on the sn-1 and C20:4 on the sn-2 positions

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. Overall design: To compare the genetic differences between AOAH-treated and vehicle-treated activated mouse CD8+ T cells: The mouse CD8+ T cells were first isolated from splenocytes of C57BL/6 mice using EasySep™ Mouse CD8+ T Cell Isolation Kit (Stem cell technologies), and pre-treated in T-cell medium (Miltenyi) with either 10 µg/mL rhAOAH-His (Wuxi Biologics) + ethanol control, or an equal volume of 10% glycerol PBS + ethanol control, or 10% glycerol PBS + 60 µM PC16:0-20:4 (Avanti Lipids) for 22 hours. Next, the pre-treated CD8+ T cells were activated by 1 µg/mL plate-bound anti-mouse CD3 antibody (Cytek Biosciences) and 0.5 µg/mL anti-mouse CD28 antibody (Cytek Biosciences) for 24 hours.