Soluble immunosuppressive checkpointome of mesenchymal stromal cells: Immunoregulatory mechanisms and a conceptual framework for potency assessment

Table 1 The classical mediator set and the soluble checkpointome compared

Property	Classical mediators (IDO, PGE2, nitric oxide)	Soluble checkpointome
Molecular class	Enzymes, lipid and gaseous messengers[5-7]	Immunoglobulin superfamily ligands, lectins and MHC class Ib isoforms[9-12]
Receptor engaged	No dedicated inhibitory receptor; act on metabolism or signaling tone	Defined inhibitory receptors: PD-1, TIM-3, ILT2 and ILT4[9,11,12]
Inducibility	Licensed by IFN-γ and TNF-α[5,8]	Mixed: PD-L1 and galectin-9 induced, galectin-1 constitutive[9-11]
Measured in the product	Indirectly, through kynurenine or nitrite	Directly, as protein in cell-free conditioned medium[9,11]
Precedent as a potency marker	IDO proposed and used[39]	Galectin-9 predicts preparation-level potency[16]; PD-L1 proposed alongside IDO1[39]
Suitability for a release assay	Enzymatic readouts are stable but reflect a single pathway	Multi-analyte panel, matches the assay matrix logic already advocated[14,15]

IDO: Indoleamine 2,3-dioxygenase; PGE2: Prostaglandin E2; MHC: Major histocompatibility complex; PD-1: Programmed death 1; TIM-3: T cell immunoglobulin and mucin domain-containing protein 3; ILT: Immunoglobulin-like transcript; IFN-γ: Interferon gamma; TNF-α: Tumor necrosis factor alpha; PD-L1: Programmed death ligand-1.