GRIK2 is a target for bladder cancer stem-like cell-targeting immunotherapy

Recent studies revealed that treatment resistant cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) can be targeted by cytotoxic T lymphocytes (CTLs). CTLs recognize antigenic peptide derived from tumor-associated antigens (TAAs), thus identification of tumor-associated antigens (TAAs) expressed in CSCs/CICs is essential. Human leucocyte antigen (HLA) ligandome analysis using mass spectrometry enables analysis of naturally expressed antigenic peptides; however, HLA ligandome analysis requires large scale of sample and it is challenging for CSCs/CICs. In this study, we established novel bladder CSC/CIC model from a bladder cancer cell line UM-UC-3 cells using ALDEFLUOR assay. CSCs/CICs were isolated as aldehyde dehydrogenase (ALDH) high cells and several ALDHhigh clone cells were established. ALDHhigh clone cells were enriched with CSCs/CICs by sphere formation and tumorigenicity in immune deficient mouse. HLA ligandome analysis and gene expression (CAGE) using ALDHhigh clone cells revealed distinctive antigenic peptide repertoire in bladder CSCs/CICs, and we identified GRIK2 derived antigenic peptide is specifically expressed in CSCs/CICs. GRIK2 peptide-specific CTL clone recognized GRIK2-overexpressed UM-UC-3 cells and ALDHhigh clone cells indicating that GRIK2 peptide can be a novel target for bladder CSCs/CICs-targeting immunotherapy. Overall design: Urothelial CSCs/CICs were isolated by an ALDEFLUOR assay fron human urothelial carcinoma cells, UM-UC-3. Aldehyde dehydrogenase-high (ALDHhigh) clone cell were isolated as urothelial CSCs/CICs. ALDHlow clone cells were isolated as urothelial non-CSCs/CICs. ALDHhigh clones (H-1, H-6, H-10) were used as CSCs/CICs clones and ALDHlow clones (L-1, L-3, L-8) were used as non-CSCs/CICs. Wild type UM-UC-3 cells were used as a control.