Bladder tumor ILC1s undergo Th17‐like differentiation in human bladder cancer

Abstract

Purpose Human innate lymphoid cells (hILCs) are lineage‐negative immune cells that do not express rearranged adaptive antigen receptors. Natural killer (NK) cells are hILCs that contribute to cancer defense. The role of non‐NK hILCs in cancer is unclear. Our study aimed to characterize non‐NK hILCs in bladder cancer. Experimental design Mass cytometry was used to characterize intratumoral non‐NK hILCs based on 35 parameters, including receptors, cytokines, and transcription factors from 21 muscle‐invasive bladder tumors. Model‐based clustering was performed on t‐distributed stochastic neighbor embedding (t‐SNE) coordinates of hILCs, and the association of hILCs with tumor stage was analyzed. Results Most frequent among intratumoral non‐NK hILCs were hILC1s, which were increased in higher compared with lower stage tumors. Intratumoral hILC1s were marked by Th17‐like phenotype with high RORγt, IL‐17, and IL‐22 compared to Th1 differentiation markers, including Tbet, perforin, and IFN‐γ. Compared with intratumoral hILC2s and hILC3s, hILC1s also had lower expression of activation markers (NKp30, NKp46, and CD69) and increased expression of exhaustion molecules (PD‐1 and Tim3). Unsupervised clustering identified nine clusters of bladder hILCs, which were not defined by the primary hILC subtypes 1–3. hILC1s featured in all the nine clusters indicating that intratumoral hILC1s displayed the highest phenotypic heterogeneity among all hILCs. Conclusions hILC1s are increased in higher stage tumors among patients with muscle‐invasive bladder cancer. These intratumoral hILC1s exhibit an exhausted phenotype and Th17‐like differentiation, identifying them as potential targets for immunotherapy.

KLRF1, a novel marker of CD56bright NK cells, predicts improved survival for patients with locally advanced bladder cancer

“…As described, 8 bladder tumors were surgically excised under sterile conditions as per standard of care. A portion of the tumor was separated and placed in Roswell Park Memorial Institute (RPMI) 1640 medium containing 1% antibiotic (Penicillin–Streptomycin) and transported on ice.…”

“…To define the phenotypic diversity of human bladder tumor innate lymphoid cells, we designed a CyTOF panel of 36 antibodies ( Table S1 ). 8 As described, 8 cells were thawed in Hank's balanced salt solution without Ca2 + or Mg2 + + 10% FBS and the number of viable cells was quantified using trypan blue. Before surface staining, cells were stained with cisplatin or discrimination of dead cells from live cells.…”

“…The NK cells were then identified based on the intensity of selected markers (CD45 + , CD3 − , CD14 − , CD19 − , ILT3 − , and CD56 bright/dim ) as described. 6 , 8 The expression levels of individual markers were visualized in tSNE plots. The comparison of expression levels of individual markers was displayed in the violin plot using the ggplot function in R.…”

“…PhenoGraph in the package. The NK cells were then identified based on the intensity of selected markers (CD45 + , CD3 − , CD14 − , CD19 − , ILT3 − , and CD56 bright/dim ) as described 6,8 . The expression levels of individual markers were visualized in tSNE plots.…”

Interleukin-17 Family Cytokines in Metabolic Disorders and Cancer

“…Although not exhaustive, the IL-17 family of cytokines have also been implicated in cancers of the lung, prostate, cervix, ovary, bladder, and skin [ 213 , 214 , 215 , 216 , 217 , 218 , 219 , 220 , 221 , 222 ]. These areas have been extensively covered by a number of review articles and will not be elaborated here.…”