KLRF1, a novel marker of CD56^bright NK cells, predicts improved survival for patients with locally advanced bladder cancer

Background: Bladder tumor-infiltrating CD56^bright NK cells are more tumor cytotoxic than their CD56^dim counterparts. Identification of NK cell subsets is labor-intensive and has limited utility in the clinical setting. Here, we sought to identify a surrogate marker of bladder CD56^bright NK cells and to test its prognostic significance. Methods: CD56^bright and CD56^dim NK cells were characterized with the multiparametric flow (n = 20) and mass cytometry (n = 21) in human bladder tumors. Transcriptome data from bladder tumors (n = 351) profiled by The Cancer Genome Atlas (TCGA) were analyzed. The expression levels of individual markers in intratumoral CD56^bright and CD56^dim NK cells were visualized in tSNE plots. Expressions of activation markers were also compared between Killer Cell Lectin-Like Receptor Subfamily F Member 1 (KLRF1)⁺ and KLRF1⁻ NK cells. Results: Intratumoral CD56^bright NK cells displayed a more activated phenotype compared to the CD56^dim subset. Multiple intratumoral cell types expressed CD56, including bladder tumor cells and nonspecific intratumoral CD56 expression was associated with worse patient survival. Thus, an alternative to CD56 as a marker of CD56^bright NK cells was sought. The activation receptor KLRF1 was significantly increased on CD56^bright but not on CD56^dim NK cells. Intratumoral KLRF1⁺ NK cells were more activated and expressed higher levels of activation molecules compared with KLRF1⁻ NK cells, analogous to the distinct effector function of NK cells across CD56 expression. High intratumoral KLRF1 was associated with improved recurrence-free survival (hazard ratio [HR] 0.53, p = 0.01), cancer-specific survival (HR 0.47, p = 0.02), and overall survival (HR 0.54, p = 0.02) on multivariable analyses that adjusted for clinical and pathologic variables. Conclusions: KLRF1 is a promising prognostic marker in bladder cancer and may guide treatment decisions upon validation.