Introduction & Objectives: Bladder cancer (BC) is a complex disease associated with high morbidity and mortality rates if not optimally treated. Currently, the major challenges in the clinical management of bladder cancer are disease recurrence and progression in non-muscle invasive bladder cancer (NMIBC), and therapy failure and severe side effects in muscle invasive bladder cancer (MIBC). The aim of our study was to identify new molecular targetable biomarkers to improve BC patients’ stratification and the outcome of current immunotherapies.
Materials & Methods: In a prospective cohort of 70 BC patients and a series of BC cell lines that mimic the four stages of bladder oncogenesis, we assessed the genetic and molecular regulation of Telomerase Reverse Transcriptase (TERT) in maintaining telomeres’ length and explored the expression of immune checkpoint at the mRNA level. Furthermore, we investigated the dysregulation of two microRNAs in patient’s biopsies and in BC cell lines and their potential role in regulating the genes of interest.
Results: We found that TERT gene expression was undetectable in healthy bladder tissues but was upregulated in invasive stages and high tumour grade. The TERT expression was linked with the combined presence of C250T promoter mutation and THOR hypermethylation, resulting in progressing tumors and maintaining of telomere length.
In the same BC cohort, we showed that PD-L1 was highest among NMIBC specimens, while PD-L2 was significantly upregulated in MIBC. Similarly, in human bladder cancer cell lines, PD-L2 was expressed in NMIBC stage and increased according to the histological MIBC stage.
We also showed that miR-100-5p and 138-5p, potential regulators of TERT and PD1-Ligands, were highly expressed in healthy bladder specimens and cell lines, while expression decreased in the BC tissues and BC cell lines. In line with the binding prediction scores for these miRNAs on target genes, miRs 100-5p and 138-5p expression strongly inverse correlated with TERT, PD-L1, PD-L2 expression, but not PD1.
Conclusions: Our work highlight a new molecular loop involving TERT, PD1-Ligands and miR-138-5p in BC, that might represent not only a useful biomarker for improved diagnosis and patients’ stratification, but also as a promising axis that can be therapeutically targeted in situ.

Immune checkpoints and telomerase: novel targetable axis in bladder cancer’s management