Preclinical evaluation of bozepinib in bladder cancer cell lines: modulation of the NPP1 enzyme.

Bladder cancer (BC) is the most common cancer of the urinary tract. Bozepinib (BZP), a purine-derived molecule, is a potential compound for the treatment of cancer. Purinergic signaling consists of the activity of nucleosides and nucleotides present in the extracellular environment, modulating a variety of biological actions. In cancer, this signaling is mainly controlled by the enzymatic cascade involving the NTPDase/E-NPP family and ecto-5'-nucleotidase/CD73, which hydrolyze extracellular adenosine triphosphate (ATP) to adenosine (ADO). The aim of this work is to evaluate the activity of BZP in the purinergic system in BC cell lines and to compare its in vitro antitumor activity with cisplatin, a chemotherapeutic drug widely used in the treatment of BC. In this study, two different BC cell lines, grade 1 RT4 and the more aggressive grade 3 T24, were used along with a human fibroblast cell line MRC-5, a cell used to predict the selectivity index (SI). BZP shows strong antitumor activity, with notable IC50 values (8.7 ± 0.9 µM for RT4; 6.7 ± 0.7 µM for T24), far from the SI for cisplatin (SI for BZP: 19.7 and 25.7 for RT4 and T24, respectively; SI for cisplatin: 1.7 for T24). BZP arrests T24 cells in the G2/M phase of the cell cycle, inducing early apoptosis. Moreover, BZP increases ATP and ADP hydrolysis and gene/protein expression of the NPP1 enzyme in the T24 cell line. In conclusion, BZP shows superior activity compared to cisplatin against BC cell lines in vitro.

LaSOM 335, active against bladder cancer cells, interferes with Let-60 (hRas) and reduces CD73 expression/activity.

Bladder cancer is the fourth most common malignancy in men. It can present along the entire continuum of severity, from mild to well-differentiated disease to extremely malignant tumors with low survival rates. Human RAS genes are the most frequently mutated oncogenes in human cancers, and the critical role of aberrant Ras protein function in carcinogenesis is well established. Therefore, considerable efforts have been devoted to the development of anti-Ras inhibitors for cancer treatment. This study presents the biphenyl dihydropyrimidinone LaSOM 335 with high activity against T24 bladder cancer cells (IC50 = 10.73 ± 0.53 µM) and selectivity of cytotoxicity for this cancer cell line compared to two non-cancer cell lines investigated. Furthermore, we also show that this compound reduced vulvar development in the mutant let-60 gene of Caenorhabditis elegans. Let-60 is a homolog of the mammalian Ras gene. In addition, we observed that LaSOM 335 inhibits the enzymatic activity of CD73 and decreases CD73 expression. Possibly, this expression decrease is due to downstream EGFR signaling via the Ras-Raf-ERK pathway, that directly regulates CD73 expression via ERK1/2. Evidence suggests that non-immunomodulating functions of CD73 play an equally important role for cancer cell survival, progression, and migration. Regarding we also notice that LaSOM 335 was safe in the in vivo model of C. elegans. The set of these findings makes this biphenyl dihydropyrimidinone a promising candidate for further investigations in the bladder cancer field.