Single-cell sequencing technologies have noteworthily improved our understanding of the genetic map and molecular characteristics of bladder cancer (BC). Here we identify CD39 as a potential therapeutic target for BC via single-cell transcriptome analysis. In a subcutaneous tumor model and orthotopic bladder cancer model, inhibition of CD39 (CD39i) by sodium polyoxotungstate is able to limit the growth of BC and improve the overall survival of tumor-bearing mice. Via single cell RNA sequencing, we find that CD39i increase the intratumor NK cells, conventional type 1 dendritic cells (cDC1) and CD8 + T cells and decrease the Treg abundance. The antitumor effect and reprogramming of the tumor microenvironment are blockaded in both the NK cells depletion model and the cDC1-deficient Batf3−/− model. In addition, a significant synergistic effect is observed between CD39i and cisplatin, but the CD39i + anti-PD-L1 (or anti-PD1) strategy does not show any synergistic effects in the BC model. Our results confirm that CD39 is a potential target for the immune therapy of BC.

The essential G1-cyclin, CCND1, is a collaborative nuclear oncogene that is frequently overexpressed in cancer. D-type cyclins bind and activate CDK4 and CDK6 thereby contributing to G1–S cell-cycle progression. In addition to the nucleus, herein cyclin D1 was also located in the cytoplasmic membrane. In contrast with the nuclear-localized form of cyclin D1 (cyclin D1NL), the cytoplasmic membrane-localized form of cyclin D1 (cyclin D1MEM) induced transwell migration and the velocity of cellular migration. The cyclin D1MEM was sufficient to induce G1–S cell-cycle progression, cellular proliferation, and colony formation. The cyclin D1MEM was sufficient to induce phosphorylation of the serine threonine kinase Akt (Ser473) and augmented extranuclear localized 17β-estradiol dendrimer conjugate (EDC)-mediated phosphorylation of Akt (Ser473). These studies suggest distinct subcellular compartments of cell cycle proteins may convey distinct functions.

Calcium/calmodulin-dependentprotein kinase II inhibitor I (CAMK2N1) as one of the tumor suppressor genes is significantly downregulated in prostate cancer (PCa). Reduced expression of CAMK2N1 is positively correlated with PCa progression. However, the mechanisms of CAMK2N1 downregulation in PCa are still unclear. The promoter region of CAMK2N1 contains a large number of CG loci, providing the possibility for DNA methylation. Consequently, we hypothesized that DNA methylation can result in the reduced expression of CAMK2N1 in PCa. In the presented study, the DNA methylation level of CAMK2N1 in prostate cells and clinical specimens was determined by bisulfite sequencing (BS), pyrosequencing, and in silico analysis. Results showed that CAMK2N1 was highly methylated in PCa cells and tissues compared to normal prostate epithelial cells and nonmalignant prostate tissues, which was associated with the clinicopathological characteristics in PCa patients. Afterwards, we explored the expression of CAMK2N1 and its DNA methylation level by qRT-PCR, western blot, BS, and methylation-specific PCR in PCa cells after 5-Aza-CdR treatment or DNMT1 genetic modification, which demonstrated that the reduced expression of CAMK2N1 can be restored by 5-Aza-CdR treatment via demethylation. Moreover, DNMT1 formed a positive feedback loop with CAMK2N1 in PCa cells. The expression of CAMK2N1 was downregulated by DNMT1-mediated DNA methylation, which reversely induced DNMT1 expression through activating AKT or ERK signaling pathway. Finally, functional assays including wound healing, invasion, and migration assay, as well as the xenograft model in nude mice indicated that CAMK2N1 inhibited the invasion, migration, and proliferation of PCa cells and these effects were reversed by DNMT1 overexpression. In conclusion, DNMT1-mediated hypermethylation of CAMK2N1 not only downregulates the gene expression but also promotes the progression of PCa.