Background: Exosomes mediated crosstalk between tumor cells and other stromal cells including tumor associated macrophages plays an essential role in reprogramming tumor microenvironment (TME) to facilitate tumor progression. However, the mechanism of tumor derived exosomes promotes bladder cancer progression have not been defined. Methods: Exosomes were extracted from bladder cancer cells MB49 conditioned medium by ultracentrifugation. The effects of MB49-derived exosomes on macrophages polarization were analyzed by qPCR, flow cytometry, and Western blot. The immunosuppressive phenotype and function of MB49-derived exosomes stimulated macrophages were verified by tumor xenograft assays and T cell co-culture experiments. Exosomal miRNAs were analyzed by microarray to identify potential targets regulating macrophage polarization. Results: MB49-derived exosomes could be ingested by macrophages, consequently promoting macrophages immunosuppressive polarization. Mechanically, the MB49-derived exosomes induced macrophage M2 polarization was mediated by down-regulation of PTEN and activation of AKT/STAT3/6 signaling. Moreover, hindrance of the generation or secretion of exosomes by GW4869 inhibited macrophages differentiation into immunosuppressive phenotype and function, thereby suppressed tumor growth in a mouse subcutaneous tumor model. Conclusion: Our study confirmed the contribution of bladder cancer derived exosomes on the establishment of immunosuppressive TME and provided a potential therapeutic target for bladder cancer treatment.

Retinoblastoma (RB) protein can exert tumor suppressor functions even when it becomes phosphorylated. It is thus essential to understand how phosphorylated RB (p-RB) expression and function are regulated. Here, we demonstrated that RING finger domain protein TRIM28 bound and promoted ubiquitination and degradation of CDK4/6-phosphorylated RB protein. SETDB1, a known TRIM28 binding partner, protected p-RB from degradation through the binding of methylated RB by its Tudor domain independent of its methyltransferase activity. SETDB1 was found to be frequently overexpressed due to gene amplification and positively correlated with p-RB in prostate cancer patient specimens. Inhibition of SETDB1 expression using a gene-specific antisense oligonucleotide (ASO) reduced tumor growth but accelerated RB protein degradation, limiting the therapeutic efficacy. However, coadministration of the CDK4/6 inhibitor palbociclib blocked ASO-induced RB degradation and resulted in a much greater cancer-inhibitory effect than each inhibitor alone both in vitro and in vivo. This study identified CDK4/6-dependent, TRIM28-mediated proteasomal degradation as a mechanism of RB inactivation and reveals SETDB1 as a key inhibitor of this process. Our findings suggest that combined targeting of SETDB1 and CDK4/6 represents a viable approach for the treatment of cancers with SETDB1 gene amplification or overexpression. Significance: The identification of a role for TRIM28 and SETDB1 in regulating CDK4/6-phosphorylated RB stability uncovers a combination strategy using CDK4/6 and SETDB1 inhibition to decrease RB degradation and inhibit cancer growth.

Uropathogenic Escherichia coli (UPEC) is the primary causative agent of urinary tract infections (UTIs) in humans. Moreover, as one of the most common bacterial pathogens, UPEC imposes a substantial burden on healthcare systems worldwide. Epithelial cells and macrophages are two major components of the innate immune system, which play critical roles in defending the bladder against UPEC invasion. Yet, the routes of communication between these cells during UTI pathogenesis are still not fully understood. In the present study, we investigated the role of membrane-bound nanovesicles (exosomes) in the communication between bladder epithelial cells and macrophages during UPEC infection, using an array of techniques such as flow cytometry, miRNA profiling, RNA sequencing, and western blotting. Moreover, our in vitro findings were validated in a mouse model of UPEC-induced cystitis. We found that UPEC infection induced the bladder epithelial MB49 cell line to secrete large numbers of exosomes (MB49-U-Exo), which were efficiently absorbed by macrophages both in vivo and in vitro. Assimilation of MB49-U-Exo induced macrophages to produce proinflammatory cytokines, including tumor necrosis factor (TNF)α. Exposure of macrophages to MB49-U-Exo reduced their phagocytic activity (by downregulating the expression of phagocytosis-related genes) and increased their rate of apoptosis. Mechanistically, we showed that MB49-U-Exo were enriched in miR-18a-5p, which induced TNFα expression in macrophages by targeting PTEN and activating the MAPK/JNK signaling pathway. Moreover, administration of the exosome secretion inhibitor GW4869 or a TNFα-neutralizing antibody alleviated UPEC-mediated tissue damage in mice with UPEC-induced cystitis by reducing the bacterial burden of the bladder and dampening the associated inflammatory response. Collectively, these findings suggest that MB49-U-Exo regulate macrophage function in a way that exacerbates UPEC-mediated tissue impairment. Thus, targeting exosomal -release or TNFα signaling during UPEC infection may represent promising non-antibiotic strategies for treating UTIs.

Background: As a novel necrosis manner, ferroptosis has been increasingly reported to play a role in tumor progression and treatment, however, the specific mechanisms underlying its development in prostate cancer remain unclear. Growing evidence showed that peroxisome plays a key role in ferroptosis. Herein, we identified a novel mechanism for the involvement of ferroptosis in prostate cancer progression, which may provide a new strategy for clinical treatment of prostate cancer. Methods: Label-Free Mass spectrometry was used to screen and identify candidate proteins after ferroptosis inducer-ML210 treatment. Immunohistochemistry was undertaken to explore the protein expression of AGPS in prostate cancer tissues compared with normal tissues. Co-immunoprecipitation and GST pull-down were used to identify the directly binding of AGPS to MDM2 in vivo and in vitro. CCK8 assay and colony formation assay were used to illustrate the key role of AGPS in the progression of prostate cancer in vitro. The xenograft model was established to verify the key role of AGPS in the progression of prostate cancer in vivo. Results: AGPS protein expression was downregulated in prostate cancer tissues compared with normal tissues from the first affiliated hospital of Zhengzhou University dataset. Lower expression was correlated with poorer overall survival of patients compared to those with high expression of AGPS. In addition, AGPS can promote ferroptosis by modulating the function of peroxisome-resulting in the lower survival of prostate cancer cells. Furthermore, it was shown that AGPS can be ubiquitinated and degraded by the E3 ligase-MDM2 through the proteasomal pathway. Meanwhile, kinase TrkA can promote the combination of AGPS and MDM2 by phosphorylating AGPS at Y451 site. It was verified that kinase TrkA inhibitor—Larotrectinib can increase the susceptibility of prostate cancer cells to ferroptosis, which leads to the inhibition of prostate cancer proliferation to a great extent in vitro and in vivo. Conclusion: Based on these findings, we proposed the combination of ferroptosis inducer and TrkA inhibitor to synergistically exert anti-tumor effects, which may provide a new strategy for the clinical treatment of prostate cancer.

• Intratumor heterogeneity is a key driver for local relapse and treatment failure. • The prostate cancer exhibited fewer mutations in CRPC compared to HSPC, maybe due to undergo the evolutionary bottleneck introduced by ADT treatment. • New candidate genes, including MYO15A, CHD6 and LZTR1, may contribute to the evolution of castration-resistant prostate cancer. • Tracking of the dynamics of subclones or mutational events could provide a novel and more effective indicator of ADT treatment response.

LHPP, a novel, recognized tumor suppressor, exerts a critical influence on the regulation of tumor cell proliferation and survival by modulating various signaling pathways with its phosphatase activity. Here, we unveil a robust correlation between reduced LHPP expression and adverse prognosis in prostate cancer. We demonstrate that LHPP interacts with AKT, thereby dampening AKT phosphorylation and subsequently inhibiting ACSL4 phosphorylation at the T624 site. This interaction impedes phosphorylation-dependent ubiquitination, thwarting SKP2 from recognizing and binding to ACSL4 at the K621 site. As a result, ACSL4 is spared from lysosomal degradation, leading to its accumulation and the promotion of lipid peroxidation, and ferroptosis. Moreover, our findings reveal that Panobinostat, a potent histone-deacetylase inhibitor, intricately regulates LHPP expression at multiple levels through the inhibition of HDAC3. This complex modulation enhances the ferroptosis pathway, offering a novel mechanism for curtailing the growth of prostate tumors and highlighting its significant translational potential for clinical application.