ASC-J9® increases the bladder cancer chemotherapy efficacy via altering the androgen receptor (AR) and NF-κB survival signals.

BACKGROUND: The current chemotherapy regimens may extend survival for patients with metastatic bladder cancer (BCa) for a few months, but eventually most patients succumb to disease because they develop resistance to their chemotherapy. METHODS: TCGA human clinical sample survey and urothelial tumor tissue microarrays (TMAs) were applied to investigate the expression of androgen receptor (AR) and NF-κB. Multiple BCa cell lines were used to test chemotherapy's efficacy via multiple assays including XTT, flow cytometry, TUNEL, and BrdU incorporation. The effects of the AR degradation enhancer, ASC-J9®, combined with various chemotherapy reagents were examined both in vivo and in vitro. RESULTS: We unexpectedly found that in muscle-invasive BCa (miBCa) the signals of both the AR and NF-κB were increased via a TCGA sample survey. Results from multiple approaches revealed that targeting these two increased signals by combining various chemotherapeutic agents, including Cisplatin, Doxorubicin or Mitomycin C, with ASC-J9® led to increase the therapeutic efficacy. The combined therapy increases the expression of the pro-apoptosis BAX gene and cell cycle inhibitor p21 gene, yet suppresses the expression of the pro-survival BCL2 gene in miBCa cells. Preclinical studies using an in vivo mouse model with xenografted miBCa cells confirmed in vitro cell line data showing that treatment with ASC-J9® combined with Cisplatin can result in suppressing miBCa progression better than Cisplatin alone. CONCLUSIONS: Together, these results support a novel therapeutic approach via combining Cisplatin with ASC-J9® to better suppress the progression of miBCa.

Antibiotic prophylaxis at the time of catheter removal after radical prostatectomy: A prospective randomized clinical trial.

OBJECTIVE: To evaluate the role of antibiotic prophylaxis with oral ciprofloxacin prior to urinary catheter removal after radical prostatectomy in preventing urinary tract infection (UTI). MATERIALS AND METHODS: Patients undergoing radical prostatectomy were prospectively enrolled and randomized to either the antibiotic prophylaxis group (2 doses of oral ciprofloxacin prior to urinary catheter removal) or the control group (no antibiotics given prior to urinary catheter removal). Neither patients nor study providers were blinded to the group. The primary objective was to assess for development of UTI. The secondary objective was to assess for development of Clostridium difficile (C diff) enterocolitis. Continuous variables were compared using a 2-sample t test. Categorical variables were compared using Pearson's chi-squared test or Fisher's exact test. RESULTS: One hundred seventy-five patients were enrolled and randomized (90 control and 85 antibiotic prophylaxis). After randomization, 4 patients were excluded and 4 patients withdrew voluntarily. One hundred sixty-seven patients (84 control and 83 antibiotic prophylaxis) completed the study and were available for analysis. There were no significant differences in baseline characteristics, perioperative data, or complications. There was no significant difference in the rate of UTI between the control group and antibiotic prophylaxis group (5.95% vs. 6.02%, P = 1). There was also no significant difference in the rates of C diff infection between the control and the antibiotic prophylaxis groups (3.57% vs. 0%, P = 0.21). CONCLUSIONS: In this prospective, randomized, controlled trial, the use of antibiotic prophylaxis with oral ciprofloxacin prior to urinary catheter removal after radical prostatectomy did not decrease the rate of UTI, and was not associated with an increased incidence of C diff enterocolitis.

Antiandrogen Therapy with Hydroxyflutamide or Androgen Receptor Degradation Enhancer ASC-J9 Enhances BCG Efficacy to Better Suppress Bladder Cancer Progression.

Recent studies suggest that the androgen receptor (AR) might play important roles in influencing bladder cancer progression, yet its clinical application remains unclear. Here, we developed a new combined therapy with Bacillus Calmette-Guérin (BCG) and the AR degradation enhancer ASC-J9 or antiandrogen hydroxyflutamide (HF) to better suppress bladder cancer progression. Mechanism dissection revealed that ASC-J9 treatment enhanced BCG efficacy to suppress bladder cancer cell proliferation via increasing the recruitment of monocytes/macrophages that involved the promotion of BCG attachment/internalization to the bladder cancer cells through increased integrin-α5ß1 expression and IL6 release. Such consequences might then enhance BCG-induced bladder cancer cell death via increased TNFα release. Interestingly, we also found that ASC-J9 treatment could directly promote BCG-induced HMGB1 release to enhance the BCG cytotoxic effects for suppression of bladder cancer cell growth. In vivo approaches also concluded that ASC-J9 could enhance the efficacy of BCG to better suppress bladder cancer progression in BBN-induced bladder cancer mouse models. Together, these results suggest that the newly developed therapy combining BCG plus ASC-J9 may become a novel therapy to better suppress bladder cancer progress.

1 alpha, 25-dihydroxylvitamin D3 promotes Bacillus Calmette-Guérin immunotherapy of bladder cancer.

Bacillus Calmette-Guérin (BCG), a vaccine against tuberculosis(TB), has been used and proven to be one of the most effective treatments for non-muscle invasive bladder cancer (BCa). However, the mechanisms of BCG action have not been completely understood, thereby limiting the improvement of BCG therapy. Vitamin D deficiency has been associated with a high risk of TB infection, and the beneficial effect of UV exposure in TB patients was proven to be mediated via activation of vitamin D signals of innate immune cells. Thus, vitamin D signals might be involved in mediating BCG immunotherapy. To test this hypothesis, we examined the impact of 1 alpha, 25-dihydroxyvitamin D3 (1,25-VD) on BCG-induced response in BCa cells and macrophage cells. Our data revealed that 1,25-VD promotes BCG-induced interleukin 8 (IL-8) secretion by BCa cells, consequently inducing the migration of macrophage, THP-1. This THP-1 cell migration promoted by 1,25-VD can be blocked by IL-8 neutralized antibody. Furthermore, 1,25-VD increased BCG-induced expression of macrophage markers in THP-1 cell, and enhanced the BCG-induced THP-1 cytotoxicity against low-grade BCa cells. Importantly, a pre-clinical trial using the N-butyl-N-(4-hydroxybutyl)-nitrosamine (BBN)-induced BCa mouse model revealed that intravesical co-treatment of 1,25-VD with BCG can prolong mice survival. These data demonstrate a novel mechanism by which 1,25-VD promotes BCG-mediated anti-BCa pathways and provides a platform for improving BCG efficacy with combination of 1,25-VD.

Vitamin E succinate inhibits the function of androgen receptor and the expression of prostate-specific antigen in prostate cancer cells.

Although epidemiological evidence indicates that a daily supplement of vitamin E may reduce the risk of prostate cancer, the detailed mechanism underlying this effect remains unclear. Here we demonstrate that alpha-tocopheryl succinate (VES) can suppress the expression of prostate-specific antigen (PSA), a marker for the progression of prostate cancer. VES can also suppress androgen receptor (AR) expression by means of transcriptional and posttranscriptional modulation, but not ligand binding, nuclear translocation, or AR dimerization. This VES-mediated inhibition of AR is selective because VES does not repress the expression of other nuclear receptors. Cell growth studies further show that VES inhibits the growth of prostate cancer LNCaP cells. In contrast, hydroxyflutamide (HF), an antiandrogen currently used to treat prostate cancer patients, only slightly inhibits LNCaP cell growth. Interestingly, simultaneous addition of HF and VES results in a more significant inhibition of LNCaP cell growth. Moreover, selenomethionine (SM), a prostate cancer treatment adjuvant, shows an inhibitory effect on LNCaP cell growth, yet has no effect on the AR/PSA pathway. Together, our data indicate that VES may suppress androgen/AR-mediated cell growth and PSA expression by inhibiting AR expression at both the transcription and translation levels. This previously undescribed mechanism may explain how VES inhibits the growth of prostate cancer cells and help us to establish new therapeutic concepts for the prevention and treatment of prostate cancer.

Ubiquitination of a novel deubiquitinating enzyme requires direct binding to von Hippel-Lindau tumor suppressor protein.

von Hippel-Lindau (VHL) disease is a hereditary cancer syndrome caused by germline mutations of the VHL gene. Recent studies suggest that VHL protein (pVHL) is a component of an E3 ubiquitin ligase, but the detailed biological function of pVHL remains to be determined. To further elucidate the biological functions of pVHL, we searched pVHL-interacting proteins using yeast two-hybrid screening. A novel protein named VHL-interacting deubiquitinating enzyme 1 (VDU1) was identified as being able to directly interact with pVHL in vitro and in vivo. We have determined the full-length cDNA of this enzyme, which includes two putative subtypes. Type I consists of 942 amino acids, and type II consists of 911 amino acids with predicted molecular masses of 107 and 103 kDa, respectively. We have also cloned a mouse homologue of this enzyme. Sequence analysis reveals that this protein is conserved between human and mouse and contains the signature motifs of the ubiquitin-specific processing protease family. Enzymatic function studies demonstrate its deubiquitinating activity. We have determined that the VDU1-interacting region in pVHL is located in its beta-domain, and several naturally occurring mutations located in this domain disrupt the interaction between pVHL and VDU1 protein. Co-immunoprecipitation demonstrates that VDU1 can be recruited into the pVHL-elongin C-elongin B complex. Finally, we demonstrate that VDU1 is able to be ubiquitinated via a pVHL-dependent pathway for proteasomal degradation, and VHL mutations that disrupt the interaction between VDU1 and pVHL abrogate the ubiquitination of VDU1. Our findings indicate that VDU1, a novel ubiquitin-specific processing protease, is a downstream target for ubiquitination and degradation by pVHL E3 ligase. Targeted degradation of VDU1 by pVHL could be crucial for regulating the ubiquitin-proteasome degradation pathway.