Inhibition of miR-20a by pterostilbene facilitates prostate cancer cells killed by NK cells via up-regulation of NKG2D ligands and TGF-ß1down-regulation.

Natural killer (NK) cells play a potent role in antitumor immunity via spontaneously eliminating tumor directly. However, some tumors such as prostate cancer constantly escape this immune response by down-regulating cell surface molecule recognition and/or secreting immune impressive cytokines. Here, we found pterostilbene, a natural agent with potent anticancer activity, could enhance expression of major histocompatibility complex class I chain-related proteins A and B (MICA/B) on prostate cancer cells surface, which are ligands of the natural killer group 2 member D (NKG2D) expressed by NK cells, and inhibit TGF-ß1 secretion by prostate cancer cells. Further, we discovered that these effects were caused by inhibition of miR-20a in prostate cancer cells by pterostilbene. MiR-20a could target the 3' untranslated region (UTR) of MICA/B, resulting in their expression down-regulation. Inhibition of TGF-ß1 function by its specific antibody attenuated its impairment to NKG2D on NK cells. Finally, we observed that pterostilbene-treated prostate cancer cells were more easily to be killed by NK cells. Taken together, our findings demonstrated inhibition of miR-20a by pterostilbene in prostate cancer cells could increase MICA/B expression and decrease TGF-ß1 secretion, which enhanced NK cell-mediated cytotoxicity againt prostate cancer cells, suggesting a potential approach for increasing anti-prostate cancer immune.

4-1BB antibody enhances cytotoxic activity of natural killer cells against prostate cancer cells via NKG2D agonist combined with IL-27.

Aims: To enhance the cytotoxicity of natural killer (NK) cells against prostate cancer cells via NKG2D agonist, with 4-1BB antibody and IL-27 combination. Materials & methods: FACS was used to detect degranulation and cell surface receptors in NK cells isolated from healthy donors. Cytokine concentrations were measured using ELISA. NK-cell cytotoxicity was analyzed using Cell Counting Kit-8. Results: NKG2D agonist, 4-1BB antibody and IL-27 combination treatment improved the activating receptor expression and IFN-γ and TNF-α secretion but decreased the suppressive receptor CD158a expression and IL-10 secretion in NK cells. The combined treatment enhanced NK-cell cytotoxicity against both PC3 and DU145 cells with concurrent enhanced STAT3 activation. Conclusion: 4-1BB antibody and IL-27 improved NKG2D agonist function in NK cells against prostate cancer cells.

Efficacy and Adverse Reaction of Tislelizumab Combined with Chemotherapy on Urothelial Carcinoma / 肿瘤防治研究

Objective To evaluate the efficacy and safety of tislelizumab combined with chemotherapy in the treatment of urothelial carcinoma in the real word. Methods We enrolled 32 patients with urothelial carcinoma who were treated with tislelizumab and chemotherapy (gemcitabine/cisplatin or paclitaxel). The incidence of treatment-related adverse reactions during treatment and the efficacy evaluation were statistically analyzed. Results All patients were divided into two groups: 15 patients in the tislelizumab combined with paclitaxel group and 17 patients in the tislelizumab combined with GC group. Among 24 efficacy-evaluable patients, the ORR was 54.2% and the DCR was 83.3%. The ORR were 50.0% and 58.3%, and the DCR were 75.0% and 91.7% in the tislelizumab combined with paclitaxel group and the tislelizumab combined with GC group respectively. Common treatment-related adverse reactions included anemia (56.3%), loss of appetite (53.1%) and skin pruritus (50.0%). The grade 3-4 treatment-related adverse events occurred in 21.8% of patients. Common immune-related adverse reactions included skin toxicity (53.1%) and immune colitis (9.4%). Conclusion Tislelizumab combined with chemotherapy on urothelial cancer has significant curative effect, safety and controllability, but attention should be paid to immune-related adverse reactions.

Prostaglandin E2 Inhibits Prostate Cancer Progression by Countervailing Tumor Microenvironment-Induced Impairment of Dendritic Cell Migration through LXRα/CCR7 Pathway.

Migration and homing of dendritic cells (DCs) to lymphoid organs are quite crucial for T cell-induced immune response against tumor. However, tumor microenvironment can make some tumor cells escape immune response by impairing DC migration. Prostaglandin E2 (PGE2) plays important roles in initiating and terminating inflammatory responses. In this study, we investigated whether PGE2 could inhibit murine prostate cancer progression by countervailing tumor microenvironment-induced impairment of dendritic cell migration. We found that murine prostate cancer cell line RM-1-conditioned medium impaired chemotactic movement of marrow-derived DCs and splenic cDCs toward CC chemokine receptor-7 (CCR7) ligand CCL19 in vitro and migration to draining lymph gland in vivo. Meanwhile, it also induced LXRα activation and CCR7 inhibition on maturing DCs. However, the treatment of PGE2 rescued this impairment of DC migration with upregulation of CCR7 and inhibition of LXRα. Further, it was observed that PGE2 also increased MMP9 expression and activated Notch1 signaling on DCs. In RM-1-bearing mouse model, PGE2 treatment was identified to inhibit tumor growth and induce more tumor-infiltrating T cells and CD11c dendritic cells in tumor sites. Therefore, our findings may demonstrate a new perspective for therapeutic interventions on prostate cancer immunoescape.

Liver X receptor activation inhibits PC-3 prostate cancer cells via the beta-catenin pathway.

BACKGROUND: Liver X receptors (LXRs) are nuclear receptors family of ligand-dependent transcription factors that play a crucial role in regulating cholesterol metabolism and inflammation. Recent studies show that LXR agonists exhibit anti-cancer activities in a variety of cancer cell lines including prostate. To further identify the potential mechanisms of LXRα activation on prostate cancer, we investigated the effect of LXR agonist T0901317 on PC3 prostate cancer cell and in which activity of beta-catenin pathway involved. METHODS: Prostate cancer PC3 cells were transfected with LXR-a siRNA and treated with LXR activator T0901317. qRT-PCR and western blot were used to detect the LXR-a expression. beta-catenin, cyclin D1 and c-MYC were analyzed by western blot. Cell apoptosis was examined by flow cytometry and Cell proliferation was assessed by Cell Counting Kit-8 assay. Cell migration was detected by Transwell chambers. RESULTS: Data showed that T0901317 significantly inhibited PC3 cell proliferation as well as invasion and increased apoptosis in vitro. Furthermore, we found that LXRα activation induced the reduction of beta-catenin expression in PC3 cells, and this inhibitory effect could be totally abolished when cells were treated with LXRα. Meanwhile, the expression of beta-catenin target gene cyclin D1 and c-MYC were also decreased. CONCLUSIONS: This study provided additional evidence that LXR activation inhibited PC-3 prostate cancer cells via suppressing beta-catenin pathway.

Potent anti-prostate cancer immune response induced by dendritic cells transduced with recombinant adenoviruses encoding 4-1BBL combined with cytokine-induced killer cells.

AIM: To test the effect of dendritic cells (DCs) transduced with recombinant adenoviruses encoding 4-1BBL combined with cytokine-induced killer cells (CIKs) against prostate cancer. METHOD: Flow cytometry was used to detect the surface markers of the co-cultured cells, and cytotoxicity against prostate cancer cells in vitro as well as antitumor activities in vivo were observed. RESULTS: Our results showed that Ad-4-1BBL-transduced DCs could increase percentage of CD3(+)CD56(+) cells in CIKs, and CIKs co-cultured with Ad-4-1BBL-transduced DCs could augment the secretion of IL-12 and IFN-γ and decrease TGF-ß production. In addition, Ad-4-1BBL-transduced DCs enhanced the cytotoxicity of CIKs against prostate cancer and resulted in inhibition of tumor growth and tumor-bearing animals' survival. CONCLUSION: These results demonstrate that 4-1BBL-engineered DCs can improve CIKs cytotoxicity against prostate cancer cells.

Effects of transplantation with marrow-derived mesenchymal stem cells modified with survivin on renal ischemia-reperfusion injury in mice.

PURPOSE: To determine whether renal injury induced by ischemia-reperfusion (I/R) could be further improved by mesenchymal stem cells (MSCs) modified with survivin. MATERIALS AND METHODS: Lentiviral vectors were used to introduce the survivin gene into MSCs and the MSCs modified with survivin were transplanted into established mice models of renal I/R injury. Seven days later, serum creatinine (Scr) and blood urea nitrogen (BUN) were measured and the survival of MSCs was determined. Hematoxylin and eosin staining was used to assess renal pathological change. The expressions of hepatocyte growth factor (HGF) and basic fibroblast growth factor (bFGF) in kidney tissue were detected by western blot. RESULTS: Mice transplanted with survivin-modified MSCs demonstrated good renal function recovery with Scr and BUN decline close to normal levels and improvement of renal I/R injury repair. Additionally, the survival of transplanted MSCs modified with survivin was enhanced and the expression of HGF and bFGF in kidney tissue was increased. CONCLUSION: Our results demonstrated that MSCs engineered to over-express survivin could enhance their therapeutic effect on renal I/R injury in mice, probably via the improved survival ability of MSCs and increased production of protective cytokines in ischemic tissue.

Enhanced function of cytotoxic T lymphocytes induced by dendritic cells modified with truncated PSMA and 4-1BBL.

Interactions between costimulatory molecules and their receptors are vital for Ag-presenting dendritic cells (DCs) to initiate T cells activation, expansion and their antitumor immune responses. Augmentation of costimulatory signal due to the interaction of DCs and T cells may amplify, sustain and drive diversity of cytotoxic T lymphocytes (CTLs) and consequently enhance the antitumor response. 4-1BBL/4-1BB is such a pair of costimulatory ligand and receptor, playing an important role in the co-stimulation of CTLs. Previously, we demonstrated that DCs transduced with recombinant adenovirus encoding truncated PSMA (tPSMA) and m4-1BBL could induce prostate cancer regression in mouse models. In the present study, we further explored the adjuvant role of 4-1BBL in modulating CTLs activation induced by tPSMA gene-pulsed DCs. The apoptosis and cytotoxicity against tPSMA expressing RM-1 cells of CTLs were determined. Results showed that tPSMA gene-pulsed DCs effectively induced T lymphocyte activation and cytotoxicity, which was enhanced by upregulated expression of 4-1BBL, displaying better cell viability, lower CTLs apoptosis, higher expression anti-apoptotic protein of Bcl-xL and phosphorylation of P38, enhanced NF-κB activation, as well as more IFN-γ production. These results demonstrated that 4-1BBL may play a significant role in the co-stimulation pathway for Ag-presenting DCs-mediated CTLs activity, which might be a beneficial adjuvant factor for DCs-based cancer immunotherapy.

4-1BB protects dendritic cells from prostate cancer-induced apoptosis.

It has been shown that human prostate cancer (PCa) cells induced apoptotic death of the most potent antigen-presenting cells, dendritic cells (DCs), which are responsible for the induction of specific antitumor immune responses. Here, we investigated the function of 4-1BB on protecting DCs from prostate cancer-induced apoptosis with an agonistic mAb to 4-1BB. RM-1 cells and DCs were co-incubated for 48 h and DC apoptosis was assessed by Annexin Vassay. TNF-α and IL-12 production were assessed by enzyme-linked immunosorbent assay (ELISA) and Bcl-2 and Bcl-xL on DCs were analyzed by Western blot. We have shown that co-incubation of RM-1 cells with DCs is accompanied by an increased level of DCs apoptosis. Triggering 4-1BB on DCs resulted in increased resistance of DCs to RM-1 cells-induced apoptosis, which was owing to the up-regulated expression of Bcl-2 and Bcl-xL, and increased secretion of TNF-αand IL-12. These results demonstrate that triggering 4-1BB on DCs could increased resistance of DCs to PCa-induced apoptosis.

Combination immunotherapy with 4-1BBL and CTLA-4 blockade for the treatment of prostate cancer.

Immune regulation has been shown to be involved in the progressive growth of some murine tumours. Interruption of immune regulatory pathways via activation of 4-1BB or cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) blockade appears to be a promising strategy for cancer immunotherapy. In this study, we examined the effectiveness of 4-1BBL-expressing tumor cell vaccine in combination with CTLA-4 blockade on rejection of murine prostate cancer RM-1. We found that the combination of both a vaccine consisting of 4-1BBL-expressing RM-1 cells and CTLA-4 blockade resulted in regression of RM-1 tumors and a significant increase in survival of the tumour cell recipients, compared to that of either treatment alone. The combined vaccination resulted in higher CTL against RM-1 cells and increased secretion of IFN-γ, TNF-α, and IL-2 in the mix-cultured supernatant. These results suggest that combining activation of 4-1BB and blockade of CTLA-4 may offer a new strategy for prostate cancer immunotherapy.

Anti-tumor effects induced by gene vaccines co-expressing truncated human prostate specific membrane antigen gene and mouse 4-1BBL / 中华泌尿外科杂志

Objective To investigate the influence of m4-1BBL on the anti-tumor effects induced by truncated human prostate specific membrane antigen (tPSMA) gene in mice. Methods A eukaryotic expression plasmid encoding tPSMA and m4-1BBL (pDC316-tPSMA-IRES-m4-1BBL), pDC316-tPSMA and pDC316 were constructed. C57BL/6 mice were vaccinated in the quadriceps femoris, respectively. The CTL activity of spleen cells from the immunized mice against prostate cancer RM-1-tPSMA was detected by CCK-8 kit in vitro. The tumor growth was then observed. Results The target cell specific cytotoxicity rate induced by pDC316-tPSMA-IRES-m4-1BBL was 42.6%, compared to 24.8% in the pDC316-tPSMA group and 10.8% in the pDC316 group. The difference was significant (P<0.05). The volume of tumor in the pDC316 group was 2657.4mm3 7 d after vaccination, compared to 1334.5 mm3 in the pDC316-tPSMA group, 9 d after vaccination. In the pDC316-tPSMA-IRES-m4-1BBL group, the tumor volume was 445.8 mm3, 12d after vaccination. The difference was significant (P<0.05). Conclusion Gene vaccines co-expressing tPSMA gene and m4-1BBL gene could significantly enhance anti-prostate cancer effects in mice.

The change of immunoactivity of dendritic cells induced by mouse 4-1BBL recombinant adenovirus.

PURPOSE: The purpose of this study is to construct a recombinant adenovirus vector carrying mouse 4-1BBL and observe its effects in dendritic cells. MATERIALS AND METHODS: Mouse 4-1BBL cDNA was taken from the plasmid pcDNA3-m4- 1BBL and subcloned into adenovirus shuttle plasmid pAdTrack-CMV, and then transformed into competent BJ5183 with plasmid pAdEasy-1. After recombination in E.coli, Ad-4-1BBL was packaged and amplified in HEK 293 cells. The expression of 4-1BBL in Ad-4-1BBL-transfected mouse prostate cancer cell line RM-1 was detected by reverse transcription polymerase chain reaction (RT-PCR) and Western blot. After the co-culture of dendritic cells (DCs) with Ad-4-1BBL-transfected RM-1 cells, interleukin (IL)-6 and IL-12 production were assessed by enzyme-linked immunosorbent assay (ELISA) and co-stimulatary molecules (CD80 and CD86) on DCs were analyzed by flow cytometry. RESULTS: The levels of IL-6 (3,960 pg/mL) and IL-12 (249 pg/mL) production in Ad-m4-1BBL-pulsed DCs were more than those in none-pulsed DCs. The differences were statistically significant (p < 0.05). The expression of co-stimulatary molecules (CD80 and CD86) was up-regulated in Ad-m4-1BBL-pulsed DCs. CONCLUSION: The results indicated the recombinant mouse 4-1BBL can effectively activate DCs.