ASC-J9(®) suppresses castration resistant prostate cancer progression via degrading the enzalutamide-induced androgen receptor mutant AR-F876L.

Androgen deprivation therapy (ADT) with the newly developed powerful anti-androgen enzalutamide (Enz, also known as MDV3100) has promising therapeutic effects to suppress castration resistant prostate cancer (CRPC) and extending patients' lives an extra 4.8 months. However, most Enz therapy eventually fails with the development of Enz resistance. The detailed mechanisms how CRPC develops Enz resistance remain unclear and may involve multiple mechanisms. Among them, the induction of the androgen receptor (AR) mutant AR-F876L in some CRPC patients may represent one driving force that confers Enz resistance. Here, we demonstrate that the AR degradation enhancer, ASC-J9(®), not only degrades wild-type AR, but also has the ability to target AR-F876L. The consequence of suppressing AR-F876L may then abrogate AR-F876L mediated CRPC cell proliferation and metastasis. Thus, developing ASC-J9(®) as a new therapeutic approach may represent a novel therapy to better suppress CRPC that has already developed Enz resistance.

Wang R ，Lin W ，Lin C ，Li L ，Sun Y ，Chang C ... -  《-》

Preclinical Study using Malat1 Small Interfering RNA or Androgen Receptor Splicing Variant 7 Degradation Enhancer ASC-J9(®) to Suppress Enzalutamide-resistant Prostate Cancer Progression.

While androgen-deprivation-therapy with the recently developed antiandrogen enzalutamide (Enz) shows promising therapeutic benefits in men with metastatic castration-resistant prostate cancer (PCa), many patients develop resistance to Enz, which may involve the induction of the androgen receptor (AR) splicing variant 7 (AR-v7). Our aim is to identify the mechanisms responsible for AR-v7 production and to develop novel preclinical approaches to suppress the Enz-resistant (EnzR) PCa. We established EnzR-PCa cell lines and examined the long noncoding RNA Malat1 (Malat1) function in conferring Enz resistance. We also examined the in vivo effects of Malat1 short interfering RNA and the AR-v7 degradation enhancer, ASC-J9®. Enz resistance and expression of Malat1 and AR-v7. All statistical comparisons were analyzed with a t-test or one way analysis of variance followed by t-test. We demonstrated that Malat1 is indispensable for Enz-induced AR-v7 production in VCaP and EnzR-C4-2 cells. We observed increased AR-v7 and Malat1 expression in our established EnzR-PCa cell lines and in some PCa patients who received Enz treatment. Targeting the Malat1/AR-v7 axis resulted in altering the PCa resistance to androgen deprivation therapy with Enz. The limitation of this study includes the small sample size from the same human patients before and after receiving Enz treatment. Targeting the Malat1/AR-v7 axis via Malat1-short interfering RNA or AR-v7 degradation enhancer ASC-J9® in EnzR-PCa cell lines and mouse models suppressed EnzR-PCa progression. Androgen deprivation therapy-enzalutamide treatment may not be the best choice for prostate cancer patients who have higher expression of the Malat1/androgen receptor splicing variant 7 axis, and new therapies using Malat1-short interfering RNA or ASC-J9® may be developed in the future to better suppress enzalutamide-resistant prostate cancer.

Wang R ，Sun Y ，Li L ，Niu Y ，Lin W ，Lin C ，Antonarakis ES ，Luo J ，Yeh S ，Chang C ... -  《-》

Natural killer cells suppress enzalutamide resistance and cell invasion in the castration resistant prostate cancer via targeting the androgen receptor splicing variant 7 (ARv7).

Despite the success of androgen-deprivation therapy (ADT) with the newly developed anti-androgen enzalutamide (Enz, also known as MDV3100) to suppress castration resistant prostate cancer (CRPC) in extending patient survival by an extra 4.8 months, eventually patients die with the development of Enz resistance that may involve the induction of the androgen receptor (AR) splicing variant ARv7. Here we identify an unrecognized role of Natural Killer (NK) cells in the prostate tumor microenvironment that can be better recruited to the CRPC cells to suppress ARv7 expression resulting in suppressing the Enz resistant CRPC cell growth and invasion. Mechanism dissection revealed that CRPC cells, compared to normal prostate epithelial cells, could recruit more NK cells that might then lead to alterations of the microRNA-34 and microRNA-449 to suppress both ARv7 expression and ARv7-induced EZH2 expression to suppress CRPC cell invasion. Together, these results identify a new potential therapy using recruited NK cells to better suppress the Enz resistance and cell invasion in CRPC at the later enzalutamide resistant stage.

Lin SJ ，Chou FJ ，Li L ，Lin CY ，Yeh S ，Chang C ... -  《-》

A novel androgen receptor antagonist JJ-450 inhibits enzalutamide-resistant mutant AR(F876L) nuclear import and function.

Castration-resistant prostate cancer can develop resistance to enzalutamide because of androgen receptor (AR) point mutations, AR overexpression, constitutively active AR splice variants, and/or elevated intratumoral androgen synthesis. The point mutation ARF876L was reported to be stimulated, instead of inhibited, by enzalutamide, thus contributing to enzalutamide resistance. We have recently developed JJ-450 as a novel AR antagonist with the potential to treat enzalutamide-resistant castration-resistant prostate cancer (CRPC). We employed several assays to determine the impact of JJ-450 and enzalutamide on prostate cancer cell lines expressing green fluorescent protein (GFP)-ARF876L . These assays include a prostate-specific antigen enhancer/promoter-based luciferase assay to determine AR transcriptional activity, a quantitative real-time polymerase chain reaction assay, and Western blot analysis to detect expression of AR-target genes at the messenger RNA and protein level, fluorescence microscopy to show AR subcellular localization, and a 5-bromo-2'-deoxyuridine assay to measure prostate cancer cell proliferation. As expected, enzalutamide inhibited wild-type (WT) AR but not ARF876L transcriptional activity in the luciferase assay. In contrast, JJ-450 inhibited both WT-AR and ARF876L transcriptional activity to a similar extent. Also, enzalutamide retarded androgen-induced nuclear import of GFP-AR, but not GFP-ARF876L , whereas JJ-450 retarded nuclear import of both GFP-AR and GFP-ARF876L . To further evaluate JJ-450 inhibition of ARF876L , we stably transfected C4-2 cells separately with GFP-AR or GFP-ARF876L . Enzalutamide inhibited endogenous AR-target gene expression in C4-2-GFP-ARWT , but not in the C4-2-GFP-ARF876L subline, whereas JJ-450 inhibited AR-target gene expression in both C4-2 sublines. More importantly, enzalutamide inhibited proliferation of C4-2-GFP-ARWT , but not of the C4-2-GFP-ARF876L subline, whereas JJ-450 inhibited proliferation of both C4-2 sublines. JJ-450 inhibits enzalutamide-resistant ARF876L mutant nuclear translocation and function. Our findings suggest that JJ-450 and its analogs should be further developed to provide a potential new approach for the treatment of enzalutamide-resistant CRPC.

Wu Z ，Wang K ，Yang Z ，Pascal LE ，Nelson JB ，Takubo K ，Wipf P ，Wang Z ... -  《-》

Androgen-deprivation therapy with enzalutamide enhances prostate cancer metastasis via decreasing the EPHB6 suppressor expression.

Early studies suggested that using ADT with the recently developed anti-androgen Enzalutamide (Enz, also named as MDV3100 could extent castration resistant prostate cancer (CRPC) patients' survival an extra 4.8 months. Yet the therapy in most patients might eventually fail due to development of Enz-resistance. Here we found Enz might also increase some unwanted side-effects via increasing the CRPC cell invasion that might involve altering the Enz-mediated androgen receptor (AR)/EPHB6 suppressor/JNK signaling. Results from multiple clinical surveys also indicated that EPHP6 might function as a suppressor of PCa metastasis. Mechanism dissection revealed that Enz-mediated AR might function via binding to the androgen-response-element (ARE) on the EPHB6 promoter to decrease EPHB6 suppressor expression, which might then activate the phosphorylation of JNK signals to increase the CRPC cell invasion. Targeting this newly identified AR/EPHB6/JNK signaling with JNK inhibitor (SP600125) may then block/reverse the Enz-increased CRPC cell invasion. Collectively, our results suggest that Enz may increase CRPC cell invasion via altering the AR/EPHB6/JNK/MMP9 signaling and targeting this newly identified signaling may help us to increase the Enz efficacy to better suppress the CRPC at the later metastatic stage.

Chen J ，Li L ，Yang Z ，Luo J ，Yeh S ，Chang C ... -  《-》