Eugene Lin

Regulation of Androgen Receptor and Prostate Cancer Growth by Cyclin-dependent Kinase 5

Prostate cancer is the most frequently diagnosed male malignancy. The normal prostate development and prostate cancer progression are mediated by androgen receptor (AR). Recently, the roles of cyclin-dependent kinase 5 (Cdk5) and its activator, p35, in cancer biology are explored one after another. We have previously demonstrated that Cdk5 may regulate proliferation of thyroid cancer cells. In addition, we also identify that Cdk5 overactivation can be triggered by drug treatments and leads to apoptosis of prostate cancer cells. The aim of this study is to investigate how Cdk5 regulates AR activation and growth of prostate cancer cells. At first, the data show that Cdk5 enables phosphorylation of AR at Ser-81 site through direct biochemical interaction and, therefore, results in the stabilization of AR proteins. The Cdk5-dependent AR stabilization causes accumulation of AR proteins and subsequent activation. Besides, the positive regulations of Cdk5-AR on cell growth are also determined in vitro and in vivo. S81A mutant of AR diminishes its interaction with Cdk5, reduces its nuclear localization, fails to stabilize its protein level, and therefore, decreases prostate cancer cell proliferation. Prostate carcinoma specimens collected from 177 AR-positive patients indicate the significant correlations between the protein levels of AR and Cdk5 or p35. These findings demonstrate that Cdk5 is an important modulator of AR and contributes to prostate cancer growth. Therefore, Cdk5-p35 may be suggested as diagnostic and therapeutic targets for prostate cancer in the near future.

Cyclin-dependent kinase 5 modulates STAT3 and androgen receptor activation through phosphorylation of Ser727 on STAT3 in prostate cancer cells

Cyclin-dependent kinase 5 (Cdk5) is known to regulate prostate cancer metastasis. Our previous results indicated that Cdk5 activates androgen receptor (AR) and supports prostate cancer growth. We also found that STAT3 is a target of Cdk5 in promoting thyroid cancer cell growth, whereas STAT3 may play a role as a regulator to AR activation under cytokine control. In this study, we investigated the regulation of Cdk5 and its activator p35 on STAT3/AR signaling in prostate cancer cells. Our results show that Cdk5 biochemically interacts with STAT3 and that this interaction depends on Cdk5 activation in prostate cancer cells. The phosphorylation of STAT3 at Ser⁷²⁷ (p-Ser⁷²⁷-STAT3) is regulated by Cdk5 in cells and xenograft tumors. The mutant of STAT3 S727A reduces its interaction with Cdk5. We further show that the nuclear distribution of p-Ser⁷²⁷-STAT3 and the expression of STAT3-regulated genes (junB, c-fos, c-myc, and survivin) are regulated by Cdk5 activation. STAT3 mutant does not further decrease cell proliferation upon Cdk5 inhibition, which implies that the role of STAT3 regulated by Cdk5 correlates to cell proliferation control. Interestingly, Cdk5 may regulate the interaction between STAT3 and AR through phosphorylation of Ser⁷²⁷-STAT3 and therefore upregulate AR protein stability and transactivation. Correspondingly, clinical evidence shows that the level of p-Ser⁷²⁷-STAT3 is significantly correlated with Gleason score and the levels of upstream regulators (Cdk5 and p35) as well as downstream protein (AR). In conclusion, this study demonstrates that Cdk5 regulates STAT3 activation through Ser⁷²⁷ phosphorylation and further promotes AR activation by protein-protein interaction in prostate cancer cells.

The significance of Her2 on androgen receptor protein stability in the transition of androgen requirement in prostate cancer cells.

Androgen ablation therapy is the most common strategy for suppressing prostate cancer progression; however, tumor cells eventually escape androgen dependence and progress to an androgen-independent phase. The androgen receptor (AR) plays a pivotal role in this transition. To address this transition mystery in prostate cancer, we established an androgen-independent prostate cancer cell line (LNCaPdcc), by long-term screening of LNCaP cells in androgen-deprived conditions, to investigate changes of molecular mechanisms before and after androgen withdrawal. We found that LNCaPdcc cells displayed a neuroendocrine morphology, less aggressive growth, and lower expression levels of cell cycle-related factors, although the cell cycle distribution was similar to parental LNCaP cells. Notably, higher protein expression of AR, phospho-Ser(81)-AR, and PSA in LNCaPdcc cells were observed. The nuclear distribution and protein stability of AR increased in LNCaPdcc cells. In addition, cell proliferation results exhibited the biphasic nature of the androgen (R1881) effect in two cell lines. On the other hand, LNCaPdcc cells expressed higher levels of Her2, phospho-Tyr(1221/1222)-Her2, ErbB3, and ErbB4 proteins than parental LNCaP cells. These two cell lines exhibited distinct responses to Her2 activation (by heregulin treatment) on Her2 phosphorylation and Her2 inhibition (by AG825 or Herceptin treatments) on proliferation. In addition, the Her2 inhibitor more effectively caused AR degradation and diminished AR Ser(81) phosphorylation in LNCaPdcc cells. Taken together, our data demonstrate that Her2 plays an important role in the support of AR protein stability in the transition of androgen requirement in prostate cancer cells. We hope these findings will provide novel insight into the treatment of hormone-refractory prostate cancer.

Retinoic Acid Induces Apoptosis of Prostate Cancer DU145 Cells through Cdk5 Overactivation

Retinoic acid (RA) has been believed to be an anticancer drug for a long history. However, the molecular mechanisms of RA actions on cancer cells remain diverse. In this study, the dose-dependent inhibition of RA on DU145 cell proliferation was identified. Interestingly, RA treatment triggered p35 cleavage (p25 formation) and Cdk5 overactivation, and all could be blocked by Calpain inhibitor, Calpeptin (CP). Subsequently, RA-triggered DU145 apoptosis detected by sub-G1 phase accumulation and Annexin V staining could also be blocked by CP treatment. Furthermore, RA-triggered caspase 3 activation and following Cdk5 over-activation were destroyed by treatments of both CP and Cdk5 knockdown. In conclusion, we report a new mechanism in which RA could cause apoptosis of androgen-independent prostate cancer cells through p35 cleavage and Cdk5 over-activation. This finding may contribute to constructing a clearer image of RA function and bring RA as a valuable chemoprevention agent for prostate cancer patients.

Inhibitory effects of Rhenium-188-labeled Herceptin on prostate cancer cell growth: A possible radioimmunotherapy to prostate carcinoma

Abstract Purpose: Herceptin is widely used in treating Her2-overexpressing breast cancer. However, the application of Herceptin in prostate cancer is still controversial. Our previous results have indicated the relevance of Her2 in the transition of the androgen requirement in prostate cancer cells. In this study, the effects of radioimmunotherapy against Her2 in prostate cancer were investigated. Materials and methods: DU145, an androgen receptor-negative prostate cancer cell line, was used in vitro and in vivo to evaluate the effects of Herceptin labeled with a beta emitter, Rhenium-188 (Re-188). Its effects on cell growth, extent of apoptosis, the bio-distribution of Re-188 labeled Herceptin (Re-H), and protein levels were determined. Results: Treatments with Re-188 and Re-H reduced the proliferation of DU145 cells in dose- and time-dependent manners compared to the Herceptin-treated group. Growth inhibition and apoptosis were induced after Re-H treatment; growth inhibition was more distinct in cells with high Her2/p-Her2 levels. Our in vivo xenograft studies revealed that Re-H treatment significantly retarded tumor growth and altered the levels of apoptosis-related proteins. The bio-distribution of Re-H in mice demonstrated a tissue-specific pattern. Importantly, the levels of p35 protein, which is related to cancer cell survival and invasion, dramatically decreased after Re-H treatment. Conclusions: Our data demonstrate that Re-188-labeled Herceptin effectively inhibited the growth of DU145 cells compared to the Herceptin- and Re-188-treated cohorts. This implies that targeting Her2 by both radio- and immuno- therapy might be a potential strategy for treating patients with androgen-independent prostate cancer.

Comparison between Monopolar and Bipolar TURP in Treating Benign Prostatic Hyperplasia: 1-Year Report

Purpose. The major complication of traditional monopolar TURP is water intoxication due to absorption of distilled water during extended resection. A newly developed bipolar electrocautery device which uses saline irrigation decreases the risk of TUR syndrome. The aim of this study was to evaluate the efficacy and safety of this bipolar system. Methods. From April to October 2003, a total of 40 patients aged 58 to 86 years (mean: 69.3 yr) were randomized into two comparable groups; one underwent bipolar TURP and the other received monopolar TURP. Pre-operative and post-operative serum sodium concentration, hemoglobin level, resection time, blood loss and weight of specimen were evaluated. Qmax, I-PSS and bladder neck contracture were reassessed at 6-month and 1-year follow-ups. Results. There were no significant differences in hemoglobin level, blood loss, resection time or weight of specimen between the two groups. However, the difference in serum sodium level after operation was significantly greater in the monopolar group (p＜0.001). Conclusions. The bipolar electrocautery device is safer than the traditional monopolar device for the TURP procedure; the bipolar device decreases the risk of TUR syndrome and produces results similar to those obtained with the monopolar device at 6-month and 1 year follow-ups.

Cdk5 Directly Targets Nuclear p21CIP1 and Promotes Cancer Cell Growth

The significance of Cdk5 in cell-cycle control and cancer biology has gained increased attention. Here we report the inverse correlation between the protein levels of Cdk5 and p21CIP1 from cell-based and clinical analysis. Mechanistically, we identify that Cdk5 overexpression triggers the proteasome-dependent degradation of p21CIP1 through a S130 phosphorylation in a Cdk2-independent manner. Besides, the evidence from cell-based and clinical analysis shows that Cdk5 primarily regulates nuclear p21CIP1 protein degradation. S130A-p21CIP1 mutant enables to block either its protein degradation or the increase of cancer cell growth caused by Cdk5. Notably, Cdk5-triggered p21CIP1 targeting primarily appears in S-phase, while Cdk5 overexpression increases the activation of Cdk2 and its interaction with DNA polymerase δ. The in vivo results show that Cdk2 might play an important role in the downstream signaling to Cdk5. In summary, these findings suggest that Cdk5 in a high expression status promotes cancer growth by directly and rapidly releasing p21CIP1-dependent cell-cycle inhibition and subsequent Cdk2 activation, which illustrates an oncogenic role of Cdk5 potentially applied for future diagnosis and therapy. Cancer Res; 76(23); 6888-900. ©2016 AACR.

Intravesical Migration of Intrauterine Device Resulting in Bladder Stone: Report of a Case

Foreign bodies may be introduced into the bladder iatrogenically, via migration from adjacent organs, as in our patient or by penetrating injuries.1 Transuterine migration of an intrauterine device into the urinary bladder is a rare complication. Herein we report a 48-year-old female who complained of intermittent hematuria with frequent urinary tract infection (UTI) for 2 months. An IUD with secondary stone formation was found by plain abdominal radiography. The bladder stone was fragmented by an electrohydraulic lithotriptor and the IUD was removed by grasping forceps. The postoperative course was uneventful.

The Efficacy of a Combined Pneumatic/Ultrasound Device in Percutaneous Nepholithotripsy

Purpose. Percutaneous nephrolithotripsy (PCNL) is widely accepted as the procedure of choice for removing large, complex renal calculi. A ”2 in 1” pneumatic/ultrasonic intracorporeal lithotriptor has been developed for PCNL procedures. It combines the stone clearing efficiency of an ultrasonic device with the fragmentation strength of a pneumatic probe into a single handheld unit. We present our clinical experience with this device in a prospective randomized comparison of the combined lithotriptor ”2 in 1” ”Swiss Lithoclast Master” with individual ultrasonic and pneumatic devices in performing lithotripsy. Methods. A total of 40 consecutive patients aged 48 to 71 years (mean: 59.9 yr; 28 men and 12 women) underwent PCNL to treat symptomatic calculi from April 2003 to August 2004. Patients were randomized into two comparable groups; 20 patients in group one underwent PCNL with the combined pneumatic/ultrasonic device while 20 patients in group two underwent PCNL with individual ultrasonic and pneumatic components. The stone burden was assessed before and after the operative procedure. The stone clearance rate in mm^2 per minute was calculated for these two groups. Complications and stone-free rates were also compared between the groups. Results. There were no significant differences in stone location or stone composition between the two groups of patients. The duration of the operation for complete stone clearance was considerably less with the combined device (40.5±10.1 vs 65.5±24.7 mm, p=0.01). Furthermore, the stone clearance rate was much higher with the new ”2-in-1” lithotriptor (23.5±10.1 vs 12.4±5.0 mm^2/min, p=0.006). Hydrothorax was noted in one patient who underwent treatment with the ”Swiss Lithoclast Master”, but it was not attributed to the device. Conclusions. The new combined lithotriptor ”2 in 1” ”Swiss Lithoclast Master” is able to disintegrate and extract stone material more rapidly than individual ultrasonic and pneumatic devices. The Swiss Lithoclast Master appears to be efficacious and safe for removing large renal calculi.