icaritin and Prostatic-Neoplasms

Prostate cancer (PCa) is one of the most common health-related issues in the male individuals of western countries. Icaritin (ICT) is a traditional Chinese herbal medicine that exhibits antitumor efficacy in variety of cancers including PCa. However, the precise function and detailed molecular mechanism of ICT in the regression of PCa remain unclear. Ubiquitin-conjugating enzyme E2C (UBE2C) is an anaphase-promoting complex/cyclosome (APC/C)-specific ubiquitin conjugating enzyme, which acts as an oncogene in PCa progression. The function of ICT in PCa was investigated in transgenic adenocarcinoma mouse prostate (TRAMP) mice using survival analysis, hematoxylin and eosin (HE) staining, TUNEL assay, and immunohistochemistry and in human PCa cell lines using various molecular techniques and functional assays including plasmid construction and transfection. Bioinformatic analyses were performed to identify the interaction between miRNA and UBE2C via the TargetScan algorithm. We demonstrated that ICT inhibited the development and progression of PCa in TRAMP mice by improving the survival rate and tumor differentiation. Furthermore, we found that ICT could significantly inhibit cell proliferation and invasion and induce apoptosis in PCa cells. Consistently, downregulation of UBE2C suppressed the proliferation and invasion of PCa cells. Moreover, a luciferase reporter assay confirmed that UBE2C was a direct target of miR-381-3p. Meanwhile, ICT simultaneously downregulated UBE2C expression and upregulated miR-381-3p levels in human PCa cells. Altogether, our findings provide a strong rationale for the clinical application of ICT as a potential oncotherapeutic agent against PCa via a novel molecular mechanism of regulating the miR-381-3p/UBE2C pathway.

Topics: Animals; Apoptosis; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Disease Models, Animal; Flavonoids; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Humans; Male; MicroRNAs; Prostatic Neoplasms; RNA Interference; Ubiquitin-Conjugating Enzymes; Xenograft Model Antitumor Assays

Neuroendocrine prostate cancer (NEPC) has a poor prognosis, with a median survival of less than 1 year after diagnosis. Following androgen deprivation therapy, prostate adenocarcinoma cells have been observed to develop an androgen receptor-negative, terminally differentiated and indolent neuroendocrine-like phenotype. However, several molecular events, including interleukin 6 (IL-6) stimulation, in the prostate microenvironment result in the appearance of aggressive, highly proliferative castrate-resistant NEPC. In this study, we examined the mechanistic effects of a natural prenylflavonoid, icaritin (ICT), on neuroendocrine differentiation in IL-6-induced LNCaP cells and NEPC development in the male transgenic adenocarcinoma of the mouse prostate (TRAMP) model. TRAMP mice received daily intraperitoneal injection of ICT or vehicle. ICT induced apoptosis in prostate tumor, suppressed NEPC development and, accordingly, improved overall survival in TRAMP mice. Expression of neuroendocrine markers (synaptophysin) and androgen receptor in TRAMP mice and neuroendocrine-like LNCaP cells were inhibited by ICT. Suppression of neuroendocrine and NEPC development by ICT was associated with dose-dependent inhibitory effects on abnormally elevated IL-6/STAT3 and Aurora kinase A in vitro and in vivo Since ICT demonstrated favorable pharmacokinetic and safety profiles with marked enrichment in prostate tissues, our study provides evidence for the development of prenylflavonoid as a multimodal therapeutic agent against NEPC.

Topics: Animals; Aurora Kinase A; Carcinoma, Neuroendocrine; Cell Differentiation; Flavonoids; Gene Expression Regulation, Neoplastic; Humans; Interleukin-6; Male; Mice; Mice, Transgenic; Prostatic Neoplasms; Receptors, Androgen; Signal Transduction; STAT3 Transcription Factor; Synaptophysin; Tumor Microenvironment

We aimed to investigate whether icaritin (ICT) would inhibit serum proinflammatory cytokines and postpone prostate cancer (PCa) development and progression in both normal diet and high-fat diet (HFD) transgenic adenocarcinoma mouse prostate (TRAMP) mice.. TRAMP mice were randomly divided into four groups: normal diet with/without ICT group and HFD with/without ICT group. Each TRAMP mouse received intraperitoneal injection of ICT solution at the dose of 30 mg/kg 5 times per week.. ICT treatment could significantly increase the survival when compared with those in normal diet group (P = 0.015, log-rank test) and HFD group (P = 0.009, log-rank test). Proinflammatory cytokine levels, including IL-1α, IL-1β, IL-6, and TNF-α, were decreased more or less in ICT-treated TRAMP mice. Moreover, significant higher inflammation scores were detected in normal diet group and HFD group compared with their relevant ICT treatment groups (P = 0.026 and P = 0.006, respectively). Meanwhile, the incidences of well-differentiated tumor tissue in two ICT treatment groups (39.13 and 31.82 %) were moderately higher than control groups (29.41 and 20.00 %, respectively), though no significant difference was observed.. Taken together, our findings indicate that ICT could inhibit the development and progression of PCa in TRAMP mice via inhibiting proinflammatory cytokines.

Topics: Adenocarcinoma; Animals; Antineoplastic Agents; Cytokines; Diet, High-Fat; Disease Models, Animal; Disease Progression; Flavonoids; Inflammation; Male; Mice; Mice, Transgenic; Prostatic Neoplasms; Signal Transduction

Human epidermal growth factor receptor type 2 (HER2) and androgen receptor (AR) are critical factors for prostate cancer (PCa) progression. These factors regulate tumor cell survival and proliferation, and remain as crucial drivers of castration-resistant PCa progression. Icaritin (ICT) is a prenyl flavonoid derived from the Epimedium genus, which has many biological and pharmacological effects. Using androgen-sensitive human prostate carcinoma LNCaP cell lines, we found that 35 μg/ml of ICT could inhibit more than 50% of cell proliferation, induce cell apoptosis, and lead to a strong G1 phase arrest by targeting cyclin-related proteins and suppressing the ability of cell invasion. Moreover, ICT exerts its potent anticancer efficacy by inducing polyomavirus enhancer activator 3 (PEA3) to inhibit the aberrantly activated HER2/AR signaling. In addition, after PEA3 expression was silenced by specific small-interference RNA, we found that both the ICT-inhibited effect on LNCaP cell proliferation and the ICT-induced cell apoptosis rate decreased. These results provide alternative mechanisms for the antitumor actions of ICT, indicating that ICT might be a promising therapeutic agent, as well as a preventive agent, for hormone therapy-resistant PCa.

Topics: Adenovirus E1A Proteins; Cell Line, Tumor; Flavonoids; Humans; Male; Prostatic Neoplasms; Prostatic Neoplasms, Castration-Resistant; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-ets; Receptor, ErbB-2; Receptors, Androgen; Signal Transduction

Persistent androgen receptor (AR) signaling is the key driving force behind progression and development of castration-resistant prostate cancer (CRPC). In many patients, AR COOH-terminal truncated splice variants (ARvs) play a critical role in contributing to the resistance against androgen depletion therapy. Unfortunately, clinically used antiandrogens like bicalutamide (BIC) and enzalutamide (MDV), which target the ligand binding domain, have failed to suppress these AR variants. Here, we report for the first time that a natural prenylflavonoid, icaritin (ICT), can co-target both persistent AR and ARvs. ICT was found to inhibit transcription of key AR-regulated genes, such as KLK3 [prostate-specific antigen (PSA)] and ARvs-regulated genes, such as UBE2C and induce apoptosis in AR-positive prostate cancer (PC) cells. Mechanistically, ICT promoted the degradation of both AR and ARvs by binding to arylhydrocarbon-receptor (AhR) to mediate ubiquitin-proteasomal degradation. Therefore, ICT impaired AR transactivation in PC cells. Knockdown of AhR gene restored AR stability and partially prevented ICT-induced growth suppression. In clinically relevant murine models orthotopically implanted with androgen-sensitive and CRPC cells, ICT was able to target AR and ARvs, to inhibit AR signaling and tumor growth with no apparent toxicity. Our results provide a mechanistic framework for the development of ICT, as a novel lead compound for AR-positive PC therapeutics, especially for those bearing AR splice variants.

Topics: Animals; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Flavonoids; Gene Expression Regulation, Neoplastic; Humans; Kallikreins; Male; Mice, Inbred NOD; Molecular Targeted Therapy; Prostate-Specific Antigen; Prostatic Neoplasms; Protein Stability; Receptors, Androgen; Receptors, Aryl Hydrocarbon; RNA Splicing; Signal Transduction; Ubiquitin; Ubiquitin-Conjugating Enzymes; Xenograft Model Antitumor Assays

Icariin and icaritin with prenyl group have been demonstrated for their selective estrogen receptor modulating activities. We screened their effects on cell growth in human prostate carcinoma PC-3 cell line (estrogen receptor positive) in vitro. PC-3 cell line was used for the measurement of anti-carcinoma activities of 0-100 micromol/l icaritin and 30 micromol/l icariin. 1 micromol/l 17-beta estradiol (E(2)) served as the estrogen positive control, and 1 micromol/l ICI 182,780 [7 alpha-[9 (4,4,5,5,5-pentafluoropentyl) sulfinyl] nonyl]-estra-1,3,5(10)-triene-3,17h-diol]] served as the specific estrogen receptor antagonist. Primary cultured rat prostate basal cells used as cell growth selective control. The growth-inhibitory effects were analyzed using MTT assay, and fluorochrome staining, flow cytometry, and immunoblotting were employed to illustrate the possible mechanisms. When treated with icaritin for 24 to 72 h, cell growth was strongly inhibited (at 48 h IC(50) was 10.74+/-1.59 micromol/l, P<0.001) companied with a mitochondrial transmembrane potential (_Psim) drop. Meanwhile, few changes in IC(50) could be observed when co-incubated with ICI 182,780. Icaritin-induced growth inhibition was associated with G(1) arrest (P<0.05), and G(2)-M arrest depending upon doses. Consistently with G(1) arrest, icaritin increased protein expressions of pRb, p27(Kip1) and p16(Ink4a), while showed decrease in phosphorylated pRb, Cyclin D1 and CDK4. Comparatively, icariin has much lower effects on PC-3 cells and showed only weak G(1) arrest, suggesting a possible structure-activity relationship. These findings suggested a novel anticancer efficacy of icaritin mediated selectively via induction of cell cycle arrest but not associated with estrogen receptors in PC-3 cells.

Topics: Antineoplastic Agents, Phytogenic; Blotting, Western; Carcinoma; Cell Cycle; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Estradiol; Flavonoids; Flow Cytometry; G1 Phase; G2 Phase; Humans; Immunoblotting; Inhibitory Concentration 50; Male; Membrane Potential, Mitochondrial; Prostatic Neoplasms