scholarly journals GREB1 amplifies androgen receptor output in prostate cancer and contributes to antiandrogen resistance

2018 ◽  
Author(s):  
Eugine Lee ◽  
John Wongvipat ◽  
Danielle Choi ◽  
Ping Wang ◽  
Deyou Zheng ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Dna Binding ◽  
Cell Lines ◽  
Target Gene ◽  
Expression Level ◽  
Genomic Alteration ◽  
Genomic Amplification ◽  
P300 Recruitment

AbstractGenomic amplification of the androgen receptor (AR) is an established mechanism of antiandrogen resistance in prostate cancer. Here we show that the magnitude of AR signaling output, independent of AR genomic alteration or expression level, also contributes to antiandrogen resistance, through upregulation of the coactivator GREB1. We demonstrate 100-fold heterogeneity in AR output within cell lines and show that cells with high AR output have reduced sensitivity to enzalutamide. Through transcriptomic and shRNA knockdown studies, together with analysis of clinical datasets, we identify GREB1 as a gene responsible for high AR output. We show that GREB1 is an AR target gene that amplifies AR output by enhancing AR DNA binding and promoting p300 recruitment. GREB1 knockdown in high AR output cells restores enzalutamide sensitivity in vivo. Thus, GREB1 is a candidate driver of enzalutamide resistance through a novel feed forward mechanism.

scholarly journals GREB1 amplifies androgen receptor output in human prostate cancer and contributes to antiandrogen resistance

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Eugine Lee ◽  
John Wongvipat ◽  
Danielle Choi ◽  
Ping Wang ◽  
Young Sun Lee ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Target Gene ◽  
Prostate Cancer Cell ◽  
Human Prostate ◽  
Genomic Alteration ◽  
Human Prostate Cancer ◽  
Genomic Amplification ◽  
Human Prostate Cancer Cell

Genomic amplification of the androgen receptor (AR) is an established mechanism of antiandrogen resistance in prostate cancer. Here, we show that the magnitude of AR signaling output, independent of AR genomic alteration or expression level, also contributes to antiandrogen resistance, through upregulation of the coactivator GREB1. We demonstrate 100-fold heterogeneity in AR output within human prostate cancer cell lines and show that cells with high AR output have reduced sensitivity to enzalutamide. Through transcriptomic and shRNA knockdown studies, together with analysis of clinical datasets, we identify GREB1 as a gene responsible for high AR output. We show that GREB1 is an AR target gene that amplifies AR output by enhancing AR DNA binding and promoting EP300 recruitment. GREB1 knockdown in high AR output cells restores enzalutamide sensitivity in vivo. Thus, GREB1 is a candidate driver of enzalutamide resistance through a novel feed forward mechanism.

scholarly journals Up-Regulation of LINC00665 Facilitates the Malignant Progression of Prostate Cancer by Epigenetically Silencing KLF2 Through EZH2 and LSD1

2021 ◽  
Vol 11 ◽  
Author(s):  
Peng Xue ◽  
Miao Yan ◽  
Kunpeng Wang ◽  
Jinbao Gu ◽  
Bing Zhong ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Cell Lines ◽  
Malignant Progression ◽  
Expression Level ◽  
Regulation Mechanism ◽  
Knock Down ◽  
Qrt Pcr ◽  
Mouse Xenograft Model ◽  
Du145 Cells

This study aimed to explore the function of LINC00665 on the proliferation and metastasis of prostate cancer (PCa), and the potential regulatory mechanisms were also investigated. The expression level of LINC00665 in 50 pairs of PCa tissues and adjacent ones was studied by qRT-PCR, and the associations between LINC00665 and clinicopathological characteristics of PCa patients were analyzed. Control group (sh-NC) and LINC00665 knock-down group (sh-LINC00665) were set in 22RV1 and DU145 cells, respectively. The biological functions of LINC00665 in PCa cell lines were assessed by CCK-8, EdU, Transwell assays, and the nude mouse xenograft model was used to evaluate the tumorigenicity in vivo. In addition, qRT-PCR, Western Blot, RIP and ChIP assays were also used to determine the regulation mechanism of LINC00665 in PCa cell lines. In this study, our results showed that LINC00665 expression level in PCa cancer tissues was significantly up-regulated, compared with that in adjacent ones. Besides, similar results were found in PCa cell lines. Knock-down of LINC00665 significantly attenuated the proliferation and migration ability in 22RV1 and DU145 cells, compared to sh-NC. Mechanically, LINC00665 could interact with EZH2 and LSD1, recruiting them to KLF2 promoter region to inhibit its transcription. Moreover, the tumor-suppressive effects mediated by sh-LINC00665 were significantly reversed through the down-regulation of KLF2. Also, the suppression of LINC00665 impaired tumor growth of PCa in vivo. In summary, LINC00665 exerted the oncogenic functions in PCa cell lines by epigenetically silencing KLF2 expression by binding to EZH2 and LSD1, illuminating a novel mechanism of LINC00665 in the malignant progression of PCa and furnishing a prospective therapeutic biomarker to combat PCa.

Effect of generation 2.5 antisense inhibitor of androgen receptor on MDV3100-resistant prostate cancer cell growth in vitro and in vivo.

2013 ◽  
Vol 31 (6_suppl) ◽  
pp. 94-94
Author(s):  
Yoshiaki Yamamoto ◽  
Eliana Beraldi ◽  
Yohann Loriot ◽  
Tianyuan Zhou ◽  
Youngsoo Kim ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Cell Growth ◽  
Cell Lines ◽  
Splice Variants ◽  
Full Length ◽  
Castration Resistant Prostate Cancer ◽  
Exon 1 ◽  
Regulated Gene Expression

94 Background: MDV3100 is a potent androgen receptor (AR) antagonist with activity in castration resistant prostate cancer (CRPC); however, progression to MDV3100-resistant (MDV-R) CRPC frequently occurs with rising serum PSA levels, implicating AR full length or variants in disease progression. We studied the activity of Generation 2.5 antisense oligonucleotide (ASO) targeting the AR full length (ARfl) and splice variants in MDV-R CRPC models. Methods: and Results: ThreeASOs targeting exon 1, intron 1, or exon 8 were designed to suppress ARfl and known AR splice variants. We generated by selection MDV-R LNCaP-derived sub-lines that uniformly expressed high levels of both ARfl and AR-V7 compared to CRPC LNCaP cell lines. MDV-3100 induced time- and dose-dependent increases in ARfl and AR-V7 protein levels; ARfl levels were ~20-fold higher than AR-V7. All 3 AR-ASO decreased ARfl and PSA expression. Exon 1 ASO decreased expression of both ARfl and AR-V7 in MDV-R-LNCaP cells; in contrast, exon 8 ASO decreased ARfl without reducing AR-V7 levels. Exon 1 ASO also most potently suppressed ARfl and splice variants in M12 cells stably overexpressing AR splice variants AR-V7 and AR-V567es. Despite these differential effects on ARfl and splice variant knockdown, the AR ASO similarly inhibited cell growth and induced apoptosis and G1 cell cycle arrest in LNCaP-derived CRPC and MDV-R cell lines. In 22RV-1 cells (which express endogenous ARfl and AR-V7), exon 1 ASO more potently suppressed ARfl and AR-V7 levels, AR transcriptional activity and AR-regulated gene expression compared to exon 8 ASO, but inhibition of cell growth did not differ significantly. Exon 1 ASO was evaluated in vivo in MDV-R49F CRPC LNCaP xenografts; mean tumor volume and serum PSA levels decreased significantly by 40% and 50%, respectively, compared to controls. Conclusions: While MDV-3100 induces both ARfl and AR-V7 levels, the biologic consequences appear cell line dependent and mainly driven by ARfl. AR-ASO knockdown of ARfl and its splice variants suppresses MDV-R LNCaP tumor growth, providing pre-clinical proof of principle to support clinical evaluation in post-AR pathway inhibitor CRPC.

ARV-110: An oral androgen receptor PROTAC degrader for prostate cancer.

2019 ◽  
Vol 37 (7_suppl) ◽  
pp. 259-259 ◽  
Author(s):  
Taavi Neklesa ◽  
Lawrence B Snyder ◽  
Ryan R Willard ◽  
Nicholas Vitale ◽  
Jennifer Pizzano ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Target Gene ◽  
Point Mutations ◽  
Genetic Alterations ◽  
Significant Inhibition ◽  
Castration Resistant Prostate Cancer ◽  
Orally Bioavailable

259 Background: The Androgen Receptor (AR) remains the principal driver of castration-resistant prostate cancer during the transition from a localized to metastatic disease. Most patients initially respond to inhibitors of the AR pathway, but the response is often relatively short-lived. The majority of patients progressing on enzalutamide or abiraterone exhibit genetic alterations in the AR locus, either in the form of amplifications or point mutations in the AR gene. Given these mechanisms of resistance, our goal is to eliminate the AR protein using the PROteolysis TArgeting Chimera (PROTAC) technology. Methods: Here we report an orally bioavailable small molecule AR PROTAC degrader, ARV-110, that promotes ubiquitination and degradation of AR. This molecule has been characterized in in vitro degradation and functional assays, and DMPK, toxicology and preclinical efficacy studies. Results: ARV-110 robustly degrades AR in all cell lines tested, with an observed half-maximal degradation concentration (DC50) of ~1 nM. ARV-110 treatment leads to highly selective AR degradation, as demonstrated by proteomic studies. In VCaP cells, PROTAC-mediated AR degradation suppresses the expression of the AR-target gene PSA, inhibits AR-dependent cell proliferation, and induces apoptosis at low nanomolar concentrations. Further, ARV-110 degrades clinically relevant mutant AR proteins and retains activity in a high androgen environment. In mouse xenograft studies, greater than 90% AR degradation is observed at a 1 mg/kg PO QD dose. Significant inhibition of tumor growth and AR signaling has been achieved in LNCaP, VCaP and prostate cancer patient derived xenograft (PDX) models. Notably, ARV-110 demonstrates in vivo efficacy and reduction of AR-target gene expression in a long term, castrate, enzalutamide-resistant VCaP tumor model. Conclusions: In summary, we report preclinical data on an orally bioavailable AR PROTAC degrader, ARV-110, that demonstrates efficacy in multiple prostate cancer models. ARV-110 has completed IND-enabling studies and FIH studies are planned for 1Q2019.

scholarly journals Development of a Transcriptional Amplification System Based on the PEG3 Promoter to Target Androgen Receptor-Positive and -Negative Prostate Cancer Cells

2019 ◽  
Vol 20 (1) ◽  
pp. 216
Author(s):  
Pallavi Jain ◽  
Pier-Luc Clermont ◽  
Francis Desmeules ◽  
Amina Zoubeidi ◽  
Bertrand Neveu ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Cell Lines ◽  
Cancer Management ◽  
Castration Resistant ◽  
Genetic Modifications ◽  
Amplification System ◽  
Transcriptional Amplification

Localized prostate cancer (PCa) is often curable, whereas metastatic disease treated by castration inevitably progresses toward castration-resistant PCa (CRPC). Most CRPC treatments target androgen receptor (AR) signaling. However, not all CRPC cells rely on AR activity for survival and proliferation. With advances in immunotherapy and fluid biopsies for cancer management, expression systems specific for both AR-positive and -negative PCa are required for virus-based vaccines and cell imaging. To target both AR-responsive and non-responsive cells, we developed a three-step transcriptional amplification (3STA) system based on the progression elevated gene-3 (PEG3) promoter named PEG3AP1-3STA. Notably, we report on different genetic modifications that significantly improved PEG3 promoter’s strength in PCa cells. Adenoviruses incorporating PEG3 promoter with and without transcriptional amplification systems were generated. The potential of PEG3AP1-3STA to target PCa cells was then evaluated in vitro and in vivo in androgen-responsive and non-responsive PCa cell lines. PEG3AP1-3STA was shown to be active in all PCa cell lines and not regulated by androgens, and its activity was amplified 97-fold compared to that of a non-amplified promoter. The PEG3AP1-3STA system can thus be used to target advanced AR+ and AR− cells for imaging or immunovirotherapy in advanced PCa.

An oral androgen receptor PROTAC degrader for prostate cancer.

2017 ◽  
Vol 35 (6_suppl) ◽  
pp. 273-273 ◽  
Author(s):  
Taavi K Neklesa ◽  
Lawrence B Snyder ◽  
Mark Bookbinder ◽  
Xin Chen ◽  
Andrew P Crew ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Cell Lines ◽  
Genetic Alterations ◽  
Tumor Growth Inhibition ◽  
Castration Resistant Prostate Cancer ◽  
Null Cell ◽  
Orally Bioavailable

273 Background: The Androgen Receptor (AR) remains the principal driver of castration-resistant prostate cancer during the transition from a localized to metastatic disease. Most patients initially respond to inhibitors of the AR pathway, but the response is often short-lived. The majority of patients progressing on enzalutamide or abiraterone exhibit genetic alterations in the AR locus, either in the form of amplifications or point mutations in the AR gene. Given these mechanisms of resistance, our goal is to eliminate the AR protein using the PROteolysis TArgeting Chimera (PROTAC) technology. Further, we sought to make an orally bioavailable AR PROTAC. Methods: Medicinal chemistry efforts yielded a small molecule AR PROTAC that simultaneously binds E3-ubiquitin ligase and AR, thus leading to ubiquitination and degradation of AR. This molecule has been characterized in in vitro and in vivo preclinical studies. Results: Our lead oral AR PROTAC degrades 92-98% of total AR in all cell lines tested, with 50% degradation concentration (DC50) < 1 nM. AR degradation suppresses the expression of AR-target gene PSA, inhibits cell proliferation, and induces potent apoptosis in VCaP cells. No activity is observed in AR-null cell lines, such as PC-3. While enzalutamide loses its activity in the presence of elevated androgens, AR PROTAC maintains its antiproliferative activity. Further, AR PROTAC is able to degrade all clinically relevant mutant AR proteins. A robust oral bioavailability is observed across multiple species and overall ADME properties are encouraging. Approximately 95% AR degradation is observed in AR-amplified VCaP xenografts at doses as low as 10 mg/kg. Congruent with AR degradation, a dose responsive tumor growth inhibition is observed in AR-dependent xenograft studies. Conclusions: In summary, we report the first orally bioavailable AR PROTAC that robustly degrades AR in vitro and in vivo.

scholarly journals Inhibition of Androgen Receptor Transcriptional Activity as a Novel Mechanism of Action of Arsenic

2009 ◽  
Vol 23 (3) ◽  
pp. 412-421 ◽  
Author(s):  
Adena E. Rosenblatt ◽  
Kerry L. Burnstein
Keyword(s):  
Prostate Cancer ◽  
Androgen Receptor ◽  
Male Infertility ◽  
Transcriptional Activity ◽  
Target Gene ◽  
Nuclear Translocation ◽  
Therapeutic Option ◽  
Protein Levels

Abstract Environmental sodium arsenite is a toxin that is associated with male infertility due to decreased and abnormal sperm production. Arsenic trioxide (ATO), another inorganic trivalent semimetal, is an effective therapy for acute promyelocytic leukemia, and there is investigation of its possible efficacy in prostate cancer. However, the mechanism of arsenic action in male urogenital tract tissues is not clear. Because the androgen receptor (AR) plays an important role in spermatogenesis and prostate cancer, we explored the possibility that trivalent arsenic regulates AR function. We found that arsenic inhibited AR transcriptional activity in prostate cancer and Sertoli cells using reporter gene assays testing several androgen response element-containing regions and by assessing native target gene expression. Arsenic inhibition of AR activity was not due to down-regulation of AR protein levels, decreased hormone binding to AR, disruption of AR nuclear translocation, or interference with AR-DNA binding in vitro. However, chromatin immunoprecipitation studies revealed that arsenic inhibited AR recruitment to an AR target gene enhancer in vivo. Consistent with a deficiency in AR-chromatin binding, arsenic disrupted AR amino and carboxyl termini interaction. Furthermore, ATO caused a significant decrease in prostate cancer cell proliferation that was more pronounced in cells expressing AR compared with cells depleted of AR. In addition, inhibition of AR activity by ATO and by the AR antagonist, bicalutamide, was additive. Thus, arsenic-induced male infertility may be due to inhibition of AR activity. Further, because AR is an important target in prostate cancer therapy, arsenic may serve as an effective therapeutic option.

scholarly journals Novel DNA Motif Binding Activity Observed In Vivo With an Estrogen Receptor α Mutant Mouse

2014 ◽  
Vol 28 (6) ◽  
pp. 899-911 ◽  
Author(s):  
Sylvia C. Hewitt ◽  
Leping Li ◽  
Sara A. Grimm ◽  
Wipawee Winuthayanon ◽  
Katherine J. Hamilton ◽  
...  
Keyword(s):  
Dna Binding ◽  
Target Gene ◽  
Mutant Mouse ◽  
Estrogen Receptor Α ◽  
Binding Activity ◽  
Estrogen Response Element ◽  
Dna Binding Domain ◽  
Estrogen Response ◽  
Uterine Growth

Abstract Estrogen receptor α (ERα) interacts with DNA directly or indirectly via other transcription factors, referred to as “tethering.” Evidence for tethering is based on in vitro studies and a widely used “KIKO” mouse model containing mutations that prevent direct estrogen response element DNA- binding. KIKO mice are infertile, due in part to the inability of estradiol (E2) to induce uterine epithelial proliferation. To elucidate the molecular events that prevent KIKO uterine growth, regulation of the pro-proliferative E2 target gene Klf4 and of Klf15, a progesterone (P4) target gene that opposes the pro-proliferative activity of KLF4, was evaluated. Klf4 induction was impaired in KIKO uteri; however, Klf15 was induced by E2 rather than by P4. Whole uterine chromatin immunoprecipitation-sequencing revealed enrichment of KIKO ERα binding to hormone response elements (HREs) motifs. KIKO binding to HRE motifs was verified using reporter gene and DNA-binding assays. Because the KIKO ERα has HRE DNA-binding activity, we evaluated the “EAAE” ERα, which has more severe DNA-binding domain mutations, and demonstrated a lack of estrogen response element or HRE reporter gene induction or DNA-binding. The EAAE mouse has an ERα null–like phenotype, with impaired uterine growth and transcriptional activity. Our findings demonstrate that the KIKO mouse model, which has been used by numerous investigators, cannot be used to establish biological functions for ERα tethering, because KIKO ERα effectively stimulates transcription using HRE motifs. The EAAE-ERα DNA-binding domain mutant mouse demonstrates that ERα DNA-binding is crucial for biological and transcriptional processes in reproductive tissues and that ERα tethering may not contribute to estrogen responsiveness in vivo.

Preclinical Quantification of Prostate Cancer-Associated Vascular Alterations in the Bone Microenvironment in vivo

2020 ◽  
Vol 61 (6) ◽  
pp. 188-200
Author(s):  
Malte Schroeder ◽  
Lennart Viezens ◽  
Jördis Sündermann ◽  
Svenja Hettenhausen ◽  
Gerrit Hauenherm ◽  
...  
Keyword(s):  
Prostate Cancer ◽  
Blood Flow ◽  
Flow Velocity ◽  
Cell Lines ◽  
Blood Flow Velocity ◽  
Tumour Growth ◽  
Prostate Cancer Cell ◽  
Bone Microenvironment ◽  
Prostate Cancer Cell Lines

Introduction: Prostate cancer has a special predilection to form bone metastases. Despite the known impact of the microvascular network on tumour growth and its dependence on the organ-specific microenvironment, the characteristics of the tumour vasculature in bone remain unknown. Methods: The cell lines LNCaP, DU145, and PC3 were implanted into the femurs of NSG mice to examine the microvascular properties of prostate cancer in bone. Tumour growth and the functional and morphological alterations of the microvasculature were analysed for 21 days in vivo using a transparent bone chamber and fluorescence microscopy. Results: Vascular density was significantly lower in tumour-bearing bone than in non-tumour-bearing bone, with a marked loss of small vessels. Accelerated blood flow velocity led to increased volumetric blood flow per vessel, but overall perfusion was not affected. All of the prostate cancer cell lines had similar vascular patterns, with more pronounced alterations in rapidly growing tumours. Despite minor differences between the prostate cancer cell lines associated with individual growth behaviours, the same overall pattern was observed and showed strong similarity to that of tumours growing in soft tissue. Discussion: The increase in blood flow velocity could be a specific characteristic of prostate cancer or the bone microenvironment.

scholarly journals Cisplatin loaded multiwalled carbon nanotubes reverse drug resistance in NSCLC by inhibiting EMT

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Yuxin Qi ◽  
Wenping Yang ◽  
Shuang Liu ◽  
Fanjie Han ◽  
Haibin Wang ◽  
...  
Keyword(s):  
Lung Cancer ◽  
Carbon Nanotubes ◽  
Drug Resistance ◽  
Western Blot ◽  
Multiwalled Carbon Nanotubes ◽  
Cell Lines ◽  
Nude Mice ◽  
Expression Level ◽  
Multiwalled Carbon

Abstract Background Lung cancer is one of the important health threats worldwide, of which 5-year survival rate is less than 15%. Non-small-cell lung cancer (NSCLC) accounts for about 80% of all lung cancer with high metastasis and mortality. Methods Cisplatin loaded multiwalled carbon nanotubes (Pt-MWNTS) were synthesized and used to evaluate the anticancer effect in our study. The NSCLC cell lines A549 (cisplatin sensitive) and A549/DDP (cisplatin resistant) were used in our in vitro assays. MTT was used to determine Cancer cells viability and invasion were measured by MTT assay and Transwell assay, respectively. Apoptosis and epithelial-mesenchymal transition related marker proteins were measured by western blot. The in vivo anti-cancer effect of Pt-MWNTs were performed in male BALB/c nude mice (4-week old). Results Pt-MWNTS were synthesized and characterized by X-ray diffraction, Raman, FT-IR spectroscopy and scan electron microscopy. No significant cytotoxicity of MWNTS was detected in both A549/DDP and A549 cell lines. However, Pt-MWNTS showed a stronger inhibition effect on cell growth than free cisplatin, especially on A549/DDP. We found Pt-MWNTS showed higher intracellular accumulation of cisplatin in A549/DDP cells than free cisplatin and resulted in enhanced the percent of apoptotic cells. Western blot showed that application of Pt-MWNTS can significantly upregulate the expression level of Bax, Bim, Bid, Caspase-3 and Caspase-9 while downregulate the expression level of Bcl-2, compared with free cisplatin. Moreover, the expression level of mesenchymal markers like Vimentin and N-cadherin was more efficiently reduced by Pt-MWNTS treatment in A549/DDP cells than free cisplatin. In vivo study in nude mice proved that Pt-MWNTS more effectively inhibited tumorigenesis compared with cisplatin, although both of them had no significant effect on body weight. Conclusion Pt-MWNT reverses the drug resistance in the A549/DDP cell line, underlying its possibility of treating NSCLC with cisplatin resistance.

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