Methyltrienolone r1881

Manufactured by PerkinElmer

Sourced in United States

Methyltrienolone (R1881) is a synthetic androgen receptor agonist commonly used in research applications. It functions as a high-affinity ligand for the androgen receptor, which plays a role in the regulation of gene expression related to male sexual characteristics and other physiological processes.

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9 protocols using methyltrienolone r1881

Molecular Reagents and Plasmids used in Study

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Methyltrienolone (R1881) (Perkin-Elmer, Waltham, MA, USA) was dissolved in ethanol and Bicalutamide (Sigma-Aldrich, St Louis, MO, USA) was dissolved in dimethyl sulfoxide and stored at −20 °C. pCMV6-XL5-PDCD4 (SC111794) was purchased from OriGene Technologies Inc. (Rockville, MD, USA). pCMV6-EV was generated by PDCD4 removal and self-ligation. FlexiTube siRNA were purchased from Qiagen (Valencia, CA, USA). Details of siRNAs, shRNAs, plasmids, and miRNAs are provided in Supplementary Table S1.

Zennami K., Choi S.M., Liao R., Li Y., Dinalankara W., Marchionni L., Rafiqi F.H., Kurozumi A., Hatano K, & Lupold S.E. (2018). PDCD4 is an androgen-repressed tumor suppressor that regulates prostate cancer growth and castration resistance. Molecular cancer research : MCR, 17(2), 618-627.

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Preparation of R1881 Compound

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All chemicals were obtained from Sigma-Aldrich and of analytical grade unless otherwise noted (MO, USA). Methyltrienolone (R1881) was obtained from PerkinElmer (MA, USA).

Narain N.R., Diers A.R., Lee A., Lao S., Chan J.Y., Schofield S., Andreazi J., Ouro-Djobo R., Jimenez J.J., Friss T., Tanna N., Dalvi A., Wang S., Bunch D., Sun Y., Wu W., Thapa K., Gesta S., Rodrigues L.O., Akmaev V.R., Vishnudas V.K, & Sarangarajan R. (2016). Identification of Filamin-A and -B as potential biomarkers for prostate cancer. Future Science OA, 3(1), FSO161.

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Molecular Analysis of NF-κB Signaling

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Anti-AR (N-20), anti-NF-κB (sc-109), and anti-GAPDH (6c5) antibodies and an anti-p-NF-κB antibody (Ser536) were purchased from Santa Cruz Biotechnology and Cell Signaling Technology, respectively. We obtained methyltrienolone (R1881) from PerkinElmer, and HF and CDDP from Sigma-Aldrich.

Kashiwagi E., Ide H., Inoue S., Kawahara T., Zheng Y., Reis L.O., Baras A.S, & Miyamoto H. (2016). Androgen receptor activity modulates responses to cisplatin treatment in bladder cancer. Oncotarget, 7(31), 49169-49179.

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Synergistic Androgen and FOXM1 Inhibition

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Specific antibodies including Anti-FOXM1 and anti-AR were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). The synergistic androgen methyltrienolone (R1881) was purchased from PerkinElmer (Walthan, MA). The antiandrogen compound Casodex and FOXM1 inhibitor Siomycin A were purchased from Sigma Aldrich (St. Louis, MO, USA).

Liu Y., Liu Y., Yuan B., Yin L., Peng Y., Yu X., Zhou W., Gong Z., Liu J., He L, & Li X. (2017). FOXM1 promotes the progression of prostate cancer by regulating PSA gene transcription. Oncotarget, 8(10), 17027-17037.

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LNCaP Cells with Regulated AR-V7 Expression

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LNCaP cells were purchased from the American Type Culture Collection (Manassas, VA). The LNCaP-AR-V7-pHage cell line is an LNCaP cell line with TET (doxycycline) regulated AR-V7 expression generated in our lab [11 (link)]. Parental LNCaP and LNCaP-AR-V7-pHage cells were maintained RPMI and 10% serum as previously described [11 (link), 47 (link)]. Methyltrienolone (R1881) was purchased from Perkin-Elmer (Boston, MA). [U-13C]-glucose, [1-13C]-glutamine, and [5-13C]-glutamine were purchased from Cambridge Isotope Laborartories, Inc (Andover, MA). Palmitic acid (1-¹⁴C) and oleic acid (1-¹⁴C) were purchased from Elmer (Waltham, MA)

Shafi A.A., Putluri V., Arnold J.M., Tsouko E., Maity S., Roberts J.M., Coarfa C., Frigo D.E., Putluri N., Sreekumar A, & Weigel N.L. (2015). Differential regulation of metabolic pathways by androgen receptor (AR) and its constitutively active splice variant, AR-V7, in prostate cancer cells. Oncotarget, 6(31), 31997-32012.

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Receptor Binding and Metal Complex Assays

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Methyltrienolone (R1881) was purchased from PerkinElmer (Waltham, MA) and dissolved in DMSO. TTM, DSF, PDTC, BCS, CuSO₄, NAC and LGR were obtained from Sigma (St Louis, MO). TTM, DSF and PDTC were dissolved in DMSO while BCS, CuSO₄, NAC and LGR in water. EUK118 was from Cayman Chemical (Ann Arbor, MI). Copper-64 was prepared at Washington University; St. Louis, MO.

Safi R., Nelson E.R., Chitneni S.K., Franz K.J., George D.J., Zalutsky M.R, & McDonnell D.P. (2014). Copper Signaling Axis as a Target for Prostate Cancer Therapeutics. Cancer research, 74(20), 5819-5831.

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Analytical Protocol for Persistent Organic Pollutants

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The following chemicals were purchased from AccuStandard: 1,1,1-Trichloro-2, 2-bis-(p-chlorophenyl)ethane (p,p’-DDT), 1,1,1-trichloro-2,4-bis-(o-chlorophenyl)ethane (o,p’-DDT), 1,1- dichloro-2,2-bis-(chlorophenyl) ethane (p,p’-DDD), 1,1-dichloro- 2,2-bis-(chlorophenyl) ethylene (p,p’-DDE), 1,1-dichloro-2, 4-bis-(chlorophenyl) ethylene (o,p’-DDE), 1,1-dichloro-2,4-bis-(chlorophenyl) ethane (o,p’-DDD), surrogate standards PCB-67 and PCB-191, and internal standards PCB-30 and PCB-82. The performance reference compounds (PRCs) ¹³C-o,p’-DDD and ¹³C- o,p’-DDE and p,p’-DDT-d₈, o,p’-DDT-d₈, p,p’-DDD-d₈, and p,p’-DDE-d₈ were purchased from Cambridge Isotope Laboratories (Tewksbury, MA) or C/D/N Isotopes (Pointe-Claire, Canada). Chemicals known to activate estrogen receptor (ER) (17β-estradiol), Aryl-hydrocarbon Receptor (AhR) (PCB126), Glucocorticoid Receptor (GR) (dexamethasone) and inhibit the Androgen Receptor (anti-AR) (cyproterone acetate) were purchased at the highest purity available from Sigma-Aldrich (St. Louis, MO). The AR ligand (methyltrienolone, R1881) was purchased from Perkin Elmer. Solvents and other chemicals were of analytical grade and purchased from Fisher Scientific. Deionized water with electrical resistivity of 18.2 M /cm was prepared using a Barnstead E-pure system.

Crago J., Xu E.G., Kupsco A., Jia F., Mehinto A.C., Lao W., Maruya K.A., Gan J, & Schlenk D. (2016). Trophic transfer and effects of DDT in male hornyhead turbot (Pleuronichthys verticalis) from Palos Verdes Superfund site, CA (USA) and comparisons to field monitoring. Environmental pollution (Barking, Essex : 1987), 213, 940-948.

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Molecular Mechanisms of DNA Repair Regulation

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Methyltrienolone (R1881) and enzalutamide were obtained from Perkin Elmer and Axon Medchem respectively. Camptothecin, dihydrotestosterone (DHT), DMSO, doxycycline, bicalutamide, hydroxyl urea and Mirin were obtained from Sigma. Olaparib was purchased from LC laboratories and ATM inhibitor KU0055399 from Calbiochem. Cell culture media, foetal bovine serum (FBS) and other cell culture reagents including antibiotics were obtained from Life Technologies. Antibodies were obtained from commercial suppliers as below and used at recommended dilutions for each application:

Phospho-Histone H2A.X (Ser139) (JBW301; cat # 05-636) from Millipore—used in foci analysis, үH2A.X (phosphoS139; cat # ab2893) from Abcam used for immuno histochemistry.

Rad51 (cat no. H-92; sc8349) from Santa Cruz Biotechnology.

Mre11 (cat no. 4895 S) from Cell Signaling Technology.

PAR antibody (cat no. GTX75054 from Source Bioscience).

AR (AR441; cat no. M356201-2 from Dako).

Beta-actin antibody (Rabbit actin: Cell Signaling Technology (cat no. 4970).

Mouse actin: Abcam (cat no. ab6276), 1:5000).

IgG-Alexa 555 from Life Technologies.

PARP1, H-250 Cat. no. sc-7150 from Santa Cruz Biotechnology.

Asim M., Tarish F., Zecchini H.I., Sanjiv K., Gelali E., Massie C.E., Baridi A., Warren A.Y., Zhao W., Ogris C., McDuffus L.A., Mascalchi P., Shaw G., Dev H., Wadhwa K., Wijnhoven P., Forment J.V., Lyons S.R., Lynch A.G., O’Neill C., Zecchini V.R., Rennie P.S., Baniahmad A., Tavaré S., Mills I.G., Galanty Y., Crosetto N., Schultz N., Neal D, & Helleday T. (2017). Synthetic lethality between androgen receptor signalling and the PARP pathway in prostate cancer. Nature Communications, 8, 374.

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G6PD Regulation and Glucose Metabolism

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6AN and tamoxifen were purchased from Sigma-Aldrich (St Louis, MO, USA). Methyltrienolone (R1881) and ¹⁴C-labeled glucose (glucose, D-[¹⁴C(U)]) were purchased from PerkinElmer (Waltham, MA, USA). Rapamycin was purchased from Cell Signaling (Danvers, MA, USA). Anti-GAPDH antibody was from Sigma-Aldrich. Anti-G6PD, anti-pS6 Ser235/236 and anti-S6 antibodies were purchased from Cell Signaling. The NADP/NADPH red fluorescence quantitation assay was from e-ENZYME (Gaithersburg, MD, USA). G6PD siRNAs and Universal siRNA control were purchased from Sigma-Aldrich. Hoechst and MitoSOX were from Life Technologies (Carlsbad, CA, USA).

Tsouko E., Khan A.S., White M.A., Han J.J., Shi Y., Merchant F.A., Sharpe M.A., Xin L, & Frigo D.E. (2014). Regulation of the pentose phosphate pathway by an androgen receptor–mTOR-mediated mechanism and its role in prostate cancer cell growth. Oncogenesis, 3(5), e103-.

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