AR/androgen receptor[1]

Racemic mixtures containing becalutamide are accessible. Approximately thirty times more binding affinity of R isomer (R-bicalutamide) to AR exists than that of S isomer [1]. In a dose-dependent manner, (R)-bicalutamide (0.02 – 20 μM) decreases the survival of naive LNCaP cells [2].

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Bicalutamide is a first generation antiandrogen used to treat prostate cancer, which was approved for clinical application in 1995. Bicalutamide is available as a racemic mixture. However the R isomer (R-bicalutamide Fig. 1) has an ≈30-fold higher binding affinity to the AR than the S isomer and was therefore used in this study. Although R-bicalutamide treatment has a good effect initially, prostate tumors will lose their sensitivity to the antiandrogen and become refractory after 2–3 years. The underlying mechanism is not clear, but mutation of the AR has frequently been reported. Some hot–spot mutations have been linked to treatment with individual anti-androgens, such as T877A, with hydroxyflutamide (HF) treatment, and W741(C/L), with bicalutamide treatment.[1]

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In order to investigate the effect of different mutations of AR on the efficacy of R-bicalutamide, MD simulations, MM-GBSA binding free energy calculations and per-residue decomposition methods were performed to analyze the interaction mechanism of R-bicalutamide with WT/mutant ARs. MD simulations show that helix H12 plays a vital role for the binding between R-bicalutamide and ARs. When the B-ring of R-bicalutamide is close to helix H12, H12 will be pushed aside, which distorts the coactivator binding site and results in the inactivation of transcription. So we in silico verified that the mutations of T877A, F876L, F876L_T877A and L701H could not switch R-bicalutamide from an AR antagonist to agonist. In addition, when the B-ring of R-bicalutamide is far away from helix H12, steric clash is reduced, which is favorable for H12 closing, resulting in effective coactivator binding site formation and promoting transcription. Our data indicate that the single mutation of W741C and double mutations of W741C_T877A can switch R-bicalutamide from AR antagonist to agonist. Furthermore, the per-residue free energy decomposition shows that, once the contribution of residue M895 increases, M895, together with H12, becomes stable, which is conductive to the formation of effective coactivator binding site to facilitate transcription.[1]

In VCaP xenograft mice, (R)-Bicalutamide (10 mg/kg; ig; daily; for 4 days) exhibits antitumor efficaciousness[3].

In order to investigate the effect of different mutations of AR on the efficacy of R-bicalutamide, MD simulations, MM-GBSA binding free energy calculations and per-residue decomposition methods were performed to analyze the interaction mechanism of R-bicalutamide with WT/mutant ARs. MD simulations show that helix H12 plays a vital role for the binding between R-bicalutamide and ARs. When the B-ring of R-bicalutamide is close to helix H12, H12 will be pushed aside, which distorts the coactivator binding site and results in the inactivation of transcription. So we in silico verified that the mutations of T877A, F876L, F876L_T877A and L701H could not switch R-bicalutamide from an AR antagonist to agonist. In addition, when the B-ring of R-bicalutamide is far away from helix H12, steric clash is reduced, which is favorable for H12 closing, resulting in effective coactivator binding site formation and promoting transcription. Our data indicate that the single mutation of W741C and double mutations of W741C_T877A can switch R-bicalutamide from AR antagonist to agonist. Furthermore, the per-residue free energy decomposition shows that, once the contribution of residue M895 increases, M895, together with H12, becomes stable, which is conductive to the formation of effective coactivator binding site to facilitate transcription.[1]

Establishment of LNCaP-Rbic subclone[2]
LNCaP cells were continuously exposed to 20 μM of (R)-bicalutamide for 8 months and after only a few days LNCaP cell proliferation was strongly hampered. Such conditions persisted for the entire 8-month period, after which cells started to proliferate again and in only a few weeks a subclone of actively proliferating cells was isolated, established and called LNCaP-Rbic. This clone exhibited a faster growth than that of the parental cell line and a substantially reduced doubling time from 55 to 37...[2]

Animal/Disease Models: CD1 male nude (nu/nu) mice, with VCaP xenografts[3]
Doses: 10 mg/kg
Route of Administration: Orally gavage, daily, for 4 consecutive weeks
Experimental Results: Suppressed tumor growth.

[1]. Molecular mechanism of R-bicalutamide switching from androgen receptor antagonist to agonist induced by amino acid mutations using molecular dynamics simulations and free energy calculation. J Comput Aided Mol Des. 2016 Dec;30(12):1189-1200.

[2]. Prolonged exposure to (R)-bicalutamide generates a LNCaP subclone with alteration of mitochondrial genome. Mol Cell Endocrinol. 2014 Jan 25;382(1):314-324.

[3]. Effect of Small Molecules Modulating Androgen Receptor (SARMs) in Human Prostate Cancer Models. PLoS One. 2013; 8(5): e62657.

(R)-bicalutamide is a N-[4-cyano-3-(trifluoromethyl)phenyl]-3-[(4-fluorophenyl)sulfonyl]-2-hydroxy-2-methylpropanamide that is the (R)-enantiomer of bicalutamide. It has a role as an androgen antagonist and an antineoplastic agent. It is an enantiomer of a (S)-bicalutamide.
See also: Bicalutamide (annotation moved to).

C18H14F4N2O4S

430.37

430.061

C, 50.23; H, 3.28; F, 17.66; N, 6.51; O, 14.87; S, 7.45

113299-40-4

Bicalutamide;90357-06-5

56069

White to off-white solid powder

1.5±0.1 g/cm3

650.3±55.0 °C at 760 mmHg

178-181ºC

347.1±31.5 °C

0.0±2.0 mmHg at 25°C

1.578

4.94

2

9

5

29

750

1

C[C@](CS(=O)(=O)C1=CC=C(C=C1)F)(C(=O)NC2=CC(=C(C=C2)C#N)C(F)(F)F)O

LKJPYSCBVHEWIU-KRWDZBQOSA-N

InChI=1S/C18H14F4N2O4S/c1-17(26,10-29(27,28)14-6-3-12(19)4-7-14)16(25)24-13-5-2-11(9-23)15(8-13)18(20,21)22/h2-8,26H,10H2,1H3,(H,24,25)/t17-/m0/s1

(2R)-N-[4-cyano-3-(trifluoromethyl)phenyl]-3-(4-fluorophenyl)sulfonyl-2-hydroxy-2-methylpropanamide

(R)-Bicalutamide; 113299-40-4; (R)-Casodex; (-)-Bicalutamide; R-BICALUTAMIDE; (2R)-N-[4-CYANO-3-(TRIFLUOROMETHYL)PHENYL]-3-[(4-FLUOROPHENYL)SULFONYL]-2-HYDROXY-2-METHYLPROPANAMIDE; Bicalutamide, (R)-; Bicalutamide, (-)-;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO: 100 mg/mL (232.36 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.81 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (5.81 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (5.81 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)