Fig. 5. (R)-9b inhibits enzalutamide-resistant CRPC xenograft tumor growth in vivo.

(A) Enzalutamide-resistant C4–2B cells were injected subcutaneously in castrated male SCID mice. Once tumors were palpable, he mice were treated subcutaneously with either vehicle (Captisol; n = 6) or (R)-9b at 12 (n = 7) or 20 mg/kg (n = 7) five times a week. Tumor volumes were measured twice a week. (B and C) Once they reached ~1000 mm³ in volume, the tumors were harvested and photographed, and their weights were recorded. Tumors 4 and 5 are from the same vehicle-treated mice. (D) Weights of vehicle- and (R)-9b–treated mice were recorded. (E and F) Prostates (E) and brains (F) of the mice were harvested, and RNA was prepared, followed by qRT-PCR (n = 4 each). (G to I) Tumors were harvested, and RNA was prepared, followed by qRT-PCR of AR (G), KLK3 (H), and TMPRSS2 (I) mRNAs (n = 3 mice each). Data are represented as means ± SEM. *P < 0.05, **P < 0.01, and ***P < 0.001. NS, not significant, Student’s t test. (J) Tumor lysates (n = 3) were IP using acK609-AR or pY267-AR antibodies, followed by immunoblotting by AR antibody (top two panels). pACK1, total AR, ACK1, and actin in the tumor lysates are shown in the bottom panels. Shown below each blot is the densitometric measurement of change in abundance relative to control.