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| Cooperative Effects of AR and mTOR Inhibition In Vitro and In Vivo (A) In vitro response of Pten null;Ar+ murine (CaP8) and human (LNCaP) prostate cancer cells to AR knockdown (sh-AR) or pharmacological inhibition of AR (MDV3100, 10 mM) with and without rapamycin (R: 1 nM) treatment (Sc, control sh oligo). (B and D) In vivo response to treatments with castration, MDV3100, rapamycin, or their combinations as measured by cell proliferation (Ki67+cells) and (C and D) tumor burden in Pb-Cre+;-PtenL/L and Pb-Cre+;PtenL/L:ArL/Y mutants. Scale bars represent 2 mm (C), 200 mm (D), and 75 mm (D, inset). Error bars represent mean ± SD. |
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Rating |
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| Source |
Cancer Cell , 2011, 19, 1–13. Rapamycin (Sirolimus) purchased from Selleck |
| Method |
Cell viability Analysis |
| Cell Lines |
Ar+murine (CaP8) and human (LNCaP) prostate cancer cells, Pb-Cre+;PtenL/L mice, Pb-Cre+;PtenL/L;ArL/Y mice |
| Concentrations |
1 nM, 4 mg/kg |
| Incubation Time |
0-4 weeks |
| Results |
These data suggest that CaPs with AR loss have greater reliance upon the PI3K/AKT/mTOR-signaling pathways and that combined AR/androgen blockage in conjunction with PI3K/AKT/mTOR inhibition (by Rapamycin) is more effective for CaPs initiated by PTEN loss or PI3K/AKT activation. |
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| A, ANOVA of IC50 values of the dual PI3K/mTOR inhibitor NVP-BEZ235 in 23 melanoma cell lines showing no significant difference in sensitivity to this compound in B-Raf mutant and Braf wild-type cells. B, decreases in pAkt and pP70S6K in a dose and time-dependent fashion in 2 melanoma cell lines treated with NVP-BEZ235. pP70S6K levels are undetectable at all concentrations and time points studied, whereas levels of pAkt start rising again after 4 hours of drug exposure in a dose-dependent fashion. C, clonogenic assays in 2 melanoma cell lines (YUVON and YUSIK) treated with NVP-BEZ235 at different concentrations. NVPBEZ235 was effective in inhibiting colony formation at low nanomolar concentrations. |
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Rating |
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| Source |
Clin Cancer Res, 2010, 16, 6029-6039. BEZ235 (NVP-BEZ235) purchased from Selleck |
| Method |
Synergism studies, Immunoblot assays, Clonogenic assays |
| Cell Lines |
Melanoma cell |
| Concentrations |
0.1-100 μM |
| Incubation Time |
1-24 h |
| Results |
One of the proposed mechanisms of resistance to PI3KIs is mutation in the Ras-Raf pathway, which are found in more than half of melanomas. By ANOVA, no association was found between the IC50 values of NVP-BEZ235 and the presence or absence of B-Raf mutations (Fig. A). The targets of NVP-BEZ235, pAkt and pP70S6K, were both decreased with exposure to the drug in a time- and dose-dependent fashion, as shown in Figure B for YUVON and YUSIK cell lines. Clonogenicity was studied in YUVON and YUSIK cells with exposure to the dual PI3K/mTOR inhibitor. As shown in Figure C, NVP-BEZ235 effectively inhibits clonogenicity at low nanomolar concentrations. |
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| Confocal microscopy images of NO formation. DAF 2 DA-loaded washed platelets (1.0109 platelets/mL) were preincubated at 378C with saline (A), and then stimulated for 1min with 0.1 (B), 1.0 (C) or 10 (D) mM AEA. In Panel (E–F–G) washed platelets were preincubated with 1 mM SR1 (E), 1 Mm MK2206 (F) or 20 mM LY294002 (G), and then stimulated for 1min with 1.0 mM AEA. In panel (H) is reported the effect of 5 mg/mL collagen, used as a positive control. All the experiments were carried out in the presence of 100 mM L-arginine. NO formation was visualized by confocal microscropy as detailed in Methods. |
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Rating |
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| Source |
J Cell Biochem, 2010, 112, 924–932. MK-2206 2HCl purchased from Selleck |
| Method |
Confocal microscopy |
| Cell Lines |
platelets |
| Concentrations |
1μM |
| Incubation Time |
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| Results |
Upon treatment with increasing concentrations of AEA from 0.1 to 10 mM (Fig. B–D) platelets were fully fluorescently marked as compared to control (Fig. A) and the increase in DAF 2 fluorescence intensity appears to be dose-dependent. In agreement with data shown in Figure 4, SR1(Fig. E) and MK2206 (Fig. F) abolished fluorescence intensity induced by AEA, while LY294002 (Fig. 5G) was less potent. |
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| Effect of ATM inhibitor in etoposide-induced cell death. The cells were pretreated with KU55933 (2 M) for 1 h before etoposide treatment. |
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Rating |
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| Source |
J Biol Chem , 2011, 286, 8394-8404. KU-55933 purchased from Selleck |
| Method |
Cell Death Assay |
| Cell Lines |
HeLa cells |
| Concentrations |
2 μM |
| Incubation Time |
1 h |
| Results |
When the HeLa cells were pretreated with a specific ATM inhibitor KU55933, the ATM inhibition additively increased the etoposide-induced cell death in the PrxII knockdown cells. |
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| Mean IL-8 concentrations determined by ELISA of the supernatants of HeLa cells infected with wild-type Salmonella. Kinase inhibitors are indicated on the x axis, and the target families of the inhibitors are indicated above each column. CEC, chelerythrine; Pim Inh, Pim-1 inhibitor 2. Inhibitors that significantly affected IL-8 production relative to the control (P < 0.05, Bonferroni post hoc test from one-way ANOVA) are indicated with an asterisk. Relative cell viability is also shown, as determined by reduction of XTT by viable cells. A450, absorbance at 450 nm. |
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Sci Signal, 2011 Sep, 4:rs9. SGI-1776 free base purchased from Selleck |
| Method |
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| Cell Lines |
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| Concentrations |
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| Incubation Time |
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| Results |
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| Three-dimensional responses of MCF7/IGF-1R cells to TAM (1 μM), E2 and IGF-1. Compared to parental MCF7 cells (a), MCF7/IGF-1R cells (b) in three-dimensional (3D) culture formed bigger acini in response to IGF-1 stimulation and displayed significant TAM resistance when treated with TAM (1 μM) + E2 + IGF-1, which was removable by kinase inhibitors BMS-536924, U0126 and BEZ235 (c). Cells (10,000/well) were seeded in 96-well plates. Acini were formed on 100% Matrigel and cultured for 14 days in starving medium containing 2% Matrigel and 5% charcoal/dextran-stripped fetal bovine serum with the treatments as indicated. Concentrations used: TAM (1 μM), E2 (1 nM) and IGF-1 (100 ng/mL). Confocal image original magnification, × 20. Red, rhodamine phalloidin (actin). Blue, Hoechst blue stain. Results are representative of two individual experiments. |
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Breast Cancer Res Treat, 2010, 124(3), 667-675. Panobinostat (LBH589) purchased from Selleck |
| Method |
Western blot/etc |
| Cell Lines |
MDA-MB-231, MDA-MB-468 cells/etc |
| Concentrations |
0.5-10 μM/etc |
| Incubation Time |
24 h/etc |
| Results |
In both MDA-MB-231 and MDA-MB-468 cell lines, exposure to LBH589 and other compounds produced significant global increase of nuclear H3K4me2, which is the specific substrate of LSD1. The enhanced level of histone methylation by HDAC inhibitors parallels the increase of acetylation of histone 3. |
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| Rapamycin (RPM) inhibits OA-induced lipogenesis. Male SD rats were divided into three groups, and then treated as follows for seven days: group I (control; Ctrl), normal diet + vehicle; group II (OA), 1% OA diet + vehicle; group III (OA + RPM), 1% OA diet + RPM. (A) Hepatic TG concentrations were determined using the serum triglyceride determination kit. (B) Fixed liver sections were subjected to HE and Oil Red O staining as well as immunohistochemistry assays with SREBP-1 antibody. Representative microphotographs of liver specimens from all groups of rats are shown. (C) The levels of mRNA of the indicated genes involved in lipogenesis were determined by real-time PCR. Each bar represents the mean ± SE of the results obtained from six rats. * P < 0.05, ** P < 0.01, *** P < 0.001. |
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Rating |
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| Source |
J Lipid Res , 2011, 52, 1617-1625. Rapamycin (Sirolimus) purchased from Selleck |
| Method |
Histological analysis, immunohistochemistry, Liver triglyceride content analysis, real-time PCR |
| Cell Lines |
Male SD rats |
| Concentrations |
|
| Incubation Time |
7 d |
| Results |
To determine the effect of rapamycin, an mTOR inhibitor, in the OA-induced fatty liver, rats were fed 1% OA and administered rapamycin for seven days. OA-induced lipid accumulation was completely inhibited by the treatment with rapamycin (Fig. A). HE and Oil Red O staining of liver sections clearly confi rmed the results (Fig. B). Immunohistochemistry analyses showed signifi cantly repressed SREBP-1 in the rapamycin-cotreatment liver ( Fig. B ). All the downstream effectors of SREBP-1, such as Lxr-α, Acc, Scd-1 and Fas, were consistently suppressed by rapamycin ( Fig. C ). |
