Lipid rafts (membrane rafts) in MDA-MB-231/IR cells
PI3K/Akt signaling pathway (specifically, the raft-resident components: p-Akt (Ser473), p-Akt (Thr308), p-PI3Kp85, p-mTOR (Ser2448), p4E-BP1) [3]

In radio-resistant triple-negative breast cancer cells (MDA-MB-231/IR), 10-gingerol suppressed proliferation in a dose- and time-dependent manner. The IC50 values at 24 and 48 hours were 121.2 μM and 101.4 μM, respectively. It showed fewer anti-proliferative effects in parental MDA-MB-231 cells and fewer cytotoxic effects in MCF-10A normal mammary epithelial cells (IC50: 327.8 μM at 24h, 219.8 μM at 48h) compared to docetaxel. [3]
10-gingerol significantly reduced the clonogenic ability of MDA-MB-231/IR cells at concentrations of 40, 20, 10, and 5 μM after a 10-day incubation. [3]
10-gingerol significantly inhibited the migration of MDA-MB-231/IR cells at concentrations of 90 and 45 μM in a wound-healing assay after 24 hours. [3]
10-gingerol significantly reduced the number of invading MDA-MB-231/IR cells in a Transwell cell invasion assay. [3]
10-gingerol induced apoptosis in MDA-MB-231/IR cells. Hoechst staining revealed visible chromatin condensation after treatment. Western blot analysis showed that 10-gingerol activated the expression of pro-apoptotic markers including the Bax/Bcl-2 ratio, cleaved caspase-7, cleaved PARP, and cleaved caspase-3 compared to untreated cells. [3]
Western blot analysis demonstrated that 10-gingerol (24h treatment) caused a dose-dependent decrease in the phosphorylation of p-Akt (Ser473), p-Akt (Thr308), p-PI3Kp85, and p4E-BP1 in MDA-MB-231/IR cells. A decreased phosphorylation of p-mTOR (Ser2448) was only observed at the two highest doses. [3]
10-gingerol (90 μM, 24h) modulated lipid rafts in MDA-MB-231/IR cells, as indicated by the shifting of the lipid raft marker caveolin-1 from raft fractions (3-5) to non-raft fractions (10-12) in a sucrose density gradient centrifugation assay. Co-treatment with cholesterol (0.1 mM) recovered caveolin-1 in the raft fractions. Immunofluorescence confirmed that the localization of caveolin-1 in the membrane was greatly affected by 10-gingerol treatment. [3]
Treatment with 10-gingerol (90 μM, 24h) caused a reduction in total cholesterol levels in the lipid raft fractions of MDA-MB-231/IR cells. [3]
Following 10-gingerol exposure, displacement of major components of the PI3K/Akt signaling pathway (activated forms of Akt, mTOR, and PI3Kp85) from the lipid raft fractions was evident. [3]
10-gingerol was found to dephosphorylate p-Akt (Ser473) and p-Akt (Thr308) in isolated lipid rafts in a dose-dependent manner. Co-treatment of 10-gingerol (90 μM) with cholesterol (0.1 mM) almost restored their phosphorylation status in the lipid rafts. [3]
10-gingerol down-regulated the Akt downstream targets GSK3β and cyclin D1 in a dose-dependent manner. Down-regulation of β-catenin was only observed at the two highest doses of 10-gingerol. [3]

Cell Culture: MDA-MB-231, MDA-MB-231/IR, and MCF-10A cells were cultured. MDA-MB-231 and MDA-MB-231/IR cells were grown in DMEM supplemented with 10% FBS and antibiotics. MCF-10A cells were cultured in mammary epithelial cell growth medium supplemented with recommended growth factors (insulin, hydrocortisone, EGF, bovine pituitary extract). All cells were incubated at 37°C in a 5% CO2 incubator. [3]
Cell Proliferation Assay (MTT): Cells were seeded in 96-well plates (5000 cells/well) and incubated for 24h. They were then exposed to 10-gingerol (200, 100, 50, 25, 12.5, 6.25 μM) for 24 and 48h. After treatment, cells were washed with PBS, and an MTT solution (0.5 mg/mL in PBS) was added to each well and incubated for 4h at 37°C. The resulting formazan crystals were dissolved in 100 μL of DMSO, and the absorbance was measured at 570 nm using a microplate reader. The percentage of cell viability was calculated using the formula: ((control - treated) ÷ control) × 100%. IC50 values were calculated using GraphPad Prism 5 software. [3]
Colony Formation Assay: MDA-MB-231/IR cells (200 cells/dish) were seeded in 60 mm dishes and incubated for 24h. Cells were then treated with 10-gingerol (40, 20, 10, 5 μM) and incubated for 10 days. After incubation, colonies were fixed, stained with crystal violet, and manually counted. Results were expressed as a percentage of the untreated control. [3]
Wound Healing Assay: MDA-MB-231/IR cells (1×10^5) were seeded in 6-well plates and grown to 95% confluence. The monolayer was scratched with a sterile pipette tip, washed with PBS, and exposed to 10-gingerol (90, 45 μM) for 24h. The gap area was quantified and compared to controls (0h). [3]
Trans-Well Cell Invasion Assay: The upper chamber of a Trans-well insert was coated with 1% Matrigel. MDA-MB-231/IR cells (1×10^5 cells/trans well) were transferred to the upper chamber with or without 10-gingerol. The lower chamber received DMEM with 10% FBS. After 24h, cells were fixed, stained with 2% crystal violet, and photographed. [3]
Western Blot: Following treatments, cell lysates were prepared using RIPA buffer. Protein concentrations were estimated (using a BCA Protein Assay Kit). Equal amounts of protein were subjected to SDS-PAGE and transferred to PVDF membranes. Membranes were blocked with 5% skim milk and incubated with primary antibodies (diluted 1:1000) overnight at 4°C. After washing, membranes were incubated with suitable HRP-conjugated secondary antibodies (1:5000) for 1h at room temperature. Bands were developed using an ECL Plus Kit and imaged using a ChemiDoc touch imaging system. [3]
Isolation of Lipid Rafts: Serum-starved MDA-MB-231/IR cells (1×10^7) were lysed in 1.5 mL of lysis buffer (0.5% Triton X-100, 15 mM Tris, 1 mM EDTA, 100 mM NaCl, 50 mM PMSF, 5% glycerol, and anti-proteases) for 30 min on ice. 1 mL of the lysate was mixed with 1 mL of lysis buffer containing 80% sucrose and transferred to 7.5 mL of lysis buffer with 30% sucrose, followed by 3 mL of buffer with 5% sucrose. This gradient was centrifuged at 20,000 rpm for 18h. Twelve 1 mL fractions were collected. Fractions 3-5 were considered lipid raft fractions, and fractions 10-12 were non-raft fractions. [3]
Immunofluorescence: Serum-starved MDA-MB-231/IR cells (5×10^4) on coverslips were fixed with 4% paraformaldehyde for 15 min, washed, and blocked with 1% BSA. Cells were incubated with caveolin-1 primary antibody overnight at 4°C, washed, and incubated with an Alexa 488-conjugated secondary antibody for 1h at room temperature. Coverslips were then counterstained with Hoechst 33342 (5 μg/mL) and observed using a digital cell imaging system. [3]
Measurement of Cholesterol Levels: Total cholesterol levels in lipid raft fractions were measured using a commercially available total cholesterol assay kit, following the manufacturer's instructions. [3]

The lipophilicity of 10-gingerol was predicted using the SwissADME web server. It was found to have high octanol-water partition coefficients, indicating strong hydrophobic interactions with cell membranes. [3]

[1]. Comparative antioxidant and anti-inflammatory effects of [6]-gingerol, [8]-gingerol, [10]-gingerol and [6]-shogaol. J Ethnopharmacol. 2010 Feb 3;127(2):515-20.

[2]. Purification and differential biological effects of ginger-derived substances on normal and tumor cell lines. J Chromatogr B Analyt Technol Biomed Life Sci. 2012 Aug 15;903:157-62.

[3]. Ediriweera MK, Moon JY, Nguyen YT, Cho SK. 10-Gingerol Targets Lipid Rafts Associated PI3K/Akt Signaling in Radio-Resistant Triple Negative Breast Cancer Cells. Molecules. 2020;25(14):3164.

(10)-Gingerol is a β-hydroxy ketone, belonging to the phenolic and monomethoxybenzene classes. It has been reported that ginger (Zingiber officinale) contains (10)-gingerol, and relevant data are available. See also: Ginger (partial).

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10-gingerol is a major phenolic lipid found in the rhizomes of ginger (Zingiber officinale). [3]
It is amphiphilic in nature due to the presence of a hydrophobic alkyl side chain attached to a hydrophilic dihydroxybenzene ring. [3]
The study concludes that 10-gingerol suppresses proliferation, migration, and invasion and induces apoptosis in radio-resistant triple-negative breast cancer cells (MDA-MB-231/IR) by targeting lipid raft-associated PI3K/Akt signaling, suggesting a new raft-mediated treatment approach for radio-resistant breast cancer. [3]

C21H34O4

350.4923

350.245

23513-15-7

168115

White to light yellow solid powder

1.0±0.1 g/cm3

499.7±35.0 °C at 760 mmHg

45 - 46 °C

166.4±19.4 °C

0.0±1.3 mmHg at 25°C

1.513

4.61

2

4

14

25

345

1

O([H])[C@]([H])(C([H])([H])C(C([H])([H])C([H])([H])C1C([H])=C([H])C(=C(C=1[H])OC([H])([H])[H])O[H])=O)C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H]

AIULWNKTYPZYAN-SFHVURJKSA-N

InChI=1S/C21H34O4/c1-3-4-5-6-7-8-9-10-18(22)16-19(23)13-11-17-12-14-20(24)21(15-17)25-2/h12,14-15,18,22,24H,3-11,13,16H2,1-2H3/t18-/m0/s1

(5S)-5-hydroxy-1-(4-hydroxy-3-methoxyphenyl)tetradecan-3-one

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light.

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 (~285.31 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (7.13 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.)