The study suggests that the therapeutic effect of Aloperine on atopic dermatitis may involve modulation of immune responses, including downregulation of Th1 and Th2 cytokines (IL-4, IL-13, IFN-γ, TNF-α, IL-1β, IL-6) and upregulation of the immunosuppressive cytokine IL-10.

Aloperine treatment exerts the strongest cytotoxic activity against human cancer cell lines of various tissue origins, including leukemia cell lines HL-60, U937, and K562, oesophageal cancer EC109 cells, lung cancer A549 cells, and hepatocellular carcinoma HepG2 cell line, with IC50 values of 0.04 mM, 0.27 mM, 0.36 mM, 1.11 mM, 1.18 mM, and 1. The strongest cytotoxic effect of aloperine on HL-60 cells is seen at 72 hours, with an inhibition rate of 94.1%. Up to 1 mM of aloperine has no discernible impact on the viability of normal PBMNCs at 72 hours, in contrast to how it affects leukaemia cells. Aloperine treatment at 20 μM for 48 hours significantly and dose-dependently induces apoptosis and autophagy in HL-60 cells.[2]

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Aloperine exhibited potent cytotoxic activity against multiple human cancer cell lines, with IC₅₀ values ranging from 0.04 to 1.36 mM after 48 hr treatment. The lowest IC₅₀ was observed in HL-60 cells (0.04 mM).
Treatment with aloperine induced apoptosis in HL-60 cells, as evidenced by DNA fragmentation and PARP cleavage into an 89 kDa fragment after 48 hr incubation with 20 μM.
Aloperine also induced autophagy in HL-60 cells, shown by the formation of acidic vacuoles detected via acridine orange staining after 18 hr exposure at 20–100 μM.
Bcl-2 protein expression slightly decreased in HL-60 cells treated with aloperine at 50 and 100 μM.
Aloperine showed selective cytotoxicity toward cancer cells compared to normal peripheral blood mononuclear cells (PBMCs), which remained viable even at 1000 μM for 72 hr.[2]

Topical administration of 1% aloperine inhibits 2, 4-dinitrofluorobenzene (DNFB)-induced ear thickness and erythema in BALB/c mice and significantly reduces the up-regulated mRNA and protein levels of tumor necrosis factor-alpha (TNF-α), interleukin-1beta (IL-1β), and interleukin-6 (IL-6) induced by DNFB in ear biopsy homogenates. [1] Aloperine, when applied topically, dose-dependently reduces the DNFB-induced dermatitis (dermatitis index and ear thickness) in NC/Nga mice on days 13 and 14. Treatment with aloperine decreases DNFB-induced lymphocyte and eosinophil infiltration in a dose-dependent way. Treatment with aloperine also decreases the dose-dependent infiltration of mast cells brought on by DNFB. In a dose-dependent manner, aloperine lowers the plasma level of IgE. Aloperine increases the level of IL-10 in a dose-dependent manner while markedly decreasing the DNFB-induced increase in IL-4, IL-13, and IFN-γ productions. TNF-α, IL-1β, and IL-6 cytokine levels in NC/Nga mouse ear biopsies are significantly and dose-dependently reduced by aloperine treatment. [3]

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Topical application of 1% Aloperine significantly reduced DNFB-induced ear swelling (thickness) at 72 h and 96 h post-treatment compared to the DNFB-only group.
Topical application of 1% Aloperine significantly reduced DNFB-induced ear erythema.
Histopathological analysis showed that Aloperine treatment suppressed DNFB-induced leukocyte infiltration into the epidermis and dermis.
Aloperine treatment (1%) significantly reduced the protein levels of cytokines TNF-α, IL-1β, and IL-6 in ear tissue homogenates, as measured by ELISA.
Aloperine treatment (1%) significantly downregulated the mRNA expression of TNF-α, IL-1β, and IL-6 in ear skin, as measured by semi-quantitative RT-PCR.
The therapeutic effect of 1% Aloperine on DNFB-induced allergic contact dermatitis was comparable to that of the positive control, 0.1% mometasone furoate (a corticosteroid).

Cells are seeded in 96-well plates in 100-μL culture medium. Aloperine or excipient control-containing experimental media are added to the appropriate wells following incubation times of 4 hours for leukemia cells and 24 hours for solid cancer cells. The in vitro IC50 growth inhibitory values of aloperine in cancer cells are calculated after treatment with five concentrations of aloperine for 48 hours. Each well receives 10 μL of MTT solution (5 mg/mL) following incubation. Following that, the plates are incubated for 4 hours at 37 °C. 100 μL of isopropanol-HCI-SDS solution are added to each well to dissolve intracellular formazan crystals. The samples' optical densities are measured at 570 nm following an overnight incubation at 37°C. The analysis of DNA fragmentation is performed using an apoptotic DNA ladder kit after DNA has been extracted from cells that have been exposed to the recommended doses of aloperine for 48 hours. Cells are gathered and incubated in PBS containing 5 μM acridine orange for 15 minutes in order to detect autophagy. In 100 μL of PBS, the cells are resuspended after the acridine orange has been removed. With an inverted fluorescent microscope, fluorescent micrographs can be taken. The average number of cells with intense red staining for three fields, each with at least 50 cells, is used to calculate the amount of autophagy occurring in each experimental condition.

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Cell viability was assessed using MTT assay. Cells were seeded in 96-well plates and treated with aloperine at indicated concentrations for 48 hr. After incubation, MTT solution was added and formazan crystals were dissolved with isopropanol-HCl-SDS solution. Absorbance was measured at 570 nm to calculate inhibition rate and IC₅₀ values.
DNA fragmentation was analyzed using an apoptotic DNA ladder kit after treating HL-60 cells with aloperine for 48 hr. DNA was separated on a 1.2% agarose gel and visualized with ethidium bromide staining.
Western blot analysis was performed to detect PARP cleavage and Bcl-2 expression. Cells were lysed, proteins were separated by SDS-PAGE, transferred to nitrocellulose membrane, and probed with specific antibodies.
Autophagy was assessed by acridine orange staining. Cells were incubated with 5 μM acridine orange for 15 min, washed, and examined under a fluorescent microscope to quantify acidic vesicle formation.[2]

Female BALB/c mice with DNFB-induced allergic contact dermatitis
1% (w/w)
Topical application

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Female BALB/c mice (8–10 weeks old) were used.
Allergic contact dermatitis was induced by sensitizing shaved abdominal skin with 100 µL of 0.5% DNFB (in acetone:olive oil = 4:1). Five days later, mice were challenged by painting both sides of the ears with 20 µL of 0.2% DNFB. A rechallenge was performed one day later to induce extensive disease.
Mice were randomly divided into four groups (n=8): Control (no treatment), DNFB (no drug), DNFB + 0.1% mometasone furoate, and DNFB + 1% Aloperine.
Aloperine was formulated as a 1% (w/w) cream in the same emollient vehicle used for the positive control. The cream was applied topically to both ears at 0.5 h, 24.5 h, 48.5 h, and 72.5 h after the rechallenge.
Ear thickness was measured before challenge and at multiple time points (0, 24, 48, 72, 96 h) after rechallenge using a dial thickness gauge. Erythema was scored on a scale of 0-4.
Mice were euthanized 96 h after rechallenge. Ear tissues were collected for histopathology, cytokine protein analysis (ELISA), and mRNA analysis (semi-quantitative RT-PCR).

Aloperine has lower acute toxicity in mice than matrine, matrine, and strychnine, as described in the references (acute toxicity ranking: matrine ≡ matrine > strychnine > Aloperine > oxidized strychnine). [2]

[1]. Eur J Pharmacol . 2010 Mar 10;629(1-3):147-52.

[2]. Basic Clin Pharmacol Toxicol . 2011 May;108(5):304-9.

[3]. Eur J Pharmacol . 2011 May 11;658(2-3):263-9.

Aloperine has been reported to be found in several organisms with available data, including Sophora jaubertii, Thinicola incana, and others. Aloperine is an alkaloid isolated from plants in the Sophora genus (such as Sophora jaubertii). In China, it has been used clinically for decades to treat allergic contact dermatitis, eczema, and other skin inflammations. This study is the first to report the inhibitory effect of Aloperine on chemically induced allergic contact dermatitis in mice. Its mechanism of action may involve the upregulation of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) at both the protein and mRNA levels. This research suggests that Aloperine may be a potential therapeutic agent for the prevention and treatment of inflammatory skin diseases and may serve as an alternative to corticosteroids.

C15H24N2

232.3645

232.193

C, 77.53; H, 10.41; N, 12.06

56293-29-9

56293-29-9

162147

White to off-white solid powder

1.1±0.1 g/cm3

367.7±37.0 °C at 760 mmHg

69 - 71ºC

155.8±17.5 °C

0.0±0.8 mmHg at 25°C

1.583

3.21

1

2

0

17

336

4

N12C([H])([H])C([H])([H])C([H])([H])C([H])([H])[C@]1([H])[C@@]1([H])C([H])=C3C([H])([H])C([H])([H])C([H])([H])N([H])[C@@]3([H])[C@@]([H])(C2([H])[H])C1([H])[H]

SKOLRLSBMUGVOY-GBJTYRQASA-N

InChI=1S/C15H24N2/c1-2-7-17-10-13-9-12(14(17)5-1)8-11-4-3-6-16-15(11)13/h8,12-16H,1-7,9-10H2/t12-,13+,14+,15+/m0/s1

(1R,2S,9R,10R)-3,15-diazatetracyclo[7.7.1.02,7.010,15]heptadec-7-ene

Aloperine

2934.99.03.00

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, avoid exposure to moisture.

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

DMSO: 5.6~46 mg/mL (23.9~198.0 mM)
Water: ~12 mg/mL (~51.6 mM)
Ethanol: ~46 mg/mL (~198.0 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (8.95 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 20.8 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: ≥ 2.08 mg/mL (8.95 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 20.8 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: ≥ 2.08 mg/mL (8.95 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 20.8 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.)