Biochemical reagent; natural alkaloid
The primary target of d-sophoridine is not fully elucidated, but the compound induces apoptosis through the intrinsic mitochondrial pathway. d-Sophoridine triggers apoptosis in cancer cells by activating the mitochondrial apoptotic cascade, leading to the release of cytochrome c and activation of caspases. The compound may also modulate various signaling pathways involved in cell survival and proliferation. As a quinolizidine alkaloid, sophoridine is thought to interact with cellular targets that regulate apoptosis and cell cycle progression. The compound's antitumor activity is associated with its ability to induce cell death selectively in cancer cells while sparing normal cells, suggesting a degree of target selectivity.

In human pancreatic, gastric, liver, colon, gallbladder, and prostate cancer cells, sophoridine (0-500 μM; 48 hours) shows substantial growth inhibitory effects with IC50 values of approximately 20 μM to 200 μM[1]. From 26.23% (control) to 38.67% in Miapaca-2 cells and from 29.56% (control) to 39.16% in PANC-1 cells, sophoridine (0 – 20 μM) increases the S phase cell population in 48 hours, almost 1.5 and 1.3 times, respectively[1]. With a considerable increase in the Bax/Bcl-2 ratio, sophoridine (0-20 μM; 48 hours) considerably raises the levels of bad and bax and, in contrast, decreases the levels of bcl-2 and bcl-xl; [1].
In vitro, d-sophoridine demonstrates potent antiproliferative activity against various cancer cell lines, including pancreatic cancer cells. The compound induces apoptosis in a dose-dependent manner, as evidenced by increased Annexin V staining, caspase activation, and DNA fragmentation. d-Sophoridine inhibits cell proliferation and colony formation in cancer cells. The compound shows selective cytotoxicity towards cancer cells compared to normal cells, suggesting a favorable therapeutic index. In addition to its direct antitumor effects, d-sophoridine may enhance the sensitivity of cancer cells to conventional chemotherapeutic agents. The compound's in vitro activity has been characterized in multiple studies using various cancer cell lines, with IC50 values typically in the micromolar range.

Sophoridine (20 or 40 mg/kg; intraperitoneal injection; 21 days) can stop pancreatic tumor growth in xenografts[1].

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Sophoridine suppressed tumor growth in vivo[1]
To further validate Sophoridine can inhibit tumor growth in vivo, 2 × 106 Miapaca-2 cells were subcutaneously inoculated in Balb/c nude mice. Sophoridine treatment was administrated from the 7th day at 20 or 40 mg/kg intraperitoneally for 21 days. We found that Sophoridine showed significant inhibitory effects on tumor volume (Fig. 7a and b). The mass of tumors under 20 or 40 mg/kg Sophoridine treatment was significantly less than that of the control group (Fig. 7c). However, there was no significant changes on body weight under Sophoridine treatment (Fig. 7d). Furthermore, the activation of ERK and JNK in xenograft tumor tissues was tested by immunohistochemistry and immunoblotting. It showed that both ERK and JNK were activated in Sophoridine-treated xenograft tumor tissues (Fig. 7e and Additional file 7: Figure S5). In addition, IHC analysis of proliferating cell nuclear antigen (PCNA), cleaved caspase-3 and tunel staining showed significantly fewer proliferative cells, and more apoptotic cells in Sophoridine-treated tumors (Fig. 7e). The liver and kidney biochemical functions were monitored, and there were no obvious difference between control and Sophoridine treatment group (Fig. 7f). The liver and kidney from control and Sophoridine group were stained with H&E to further evaluate the toxicity after treatment. The histological structure of liver and kidney were observed and compared microscopically, and there were almost no histological changes after Sophoridine treatment (Fig. 7g). These results suggest that Sophoridine was an effective agent that can inhibit the growth of xenograft pancreatic tumors in vivo with well-tolerated toxicity[1].
In vivo, d-sophoridine has demonstrated significant antitumor activity in mouse models. Intraperitoneal administration of sophoridine at doses of 20 or 40 mg/kg for 21 days can inhibit the growth of xenograft pancreatic tumors in mice. The compound effectively suppresses tumor progression and may prolong survival in animal models. Pharmacodynamic studies show that d-sophoridine treatment leads to increased apoptosis in tumor tissues, confirming the in vitro findings. The compound is well-tolerated at efficacious doses, with no significant weight loss or organ toxicity observed. d-Sophoridine has the potential to be a novel, potent, and selective antitumor agent for pancreatic cancer with well-tolerated toxicity. Further in vivo studies are warranted to fully characterize its efficacy and safety.

In vitro binding assays for d-sophoridine are not extensively documented. However, typical approaches include surface plasmon resonance (SPR) or isothermal titration calorimetry (ITC) to measure the compound's affinity for potential target proteins. Radioligand binding assays may be used to assess interactions with specific receptors or enzymes. The compound's ability to modulate apoptosis-related proteins (e.g., Bcl-2, Bax, caspases) can be assessed by Western blotting after cell treatment. Activity-based protein profiling (ABPP) could be employed to identify direct protein targets of d-sophoridine. Given the limited information available, standard biochemical assays for studying alkaloid-protein interactions may be adapted for this compound.

Cell viability assay [1]
Cell viability was measured with CCK-8 kit, followed the manufacturer. Briefly, cancer cells seeded in 96-well plates were either treated with Sophoridine at serial concentrations for 48 h, or were treated for various time points (0, 24, 48, or 72 h). After treatment for indicated time, CCK- 8 solution was added and incubated with cancer cells for 4 h. The percentages of cell survival was measured by SpectraMax190 microplate reader based on the absorbance.

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Cell cycle and apoptosis analysis [1]
Propidium iodide (PI) staining assay was used to analyze the cell cycle distribution. After exposed to different concentrations of Sophoridine for 48 h, cancer cells were harvested and fixed with 70% ethanol, followed by centrifugation (3000 rpm, 5 min), incubation with 100 mg/mL RNase in PBS for 30 min at 37 °C, and then staining with 50 mg/mL PI in PBS. The cell cycle distribution were analyzed by a Cell Lab Quanta SC flow cytometer. The Annexin V–FITC Apoptosis Detection Kit (BioVision) was used for the apoptosis analysis. Cells (5 × 105) were exposed to different concentrations of Sophoridine for 48 h. After resuspended in 500 ml binding buffer, cells were incubated with Annexin V– fluorescein isothiocyanate (FITC; 5 ml) and PI (5 ml). After 30-min incubation, cells were analyzed by fluorescence-activated cell sorting (FACS) by flow cytometer. Annexin V–FITC–stained only cells which indicated early apoptosis and cells with Annexin V–FITC- and PI double positive signals were combined for analysis.

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Colony formation assay [1]
Five hundred cancer cells per well were seeded into 6-well plates, and then treated with Sophoridine at different concentrations for 48 h. After treatment, the cells were allowed to form cell colonies for another 7 days. Then cell colonies were fixed with 4% paraformaldehyde and stained with 0.1% crystal violet. After 3 times washing and air-dried, the stained colonies were counted and photographed under microscope.
In vitro cell-based assays for d-sophoridine are performed using various cancer cell lines, with a focus on pancreatic cancer cells (e.g., PANC-1, MIA PaCa-2). Cells are seeded in 96-well plates and treated with serial dilutions of d-sophoridine for 48-72 hours. Cell viability is assessed using MTT, CCK-8, or CellTiter-Glo assays to determine the IC50. Apoptosis is evaluated by flow cytometry using Annexin V/PI staining or by detecting caspase-3/7 activity. Cell cycle analysis is performed using propidium iodide staining to assess cell cycle arrest. The effect of d-sophoridine on apoptosis-related protein expression (Bcl-2, Bax, cytochrome c, caspases) is assessed by Western blotting. Combination studies with other anticancer agents are performed to evaluate synergistic effects.

Animal treatment with Sophoridine [1]
BALB/c homozygous (nu/nu) nude mice (6 weeks, 18 g) were used. The mice were maintained in pathogen-free environment for one week after arrival. 2 × 106 Miapca-2 cells in 100 μl PBS were inoculated in the right flank of nude mice. One week later, mice were randomly divided into 3 groups (7 mice/group). 3 groups received relatively an intraperitoneally (i.p.) injection of PBS as control, 20 mg/kg of Sophoridine, or 40 mg/kg of Sophoridine daily. After 3 weeks treatment, mice were sacrificed to weigh the tumors.
In vivo animal studies with d-sophoridine are conducted in mouse xenograft models of pancreatic cancer. Immunodeficient mice are implanted subcutaneously with human pancreatic cancer cells. When tumors reach a certain size, mice are randomized to receive d-sophoridine or vehicle control. The compound is typically administered via intraperitoneal injection at doses of 20 or 40 mg/kg daily or on a scheduled regimen. Tumor volume is measured using calipers every 2-3 days. Body weight is monitored as a measure of tolerability. At the end of the study, tumors are collected for histopathological analysis, immunohistochemistry (e.g., Ki-67 for proliferation, TUNEL for apoptosis), and pharmacodynamic analysis of apoptosis markers. Survival studies may be conducted in orthotopic models.

Pharmacokinetic properties of d-sophoridine have not been extensively characterized in the literature. As a small-molecule alkaloid, the compound is expected to have moderate lipophilicity and good membrane permeability. Following administration, d-sophoridine is likely distributed to various tissues, with potential accumulation in tumors due to enhanced permeability and retention (EPR) effects. The compound is probably metabolized in the liver and excreted via the kidneys and bile. Pharmacokinetic parameters such as half-life, clearance, and volume of distribution would need to be determined experimentally. For research use, d-sophoridine is typically formulated in saline or other suitable vehicles for administration in preclinical studies.

Toxicity Summary
The primary target organ for matrine toxicity is the nervous system. Morphological observations have shown degenerative changes in nerve cells in mouse brain tissue. (A15436) Matrine possesses various pharmacological effects, including anticancer activity and action as a κ-opioid and μ-receptor agonist. Matrine exhibits strong antitumor activity both in vitro and in vivo. Inhibition of cell proliferation and induction of apoptosis may be the mechanisms by which matrine exerts its antitumor effects. (Wikipedia)
d-Sophoridine has been reported to have well-tolerated toxicity in preclinical studies. At therapeutic doses, the compound does not cause significant weight loss, organ toxicity, or hematological abnormalities in animal models. This favorable toxicity profile is an important attribute for a potential anticancer agent. No specific toxicological studies have been published in detail. As with all research chemicals, standard safety precautions should be followed when handling d-sophoridine. The compound should be stored appropriately and used only for research purposes. Further toxicological studies are needed to fully characterize the safety profile of d-sophoridine for potential clinical development.

[1]. Sophoridine induces apoptosis and S phase arrest via ROS-dependent JNK and ERK activation in human pancreatic cancer cells. J Exp Clin Cancer Res. 2017 Sep 11;36(1):124.

(+)-Matrine has been reported to exist in Euchresta horsfieldii, Gymnospermium darwasicum, and several other organisms with relevant data. See also: Matrine (note moved to). Background: Pancreatic cancer is widely recognized as the most common primary malignant tumor and is known to be resistant to conventional chemotherapy. Therefore, there is an urgent need for novel, selective antitumor drugs. Methods: Cell growth was studied using CCK-8 and colony formation assays. Cell cycle and apoptosis were assessed using flow cytometry. Intracellular reactive oxygen species (ROS) levels were detected using the peroxidase-sensitive fluorescent probe DCFH-DA. The expression levels of cell cycle and apoptosis-related proteins were detected using Western blotting. The effects of matrine on pancreatic cancer cells in vivo were evaluated using a nude mouse xenograft model. Results: Matrine killed cancer cells but exhibited low cytotoxicity to normal cells. Pancreatic cancer cells were particularly sensitive. Matrine inhibited the proliferation of pancreatic cancer cells, induced cell cycle arrest in the S phase, and caused mitochondrial-related apoptosis. In addition, sophoridine can continuously activate the phosphorylation of ERK and JNK. Furthermore, sophoridine can promote the production of reactive oxygen species (ROS) in pancreatic cancer cells. Finally, in vivo experiments showed that sophoridine can inhibit tumor growth in a mouse xenograft model. Conclusion: These findings suggest that sophoridine is a promising candidate drug for novel, highly effective and selective antitumor therapy in pancreatic cancer. [1]
d-Sophoridine is a research compound derived from the traditional Chinese medicinal plant Sophora flavescens, which has been used in traditional medicine for various conditions. The compound's potential as a selective and well-tolerated antitumor agent for pancreatic cancer has generated significant research interest. Pancreatic cancer is one of the most lethal malignancies with limited treatment options, and the development of novel targeted therapies is urgently needed. d-Sophoridine's ability to induce apoptosis selectively in cancer cells while sparing normal cells makes it an attractive candidate for further development. Ongoing research is focused on elucidating the compound's mechanism of action, identifying its molecular targets, and evaluating its efficacy in combination with other anticancer agents.

C15H24N2O

248.36

248.189

83148-91-8

Sophoridine;6882-68-4

285698

Typically exists as solid at room temperature

1.164 g/cm3

396.738ºC at 760 mmHg

172.748ºC

1.747

0

2

0

18

356

0

O=C1CCC[C@H]2[C@@H]3CCCN4[C@@H]3[C@@H](CCC4)CN12

ZSBXGIUJOOQZMP-UHFFFAOYSA-N

InChI=1S/C15H24N2O/c18-14-7-1-6-13-12-5-3-9-16-8-2-4-11(15(12)16)10-17(13)14/h11-13,15H,1-10H2

7,13-diazatetracyclo[7.7.1.02,7.013,17]heptadecan-6-one

83148-91-8; Matridin-15-one; (+)-Matrine; (+)-Sophoridine; Matridin-15-one, (6b,7b,11a)- (9CI); 7,13-diazatetracyclo[7.7.1.02,7.013,17]heptadecan-6-one; .alpha.-Matrine; Matrene, (+)-;

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)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

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Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

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Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*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.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

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Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

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Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

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Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

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