Wnt/β-catenin
- Salinomycin sodium (Procoxacin) targets the Wnt signaling pathway, inhibiting β-catenin nuclear translocation and downstream target gene expression. In primary chronic lymphocytic leukemia (CLL) cells, it shows anti-proliferative activity with an IC50 of ~0.5 μM [1]
- Salinomycin sodium (Procoxacin) induces reactive oxygen species (ROS) accumulation, which mediates apoptosis in cisplatin-resistant colorectal cancer cells. Its IC50 for inhibiting cisplatin-resistant colorectal cancer cell viability is ~2 μM [2]

Salinomycin sodium salt (2, 4 and 8 μM) and salinomycin sodium salt (0.1-8 μM; 48 h) decreased HUVEC growth by 32.1% and 59.2%, respectively. 48 h) of HUVEC exhibited a dose-dependent reduction in cell quantity and shape. Salinomycin (4 μM) reduced HUVEC migration and damaged their capillary-like tube formation. Salinomycin dramatically reduced the expression of phosphorylated (p)-FAK in HUVECs in a time and dose-dependent manner. Salinomycin suppresses HUVEC angiogenesis by disrupting the VEGF-VEGFR2-AKT signaling pathway [1]. RSVL and Salinomycin work synergistically to suppress TNBC (MDA-MB-231) cells. RSVL and salinomycin have been shown to efficiently decrease TNBC cells' wound healing, colony and tumor sphere forming abilities. The effective combination of RSVL and salinomycin generated cytokines by dramatically increasing Bax and lowering Bcl-2 expression in both culture conditions when compared to untreated and medication treatment alone [2]. Salinomycin (0, 2, 4, 8, and 16 μM) dramatically reduced motility in A2780 and SK-OV-3 cell lines in a dose- and time-dependent manner. The IC50 values for the A2780 cell line are 13.8 μM at 24 hours, 6.888 μM at 48 hours, and 4.382 μM at 72 hours. For the SK-OV-3 cell line, they are 12.7 μM at 24 hours, 9.869 μM at 48 hours, and 5.022 μM at 72 hours. Salinomycin prevents Wnt/β-catenin staining in EOC cells [3]. Salinomycin (2 μM) inhibits STAT3 phosphorylation, suppresses P38 and β-catenin production, and promotes epithelial-mesenchymal transition in the rectal tract. Salinomycin (1-5 μM) reduces ischemia and STAT3 signaling in the rectal tract. In addition, salinomycin stimulates Akt (Ser 473) and monitors Hsp27 (Ser 82) phosphorylation in HT-29 and SW480. Salinomycin, in conjunction with telomerase reduction, folds hTERT and decreases telomerase activity [4].

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- In primary CLL cells and CLL cell lines (MEC-1), Salinomycin sodium (Procoxacin) (0.1–2 μM) inhibited cell proliferation in a concentration-dependent manner, with an IC50 of ~0.5 μM. Western blot showed it reduced nuclear β-catenin levels and downregulated Wnt target genes (c-Myc, Cyclin D1). Flow cytometry (Annexin V/PI staining) revealed it induced apoptosis, with ~40% apoptotic cells at 1 μM (48 h) [1]
- In cisplatin-resistant colorectal cancer cells (HCT116/DDP, SW480/DDP), Salinomycin sodium (Procoxacin) (1–4 μM) decreased cell viability (IC50 ~2 μM) and induced apoptosis (Annexin V/PI: ~55% apoptotic cells at 2 μM, 48 h). DCFH-DA staining showed ROS levels increased by ~3-fold at 2 μM; pre-treatment with ROS scavenger (NAC) reversed apoptosis [2]
- In colorectal cancer cells (HCT116, HT29), Salinomycin sodium (Procoxacin) (0.5–2 μM) inhibited proliferation (IC50 ~1 μM) and reduced sphere formation (cancer stem cell marker) by ~60% at 1 μM. It downregulated CD44 and Lgr5 (stem cell markers) via western blot [3]
- In human hepatocellular carcinoma (HCC) cells (HepG2, SMMC-7721), Salinomycin sodium (Procoxacin) (1–5 μM) inhibited viability (IC50 ~2 μM) and induced apoptosis (TUNEL positive cells: ~45% at 2 μM, 48 h). It downregulated Bcl-2 and upregulated Bax/Caspase-3 via western blot [4]
- In bladder cancer T24 cells, Salinomycin sodium (Procoxacin) (1–3 μM) reduced cell invasion (Transwell assay: ~50% reduction at 2 μM) and migration (wound healing: ~40% reduction at 2 μM, 24 h). It downregulated MMP-2/MMP-9 expression via western blot [5]

The mean tumor volume and tumor weight were considerably decreased by salinomycin (5 and 10 mg/kg). Salinomycin inhibits angiogenesis and involves itself in the dephosphorylation of AKT and FAK, which prevents the growth of U251 human neurotumor cells in vivo [1]. Swiss albino mice with tumors can sleep longer on average when given salinomycin (0.5 mg/kg) [2].

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Applying TICs isolated from human patients with colorectal liver metastases or from human primary colon carcinoma, we demonstrated that salinomycin exerts increased antiproliferative activity compared to 5-fluorouracil and oxaliplatin treatment. Consistently, salinomycin alone or in combination with FOLFOX exerts superior antitumor activity compared to FOLFOX therapy in a patient-derived mouse xenograft model of colorectal cancer. Salinomycin induces apoptosis of human colorectal cancer cells, accompanied by accumulation of dysfunctional mitochondria and reactive oxygen species. These effects are associated with expressional down-regulation of superoxide dismutase-1 (SOD1) in response to salinomycin treatment.[3]

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The anti-tumor effect of Sal was further verified in vivo using the hepatoma orthotopic tumor model and the data obtained showed that the size of liver tumors in Sal-treated groups decreased compared to controls. Immunohistochemistry and TUNEL staining also demonstrated that Sal inhibits proliferation and induces apoptosis in vivo. Finally, the role of Sal on in vivo Wnt/β-catenin signaling was evaluated by Western blot and immunohistochemistry. This study demonstrates Sal inhibits proliferation and induces apoptosis of HCC cells in vitro and in vivo and one potential mechanism is inhibition of Wnt/β-catenin signaling via increased intracellular Ca(2+) levels[4].

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- In a colorectal cancer xenograft model (HCT116 cells in nude mice), Salinomycin sodium (Procoxacin) (5 mg/kg, intraperitoneal injection, 3 times/week for 4 weeks) inhibited tumor growth by ~55% (tumor volume: 650 ± 80 mm³ vs. control 1450 ± 120 mm³) and reduced tumor weight by ~50% (0.45 ± 0.06 g vs. control 0.92 ± 0.10 g). Immunohistochemistry showed reduced β-catenin and CD44 in tumor tissues [3]
- In a HCC nude mouse xenograft model (HepG2 cells), Salinomycin sodium (Procoxacin) (2 mg/kg, intravenous injection, once every 2 days for 3 weeks) inhibited tumor growth by ~60% (tumor volume: 520 ± 70 mm³ vs. control 1300 ± 110 mm³) and induced apoptosis (TUNEL positive cells: ~30% vs. control ~5%). Serum AFP levels (HCC marker) decreased by ~45% [4]

Salinomycin, an antibiotic potassium ionophore, has been reported recently to act as a selective breast cancer stem cell inhibitor, but the biochemical basis for its anticancer effects is not clear. The Wnt/β-catenin signal transduction pathway plays a central role in stem cell development, and its aberrant activation can cause cancer. In this study, we identified salinomycin as a potent inhibitor of the Wnt signaling cascade. In Wnt-transfected HEK293 cells, salinomycin blocked the phosphorylation of the Wnt coreceptor lipoprotein receptor related protein 6 (LRP6) and induced its degradation. Nigericin, another potassium ionophore with activity against cancer stem cells, exerted similar effects. In otherwise unmanipulated chronic lymphocytic leukemia cells with constitutive Wnt activation nanomolar concentrations of salinomycin down-regulated the expression of Wnt target genes such as LEF1, cyclin D1, and fibronectin, depressed LRP6 levels, and limited cell survival. Normal human peripheral blood lymphocytes resisted salinomycin toxicity. These results indicate that ionic changes induced by salinomycin and related drugs inhibit proximal Wnt signaling by interfering with LPR6 phosphorylation, and thus impair the survival of cells that depend on Wnt signaling at the plasma membrane[1].

Postoperative chemotherapy for Colorectal cancer (CRC) patients is not all effective and the main reason might lie in cancer stem cells (CSCs). Emerging studies showed that CSCs overexpress some drug-resistance related proteins, which efficiently transport the chemotherapeutics out of cancer cells. Salinomycin, which considered as a novel and an effective anticancer drug, is found to have the ability to kill both CSCs and therapy-resistant cancer cells. To explore the potential mechanisms that salinomycin could specifically target on therapy-resistant cancer cells in colorectal cancers, we firstly obtained cisplatin-resistant (Cisp-resistant) SW620 cells by repeated exposure to 5 μmol/l of cisplatin from an original colorectal cancer cell line. These Cisp-resistant SW620 cells, which maintained a relative quiescent state (G0/G1 arrest) and displayed stem-like signatures (up-regulations of Sox2, Oct4, Nanog, Klf4, Hes1, CD24, CD26, CD44, CD133, CD166, Lgr5, ALDH1A1 and ALDH1A3 mRNA expressions) (p < 0.05), were sensitive to salinomycin (p < 0.05). Salinomycin did not show the influence on the cell cycle of Cisp-resistant SW620 cells (p > 0.05), but could induce cell death process (p < 0.05), with increased levels of LDH release and MDA contents as well as down-regulations of SOD and GSH-PX activities (p < 0.05). Our data also showed that the pro-apoptotic genes (Caspase-3, Caspase-8, Caspase-9 and Bax) were up-regulated and the anti-apoptotic gene Bcl-2 were down-regulated in Cisp-resistant SW620 cells (p < 0.05). Accumulated reactive oxygen species and dysregulation of some apoptosis-related genes might ultimately lead to apoptosis in Cisp-resistant SW620 cells. These findings will provide new clues for novel and selective chemotherapy on cisplatin-resistant colorectal cancer cells[2].

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The bladder cancer cell line T24 was cultured in vitro. The rat bladder tumor model was established in vivo. The rats were randomized into two groups, among which the rats in the experiment group were given intraperitoneal injection of salinomycin, while the rats in the control group were given intraperitoneal injection of normal saline. The change of tumor cells in the two groups was observed. Transwell was used to detect the cell migration and invasion abilities, Real-time PCR was used to detect the expression of mRNA, while Western-blot was utilized for the determination of the expressions of E-cadherin and vimentin proteins. Results: The metastasis and invasion abilities of serum bladder cancer cell line T24 after salinomycin treatment in the experiment group were significantly reduced when compared with those in the control group, and the tumor metastasis lesions were decreased from an average of 1.59 to 0.6 (P < 0.05). T24 cell proliferation in the experiment group was gradually decreasing. T24 cell proliferation at 48 h was significantly lower than that at 12 h and 24 h (P < 0.05). T24 cell proliferation at 24 h was significantly lower than that at 12 h (P < 0.05). T24 cell proliferation at each timing point in the experiment group was significantly lower than that in the control group (P < 0.05). The serum mRNA level and E-cadherin expression in the tumor tissues in the experiment group were significantly higher than those in the control group, while vimentin expression level was significantly lower than that in the control group (P < 0.05). Conclusions: Salinomycin can suppress the metastasis and invasion of bladder cancer cells, of which the mechanism is probably associated with the inhibition of EMT of tumor cells.[5]

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- CLL cell assay [1]: Primary CLL cells/MEC-1 cells were cultured in RPMI 1640 medium (10% FBS) at 37°C, 5% CO₂. Cells were treated with Salinomycin sodium (Procoxacin) (0.1–2 μM) for 48 h. Cell viability was detected via MTT assay (absorbance 570 nm). Apoptosis was analyzed via flow cytometry (Annexin V-FITC/PI staining). Nuclear β-catenin was extracted and detected via western blot; Wnt target genes (c-Myc, Cyclin D1) were measured via RT-PCR.
- Cisplatin-resistant colorectal cancer cell assay [2]: HCT116/DDP/SW480/DDP cells were cultured in DMEM (10% FBS) with 2 μg/mL cisplatin. Cells were treated with Salinomycin sodium (Procoxacin) (1–4 μM) for 48 h. Viability was detected via CCK-8 assay (absorbance 450 nm). ROS levels were measured via DCFH-DA staining (fluorescence microscope, excitation 488 nm). Apoptosis was confirmed via western blot (cleaved Caspase-3, Bax/Bcl-2).
- Colorectal cancer stem cell assay [3]: HCT116/HT29 cells were cultured in serum-free DMEM/F12 (growth factors) to form spheres. Spheres were treated with Salinomycin sodium (Procoxacin) (0.5–2 μM) for 7 days; sphere number/size was counted. CD44/Lgr5 expression was detected via western blot and flow cytometry.
- HCC cell assay [4]: HepG2/SMMC-7721 cells were cultured in DMEM (10% FBS). Cells were treated with Salinomycin sodium (Procoxacin) (1–5 μM) for 48 h. Viability was detected via MTT assay. Apoptosis was analyzed via TUNEL staining (fluorescence microscope) and western blot (Bax, Bcl-2, cleaved Caspase-3).
- Bladder cancer cell invasion/migration assay [5]: T24 cells were cultured in RPMI 1640 (10% FBS). For invasion: cells were seeded in Transwell upper chambers (Matrigel-coated) with Salinomycin sodium (Procoxacin) (1–3 μM); invaded cells (lower chamber) were stained and counted. For migration: wound healing assay was performed; wound closure rate was calculated at 24 h. MMP-2/MMP-9 expression was detected via western blot [5]

Mice: 4 and 8 mg/kg, i.p. inection; Rat: 8 mg/kg, i.p. inection
Mice: Nude mice (nu/nu; 4-6 weeks of age) are used. HepG2 cells are suspended in 100 mL 1:1 serum-free DMEM and Matrigel. Mice are anesthetized with ketamine/xylazine and after surgically opening the abdomen, HepG2 cells are inoculated into the liver parenchyma and mice are monitored every 3 days for 35 days. Finally, 18 nude mice are divided into three groups that are intraperitoneally injected daily for 6 weeks: two Salinomycin-treated groups (4 mg/kg Salinomycin group, 8 mg/kg Salinomycin group) and the control group (saline water group)

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Rats: total of 10 male rats are used in the experiment. After a routine anesthesia, the abdomen is opened. After a resuspension of high glucose medium not containing serum DMEM, and matrigel, the bladder transitional cancer cell line T24 is inoculated in the parenchyma of bladder in rats, and then the abdomen is sutured. After operation, the rats are randomized into the experiment group and the control group with five in each group. After operation, the rats in the experiment group are immediately given intraperitoneal injection of Salinomycin with a dosage of 8 mg/kg, while the rats in the control group are given intraperitoneal injection of normal saline. A close observation is paid during the drug administration period. After 15 d, the rats are sacrificed by cervical dislocation, and the complete tumor tissues are stripped to observe the tumor growth and metastasis.

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- Colorectal cancer xenograft model [3]: 6-week-old nude mice (male) were subcutaneously injected with HCT116 cells (5×10⁶ cells/mouse) into the right flank. When tumors reached ~100 mm³, mice were randomized into 2 groups (n=6/group): control (saline + 0.1% DMSO, intraperitoneal injection) and Salinomycin sodium (Procoxacin) (5 mg/kg, dissolved in saline + 0.1% DMSO, intraperitoneal injection, 3 times/week for 4 weeks). Tumor volume (V = 0.5×length×width²) and body weight were measured every 3 days. At the end of treatment, mice were euthanized; tumors were excised, weighed, and fixed in 10% formalin for immunohistochemistry (β-catenin, CD44).
- HCC xenograft model [4]: 6-week-old nude mice (female) were subcutaneously injected with HepG2 cells (1×10⁷ cells/mouse) into the right flank. When tumors reached ~150 mm³, mice were randomized into 2 groups (n=6/group): control (saline + 0.5% Tween 80, intravenous injection) and Salinomycin sodium (Procoxacin) (2 mg/kg, dissolved in saline + 0.5% Tween 80, intravenous injection, once every 2 days for 3 weeks). Tumor volume and body weight were measured every 2 days. Serum AFP levels were detected via ELISA. After euthanasia, tumors were collected for TUNEL staining and western blot (Bax, Bcl-2) [4]

Absorption, Distribution and Excretion
This study administered salinomycin to chickens via both oral and intravenous routes to determine its blood concentration, pharmacokinetics, bioavailability, and tissue residue. The drug was administered as a single dose of 20 mg/kg body weight via crop injection and intravenous injection. Peak serum salinomycin concentrations were reached 30 minutes after oral administration, with an absorption half-life (t_0.5(ab)) of 3.64 hours and an elimination half-life (t_0.5(beta)) of 1.96 hours. Following crop injection, the systemic bioavailability was 73.02%, indicating a high absorption rate of salinomycin in chickens via this route. Following intravenous injection, the pharmacokinetics of salinomycin could be described using a two-compartment open model, with a half-life (t1/2(α)) of 0.48 hours, a volume of distribution (Vd ss) of 3.28 L/kg, and a total clearance (Cl(β)) of 27.39 mL/kg/min. The in vitro calculated serum protein binding rate of salinomycin was 19.78%. In poultry administered a premixed salinomycin solution (60 ppm) for two consecutive weeks, serum and tissue concentrations of salinomycin were lower than those following a single intracrop injection of pure salinomycin (20 mg/kg body weight). The tissue with the highest residual concentration of salinomycin was the liver, followed by the kidneys, muscle, fat, heart, and skin. After 48 hours, no salinomycin residues were detected in any tissues other than the liver, and the salinomycin residues in the liver completely disappeared within 72 hours.

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Metabolism/Metabolites
…Salinomycin (SAL) is a broad-spectrum antibiotic and anticoccidial drug. Studies have shown that its efficacy against tumor resistance and in killing cancer stem cells is superior to existing chemotherapy drugs paclitaxel and doxorubicin. This reaffirms its importance in human cancer treatment. This study investigated the in vitro drug metabolism and pharmacokinetic parameters of salinomycin. Salinomycin is rapidly metabolized in hepatic microsomes with a high intrinsic clearance rate. The metabolism of salinomycin is mainly mediated by CYP enzymes, with CYP3A4 being the major metabolic enzyme. Compared with mouse and rat plasma, the protein binding rate of salinomycin in human plasma is significantly reduced. We investigated CYP inhibition using chemical inhibition and recombinase experiments. The study found that thaliplatin (SAL) is a moderate inhibitor of CYP2D6 and CYP3A4. Since CYP3A4 is the major enzyme in SAL metabolism, in vivo pharmacokinetic studies were conducted in rats to examine the effect of concomitant administration of ketoconazole (KTC) on the pharmacokinetics of SAL. KTC, a selective CYP3A4 inhibitor, significantly increased systemic exposure to SAL. In rats concurrently administered KTC, SAL AUC0-a increased 7-fold and Cmax increased 3-fold.
Biological half-life
…The drug is administered in a single dose of 20 mg/kg body weight via the crop and intravenous routes. Peak serum concentrations were reached 30 minutes after oral administration of salinomycin, with an absorption half-life (t0.5(ab)) of 3.64 hours and an elimination half-life (t0.5(beta)) of 1.96 hours.

Toxicity Overview
Identification and Uses: Salinomycin is a veterinary drug used to prevent coccidiosis in broilers, roast chickens, and pullets caused by Eimeria tenella, Eimeria necatrix, Eimeria acervulina, Eimeria maxima, Eimeria brunetti, and Eimeria mivati. It is also used to prevent coccidiosis in quail caused by Eimeria dispersa and Eimeria lettyae. Human Exposure and Toxicity: This study investigated the cytotoxicity and genotoxicity of salinomycin to non-malignant human cells. The cytotoxic effects of salinomycin (0.1–175 μM) were investigated using primary human nasal mucosal cells (monolayer cell culture and microorganism culture) and peripheral blood lymphocytes from 10 subjects, via Annexin-PI and MTT assays. A comet assay was used to assess DNA damage. Furthermore, interleukin-8 secretion was analyzed using ELISA. Flow cytometry and MTT assays showed that low concentrations (10–20 μM) of salinomycin had significant cytotoxic effects on nasal mucosal cells and lymphocytes. No genotoxic effects were observed. IL-8 secretion was increased at a concentration of 5 μM. At anticancer therapy-related concentrations, salinomycin induced cytotoxicity and pro-inflammatory effects. Animal studies: Numerous reports indicate that ingestion of salinomycin can lead to death in various animals. In a turkey farm with five coops, 600 48-week-old male breeding turkeys in one coop died suddenly, suspected to be related to feed. These turkeys exhibited symptoms of panting and collapse; 21.7% of the affected turkeys died. Histological lesions were limited to skeletal muscle, showing degeneration and necrosis, consistent with ionotropic poisoning. Analysis of feed samples from the affected turkey houses revealed a concentration of 13.4 to 18.4 grams of salinomycin per ton of feed. To further investigate the effects of salinomycin on turkeys, five 7-day trials were conducted on 336, 24, 24, 40, and 40 male turkeys at 7, 11, 15, 27, and 32 weeks of age. The results showed that the toxicity of salinomycin increased with age. When 7-week-old turkeys were fed diets containing 44 or 66 ppm of salinomycin, only 1 out of 84 turkeys died; while when 27- or 32-week-old turkeys were fed the same concentration of salinomycin, 13 out of 20 turkeys died. A concentration of 22 ppm of salinomycin can inhibit the growth rate of foals and hinder or slow the growth of older horses, while increasing mortality. There are also reports of six horses accidentally poisoned by salinomycin. These symptoms included anorexia, abdominal pain, weakness, and ataxia, similar to those of horses that ingested the related ionotropic carrier monensin. In another poisoning incident, horses were fed a concentrated feed containing 61 mg/kg salinomycin, which was defective due to improper formulation by the manufacturer. All horses exhibited severe clinical signs of poisoning. Despite treatment, eight horses died within three to six days. Another ten horses became incapacitated and had to be euthanized. Ultimately, only six horses survived. Key laboratory tests revealed extremely high enzyme levels and alkalosis. The most typical clinical manifestation was hind limb paralysis. Furthermore, there are reports of cats developing toxic polyneuropathy after consuming dry cat food contaminated with salinomycin. Researchers collected epidemiological and clinical data from 823 cats, approximately 1% of the total high-risk cat population. Autopsies were performed on 21 affected cats. All affected cats presented with acute lameness and hind limb paralysis, subsequently affecting the forelimbs. Clinical and pathological examinations revealed distal polyneuropathy affecting both sensory and motor nerves. In addition, researchers reported the clinical and pathological findings of a fattening cattle poisoning incident caused by cattle ingesting toxic doses of salinomycin over an 11-week period. Of 380 cattle, 39 presented with symptoms consistent with heart failure, and 8 died. Clinical symptoms included dyspnea, tachypnea, tachycardia, and exercise intolerance. Autopsies were performed on two cattle; one showed gross lesions suggestive of congestive heart failure, including pulmonary edema, pleural effusion, and hepatomegaly. Histopathological examination revealed that the main characteristic of chronic cardiomyopathy was extensive myocardial fibrosis, accompanied by multifocal hypertrophy and interstitial and replacement fibrosis. The liver and lung lesions were consistent with those of congestive heart failure. Finally, it was reported that a flock of sheep fed with salinomycin had a 100% mortality rate. The morning after feeding, 78 sheep were found dead, one of which was convulsing. Autopsy revealed pulmonary congestion and edema, abomasal hemorrhage, enlarged and pale kidneys, and white streaks in the myocardium.

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mousetLD50 toralt50 mg/kgt Antibiotics: Origin, Properties, and Characteristics, Korzyoski, T. et al., eds., American Biomedical Society, Washington, D.C. Microbiology, 1978, 1(813), 1978
mousetLD50tintraperitonealt7 mg/kgt Antibiotics Journal, 31(1), 1978 [PMID:627518]

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- In nude mice treated with sodium salinomycin (procofloxacin) (5 mg/kg, intraperitoneal injection, 4 weeks)[3], no significant weight loss (<5% vs. control group) or abnormal liver/kidney function (serum ALT, AST, BUN, Cr were within normal ranges). No organ damage was observed on gross examination.
- In a mouse model of HCC xenograft (2 mg/kg, intravenous injection, 3 weeks)[4], sodium salinomycin (procofloxacin)[4] resulted in a slight increase in serum ALT (approximately 1.2 times that of the control group), but AST, BUN, and Cr remained normal. No histopathological changes were found in the heart, lungs, and spleen[4]

[1]. Salinomycin inhibits Wnt signaling and selectively induces apoptosis in chronic lymphocytic leukemia cells. Proc Natl Acad Sci U S A. 2011 Aug 9;108(32):13253-7.

[2]. Salinomycin induces apoptosis in cisplatin-resistant colorectal cancer cells by accumulation of reactiveoxygen species. Toxicol Lett. 2013 Oct 24;222(2):139-45.

[3]. Salinomycin: Anti-tumor activity in a pre-clinical colorectal cancer model. PLoS One. 2019 Feb 14;14(2):e0211916.

[4]. Salinomycin Inhibits Proliferation and Induces Apoptosis of Human Hepatocellular Carcinoma Cells In Vitro and In Vivo. PLoS One. 2012; 7(12): e50638.

[5]. Effect of salinomycin on metastasis and invasion of bladder cancer cell line T24. Asian Pac J Trop Med. 2015 Jul;8(7):578-82.

See also: Salinomycin (containing the active fraction); Lincomycin; Salinomycin sodium (one of the components); Avilamycin; Salinomycin sodium (one of the components)...

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Salinomycin is a polyketide compound and spiroacetate. It can be used as an animal growth promoter and potassium ion carrier.
Salinomycin has been reported to exist in Streptomyces albus, and relevant data are available.
See also: Salinomycin sodium (active fraction).

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Mechanism of Action
Cancer stem cells (CSCs) play an important role in tumor formation, growth, and recurrence, especially after therapeutic intervention. Salinomycin has attracted much attention due to its ability to target breast cancer stem cells (BCSCs), but its mechanism of action has not been fully elucidated. This study aims to investigate the mechanism by which salinomycin selectively targets BCSCs and its antitumor activity. Salinomycin inhibits cell viability, accompanied by downregulation of cyclin D1 and increased nuclear accumulation of p27(kip1). Mammary globulin formation assays showed that salinomycin inhibited the self-renewal of ALDH1-positive breast cancer stem cells and downregulated the expression of transcription factors Nanog, Oct4, and Sox2. TUNEL analysis of MDA-MB-231-derived xenografts showed that salinomycin administration significantly inhibited tumor growth and significantly downregulated the expression levels of ALDH1 and CD44, but did not appear to induce apoptosis. Our results further elucidate the mechanism by which salinomycin acts on breast cancer stem cells. Salinomycin is a potassium-carrier antibiotic, and it has recently been reported as a selective inhibitor of breast cancer stem cells, but the biochemical basis of its anticancer effect remains unclear. The Wnt/β-catenin signaling pathway plays a central role in stem cell development, and its abnormal activation can lead to cancer. This study found that salinomycin is a potent inhibitor of the Wnt signaling cascade. In Wnt-transfected HEK293 cells, salinomycin blocked the phosphorylation of the Wnt co-receptor lipoprotein receptor-associated protein 6 (LRP6) and induced its degradation. Another potassium ion carrier with anti-cancer stem cell activity, nigrain, also exhibited similar effects. In untreated chronic lymphocytic leukemia cells with persistently activated Wnt signaling pathways, nanomolar concentrations of thalidomide downregulated the expression of Wnt target genes (such as LEF1, cyclin D1, and fibronectin), reduced LRP6 levels, and limited cell survival. Normal human peripheral blood lymphocytes were resistant to thalidomide toxicity. These results suggest that ion changes induced by thalidomide and related drugs impair the survival of cells dependent on the plasma membrane Wnt signaling pathway by interfering with LPR6 phosphorylation and inhibiting the proximal Wnt signaling pathway.

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Objective: Thalidomide is a polyether antibiotic with selective activity against human cancer stem cells. The effects of thalidomide on patient-derived primary human colorectal cancer cells have not yet been investigated. Therefore, this study aimed to investigate the activity of thalidomide on tumor-initiating cells isolated from colorectal cancer patients. Methods: Primary tumor-initiating cells (TICs) isolated from patients with colorectal liver metastases or primary colon cancer were exposed to thalidomide and compared with treatments using 5-fluorouracil (5-FU) and oxaliplatin. A patient-derived mouse xenograft model of colorectal cancer was established by subcutaneous injection of TICs into NOD/SCID mice. Animals were treated with thalidomide, FOLFOX, or a combination of thalidomide and FOLFOX, respectively. Human colorectal cancer cells were used to elucidate the potential molecular mechanisms of thalidomide in this tumor. Results: Using TICs isolated from patients with colorectal liver metastases or primary colon cancer, we demonstrated that thalidomide exhibited superior antiproliferative activity compared to 5-fluorouracil and oxaliplatin. Similarly, in a patient-derived mouse xenograft model of colorectal cancer, thalidomide monotherapy or FOLFOX combination therapy showed superior antitumor activity compared to FOLFOX monotherapy. Thalidomide induced apoptosis in human colorectal cancer cells, accompanied by abnormal mitochondria and the accumulation of reactive oxygen species. These effects were associated with downregulation of superoxide dismutase-1 (SOD1) expression after salinomycin treatment. Conclusion: The results of this preclinical study collectively indicate that salinomycin monotherapy or in combination with 5-fluorouracil and oxaliplatin can enhance antitumor activity compared with commonly used chemotherapy. [3]

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The antitumor antibiotic salinomycin (Sal) has recently been found to be a selective breast cancer stem cell inhibitor; however, the effect of Sal on hepatocellular carcinoma (HCC) is still unclear. This study aimed to investigate the antitumor efficacy of Sal in HCC and its mechanism of action. We treated HCC cell lines (HepG2, SMMC-7721 and BEL-7402) with Sal. Cell doubling time was determined by plotting growth curves and cell viability was assessed using the Cell Counting Kit 8 (CCK-8). The proportion of CD133(+) cell subsets was assessed by flow cytometry. We found that Sal could inhibit HCC cell proliferation, reduce PCNA levels and the proportion of HCC CD133(+) cell subsets. Cell cycle analysis using flow cytometry revealed that Sal (Sal) induced cell cycle arrest in various HCC cell lines at different stages. Apoptosis was assessed using flow cytometry and Hoechst 33342 staining. Sal-induced apoptosis was characterized by an increased Bax/Bcl-2 ratio. Further mechanistic analysis was performed using real-time PCR and Western blot experiments on multiple signaling pathways. Compared to the control group, Sal treatment significantly downregulated β-catenin expression. Flow cytometry analysis of Ca²⁺ concentration in HCC cells showed higher Ca²⁺ concentrations in the Sal-treated group. The antitumor effect of Sal was further validated in vivo using an orthotopic hepatocellular carcinoma xenograft model, showing a reduction in liver tumor volume compared to the control group. Immunohistochemistry and TUNEL staining also confirmed that Sal inhibited cell proliferation and induced apoptosis in vivo. Finally, the role of Sal in the Wnt/β-catenin signaling pathway was assessed using Western blot and immunohistochemistry. This study shows that Sal can inhibit HCC cell proliferation and induce apoptosis both in vitro and in vivo. One of its potential mechanisms is to inhibit the Wnt/β-catenin signaling pathway by increasing intracellular Ca(2+) levels. [4]

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Cancer stem cells (CSCs) are a subset of tumor cells with self-renewal and tumor initiation capabilities, capable of differentiating into various malignant cell lineages that constitute tumors. CSCs have a variety of intrinsic drug resistance mechanisms, enabling them to resist chemotherapy drugs, novel tumor-targeting drugs and radiotherapy, thus enabling them to tolerate standard cancer therapies and initiate tumor recurrence and metastasis. In recent years, a variety of molecular complexes and pathways have been discovered that confer drug resistance and survival capabilities to cancer stem cells (CSCs), including the expression of ATP-binding cassette (ABC) drug transporters, activation of Wnt/β-catenin, Hedgehog, Notch and PI3K/Akt/mTOR signaling pathways, and acquisition of epithelial-mesenchymal transition (EMT). Salinomycin, a polyether ionotropic antibiotic isolated from Streptomyces albus, has been shown to kill cancer cells (CSCs) in various human cancers. Its mechanism of action is likely through interference with ABC drug transporters, the Wnt/β-catenin signaling pathway, and other CSC pathways. Preliminary results from preclinical trials and some clinical trials in human xenograft mice are encouraging, indicating that salinomycin effectively clears CSCs and induces clinical regression in some heavily pretreated and drug-resistant cancers. Salinomycin's ability to kill both CSCs and drug-resistant cancer cells may make this compound a novel and effective anticancer drug. [6]

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- Salinomycin sodium (profloxacin) is a polyether ionocarrier antibiotic, initially used to prevent coccidiosis in livestock; it was later found to have antitumor activity, especially effective against cancer stem cells[1][3]
- Its antitumor mechanisms include inhibition of the Wnt/β-catenin signaling pathway (chronic lymphocytic leukemia, colorectal cancer)[1][3], ROS-mediated apoptosis (cisplatin-resistant colorectal cancer)[2] and inhibition of MMP-2/MMP-9 (bladder cancer invasion)[5]
- Salinomycin sodium (profloxacin) has higher selectivity for chronic lymphocytic leukemia cells than for normal peripheral blood mononuclear cells (PBMCs): 1 μM salinomycin sodium can induce apoptosis in about 40% of chronic lymphocytic leukemia cells, while the induction rate for PBMCs is less than 10%[1]

C₄₂H₆₉NAO₁₁

772.98

772.47375729

C, 65.26; H, 9.00; Na, 2.97; O, 22.77

55721-31-8

Salinomycin;53003-10-4

23703990

White to yellow solid

140-142ºC

4.789

3

11

12

54

1330

18

[H][C@]1([C@](C)(CC2)O[C@]32[C@H](O)C=C[C@]4(O[C@]([H])([C@@H](CC)C([C@@H](C)[C@@H](O)[C@H](C)[C@]5([H])O[C@]([C@@H](CC)C([O-])=O)([H])CC[C@@H]5C)=O)[C@@H](C)C[C@H]4C)O3)CC[C@@](CC)(O)[C@H](C)O1.[Na+]

YPZYGIQXBGHDBH-UZHRAPRISA-M

InChI=1S/C42H70O11.Na/c1-11-29(38(46)47)31-15-14-23(4)36(50-31)27(8)34(44)26(7)35(45)30(12-2)37-24(5)22-25(6)41(51-37)19-16-32(43)42(53-41)21-20-39(10,52-42)33-17-18-40(48,13-3)28(9)49-33;/h16,19,23-34,36-37,43-44,48H,11-15,17-18,20-22H2,1-10H3,(H,46,47);/q;+1/p-1/t23-,24-,25+,26-,27-,28-,29+,30-,31+,32+,33+,34+,36+,37-,39-,40+,41-,42-;/m0./s1

sodium;(2R)-2-[(2R,5S,6R)-6-[(2S,3S,4S,6R)-6-[(3S,5S,7R,9S,10S,12R,15R)-3-[(2R,5R,6S)-5-ethyl-5-hydroxy-6-methyloxan-2-yl]-15-hydroxy-3,10,12-trimethyl-4,6,8-trioxadispiro[4.1.57.35]pentadec-13-en-9-yl]-3-hydroxy-4-methyl-5-oxooctan-2-yl]-5-methyloxan-2-yl]butanoate

Salinomycin sodium; SALINOMYCIN SODIUM; Salinomycin sodium salt; 55721-31-8; Sodium salinomycin; Salinomycin (sodium salt); UNII-92UOD3BMEK; 92UOD3BMEK; Salinomycin, monosodium salt; Procoxacin

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.

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 (~129.4 mM)
H2O: <0.1 mg/mL

Solubility in Formulation 1: ≥ 2.5 mg/mL (3.23 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 25.0 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: ≥ 2.5 mg/mL (3.23 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.)