Eprinomectin (2.5-50 μM; 0-48 h) significantly inhibits DU145 cell viability (IC50: 12.5 μM), colony formation and migration ability, and induces G0/G1 phase cell cycle arrest [1].
Eprinomectin (10-50 μM; 48 h) induces apoptosis and autophagy, increases reactive oxygen species production and endoplasmic reticulum stress, and promotes β-catenin nucleocytoplasmic translocation in DU145 cells [1].

Eprinomectin (2.5-50 μM; 0-48 h) significantly inhibits DU145 cell viability (IC50: 12.5 μM), colony formation and migration ability, and induces G0/G1 phase cell cycle arrest [1].
Eprinomectin (10-50 μM; 48 h) induces apoptosis and autophagy, increases reactive oxygen species production and endoplasmic reticulum stress, and promotes β-catenin nucleocytoplasmic translocation in DU145 cells [1].

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Eprinomectin significantly inhibits the viability of prostate cancer (PCa) cell lines (DU145, LNCaP, VCaP, 22RV1) in a concentration-dependent manner. The half maximal inhibitory concentration (IC50) in DU145 cells is 12.5 µM. Maximum inhibition of viability was observed at 50 µM. DU145 cells (hormone-negative) showed higher sensitivity compared to other cell lines. [1]
Eprinomectin robustly reduces both the number and size of colonies formed by DU145 cells in a colony formation assay, showing approximately 50% reduction at 5 µM. [1]
Eprinomectin inhibits the migratory capacity of DU145 cells in a wound-healing assay in a concentration- and time-dependent manner at concentrations of 10 µM and 25 µM. [1]
Eprinomectin induces cell cycle arrest at the G0/G1 phase in DU145 cells, with the percentage of cells in G1 increasing from 49.68% (control) to 71.7% (50 µM EP). It downregulates the expression of cell cycle markers cyclin D1, cyclin D3, CDK4, and c-Myc. [1]
Eprinomectin downregulates the expression of cancer stem cell (CSC) markers (Oct3/4, Nanog, Sox-2, ALDH1, CD44) and genes regulating CSC properties (β-catenin, c-Myc, cyclin D1) in DU145 cells as shown by real-time PCR. Immunofluorescence confirms the delocalization of β-catenin from the nucleus to the cytoplasm. [1]
Eprinomectin inhibits the activity of the alkaline phosphatase (ALP) enzyme in DU145 cells in a concentration-dependent manner. [1]
Eprinomectin induces apoptosis in DU145 cells in a concentration-dependent manner, as shown by Annexin V/PI staining and flow cytometry. It activates cleaved caspase-3, increases pro-apoptotic BAD and DNA damage marker pH2AX, decreases PARP-1, and downregulates anti-apoptotic markers (MCL-1, c-IAP-1, XIAP, survivin, TCTP). The antiproliferative effect is attenuated by a pan-caspase inhibitor. [1]
Eprinomectin induces reactive oxygen species (ROS) production in DU145 cells in a concentration-dependent manner. The antiproliferative effect is attenuated by the antioxidant N-acetyl cysteine (NAC). [1]
Eprinomectin induces autophagy in DU145 cells, evidenced by increased expression of autophagy markers Beclin-1, Atg-5, LC-3II, and p62 via immunofluorescence and western blot. [1]
Eprinomectin induces endoplasmic reticulum (ER) stress in DU145 cells, shown by increased expression of ER stress markers PERK, Ero1-α, and PDI. This ER stress is suggested to contribute to the induction of autophagy. [1]

Eprinomectin (0.08-0.5 mg/kg; Local pipette delivery; 14 days) has insecticidal, insecticidal and acaricidal activities in cattle[2]. Pharmacokinetic Analysis in cattle[1]

Cell Viability Assay[1]
Cell Line: DU145 cells
Concentration: 2.5, 5, 10, 25 and 50 μM
Incubation Time: 48 h
Result: Significantly inhibited the cell viability.

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Western Blot Analysis[1]
Cell Line: DU145 cells
Concentration: 10, 25 and 50 μM
Incubation Time: 48 h
Result: Promotes the protein expression of pro-apoptotic marker BAD and DNA damage marker pH2AX and caspase-9 in a concentration-dependent manner. Inhibits DNA repair by reducing PARP-1.

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Cell Viability Assay (MTT): Prostate cancer cells (DU145, LNCaP, VCaP, 22RV1) were seeded in 96-well plates and allowed to reach 60-70% confluency. Cells were treated with various concentrations of Eprinomectin (2.5 to 100 µM) for 48 hours. MTT dye was then added and incubated for 3 hours. After solubilizing the formed crystals, absorbance was measured at 590 nm to calculate the percentage of cell viability relative to untreated controls. [1]
Colony Formation Assay: Approximately 500 DU145 cells were seeded in a six-well plate. After attachment, cells were treated with various concentrations of Eprinomectin (5 to 100 µM) and cultured for 2 weeks with regular media changes. Colonies were then fixed, stained with crystal violet, rinsed, dried, and counted under a light microscope. [1]
Wound-Healing/Migration Assay: DU145 cells were grown to 100% confluency in 24-well plates. A scratch was made using a pipette tip, and the detached cells were removed. Fresh media containing Eprinomectin (10 or 25 µM) was added. Images of the wound were taken at 0, 24, and 48 hours using a light microscope. The percentage of open wound area was calculated using software to assess migration inhibition. [1]
Cell Cycle Analysis by Flow Cytometry: DU145 cells treated with Eprinomectin (10, 25, 50 µM) for 48 hours were harvested, fixed, treated with RNase A, and stained with propidium iodide (PI). The DNA content was analyzed using a flow cytometer to determine the distribution of cells in different cell cycle phases. [1]
Gene Expression Analysis (Real-time PCR): Total RNA was extracted from DU145 cells treated with or without Eprinomectin (25 µM, 48h) using a triazole-chloroform method. cDNA was synthesized and used for real-time PCR to analyze the expression levels of target genes (e.g., CSC markers, β-catenin pathway genes). Relative gene expression was calculated using the 2^(-ΔΔCt) method. [1]
Alkaline Phosphatase (ALP) Assay: DU145 cells treated with Eprinomectin (10, 25, 50 µM) for 48 hours were fixed. A freshly prepared ALP staining solution was added and incubated in the dark. After removing the stain, cells were observed under a light microscope, and staining intensity was quantified using image analysis software. [1]
Apoptosis Assay (Annexin V/PI Staining & Flow Cytometry): For microscopy, DU145 cells treated with Eprinomectin were stained with Annexin V-FITC and PI and analyzed using a confocal microscope. For flow cytometry, treated cells were stained with Annexin V and PI, and at least 10,000 events were analyzed to quantify the percentage of cells in early and late apoptosis. [1]
Immunofluorescence: DU145 cells grown on chamber slides were treated with Eprinomectin, fixed, permeabilized, and blocked. They were then incubated with primary antibodies against target proteins (e.g., β-catenin, cleaved caspase-3, Beclin-1, LC-3, PERK, PDI), followed by incubation with FITC-labeled secondary antibodies. Nuclei were counterstained with DAPI. Fluorescence images were captured using a confocal microscope and analyzed for intensity. [1]
Reactive Oxygen Species (ROS) Measurement: DU145 cells treated with Eprinomectin were incubated with the fluorescent probe DCFDA. After washing, cells were viewed under a fluorescent microscope. The fluorescence intensity, proportional to ROS levels, was quantified using image analysis software. [1]
Western Blotting: Protein lysates were prepared from DU145 cells treated with Eprinomectin. Equal amounts of protein were separated by SDS-PAGE and transferred to a nitrocellulose membrane. Membranes were blocked, probed with specific primary antibodies overnight, followed by appropriate secondary antibodies. Protein bands were visualized using enhanced chemiluminescence (ECL) and quantified using image analysis software. β-actin was used as a loading control. [1]

Animal Model: Infective nematode or chorioptes bovis treated male Jersey calves[2]
Dosage: 0.08, 0.16, 0.2, 0.24 and 0.5mg/kg
Administration: Delivered topically from the tailhead to the withers at skin level via a pipette; 14 days
Result:At the lowest dose (0.08 mg/kg), maximal or near-maximal efficacy was achieved against most adult endoparasites, with the exception of Capillaria serpiginosa (87%) and Capillaria cysticercus (88%).
Control was ≥ 95% at all dose levels at day 14 post-treatment and was maintained at or near this efficacy level over the 6-week trial period.

[1]. Eprinomectin: a derivative of ivermectin suppresses growth and metastatic phenotypes of prostate cancer cells by targeting the β-catenin signaling pathway. J Cancer Res Clin Oncol. 2023 Sep;149(11):9085-9104.

[2]. Eprinomectin: a novel avermectin for use as a topical endectocide for cattle. Int J Parasitol. 1996 Nov;26(11):1237-42.

[3]. Injectable eprinomectin for cattle: Tick efficacy and pharmacokinetics. J Vet Pharmacol Ther. 2020 Mar;43(2):171-178.

Eprinomectin is a semi-synthetic macrolide derivative of ivermectin and belongs to the avermectin class of drugs. It was originally used as a broad-spectrum antiparasitic drug for cattle. [1]
This study proposes a multifaceted mechanism of action of eprinnomectin against metastatic castration-resistant prostate cancer (mCRPC) cells: it induces cell cycle arrest, apoptosis (via caspase activation and reactive oxygen species (ROS), autophagy (via endoplasmic reticulum stress), and inhibits cancer stem cell-like characteristics by disrupting the β-catenin signaling pathway. [1]
This study suggests that eprinnomectin has the potential to be used as a novel therapeutic agent for advanced prostate cancer, especially metastatic and drug-resistant phenotypes. [1]

C50H75NO14;C49H73NO14

914.13

123997-26-2

Eprinomectin-d3

White to off-white solid powder

1.23

163-166 ºC

2 °C

5.469

C[C@H]1[C@@H](NC(C)=O)[C@@H](OC)C[C@H](O[C@@H]2[C@H](C)O[C@@H](O[C@@H](/C(C)=C/C[C@@H](C3)OC4(O[C@@H]([C@@H](CC)C)[C@H](C)C=C4)C[C@H]3OC5=O)[C@@H](/C=C/C=C6CO[C@@H]7[C@@]\6(O)[C@H]5C=C(C)[C@H]7O)C)C[C@@H]2OC)O1

WPNHOHPRXXCPRA-TVXIRPTOSA-N

InChI=1S/C49H73NO14/c1-25(2)43-28(5)17-18-48(64-43)23-35-20-34(63-48)16-15-27(4)44(26(3)13-12-14-33-24-57-46-42(52)29(6)19-36(47(53)60-35)49(33,46)54)61-40-22-38(56-11)45(31(8)59-40)62-39-21-37(55-10)41(30(7)58-39)50-32(9)51/h12-15,17-19,25-26,28,30-31,34-46,52,54H,16,20-24H2,1-11H3,(H,50,51)/b13-12-,27-15-,33-14-/t26-,28-,30-,31-,34+,35-,36-,37+,38-,39-,40-,41+,42+,43+,44-,45+,46+,48+,49+/m0/s1

Avermectin B1, 4''-(acetylamino)-4''-deoxy-, (4''R)-

MK 397; MK-397; MK397; Eprinex; 4'-epi-acetylamino-4'-deoxy-avermectin B1a

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)

DMSO : 100~125 mg/mL (~136.74 )
Ethanol :~ 100 mg/mL

Solubility in Formulation 1: ≥ 2.5 mg/mL (2.73 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 (2.73 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication.
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 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: ≥ 2.5 mg/mL (2.73 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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Solubility in Formulation 4: 10% DMSO+40% PEG300+5% Tween-80+45% Saline: ≥ 2.5 mg/mL (2.73 mM)

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