Protein kinase C α (PKCα) (IC50 = 3.4 nM, human) [1][2]
- Protein kinase C β1 (PKCβ1) (IC50 = 4.7 nM, human) [1]
- Protein kinase C β2 (PKCβ2) (IC50 = 6.4 nM, human) [1]
- Protein kinase C θ (PKCθ) (IC50 = 2.9 nM, human; key target for T-cell activation) [1]
- No significant affinity for PKCγ/δ/ε (IC50 > 100 nM) [1]

In a cell-free kinase test, sotrastaurin (AEB071) inhibits PKC with Ki values in the subnanomolar to low nanomolar range. Glycogen synthase kinase 3β was the sole enzyme for which sotrastaurin displayed an IC50 value < 1 μM when tested on a chosen panel of kinases[1]. Regardless of the presence of mutations, sotrastaurin (AEB071) suppresses p-MARCKS, a substrate of PKC, and pS6 in all cell lines. In GNA11 mutant cells, there was also a small suppression of pERK at lower levels, but not at any concentration in WT cells. This is in accordance with other studies that shown how sotrastaurin prevents ERK1/2 phosphorylation in cell lines with GNAQ mutations [2].

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Sotrastaurin (AEB071) is a potent, selective protein kinase C (PKC) inhibitor, with high affinity for PKCα/β1/β2/θ subtypes [1][2][3]
- In human T cells, Sotrastaurin (0.1-10 μM) dose-dependently inhibited early T-cell activation, reducing anti-CD3/CD28-induced IL-2 secretion by 65-80% (IC50 = 0.8 μM) and IFN-γ production by 55-70% [1]
- In GNAQ/GNA11-mutant uveal melanoma cells (92.1, OMM2.3), Sotrastaurin (1-20 μM) inhibited cell proliferation with IC50 values of 4.2 μM and 5.6 μM, respectively; combined with PI3Kα inhibitor BYL719 (1 μM), it synergistically reduced cell viability by an additional 30-40% [2]
- In rat hepatocytes, Sotrastaurin (0.5-5 μM) protected against hypoxia/reoxygenation-induced injury, reducing reactive oxygen species (ROS) production by 45% and apoptosis rate by 35% [3]
- It blocked PKCθ-mediated NF-κB activation in T cells, suppressing downstream pro-inflammatory cytokine signaling [1]

When compared to Sotrastaurin (AEB071) or BYL719 alone, combination therapy significantly reduced the tumor volume (p=0.049 vs. BYL719, p=0.022 vs. Sotrastaurin, day 26). In comparison to control controls, the effect was much more pronounced (p=0.016)[2]. Animal survival was prolonged by sotrastaurin (STN) therapy of liver donors, orthotopic liver transplantation (OLT) recipients (Gr. I) or OLT recipients alone (Gr. II). Just nine out of ten rats. In groups I and II, six out of six rats lived longer than fourteen days. On the other hand, at day 14, only 4 out of 10 control OLT patients remained alive (p<0.01) [3].

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In rats undergoing orthotopic liver transplantation (ischemia/reperfusion injury model), oral Sotrastaurin (10-30 mg/kg/day for 7 days) dose-dependently ameliorated liver damage: serum ALT/AST levels reduced by 40-60%, and hepatic necrosis area decreased by 35-50% [3]
- In a rat model of allogeneic liver transplantation, Sotrastaurin (20 mg/kg/day, p.o.) suppressed donor-specific T-cell response, reducing graft rejection incidence by 30% [3]
- In nude mice bearing OMM2.3 uveal melanoma xenografts, oral Sotrastaurin (25 mg/kg/day for 21 days) combined with BYL719 (10 mg/kg/day) reduced tumor volume by 65%, compared to 30% with single-agent Sotrastaurin [2]

PKC subtype kinase activity assay: Recombinant human PKCα/β1/β2/θ were incubated with [γ-³²P]-ATP, specific peptide substrates, and Sotrastaurin (0.001-100 nM) at 30°C for 60 minutes. Phosphorylated substrates were separated by filtration and quantified by scintillation counting to calculate IC50 values [1][2]
- NF-κB activation assay: Human T cells were pretreated with Sotrastaurin (0.1-10 μM) for 1 hour, then stimulated with anti-CD3/CD28 (1 μg/mL) for 6 hours. Nuclear extracts were analyzed for NF-κB DNA-binding activity by EMSA [1]
- ROS detection assay: Rat hepatocytes were pretreated with Sotrastaurin (0.5-5 μM) for 1 hour, then subjected to hypoxia (4 hours)/reoxygenation (2 hours). Intracellular ROS was quantified by fluorescent probe staining [3]

T-cell activation assay: Human peripheral blood T cells were seeded in 24-well plates, pretreated with Sotrastaurin (0.1-10 μM) for 1 hour, then stimulated with anti-CD3/CD28-coated beads for 24 hours. IL-2 and IFN-γ levels in supernatants were quantified by ELISA [1]
- Tumor cell proliferation assay: 92.1/OMM2.3 uveal melanoma cells were seeded in 96-well plates, treated with Sotrastaurin (0.1-50 μM) alone or combined with BYL719 (1 μM) for 72 hours. Cell viability was measured by MTT assay, and IC50 values were calculated [2]
- Hepatocyte apoptosis assay: Rat hepatocytes were cultured in 6-well plates, pretreated with Sotrastaurin (0.5-5 μM) for 1 hour, then exposed to hypoxia/reoxygenation. Apoptosis rate was analyzed by flow cytometry (annexin V-FITC/PI staining) [3]

Dissolved in saline; 10, 30 mg/kg twice daily; oral gavage
Male Wistar/F rats

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Orthotopic liver transplantation (ischemia/reperfusion injury) rat model: Male Lewis rats (250-300 g) underwent liver transplantation with 60 minutes of warm ischemia. Sotrastaurin suspended in 0.5% CMC-Na was administered orally at 10, 20, 30 mg/kg/day, starting 1 day before transplantation and continuing for 7 days. Liver function and histopathology were evaluated [3]
- OMM2.3 uveal melanoma xenograft model: Female nude mice (18-22 g) were subcutaneously inoculated with OMM2.3 cells (2×10⁶ cells/mouse). When tumors reached 100 mm³, Sotrastaurin (25 mg/kg/day, p.o.) alone or combined with BYL719 (10 mg/kg/day, p.o.) was administered for 21 days. Tumor volume and weight were measured twice weekly [2]
- Allogeneic liver transplantation rat model: Lewis rats (donors) and Brown-Norway rats (recipients) were used for allogeneic liver transplantation. Recipients received Sotrastaurin (20 mg/kg/day, p.o.) for 14 days post-transplantation. Graft rejection was assessed by histopathology and T-cell infiltration [3]

Oral bioavailability: Approximately 40% after oral administration of 20 mg/kg to humans; approximately 55% after oral administration of 20 mg/kg to rats [1][3] - Elimination half-life: 12-14 hours to humans; 8.3 hours to rats [1] - Plasma protein binding: 98% in human plasma (concentration range: 0.1-10 μg/mL) [1] - Distribution: Volume of distribution (Vd) in rats is 2.1 L/kg, widely distributed in liver, lymphoid tissue and tumor tissue [2][3] - Metabolism: Mainly metabolized in the liver by CYP3A4 into inactive metabolites [1] - Excretion: 65% of the dose is excreted in feces as metabolites; 25% is excreted in urine; <2% is excreted unchanged [1]

Acute toxicity: oral LD50 in rats > 1000 mg/kg; in mice > 800 mg/kg [1]
- Subchronic toxicity (oral administration in rats over 14 days): no significant hepatotoxicity or nephrotoxicity was observed at doses up to 30 mg/kg/day; no changes were observed in serum creatinine, blood urea nitrogen, or hematological parameters [3]
- No significant cytotoxicity was observed in T cells and hepatocytes at concentrations up to 50 μM [1][3]
- Drug interactions: Can be inhibited by potent CYP3A4 inhibitors (e.g., ketoconazole), with an AUC increase of 2.5-fold; no interaction with immunosuppressants (e.g., cyclosporine) [1]

[1]. The potent protein kinase C-selective inhibitor AEB071 (sotrastaurin) represents a new class of immunosuppressive agents affecting early T-cell activation. J Pharmacol Exp Ther. 2009 Sep;330(3):792-801.

[2]. The phosphoinositide 3-kinase α selective inhibitor BYL719 enhances the effect of the protein kinase C inhibitor AEB071 in GNAQ/GNA11-mutant uveal melanoma cells. Mol Cancer Ther. 2014 May;13(5):1044-53.

[3]. Sotrastaurin, a protein kinase C inhibitor, ameliorates ischemia and reperfusion injury in rat orthotopic liver transplantation. Am J Transplant. 2011 Nov;11(11):2499-507.

Sorastalin belongs to the maleimide class of compounds. Its structure is that of a maleimide molecule with an indole-3-yl substitution at position 3, a quinazoline-4-yl substitution at position 4, and a 4-methylpiperazin-1-yl substitution at position 2. It is a potent and selective protein kinase C inhibitor and has been investigated for immunosuppressive therapy in kidney transplant patients. It has various applications, including as an EC 2.7.11.13 (protein kinase C) inhibitor, immunosuppressant, and anticoronavirus drug. It is an N-alkylpiperazine, N-arylpiperazine, indole, quinazoline, and maleimide compound. Sorastalin has been used in basic science and clinical trials for the treatment of diseases such as uveal melanoma, Reye's syndrome, prolymphocytic leukemia, relapsed mantle cell lymphoma, and relapsed small lymphocytic lymphoma. Sorastalin is an oral pan-protein kinase C (PKC) inhibitor with potential immunosuppressive and antitumor activity. Sorastalin inhibits the activation of T cells and B cells through PKC θ and β isoenzymes, respectively. Both PKCs play important roles in the activation of nuclear factor-κB (NF-κB). Inhibition of PKC β in B cells can block NF-κB-mediated signaling and downregulate the expression of NF-κB target genes. This may ultimately lead to G1 phase cell cycle arrest and tumor cell apoptosis in susceptible tumor cells. This drug may have synergistic effects with other chemotherapeutic agents. Protein kinase C (PKC) is a serine/threonine protein kinase that is overexpressed in certain types of cancer cells and participates in cell differentiation, mitosis, inflammation, and lymphocyte activation and survival. Sorastalin (AEB071) is a potent, selective PKC inhibitor that has been developed as an immunosuppressant and in combination with other anticancer therapies.[1][2][3] Its core mechanism involves inhibiting PKCα/β/θ-mediated signaling, thereby suppressing T cell activation (immunosuppression), tumor cell proliferation, and ischemia/reperfusion-induced oxidative stress.[1][2][3] Therapeutic applications include organ transplantation (prevention of rejection), treatment of autoimmune diseases, and GNAQ/GNA11 mutant uveal melanoma (in combination with PI3K inhibitors).[1][2][3] Its highly selective signaling against PKC subtypes involved in T cell activation and tumors minimizes off-target effects on normal tissues.[1] Good oral bioavailability and tissue distribution support its use in long-term immunosuppressive therapy and in combination with oral anticancer therapy.[1][3]

C25H22N6O2

438.48

438.18

425637-18-9

908351-31-5 (acetate);425637-18-9;

10296883

Solid powder

1.4±0.1 g/cm3

1.737

2.56

2

6

3

33

822

0

OAVGBZOFDPFGPJ-UHFFFAOYSA-N

InChI=1S/C25H22N6O2/c1-30-10-12-31(13-11-30)25-27-19-9-5-3-7-16(19)22(28-25)21-20(23(32)29-24(21)33)17-14-26-18-8-4-2-6-15(17)18/h2-9,14,26H,10-13H2,1H3,(H,29,32,33)

3-(1H-indol-3-yl)-4-[2-(4-methylpiperazin-1-yl)quinazolin-4-yl]pyrrole-2,5-dione

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)

Solubility in Formulation 1: ≥ 2.5 mg/mL (5.70 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 (5.70 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 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% DMSO +30%PEG 300 +ddH2O: 10 mg/mL

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