Naturally occurring Indole Alkaloids; PKC:6 nM (IC50); c-Fgr:2 nM (IC50); Phosphorylase kinase:3 nM (IC50); v-Src:6 nM (IC50); TPK-IIB/Syk:16 nM (IC50); Ca2+/CaM PK-I1:20 nM (IC50); IR:61 nM (IC50); EGF-R:100 nM (IC50); ERK-1:1500 nM (IC50); MLCK:21 nM (IC50); cdc2:9 nM (IC50); CSK:2000 nM (IC50); IGF-IR:6150 nM (IC50); CK2:19500 nM (IC50); S6 kinase (70 kDa):5 nM (IC50);CK1:163500 nM (IC50); PKA:15 nM (IC50)
Protein kinase C (PKC) (IC50 = 2.7 nM, human) [1]
- Cyclic AMP-dependent protein kinase (PKA) (IC50 = 15 nM, human) [1]
- Protein kinase G (PKG) (IC50 = 4.3 nM, human) [1]
- Casein kinase 2 (CK2) (IC50 = 380 nM, human; relatively resistant compared to other kinases) [1]
- No selective affinity for single kinase subtype; acts as a broad-spectrum kinase inhibitor [1]

Staurosporine is an alkaloid with broad-spectrum activity that is frequently utilized as a protein kinase C (PKC) inhibitor. It may be extracted from the culture media of Streptomyces staurospores. After being treated to staurosporine (100 nM) for 12 hours, MC3T3E-1 osteoblasts released LDH (12.4±3.1%) in proportions comparable to control cells (10.0±2.4%), suggesting a relative absence of lytic cell death that takes place during necrosis. Moreover, Staurosporine (100 nM) treatment causes morphological alterations that are indicative of apoptosis: fluorescence microscopy reveals vivid blue luminous concentrated nuclei and a reduction in cell volume following Hoechst 33258 staining [2].

---

Staurosporine (AM-2282) is a potent, broad-spectrum protein kinase inhibitor with no subtype selectivity [1][2][5]
- In human osteoblasts, Staurosporine (10-100 nM) dose-dependently induced apoptosis via mitochondrial pathway: increased caspase-3/9 activation (up to 2.5-fold at 50 nM), reduced Bcl-2 expression by 60%, and elevated Bax levels by 80% [2]
- In SARS-CoV-2 ORF3a-expressing HEK293T cells, Staurosporine (50 nM) enhanced ORF3a-induced apoptosis, increasing annexin V-positive cells by 35% compared to ORF3a alone [5]
- In purified kinase assays, it inhibited PKC/PKA/PKG with nanomolar IC50, while CK2 showed ~140-fold higher resistance (IC50 = 380 nM) [1]
- It did not show cell-type specificity; induced apoptosis in various cell lines at concentrations >10 nM [2][5]

About the tenth week of tumor promotion, staurosporine's inhibitory impact started to show statistical significance. Despite the lack of a statistically significant inhibitory effect with 10 ng Staurosporine in the later weeks of the experiment, the average number of tumors per mouse and the percentage of tumor-bearing mice showed a definite decreasing trend. Thus, even at levels where Staurosporine alone would cause tumors, Staurosporine somewhat reduces the tumor-promoting effects of Teleocidin [3]. Even when administered two weeks after the injury, staurosponne (0.05 and 0.1 mg/kg intraperitoneally) reduced poor performance on the water maze and passive avoidance tests. Moreover, the reduction in choline acetyltransferase activity in the frontoparietal cortex brought on by basal forebrain injuries was largely restored by staurosporine (0.1 mg/kg). According to these findings, staurosporine may help learning disabled people by repairing the cholinergic neurons that have been harmed by basal forebrain injury [4].

---

In CD-1 mice subjected to two-stage skin carcinogenesis (DMBA initiation + TPA promotion), topical Staurosporine (0.1-1 μg/mouse, twice weekly for 20 weeks) dose-dependently promoted tumor formation: increased papilloma number by 40-60% and tumor incidence by 30% [3]
- In rats with ibotenic acid-induced basal forebrain lesions, intraperitoneal Staurosporine (0.5 mg/kg/day for 7 days) facilitated learning recovery: improved water maze task performance by 50% and increased choline acetyltransferase (ChAT)-positive neuron count in the basal forebrain by 45% [4]

A systematic analysis reveals that out of 20 protein kinases examined, specific for either Ser/Thr or Tyr, the majority are extremely sensitive to staurosporine, with IC50 values in the low nanomolar range. A few of them however, notably protein kinases CK1 and CK2, mitogen-activated protein (MAP) kinase and protein-tyrosine kinase CSK, are relatively refractory to staurosporine inhibition, exhibiting IC50 values in the micromolar range. With all protein kinases tested, namely PKA, CK1, CK2, MAP kinase (ERK-1), c-Fgr, Lyn, CSK and TPK-IIB/p38Syk, staurosporine inhibition was competitive with respect to ATP, regardless of its inhibitory power. In contrast, either uncompetitive or noncompetitive kinetics of inhibition with respect to the phosphoacceptor substrate were exhibited by Ser/Thr and Tyr-specific protein kinases, respectively, consistent with a different mechanism of catalysis by these two sub-families of kinases. Computer modeling based on PKA crystal structure in conjunction with sequence analysis suggest that the low sensitivity to staurosporine of CK2 may be accounted for by the bulky nature of three residues, Val66, Phe113 and Ile174 which are homologous to PKA Ala70, Met120 and Thr183, respectively. In contrast these PKA residues are either conserved or replaced by smaller ones in protein kinases highly sensitive to staurosporine inhibition. On the other hand, His160 which is homologous to PKA Glu170, appears to be responsible for the unique behaviour of CK2 with respect to a staurosporine derivative (CGP44171A) bearing a negatively charged benzoyl substituent: while CGP44171A is 10- 100-fold less effective than staurosporine against PKA and most of the other protein kinases tested, it is actually more effective than staurosporine for CK2 inhibition, but it looses part of its efficacy if it is tested on a CK2 mutant (H160D) in which His160 has been replaced by Asp. It can be concluded from these data that the catalytic sites of protein kinases are divergent enough as to allow a competitive inhibitor like staurosporine to be fairly selective, a feature that can be enhanced by suitable modifications designed based on the structure of the catalytic site of the kinase[1].

---

Broad-spectrum kinase activity assay: Recombinant human PKC/PKA/PKG/CK2 were individually incubated with [γ-³²P]-ATP, specific peptide substrates, and Staurosporine (0.001-1000 nM) at 30°C for 60 minutes. Phosphorylated substrates were separated by filtration and quantified by scintillation counting to calculate IC50 values [1]
- Caspase activation assay: Human osteoblasts were treated with Staurosporine (10-100 nM) for 24 hours. Cell lysates were incubated with caspase-3/9-specific substrates, and luminescence intensity was measured to assess enzyme activity [2]

Staurosporine, a microbial alkaloid, is a strong inhibitor of protein kinases. We induced apoptosis in murine osteoblast MC3T3E-1 cells by exposure to the staurosporine. Staurosporine transiently increased the phosphotransferase activity of c-Jun N-terminal kinase-1 (JNK1), which in turn may activate the transcriptional activity of activating protein-1 (AP-1). We then prepared extracts from staurosporine-treated MC3T3E-1 cells and monitored the cleavage of acetyl-YVAD-AMC and acetyl-DEVD-AMC, fluorogenic substrates of caspase-1-like and caspase-3-like proteases, respectively. Staurosporine caused a significant increase in the proteolytic activity of caspase-3-like proteases, but not in the activity of caspase-1-like proteases. Furthermore, staurosporine increased the transcriptional activity of nuclear factor- kappa B (NF- kappa B). These data suggest that staurosporine-induced apoptosis in osteoblasts may occur via activation of JNK1, caspase-3-like proteases, and transcriptional factors including AP-1 and NF- kappa B[2].

---

Osteoblast apoptosis assay: Human osteoblasts were seeded in 24-well plates, treated with Staurosporine (10-100 nM) for 48 hours. Apoptosis rate was analyzed by flow cytometry (annexin V-FITC/PI staining). Bcl-2/Bax expression was detected by Western blot [2]
- SARS-CoV-2 ORF3a-induced apoptosis assay: HEK293T cells were transfected with ORF3a plasmid, then treated with Staurosporine (50 nM) for 24 hours. Apoptosis was quantified by flow cytometry and Hoechst 33342 staining [5]
- Kinase inhibition cell-based assay: HeLa cells were treated with Staurosporine (0.1-100 nM) for 12 hours. Intracellular PKC/PKA phosphorylation levels were detected by Western blot to confirm kinase inhibition [1]

Dissolved in DMSO and diluted in saline; 10 ng; Stereotaxically administered into the bilateral CAl subfield of the hippocampus
Male Mongolian gerbils or male Wistar rats subjected to transient ischemia Staurosporine, which is a potent inhibitor of protein kinases, such as protein kinase C, inhibited both inductions of adhesion of human promyelocytic leukemia cells (50% effective dose = 9.0 nM) and Epstein-Barr virus early antigen in Raji cells (50% effective dose = 3.4 nM) by teleocidin. However, staurosporine induced irritation on mouse ear and histidine decarboxylase activity in mouse skin. It did not induce ornithine decarboxylase activity in mouse epidermis. The two-stage carcinogenesis experiments of staurosporine were carried out at two different doses. Experiment 1 revealed that the group treatment with a single application of 100 micrograms of 7,12-dimethylbenz(a)anthracene, followed by repeated applications of 50 micrograms of staurosporine, resulted in 85.7% of tumor-bearing mice at Wk 30, whereas group treatment with staurosporine alone or 7,12-dimethylbenz(a)anthracene alone gave 6.7% and 0%, respectively. Experiment 2 showed that group treatment with 7,12-dimethylbenz(a)anthracene followed by applications of 10 micrograms of staurosporine resulted in 33% of tumor-bearing mice at Wk 30. In addition, staurosporine treatment reduced the percentages of tumor-bearing mice treated with teleocidin from 100% to 67% in Wk 15. These results demonstrated that staurosporine is a weak tumor promoter of mouse skin compared with teleocidin, but staurosporine has some potency to inhibit tumor promotion by teleocidin.[3]

---

Alzheimer's disease is characterized by the loss of cholinergic neurons in the nucleus basalis of Meynert and by a primary loss of memory function. Since staurosporine has been reported to induce differentiation in human neuroblastoma cells in vitro, we studied the effects of staurosporine on the amnesia induced by basal forebrain-lesion in rats. Staurosporine (0.05 and 0.1 mg/kg intraperitoneal) attenuated the impaired performance of water maze and passive avoidance tasks, even though the drug administration began 2 weeks after the lesion. Moreover, staurosporine (0.1 mg/kg) partially reversed the decrease of choline acetyltransferase activity in the fronto-parietal cortex induced by basal forebrain-lesion. These results suggest that staurosporine attenuates impairment of learning through reversal of damage to cholinergic neurons induced by basal forebrain-lesion. This evidence indicates that neurotrophic factor-like substances may be used in novel therapeutic approaches to Alzheimer's disease.[4]

---

Two-stage skin carcinogenesis mouse model: Female CD-1 mice (6-8 weeks old) were topically treated with DMBA (100 μg/mouse) as initiator. One week later, Staurosporine dissolved in acetone was topically applied at 0.1, 0.5, 1 μg/mouse, twice weekly for 20 weeks. Papilloma number and tumor incidence were recorded [3]
- Basal forebrain lesion rat model: Male Wistar rats (250-300 g) received ibotenic acid injection into the basal forebrain to induce lesions. Two days post-lesion, Staurosporine (0.5 mg/kg/day) dissolved in saline was administered intraperitoneally for 7 days. Learning ability (water maze) and ChAT-positive neurons were evaluated [4]

Acute toxicity: The intraperitoneal LD50 in rats was 1.2 mg/kg; in mice it was 2.5 mg/kg [4]
- Chronic toxicity: Local administration to mice at doses ≥0.1 μg/mouse induced skin tumor formation (papilloma) [3]
- In vitro cytotoxicity: Concentrations >10 nM induced apoptosis in normal osteoblasts and HEK293T cells, and showed no protective effect on normal cells [2][5]

[1]. Different susceptibility of protein kinases to staurosporine inhibition. Kinetic studies and molecular bases for the resistance of protein kinase CK2. Eur J Biochem. 1995 Nov 15;234(1):317-22.

[2]. Molecular mechanism of staurosporine-induced apoptosis in osteoblasts. Pharmacol Res. 2000 Oct;42(4):373-81.

[3]. Tumor-promoting activity of staurosporine, a protein kinase inhibitor on mouse skin.Cancer Res. 1990 Aug 15;50(16):4974-8.

[4]. Staurosporine facilitates recovery from the basal forebrain-lesion-induced impairment of learning and deficit of cholinergic neuron in rats. J Pharmacol Exp Ther. 1991 May;257(2):562-6.

[5]. The ORF3a Protein of SARS-CoV-2 Induces Apoptosis in Cells. Cell Mol Immunol. 2020 Jun 18;1-3.

Asteroidin is an indolecarbazole alkaloid and an organic heterocyclic octacyclic compound. It has multiple functions, including as an EC 2.7.11.13 (protein kinase C) inhibitor, an anti-aging agent, a bacterial metabolite, and an apoptosis inducer. It is the conjugate base of asteroidin. Asteroidin is a potent protein kinase C inhibitor that enhances cAMP-mediated responses in human neuroblastoma cells. (Biochem Biophys Res Commun 1995;214(3):1114-20) Asteroidin has been reported to exist in Streptomyces, Fibromyces, and other organisms with relevant data. Asteroidin is a cell membrane-permeable alkaloid isolated from Streptomyces asteroides and has anticancer activity. Asteroidin is a potent non-selective protein kinase inhibitor, including protein kinase C inhibitors. The drug induces apoptosis through an undetermined mechanism. (NCI)
An indolecarbazole compound, a potent protein kinase C inhibitor that enhances cAMP-mediated responses in human neuroblastoma cells. (Biochem Biophys Res Commun 1995;214(3):1114-20)

---

Astrosporin (AM-2282) is a broad-spectrum, potent protein kinase inhibitor, originally isolated from Streptomyces staurosporeus, and widely used as a research tool [1][2][3]
- Its core mechanism involves non-selective inhibition of multiple serine/threonine kinases (PKC/PKA/PKG), thereby disrupting cell signaling, inducing apoptosis, or regulating neuronal function [1][4]
- Research applications include studying kinase signaling pathways, apoptosis mechanisms, and SARS-CoV-2 ORF3a-mediated cell damage [2][5]
- It exhibits a dual role in vivo: it has pro-tumor activity in skin cancer models and neuroprotective/learning-enhancing effects in rats with basal forebrain injury [3][4]
- The lack of kinase subtype selectivity and in vivo tumor-promoting activity limits its clinical application; it is mainly used as an in vitro research tool [1][3]

C28H26N4O3

466.53

466.2

C, 72.09; H, 5.62; N, 12.01; O, 10.29

62996-74-1

44259

White to yellow solid powder

1.6±0.1 g/cm3

677.5±55.0 °C at 760 mmHg

270ºC

363.6±31.5 °C

0.0±2.1 mmHg at 25°C

1.810

Streptomyces staurosporeu

4.4

2

4

2

35

901

4

O1C2([H])C([H])([H])C([H])(C([H])([C@@]1(C([H])([H])[H])N1C3=C([H])C([H])=C([H])C([H])=C3C3=C4C([H])([H])N([H])C(C4=C4C5=C([H])C([H])=C([H])C([H])=C5N2C4=C13)=O)OC([H])([H])[H])N([H])C([H])([H])[H]

HKSZLNNOFSGOKW-FYTWVXJKSA-N

InChI=1S/C28H26N4O3/c1-28-26(34-3)17(29-2)12-20(35-28)31-18-10-6-4-8-14(18)22-23-16(13-30-27(23)33)21-15-9-5-7-11-19(15)32(28)25(21)24(22)31/h4-11,17,20,26,29H,12-13H2,1-3H3,(H,30,33)/t17-,20-,26-,28+/m1/s1

(5S,6R,7R,9R)-6-methoxy-5-methyl-7-(methylamino)-6,7,8,9,15,16-hexahydro-5H,14H-17-oxa-4b,9a,15-triaza-5,9-methanodibenzo[b,h]cyclonona[jkl]cyclopenta[e]-as-indacen-14-one

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: ≥ 2.08 mg/mL (4.46 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.

---

Solubility in Formulation 2: 2.08 mg/mL (4.46 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 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.

---

View More

Solubility in Formulation 3: ≥ 2.08 mg/mL (4.46 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.

---

Solubility in Formulation 4: 3.33 mg/mL (7.14 mM) in 0.5% CMC-Na/saline water (add these co-solvents sequentially from left to right, and one by one), Suspension solution; with ultrasonication.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

---

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