EGFR (IC50 = 8 μM); ErbB2 (IC50 = 49 nM); ErbB3 (IC50 = 52 nM); ErbB4 (IC50 = 52 nM)

At an IC50 of 13 nM, PD158780 inhibits the autophosphorylation of the EGF receptor in human epidermoid carcinoma A431. PD158780 exhibits a high degree of specificity towards the EGF receptor in Swiss 3T3 fibroblasts. It blocks the processes that are dependent on EGF, such as receptor autophosphorylation and thymidine incorporation, at low nanomolar concentrations. Conversely, it is necessary at micromolar levels for processes that are dependent on platelet-derived growth factor and basic fibroblast growth factor. With an IC50 of 49 and 52 nM, respectively, PD158780 suppresses heregulin-stimulated phosphorylation in SK-BR-3 and MDAMB-453 breast tumors, suggesting that the substance is effective against additional EGF receptor family members [1].

PD158780 exhibits anti-clonogenicity activity in a variety of breast tumor types with unique expression patterns of the ErbB family. PD158780, whether given intraperitoneally or orally, exhibits a good therapeutic impact on A431 epidermoid carcinoma. When human EGFR is transfected into mouse fibroblasts, PD158780 has quantifiably important effects. At isotoxic dose levels, PD158780 has notable therapeutic effects on estrogen-dependent MCF-7 breast cancer [1].

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In vivo intrahippocampal injection of the ErbB inhibitor, PD158780, impairs mGluRI-LTD at CA3-CA1 synapses and affects the exploratory behavior in the object recognition test. [2]

The enzymatic reaction is conducted in 96-well filter plates with a final volume of 0.1 mL. The reaction mixture consists of 20 mM HEPES (pH 7.4), 50 μM sodium vanadate, 40 mM magnesium chloride, 10 μM ATP (including 0.5 μCi of [³²P]ATP), 20 μg of polyglutamic acid/tyrosine, 1 ng of EGF receptor tyrosine kinase, and varying concentrations of the inhibitor PD158780 and/or ATP. All components except ATP are first added to the wells, followed by a 10-minute pre-incubation with shaking at 25°C. The reaction is initiated by adding [³²P]ATP, and the plate is incubated with shaking at 25°C for an additional 10 minutes. The reaction is stopped by adding 0.1 mL of 20% trichloroacetic acid (TCA), and the plate is chilled at 4°C for at least 15 minutes to ensure substrate precipitation. Finally, the wells are washed five times with 0.125 mL of 10% TCA, and [³²P] incorporation is measured[1].

All cell lines are cultured as monolayers in a 1:1 mixture of DMEM/F12 supplemented with 10% fetal bovine serum. For growth inhibition assays, serial dilutions of the test compound (PD158780) in 10 μL are dispensed into 24-well plates, followed by the addition of cells suspended in 2 mL of medium. The plates are then incubated for 72 hours at 37°C in a humidified environment. After incubation, cell growth is quantified by direct cell counting[1].

On day 0, tumor fragments were subcutaneously implanted into the right axillary region of mice. PD158780 was then administered via intraperitoneal injection or oral gavage. Tumor progression was subsequently monitored[1].

[1]. Biochemical and antiproliferative properties of 4-[ar(alk)ylamino]pyridopyrimidines, a new chemical class of potent and specific epidermal growth factor receptor tyrosine kinase inhibitor. Biochem Pharmacol. 1997 Oct 15;54(8):877-87.

[2]. Neuregulin 1/ErbB signalling modulates hippocampal mGluRI-dependent LTD and object recognition memory. Pharmacol Res. 2018 Apr:130:12-24.

PD158780 is a pyridopyrimidine that is pyrido[3,4-d]pyrimidine-4,6-diamine in which the amino groups at positions 4 and 6 are substituted by a m-bromophenyl group and a methyl group, respectively. It is a potent, cell-permeable, reversible ATP-competitive inhibitor of EGFR tyrosine kinase activity [IC50 values of 0.008, 49 and 52 nM for EGFR, ErbB2 (HER2) and Erb4 (HER4)]. It does not inhibit FGF or PDGF-mediated tyrosine phosphorylation. Induces G1 cell cycle arrest in MCF10A cells and is antiproliferative in A431 human epidermal carcinoma cells. It has a role as an antineoplastic agent and an EC 2.7.10.1 (receptor protein-tyrosine kinase) inhibitor. It is a pyridopyrimidine, a secondary amino compound, a member of bromobenzenes, a diamine and an aromatic amine.

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The tyrosine kinase inhibitors PD 69896, 153717, and 158780, which belong to the chemical class 4-[ar(alk)ylamino]pyridopyrimidines, have been characterized with respect to enzymology, target specificity, and antiproliferative effects in tumor cells. These compounds were competitive inhibitors with respect to ATP against purified epidermal growth factor (EGF) receptor tyrosine kinase and inhibited EGF receptor autophosphorylation in A431 human epidermoid carcinoma with IC50 values of 2085, 110, and 13 nM, respectively. Onset of inhibition was immediate once cells were exposed to these compounds, whereas recovery of receptor autophosphorylation activity after the cells were washed free of the compound was dependent on inhibitory potency. Thus, full activity returned immediately after removal of PD 69896 but required 8 hr after exposure to PD158780. PD 158780 was highly specific for the EGF receptor in Swiss 3T3 fibroblasts, inhibiting EGF-dependent receptor autophosphorylation and thymidine incorporation at low nanomolar concentrations while requiring micromolar levels for platelet-derived growth factor- and basic fibroblast growth factor-dependent processes. PD 158780 inhibited heregulin-stimulated phosphorylation in the SK-BR-3 and MDA-MB-453 breast carcinomas with IC50 values of 49 and 52 nM, respectively, suggesting that the compound was active against other members of the EGF receptor family. The antiproliferative effects of this series of compounds against A431 cells correlated precisely with the inhibitory potency against EGF receptor autophosphorylation. PD158780 reduced clone formation in soft agar of fibroblasts transformed by EGF, EGF receptor, or the neu oncogene but not ras or raf, further demonstrating its high degree of specificity. Finally, this compound was active against clone formation in several breast tumors having different expression patterns of the erbB family, indicating an anticancer utility in tumors expressing these receptors.[1]

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The neurotrophic factors neuregulins (NRGs) and their receptors, ErbB tyrosine kinases, regulate neurotransmission, synaptic plasticity and cognitive functions and their alterations have been associated to different neuropsychiatric disorders. Group 1 metabotropic glutamate receptors (mGluRI)-dependent mechanisms are also altered in animal models of neuropsychiatric diseases, especially mGluRI-induced glutamatergic long-term depression (mGluRI-LTD), a form of synaptic plasticity critically involved in learning and memory. Despite this evidence, a potential link between NRGs/ErbB signalling and mGluRI-LTD has never been considered. Here, we aimed to test the hypothesis that NRGs/ErbB signalling regulates mGluRI functions in the hippocampus, thus controlling CA1 pyramidal neurons excitability and synaptic plasticity as well as mGluRI-dependent behaviors. We investigated the functional interaction between NRG1/ErbB signalling and mGluRI in hippocampal CA1 pyramidal neurons, by analyzing the effect of a pharmacological modulation of NRG1/ErbB signalling on the excitation of pyramidal neurons and on the LTD at CA3-CA1 synapses induced by an mGluRI agonist. Furthermore, we verified the involvement of ErbB signalling in mGluRI-dependent learning processes, by evaluating the consequence of an intrahippocampal in vivo injection of a pan-ErbB inhibitor in the object recognition test in mice, a learning task dependent on hippocampal mGluRI. We found that NRG1 potentiates mGluRI-dependent functions on pyramidal neurons excitability and synaptic plasticity at CA3-CA1 synapses. Further, endogenous ErbB signalling per se regulates, through mGluRI, neuronal excitability and LTD in CA1 pyramidal neurons, since ErbB inhibition reduces mGluRI-induced neuronal excitation and mGluRI-LTD. In vivo intrahippocampal injection of the ErbB inhibitor, PD158780, impairs mGluRI-LTD at CA3-CA1 synapses and affects the exploratory behavior in the object recognition test. Thus, our results identify a key role for NRG1/ErbB signalling in the regulation of hippocampal mGluRI-dependent synaptic and cognitive functions, whose alteration might contribute to the pathogenesis of different brain diseases.[2]

C14H12BRN5

330.1826

329.028

C, 50.93; H, 3.66; Br, 24.20; N, 21.21

171179-06-9

4707

Light yellow to yellow solid powder

1.611g/cm3

499.6ºC at 760 mmHg

176 °C

255.9ºC

4.1E-10mmHg at 25°C

1.768

3.718

2

5

3

20

316

0

CNC1=NC=C2C(=C1)C(=NC=N2)NC3=CC(=CC=C3)Br

KFHMLBXBRCITHF-UHFFFAOYSA-N

InChI=1S/C14H12BrN5/c1-16-13-6-11-12(7-17-13)18-8-19-14(11)20-10-4-2-3-9(15)5-10/h2-8H,1H3,(H,16,17)(H,18,19,20)

4-N-(3-bromophenyl)-6-N-methylpyrido[3,4-d]pyrimidine-4,6-diamine

PD 158780; 171179-06-9; PD158,780; pd 158,780; N4-(3-bromophenyl)-N6-methylpyrido[3,4-d]pyrimidine-4,6-diamine; PD-158,780; AC1L1IRV; CHEBI:92843; DTXSID20274443; PD158780; PD-158780.

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 : ~16.67 mg/mL (~50.49 mM)

Solubility in Formulation 1: 1.67 mg/mL (5.06 mM) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 16.7 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: 1.67 mg/mL (5.06 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 16.7 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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 (Please use freshly prepared in vivo formulations for optimal results.)