Naturally Occurring Homoisoflavonoids Function as Potent Protein Tyrosine Kinase Inhibitors by c-Src-Based High-Throughput Screening

Abstract

Protein tyrosine kinase (PTK) inhibitors represent emerging therapeutics for cancer chemoprevention. In our study, hematoxylin (26) was identified as one of the most remarkable c-Src inhibitors in an orthogonal compound-mixing library (32200 compounds) by using an ELISA-based automated high-throughput screening (HTS) strategy. Interestingly, hematoxylin was found to be an ATP competitive broad-spectrum PTK inhibitor in vitro, with IC50 values ranging from nanomolar to micromolar level. Further studies showed that such inhibition was associated with the PTK phosphorylation and subsequent downstream signaling pathways. The structure-activity relationship assessment of the PTK inhibitory potency of hematoxylin analogues isolated from Heamatoxylon campechianum was in good agreement with the result of concurrent molecular docking simulation: the catechol moiety in ring A and the hematoxylin-like three-dimensional structure were essential for c-Src-targeted activities. Hematoxylin and its natural analogues were substantially validated to function as a new class of PTK inhibitors.

Establishment of platform for screening insulin-like growth factor-1 receptor inhibitors and evaluation of novel inhibitors

“…Some multi-targeted PTK inhibitors approved by the FDA/ EMEA, such as sorafenib [27] and sunitinib [28] , are marked as a new generation of PTK inhibitor drugs. We have previously reported that hematoxylin is a multi-targeted inhibitor of several tyrosine kinases, including c-Src, c-Met, FGFR1, EGFR, VEGFR, PDGFR, and c-Kit [29] . These data, together with the newly recognized function of hematoxylin as a potent inhibitor of IGF1R, suggest that hematoxylin may be a promising therapeutic agent for the treatment of cancer.…”

Quantitative Structure-cytotoxicity Relationship of 3-Benzylidenechromanones

“…(3E)-2,3-Dihydro-3-(phenylmethylene)-4H-1-benzopyran-4-one [1], (3E)-2,3-dihydro-3-[(4-hydroxyphenyl) methylene]-4H-1-benzopyran-4-one [2], (3E)-2,3-dihydro-3-[(3,4-dihydroxyphenyl)methylene]-4H-1-benzopyran-4-one [3], (3E)-2,3-dihydro-3-[(4-methoxyphenyl)methylene]-4H-1-benzopyran-4-one [4], (3E)-2,3-dihydro-3-[(3,4-dimethoxyphenyl)methylene]-4H-1-benzopyran-4-one [5], (3E)-2,3-dihydro-3-[(4-dimethylaminophenyl)methylene]-4H-1-benzopyran-4-one [6], (3E)-2,3-dihydro-3-[(4-fluorophenyl)methylene]-4H-1-benzopyran-4-one [7], (3E)-3-[(4-chlorophenyl)methylene]-2,3-dihydro-4H-1-benzopyran-4-one [8], (3E)-2,3-dihydro-7-hydroxy-3-[(4-hydroxyphenyl)methylene]-4H-1-benzopyran-4-one [9], (3E)-2,3-dihydro-3-[(3,4-dihydroxyphenyl)methylene]-7-hydroxy-4H-1-benzopyran-4-one [10], (3E)-2,3-dihydro-7-hydroxy-3-[(4-methoxyphenyl)methylene]-4H-1-benzopyran-4-one [11], (3E)-2,3-dihydro-3-[(4-dimethylaminophenyl)methylene]-7-hydroxy-4H-1-benzopyran-4-one [12], (3E)-2,3-dihydro-7-methoxy-3-(phenylmethylene)-4H-1-benzopyran-4-one [13], (3E)-2,3-dihydro-3-[(4-hydroxyphenyl)methylene]-7-methoxy-4H-1-benzopyran-4-one [14], (3E)-2,3-dihydro-3-[(3,4-dihydroxyphenyl)methylene]-7-methoxy-4H-1-benzopyran-4-one [15], (3E)-2,3-dihydro-7-methoxy-3-[(4-methoxyphenyl)methylene]-4H-1-benzopyran-4-one [16] and (3E)-2,3-dihydro-3-[(4-dimethylaminophenyl)methylene]-7-methoxy-4H-1-benzopyran-4-one [17] (structures shown in Figure 1) were synthesized by base-catalyzed condensation of appropriate 4-chromanone with substituted benzaldehyde derivatives according to previous methods (19,20). All compounds were dissolved in DMSO at 40 mM and stored at -20˚C before use.…”

Chlorogenic acid inhibits hypoxia-induced pulmonary artery smooth muscle cells proliferation via c-Src and Shc/Grb2/ERK2 signaling pathway

“…The tyrosine kinase activities of the purified c-Src kinase domains were determined by ELISA method as described previously (Lin et al, 2008). 96-well ELISA plates (Corning) were precoated with 20 μg/ml Poly (Glu, Tyr) 4:1 (Sigma-Aldorich).…”

“…Finally, 100 μl of a solution containing 0.03% H 2 O 2 and 2 mg/ml Ophenylenediamine in 0.1 mol/l citrate buffer, pH 5.5, was added and samples were incubated at room temperature until color emerged. The reaction was terminated by the addition of 50 μl of 2 M H 2 SO 4 , and the plate was read using Spectramax M2 microplate Reader (Molecular Devices) at 490 nm (Lin et al, 2008). IC 50 values were calculated by Graphpad Prism 5.0 software (Logit method).…”