I would add tannic acid, quercetin, genistein and luteolin to your experiements.
http://jb.oxfordjournals.org/cgi/content/abstract/139/3/495
Tannic Acid, a Potent Inhibitor of Epidermal Growth Factor Receptor Tyrosine Kinase
Er Bin Yang1,*, Liu Wei2, Kai Zhang3, Yu Zong Chen2 and Wei Ning Chen1
1 Hepatitis Viruses and Liver Cancer Research Laboratory, School of Chemical and Biomedical Engineering, Nanyang Technological University, 19 Nanyang Drive, Singapore 637551; 2 Department of Computational Science, National University of Singapore, 10 Kent Ridge Crescent, Singapore 119260; and 3 Department of Experimental Surgery, Singapore General Hospital, Outram Road, Singapore 169608
Increasing evidence supports the hypothesis that tannic acid, a plant polyphenol, exerts anticarcinogenic activity in chemically induced cancers. In the present study, tannic acid was found to strongly inhibit tyrosine kinase activity of epidermal growth factor receptor (EGFr) in vitro (IC50 = 323 nM). In contrast, the inhibition by tannic acid of p60c-src tyrosine kinase (IC50 = 14 µM) and insulin receptor tyrosine kinase (IC50 = 5 µM) was much weaker. The inhibition of EGFr tyrosine kinase by tannic acid was competitive with respect to ATP and non-competitive with respect to peptide substrate. In cultured cells, growth factor–induced tyrosine phosphorylation of growth factor receptors, including EGFr, platelet-derived growth factor receptor, and basic fibroblast growth factor receptor, was inhibited by tannic acid. No inhibition of insulin-induced tyrosine phosphorylation of insulin receptor and insulin-receptor substrate-1 was observed. EGF-stimulated growth of HepG2 cells was inhibited in the presence of tannic acid. The inhibition of serine/threonine-specific protein kinases, including cAMP-dependent protein kinase, protein kinase C and mitogen-activated protein kinase, by tannic acid was only detected at relatively high concentration, IC50 being 3, 325 and 142 µM respectively. The molecular modeling study suggested that tannic acid could be docked into the ATP binding pockets of either EGFr or insulin receptor. These results demonstrate that tannic acid is an in vitro potent inhibitor of EGFr tyrosine kinase.
http://www.sciencedirect.com/scienc...serid=10&md5=53264ec065d7f005e86df86789c2ee60
Genistein, a Tyrosine Kinase Inhibitor, Reduces EGF-Induced EGF Receptor Internalization and Degradation in Human Hepatoma HepG2 Cells
Er-Bin Yangb, a, Dong-Fang Wanga, Peter Mackb and Li-Yao Cheng a, 1
a Department of Biochemistry, National University of Singapore, 10 Kent Ridge Crescent, Singapore, 119260
b Department of Surgery, Singapore General Hospital, Outram Road, Singapore, 169068
Abstract
In this work, using the ECL Western blotting assay system, it was found that genistein, a specific tyrosine kinase inhibitor, was able to inhibit EGF-induced EGF receptor degradation and tyrosine phosphorylation in human hepatoma HepG2 cells. This inhibition was increased with increasing genistein concentration. With treatment of HepG2 cells with genistein at 37 °C for 30 min, the amount of internalized EGF, which was measured by the detection of the sorting of125I-EGF in the cells, was remarkably decreased. Under the same conditions, in cells untreated with genistein, the degradation of EGF was significantly increased. After preincubation of HepG2 cells with and without genistein for 120 min at 37 °C, the ratio between degraded and released EGF was 16 and 24, respectively. These results suggest that EGF-induced internalization and degradation of EGF-EGF receptor complexes in HepG2 cells depend on EGF receptor tyrosine kinase activity.
http://www.ncbi.nlm.nih.gov/pubmed/12168845
Anticancer Res. 2002 May-Jun;22(3):1615-27.Links
Blockade of the epidermal growth factor receptor tyrosine kinase activity by quercetin and luteolin leads to growth inhibition and apoptosis of pancreatic tumor cells.
Lee LT, Huang YT, Hwang JJ, Lee PP, Ke FC, Nair MP, Kanadaswam C, Lee MT.
Institute of Biological Chemistry Academia Sinica, Taipei, Taiwan.
To glean insights into the mechanism of their action, we assessed the effects of two flavonoids, quercetin (Qu) and luteolin (Lu), on the growth and epidermal growth factor receptor (EGFR) tyrosine kinase activity of MiaPaCa-2 cancer cells. Exposure of these EGFR-expressing cells to 20 microM Qu or Lu resulted in concomitant decreases in cellular protein phosphorylation and growth. On the cellular level, Qu and Lu sensitivity correlated with EGFR levels and rapid cell proliferation, indicating the possibility of targeting those cells most prone to neoplastic progression. Cell treatment with the flavonoids markedly diminished the extent of cellular protein phosphorylation, by effectively modulating protein tyrosine kinase (PTK) activities, including that of EGFR. Immunocomplex kinase assay revealed that both Qu and Lu inhibited the PTK activities responsible for the autophosphorylation of EGFR as well as for the transphosphorylation of enolase. Treatment of the cells with Qu or Lu also reduced the phosphotyrosyl levels of 170-, 125-, 110-, 65-, 60-, 44-, 30- and 25-kDa proteins. We identified the 170-kDa phosphotyrosylprotein as EGFR. Qu and Lu exhibited a specific action in hampering the levels of phosphorylation of this and the aforementioned proteins, while having no discernible effect on their synthesis. A time-dependent attenuation of the phosphorylation of the above proteins was demonstrable. Treatment of the cells with Qu or Lu for 6 hours showed little inhibition, but prolonging the cell treatment for 24 hours caused the suppression of phosphorylation. Further continuation of the cell treatment culminated in the induction of apoptosis, characteristically exhibiting shrinkage of the cell morphology, DNA fragmentation and poly(ADP-ribose)polymerase (PARP) degradation. The onset of apoptosis and associated events occurred in a time-dependent fashion. The data clearly demonstrate that MiaPaCa-2 cells respond to Qu and Lu by a parallel reduction in cellular protein phosphorylation and cellular proliferation. The flavonoid-evoked attenuation of the phosphorylation of EFGR and of other proteins appeared to be transient, since removal of the flavonoid from the cell growth medium after 24 hours of incubation followed by exposure to 10 nm EGF, restored protein phosphorylation and cellular proliferation. Such an addition of EGF was also able to reverse Qu- or Lu-induced cell growth inhibition and diminish nuclear digestion evoked by 20 microM Qu or Lu. Both Qu and Lu were able to reverse the effect of EGF biochemically as well as functionally. Based on the evidence accrued, the above proteins could be implicated in growth signal transduction and the subtle changes in their phosphorylation, as effected by flavonoids, utilized as a reliable guide to predict growth response. The antiproliferative effect of flavonoids might result, at least in part, from the modulation of the EGF-mediated signaling pathway. The results indicate that the blockade of the EGFR-signaling pathway by the PTK inhibitors Qu and Lu significantly inhibits the growth of MiaPaCa-2 cells and induces apoptosis. The modulation of EGFR kinase appears to be a critically important, intrinsic component of Qu- and Lu-induced growth suppression, even though other mechanisms could also have contributed to the net effect.
http://www.nature.com/bjp/journal/v128/n5/full/0702879a.html
British Journal of Pharmacology (1999) 128, 999–1010; doi:10.1038/sj.bjp.0702879
Effects of luteolin and quercetin, inhibitors of tyrosine kinase, on cell growth and metastasis-associated properties in A431 cells overexpressing epidermal growth factor receptor
Y -T Huang1, J -J Hwang2, P -P Lee3, F -C Ke4, J -H Huang1, C -J Huang1, C Kandaswami5, E Middleton Jr5 and M -T Lee1
1Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan
2Institute of Physiology, National Yang Ming University, Taipei, Taiwan
3Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, NY 14263, U.S.A.
4Department of Zoology, National Taiwan University, Taipei, Taiwan
5Department of Medicine, State University of New York at Buffalo, NY 14263, U.S.A.
Correspondence: M -T Lee, Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan
Abstract
Flavonoids display a wide range of pharmacological properties including anti-inflammatory. Anti-mutagenic, anti-carcinogenic and anti-cancer effects. Here, we evaluated the effects of eight flavonoids on the tumour cell proliferation, cellular protein phosphorylation, and matrix metalloproteinase (MMPs) secretion.
Of the flavonoids examined, luteolin (Lu) and quercetin (Qu) were the two most potent agents, and significantly inhibited A431 cell proliferation with IC50 values of 19 and 21 M, respectively.
The epidermal growth factor (EGF) (10 nM) promoted growth of A431 cells (+254.6%) and mediated epidermal growth factor receptor (EGFR) tyrosine kinase activity and autophosphorylation of EGFR were inhibited by Lu and Qu. At concentration of 20 M, both Lu and Qu markedly decreased the levels of phosphorylation of A431 cellular proteins, including EGFR.
A431 cells treated with Lu or Qu exhibited protuberant cytoplasmic blebs and progressive shrinkage morphology. Lu and Qu also time-dependently induced the appearance of a ladder pattern of DNA fragmentation, and this effect was abolished by EGF treatment.
The addition of EGF only marginally diminished the inhibitory effect of luteolin and quercetin on the growth rate of A431 cells, treatment of cellular proteins with EGF and luteolin or quercetin greatly reduced protein phosphorylation, indicating Lu and Qu may act effectively to inhibit a wide range of protein kinases, including EGFR tyrosine kinase.
EGF increased the levels of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9), while Lu and Qu appeared to suppress the secretion of these two MMPs in A431 cells.
Examination of the relationship between the chemical structure and inhibitory effects of eight flavonoids reveal that the double bond between C2 and C3 in ring C and the OH groups on C3' and C4' in ring B are critical for the biological activities.
This study demonstrates that the inhibitory effects of Lu and Qu, and the stimulatory effects of EGF, on tumour cell proliferation, cellular protein phosphorylation, and MMP secretion may be mediated at least partly through EGFR. This study supports the idea that Lu and Qu may have potential as anti-cancer and anti-metastasis agents.
These are just a few of many published studies
bug
http://jb.oxfordjournals.org/cgi/content/abstract/139/3/495
Tannic Acid, a Potent Inhibitor of Epidermal Growth Factor Receptor Tyrosine Kinase
Er Bin Yang1,*, Liu Wei2, Kai Zhang3, Yu Zong Chen2 and Wei Ning Chen1
1 Hepatitis Viruses and Liver Cancer Research Laboratory, School of Chemical and Biomedical Engineering, Nanyang Technological University, 19 Nanyang Drive, Singapore 637551; 2 Department of Computational Science, National University of Singapore, 10 Kent Ridge Crescent, Singapore 119260; and 3 Department of Experimental Surgery, Singapore General Hospital, Outram Road, Singapore 169608
Increasing evidence supports the hypothesis that tannic acid, a plant polyphenol, exerts anticarcinogenic activity in chemically induced cancers. In the present study, tannic acid was found to strongly inhibit tyrosine kinase activity of epidermal growth factor receptor (EGFr) in vitro (IC50 = 323 nM). In contrast, the inhibition by tannic acid of p60c-src tyrosine kinase (IC50 = 14 µM) and insulin receptor tyrosine kinase (IC50 = 5 µM) was much weaker. The inhibition of EGFr tyrosine kinase by tannic acid was competitive with respect to ATP and non-competitive with respect to peptide substrate. In cultured cells, growth factor–induced tyrosine phosphorylation of growth factor receptors, including EGFr, platelet-derived growth factor receptor, and basic fibroblast growth factor receptor, was inhibited by tannic acid. No inhibition of insulin-induced tyrosine phosphorylation of insulin receptor and insulin-receptor substrate-1 was observed. EGF-stimulated growth of HepG2 cells was inhibited in the presence of tannic acid. The inhibition of serine/threonine-specific protein kinases, including cAMP-dependent protein kinase, protein kinase C and mitogen-activated protein kinase, by tannic acid was only detected at relatively high concentration, IC50 being 3, 325 and 142 µM respectively. The molecular modeling study suggested that tannic acid could be docked into the ATP binding pockets of either EGFr or insulin receptor. These results demonstrate that tannic acid is an in vitro potent inhibitor of EGFr tyrosine kinase.
http://www.sciencedirect.com/scienc...serid=10&md5=53264ec065d7f005e86df86789c2ee60
Genistein, a Tyrosine Kinase Inhibitor, Reduces EGF-Induced EGF Receptor Internalization and Degradation in Human Hepatoma HepG2 Cells
Er-Bin Yangb, a, Dong-Fang Wanga, Peter Mackb and Li-Yao Cheng a, 1
a Department of Biochemistry, National University of Singapore, 10 Kent Ridge Crescent, Singapore, 119260
b Department of Surgery, Singapore General Hospital, Outram Road, Singapore, 169068
Abstract
In this work, using the ECL Western blotting assay system, it was found that genistein, a specific tyrosine kinase inhibitor, was able to inhibit EGF-induced EGF receptor degradation and tyrosine phosphorylation in human hepatoma HepG2 cells. This inhibition was increased with increasing genistein concentration. With treatment of HepG2 cells with genistein at 37 °C for 30 min, the amount of internalized EGF, which was measured by the detection of the sorting of125I-EGF in the cells, was remarkably decreased. Under the same conditions, in cells untreated with genistein, the degradation of EGF was significantly increased. After preincubation of HepG2 cells with and without genistein for 120 min at 37 °C, the ratio between degraded and released EGF was 16 and 24, respectively. These results suggest that EGF-induced internalization and degradation of EGF-EGF receptor complexes in HepG2 cells depend on EGF receptor tyrosine kinase activity.
http://www.ncbi.nlm.nih.gov/pubmed/12168845
Anticancer Res. 2002 May-Jun;22(3):1615-27.Links
Blockade of the epidermal growth factor receptor tyrosine kinase activity by quercetin and luteolin leads to growth inhibition and apoptosis of pancreatic tumor cells.
Lee LT, Huang YT, Hwang JJ, Lee PP, Ke FC, Nair MP, Kanadaswam C, Lee MT.
Institute of Biological Chemistry Academia Sinica, Taipei, Taiwan.
To glean insights into the mechanism of their action, we assessed the effects of two flavonoids, quercetin (Qu) and luteolin (Lu), on the growth and epidermal growth factor receptor (EGFR) tyrosine kinase activity of MiaPaCa-2 cancer cells. Exposure of these EGFR-expressing cells to 20 microM Qu or Lu resulted in concomitant decreases in cellular protein phosphorylation and growth. On the cellular level, Qu and Lu sensitivity correlated with EGFR levels and rapid cell proliferation, indicating the possibility of targeting those cells most prone to neoplastic progression. Cell treatment with the flavonoids markedly diminished the extent of cellular protein phosphorylation, by effectively modulating protein tyrosine kinase (PTK) activities, including that of EGFR. Immunocomplex kinase assay revealed that both Qu and Lu inhibited the PTK activities responsible for the autophosphorylation of EGFR as well as for the transphosphorylation of enolase. Treatment of the cells with Qu or Lu also reduced the phosphotyrosyl levels of 170-, 125-, 110-, 65-, 60-, 44-, 30- and 25-kDa proteins. We identified the 170-kDa phosphotyrosylprotein as EGFR. Qu and Lu exhibited a specific action in hampering the levels of phosphorylation of this and the aforementioned proteins, while having no discernible effect on their synthesis. A time-dependent attenuation of the phosphorylation of the above proteins was demonstrable. Treatment of the cells with Qu or Lu for 6 hours showed little inhibition, but prolonging the cell treatment for 24 hours caused the suppression of phosphorylation. Further continuation of the cell treatment culminated in the induction of apoptosis, characteristically exhibiting shrinkage of the cell morphology, DNA fragmentation and poly(ADP-ribose)polymerase (PARP) degradation. The onset of apoptosis and associated events occurred in a time-dependent fashion. The data clearly demonstrate that MiaPaCa-2 cells respond to Qu and Lu by a parallel reduction in cellular protein phosphorylation and cellular proliferation. The flavonoid-evoked attenuation of the phosphorylation of EFGR and of other proteins appeared to be transient, since removal of the flavonoid from the cell growth medium after 24 hours of incubation followed by exposure to 10 nm EGF, restored protein phosphorylation and cellular proliferation. Such an addition of EGF was also able to reverse Qu- or Lu-induced cell growth inhibition and diminish nuclear digestion evoked by 20 microM Qu or Lu. Both Qu and Lu were able to reverse the effect of EGF biochemically as well as functionally. Based on the evidence accrued, the above proteins could be implicated in growth signal transduction and the subtle changes in their phosphorylation, as effected by flavonoids, utilized as a reliable guide to predict growth response. The antiproliferative effect of flavonoids might result, at least in part, from the modulation of the EGF-mediated signaling pathway. The results indicate that the blockade of the EGFR-signaling pathway by the PTK inhibitors Qu and Lu significantly inhibits the growth of MiaPaCa-2 cells and induces apoptosis. The modulation of EGFR kinase appears to be a critically important, intrinsic component of Qu- and Lu-induced growth suppression, even though other mechanisms could also have contributed to the net effect.
http://www.nature.com/bjp/journal/v128/n5/full/0702879a.html
British Journal of Pharmacology (1999) 128, 999–1010; doi:10.1038/sj.bjp.0702879
Effects of luteolin and quercetin, inhibitors of tyrosine kinase, on cell growth and metastasis-associated properties in A431 cells overexpressing epidermal growth factor receptor
Y -T Huang1, J -J Hwang2, P -P Lee3, F -C Ke4, J -H Huang1, C -J Huang1, C Kandaswami5, E Middleton Jr5 and M -T Lee1
1Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan
2Institute of Physiology, National Yang Ming University, Taipei, Taiwan
3Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, NY 14263, U.S.A.
4Department of Zoology, National Taiwan University, Taipei, Taiwan
5Department of Medicine, State University of New York at Buffalo, NY 14263, U.S.A.
Correspondence: M -T Lee, Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan
Abstract
Flavonoids display a wide range of pharmacological properties including anti-inflammatory. Anti-mutagenic, anti-carcinogenic and anti-cancer effects. Here, we evaluated the effects of eight flavonoids on the tumour cell proliferation, cellular protein phosphorylation, and matrix metalloproteinase (MMPs) secretion.
Of the flavonoids examined, luteolin (Lu) and quercetin (Qu) were the two most potent agents, and significantly inhibited A431 cell proliferation with IC50 values of 19 and 21 M, respectively.
The epidermal growth factor (EGF) (10 nM) promoted growth of A431 cells (+254.6%) and mediated epidermal growth factor receptor (EGFR) tyrosine kinase activity and autophosphorylation of EGFR were inhibited by Lu and Qu. At concentration of 20 M, both Lu and Qu markedly decreased the levels of phosphorylation of A431 cellular proteins, including EGFR.
A431 cells treated with Lu or Qu exhibited protuberant cytoplasmic blebs and progressive shrinkage morphology. Lu and Qu also time-dependently induced the appearance of a ladder pattern of DNA fragmentation, and this effect was abolished by EGF treatment.
The addition of EGF only marginally diminished the inhibitory effect of luteolin and quercetin on the growth rate of A431 cells, treatment of cellular proteins with EGF and luteolin or quercetin greatly reduced protein phosphorylation, indicating Lu and Qu may act effectively to inhibit a wide range of protein kinases, including EGFR tyrosine kinase.
EGF increased the levels of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9), while Lu and Qu appeared to suppress the secretion of these two MMPs in A431 cells.
Examination of the relationship between the chemical structure and inhibitory effects of eight flavonoids reveal that the double bond between C2 and C3 in ring C and the OH groups on C3' and C4' in ring B are critical for the biological activities.
This study demonstrates that the inhibitory effects of Lu and Qu, and the stimulatory effects of EGF, on tumour cell proliferation, cellular protein phosphorylation, and MMP secretion may be mediated at least partly through EGFR. This study supports the idea that Lu and Qu may have potential as anti-cancer and anti-metastasis agents.
These are just a few of many published studies
bug
