In order to elucidate the role of the mitogen-activated protein kinases, including JNK, p38 MAPK and ERK, as well as the survival-associated PI3K/Akt signaling pathway, in the response to chemotherapy, we have conducted a comparative study regarding the effects of doxorubicin on these pathways. Doxorubicin was determined to elicit the apoptosis of NIH3T3 cells in a dose-dependent manner. Prior to cell death, both Akt and p38 MAPK were transiently activated, and subsequently inactivated almost wholly, whereas ERK and JNK evidenced sustained activations in response to the drug treatment. The inhibition of PI3K/Akt and p38 MAPK both accelerated and enhanced doxorubicin-induced apoptosis and ERK inhibition apparently exerted negative effect on apoptosis. The modulation of PI3K/Akt activation by treatment of LY294002 or expression of Akt mutants such as Akt-DN or Myr-Akt exerted a significant effect on the activation of ERK1/2. We also observed that PI3K/Akt and sustained ERK activation were associated intimately with the etoposide-induced apoptosis. Taken together, our results clearly suggest that the differential regulation of the PI3K/Akt, ERK1/2, and p38 MAPK signaling pathways are crucial in the context of DNA-damaging drug-induced apoptosis, and this has compelled us to propose that the sustained activation of ERK1/2 pathway may be generally involved in the apoptosis induced by anticancer DNA-damaging drugs, including doxorubicin and etoposide.

The exact molecular mechanisms underlying the cellular effects associated with various flavonoids have yet to be fully explained. In the present study, we have administered several flavonoids to human HaCaT keratinocytes and determined that 3,4-dihydroxy flavone (3,4-DHF) exerts a slight stimulatory effect on cell growth, although other flavonoids, including kaempferol, quercetin, and isorhamnetin, exhibited growth inhibitory properties. 3,4-DHF was found to exert an anti-apoptotic effect on etoposide-induced cell death of HaCaT keratinocytes. We were also able to determine that sustained ERK activation was intimately associated with the etoposide-induced apoptosis of HaCaT cells, and treatment with 3,4-DHF induced a significant suppression of etoposide-induced ERK activation, concomitant with the repression of poly(ADP-ribose) polymerase or the cleavage of pro-caspase 3. ERK overexpression significantly overrode the anti-apoptotic function of 3,4-DHF, but this was not true of ERK-DN. Moreover, treatment with 3,4-DHF resulted in the protection of cells from H 2 O 2 -induced cell death and exerted an apparent suppressive effect on the stress-induced generation of reactive oxygen species (ROS). Finally, we showed that 3,4-DHF almost completely abolished kaempferol-induced apoptosis, coupled with a concomitant suppression of both intracellular ROS generation and the activation of ERK. Taken together, our data clearly indicate that a host of phytochemicals, including etoposide and a variety of flavonoids, differentially regulate the apoptosis of human HaCaT keratinocytes via the differential modulation of intracellular ROS production, coupled with the concomitant activation of the ERK signaling pathway. According to these results, we are able to conclude the distinct structureactivity relationship between several flavonoids.

Recently, considerable scientific and therapeutic interest has focused on the structure and functions of the flavonoids. In a previous study, we suggested that hydroxyl (OH) substitutions on specific carbons in the skeleton of the flavonoids might significantly affect their apoptosis-modulating properties. Here, to investigate the effect of various OH substitutions on their diphenylpropane (C6C3C6) skeleton carbons, we selected 10 different flavonoids and assessed their role on UV-induced apoptosis of human keratinocytes, the principal cell type of epidermis. The results showed that 5,7,3,4-tetrahydroxylflavanone (eriodictyol) and 3,4-dihydroxy flavone (3,4-DHF) had a positive effect on cell proliferation of human HaCaT keratinocytes. Treatment with eriodictyol in particular resulted in significant suppression of cell death induced by ultraviolet (UV) light, a major skin-damaging agent. We found that eriodictyol treatment apparently reduced the percentage of apoptotic cells and the cleavage of poly(ADP-ribose) polymerase, concomitant with the repression of caspase-3 activation and reactive oxygen species (ROS) generation. The anti-apoptotic and anti-oxidant effects of eriodictyol were also confirmed in UV-induced cell death of normal human epidermal keratinocyte (NHEK) cells. Taken together, these findings suggest that eriodictyol can be used to protect keratinocytes from UV-induced damage, implying the presence of a complex structure-activity relationship (SAR) in the differential apoptosis-modulating activities of various flavonoids.

Lung cancer is one of the most common cancers in many countries and the leading cause of cancer deaths in the world accounting for 28% of all cancer death.1,2) The high mortality of this disease is due to the difficulty of early diagnosis and its high potential to invade locally and metastasize to distant organs. Therefore, there is need for novel diagnosis, treatment, and prevention, against lung cancer.As the functional compounds of the plants can serve as a starting point for the development of optimal derivatives, scientists are always searching for new medical herbs. Flavonoids are plant pigments which have been detected in all parts of plants and major functional components of many herbal preparations used in traditional medical protocols. [3][4][5][6] Fruits and vegetables, as well as popular beverages such as wine, tea, and coffee, are the main dietary sources of flavonoids. It has been reported that flavonoids show pharmacological effects such as antiviral, 7) antitumor, 8) antioxidant, 9,10) and anti-inflammatory activities. 11) They are low molecular weight polyphenolic compounds, which possess a basic 2-phenyl-benzo-gamma-pyrone structure harboring one or more hydroxyl groups. As the result of this basic chemical structure, one of the most obvious features of flavonoids is their ability to quench free radicals via the formation of resonance-stabilized phenoxyl radicals.12) Recently, They have attracted much attention because of their broad pharmacological activities, in particular antioxidant activities, and anti-tumorigenic activities. 13,14) Flavonoids exert protective effects, which appear to be related to specific structural characteristics.15) Recently, several studies have reported that the differential effects of flavonoids are attributable to substituted functional groups. 16,17) We also suggested that the OH group of the -5 or carbon-7 of the C6C3C6 skeleton is the key determinant of the anti-oxidant and anti-apoptotic activities associated with flavonoids.18,19) Therefore, investigations into the structurally related activities of flavonoids are important in terms of our understanding of their differential activities. The association between flavonoid intake and reduced disease risk was originally believed to be the consequence of the anti-oxidant effects of these compounds and recent evidence appears to indicate that flavonoids and their metabolites exert other intracellular effects, including the direct modulation of cell signaling pathways, including the MAPK( Mitogen activated protein kinase) cascade. [20][21][22][23] In this study, we describe the apoptotic cell death of human lung epithelial carcinoma A549 cells as the result of treatment with synthetic naringenin derivates, including 7-Obenzyl naringenin (KUF-1), 7-O-(m-metoxybenzyl) naringenin (KUF-2), 7-O-(2-naphtylmethyl) naringenin (KUF-5), 7-O-benzoxycarbonylmethyl naringenin (KUF-6), 7-O-(MeO-L-Leu-D-Pro-carbonylmethyl) naringenin (KUF-7), and 7-O-(MeO-Gly-D-Pro-carbonylmethyl) naringenin (KUF-8) (Fig. 1). The synthetic naring...

Flavonoids are micronutrients that are widely detected in foods of plant origin and have been ascribed pharmacological properties. Several biological functions of flavonoids have been thus far identified, whereas there currently exists a lack of evidence to support the relationship between the structure-activity relationship and apoptosis-inducing activity. In an attempt to determine the importance of the OH group or substitution of the 5- or 7-carbon in the diphenylpropane skeleton of flavonoids, we selected 14 different flavonoids with different structures, particularly with regard to the 5- or 7-carbon, and found that naringenin treatment caused a slight decrease in the cell viability of the human colorectal carcinoma RKO cells. Next, in order to characterize the effects of specific substitutions of the 7-carbon of naringenin on apoptosis-regulatory activities, and in an attempt to develop anti-proliferative flavonoid derivatives that would be more effective against colon cancer, we originally synthesized several modified naringenin derivatives (MNDs) including 7-O-benzyl naringenin (KUF-1) and 7-O-(m-metoxybenzyl) naringenin (KUF-2). Treatment with KUF-1 or KUF-2 resulted in significant apoptosis-inducing effects concomitant with losses in mitochondrial membrane potential, caspase activation, intracellular ROS production, and sustained ERK activation. Our data show that KUF-1 or KUF-2 regulate the apoptosis of RKO cells via intracellular ROS production coupled with the concomitant activation of the ERK signaling pathway, thereby implying that hydroxylation or substitution at C7 is critical for the apoptosis-inducing activity of flavonoids.