Insulin-like growth factor-binding protein-3 suppresses tumor growth via activation of caspase-dependent apoptosis and cross-talk with NF-κB signaling

Nuclear factor-kappaB (NF-κB) is constitutively activated in a variety of human cancers including prostate cancer and involved in tumorigenesis, tumor progression and chemo-resistance. Insulin-like growth factor-binding protein-3 (IGFBP-3) is a potent tumor suppressor and is significantly suppressed in a variety of cancers. Diverse biological effects of IGFBP-3 have been reported to be both dependent and independent of the IGF/IGF-I receptor (IGF-IR) axis. The precise underlying mechanisms of IGF/IGF-IR-independent, antiproliferative actions of IGFBP-3 are yet to be elucidated. We found an inverse correlation between NF-κB activity and IGFBP-3 expression during prostate cancer progression using an in vitro prostate cancer progression model. Restoration of IGFBP-3 resulted in significant inhibition of constitutively elevated NF-κB activity in prostate cancer cells. IGFBP-3 further inhibited the expression of NF-κB-regulated angiogenic factors such as VEGF and IL-8, and cell adhesion molecules, ICAM-1 and VCAM-1. This inhibitory action of IGFBP-3 was IGF/IGF-IR-independent since IGFBP-3 mutant devoid of IGF binding affinity had a similar inhibitory effect. We identified that IGFBP-3 degrades the key NF-κB regulatory molecules-IκBα and p65-NF-κB proteins through activation of caspase-8 and -3/-7, thereby inhibiting elevated NF-κB activity in prostate cancer. Finally intratumoral administration of IGFBP-3 resulted in significant tumor suppression as well as sensitization of antitumor effect of doxorubicin. Our findings indicate that IGFBP-3 exerts antitumor effects via IGF-independent mechanisms which involve activation of caspase-dependent apoptosis and cross-talk with NF-κB signaling. The use of IGFBP-3 as a cancer therapeutic with this distinctive suppression mechanism may offer alternate means to treat chemotherapy resistant tumors.     

The roles of CYP450 epoxygenases and metabolites, epoxyeicosatrienoic acids, in cardiovascular and malignant diseases

Cytochrome P450 (CYP) epoxygenases metabolize arachidonic acid to biologically active eicosanoids. The primary epoxidation products are four regioisomers of cis-epoxyeicosatrienoic acid (EET): 5,6-, 8,9-, 11,12-, and 14,15-EET. CYP2J2, CYP2C8, and CYP2C9 are the predominant epoxygenase isoforms involved in EET formation. CYP2J and CYP2C gene families in humans are abundantly expressed in the endothelium, myocardium, and kidney. The cardiovascular effects of CYP epoxygenases and EETs range from vasodilation, anti-hypertension, pro-angiogenesis, anti-atherosclerosis, and anti-inflammation to anti-injury caused by ischemia-reperfusion. Using transgenic animals for in vivo analyses of CYP epoxygenases revealed comprehensive and marked cardiovascular protective effects. In contrast, CYP epoxygenases and their metabolites, EETs, are upregulated in human tumors and promote tumor progression and metastasis. These biological effects result from the anti-apoptosis, pro-mitogenesis, and anti-migration roles of CYP epoxygenases and EETs at the cellular level. Importantly, soluble epoxide hydrolase (sEH) inhibitors are anti-hypertensive and anti-inflammatory and, therefore, protect the heart from damage, whereas the terfenadine-related, specific inhibitors of CYP2J2 exhibit strong anti-tumor activity in vitro and in vivo. Thus, CYP2J2 and arachidonic acid-derived metabolites likely play important roles in regulating cardiovascular functions and malignancy under physiological and/or pathological conditions. Moreover, although challenges remain to improving the drug-like properties of sEH inhibitors and identifying efficient ways to deliver sEH inhibitors, sEH will likely become an important therapeutic target for cardiovascular diseases. In addition, CYP2J2 may be a therapeutic target for treating human cancers and leukemia.     

Induction of apoptosis by phloroglucinol derivative from Ecklonia Cava in MCF-7 human breast cancer cells

Phloroglucinol derivatives, dioxinodehydroeckol (1) and 1-(3′,5′-dihydroxyphenoxy)-7-(2′′,4′′,6-trihydroxyphenoxy)-2,4,9-trihydroxydibenzo-1,4-dioxin (2), were isolated from Ecklonia Cava. Their ability to inhibit the proliferation of human breast cancer cells were evaluated by measuring cell death via induction of apoptosis. Compound 1 exerted a higher anti-proliferative activity in human breast cancer cells compared with compound 2. Furthermore, compound 1 induced a significant proliferative inhibition and apoptosis in a dose-dependent manner on MCF-7 human cancer cells. Treatment with compound 1 also induced the increase in caspase (-3 and -9) activity, DNA repair enzyme poly-(ADP-ribose) polymerase (PARP) cleavage, and pro-apoptotic gene and the decrease in anti-apoptotic gene. In addition, NF-κB family and -dependent activated genes were down-regulated by compound 1. These results indicated that the potential inhibitory effect of compound 1 against growth of MCF-7 human breast cancer cells might be associated with induction of apoptosis through NF-κB family and NF-κB dependent pathway. The present results suggest that compound 1 has a promising potential to be used as a valuable chemopreventive agent.     

Synergistic drug-cytokine induction of hepatocellular death as an in vitro approach for the study of inflammation-associated idiosyncratic drug hepatotoxicity

Idiosyncratic drug hepatotoxicity represents a major problem in drug development due to inadequacy of current preclinical screening assays, but recently established rodent models utilizing bacterial LPS co-administration to induce an inflammatory background have successfully reproduced idiosyncratic hepatotoxicity signatures for certain drugs. However, the low-throughput nature of these models renders them problematic for employment as preclinical screening assays. Here, we present an analogous, but high-throughput, in vitro approach in which drugs are administered to a variety of cell types (primary human and rat hepatocytes and the human HepG2 cell line) across a landscape of inflammatory contexts containing LPS and cytokines TNF, IFNγ, IL-1α, and IL-6. Using this assay, we observed drug-cytokine hepatotoxicity synergies for multiple idiosyncratic hepatotoxicants (ranitidine, trovafloxacin, nefazodone, nimesulide, clarithromycin, and telithromycin) but not for their corresponding non-toxic control compounds (famotidine, levofloxacin, buspirone, and aspirin). A larger compendium of drug-cytokine mix hepatotoxicity data demonstrated that hepatotoxicity synergies were largely potentiated by TNF, IL-1α, and LPS within the context of multi-cytokine mixes. Then, we screened 90 drugs for cytokine synergy in human hepatocytes and found that a significantly larger fraction of the idiosyncratic hepatotoxicants (19%) synergized with a single cytokine mix than did the non-hepatotoxic drugs (3%). Finally, we used an information theoretic approach to ascertain especially informative subsets of cytokine treatments for most highly effective construction of regression models for drug- and cytokine mix-induced hepatotoxicities across these cell systems. Our results suggest that this drug-cytokine co-treatment approach could provide a useful preclinical tool for investigating inflammation-associated idiosyncratic drug hepatotoxicity.