So Y. Kim

MEKK1/MEKK4 are responsible for TRAIL-induced JNK/p38 phosphorylation

The tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has been shown to activate mitogen-activated protein kinases (MAPKs) depending on caspase and mammalian sterile 20-like kinase 1 activations. However, the upstream molecule of MAPKs has not yet been identified. The mitogen-activated protein kinase kinase 1 (MEKK1) and the apoptosis signal-regulating kinase 1 (ASK1) are considered to be possible candidates for the action of MAPKKKs induced by TRAIL and the possibility of reactive oxygen species involvement has also been investigated. We found that MEKK1/MEKK4 as opposed to ASK1, are responsible for TRAIL-induced c-Jun NH2-terminal kinase (JNK) or p38 activation, and that their catalytic activity is repressed by the caspase-8 inhibitor, suggesting that the caspase-8 activation induced by TRAIL is indispensible for MEKK activation. The 14-3-3 θ was also shown to interact with and to dissociate from MEKK1 by TRAIL treatment, thus implicating the 14-3-3 protein as a negative regulator of MEKK1 activation. Taken together, we show herein that the upstream molecule of the TRAIL-induced MAPK activation is MEKK, as opposed to ASK1, via the mediation of its signal through JNK/p38 in a caspase-8-dependent manner.

The effectiveness of the oncolytic activity induced by Ad5/F35 adenoviral vector is dependent on the cumulative cellular conditions of survival and autophagy.

To overcome the poor tumor transduction efficiency of adenovirus serotype 5 (Ad5) observed in several types of cancer, the fiber region of Ad5, apart from its tail, was replaced by adenovirus serotype 35 (Ad35). The chimeric Ad5/F35 adenoviral vector did not exhibit any significant enhancement of transduction efficiency. CD46, a receptor for Ad35, was expressed in relatively small amounts in most of the cancer cells examined. Therefore, we investigated the pivotal factor(s) that render cancer cells susceptible to transduction. We discovered that the tumor transduction efficiency of Ad5/F35 was enhanced in the presence of rapamycin, an autophagy inducer, in some cancer cells. Analysis of survival potential and cell proliferation rates revealed that Ad5/F35 exerted a more pronounced oncolytic effect in cancer cells with higher survival potential in the presence of rapamycin.

HSP27 modulates survival signaling networks in cells treated with curcumin and TRAIL.

The combination of curcumin and TRAIL and their role in enhancing apoptotic cell death has been reported by many studies. However, the exact molecular mechanism of apoptosis mediated by curcumin and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is not yet completely understood. In this study, we observed a close connection between dephosphorylated Akt and an increase in phosphorylated heat shock protein 27 (HSP27) during combined treatment with curcumin and TRAIL. Akt dephosphorylation was cumulatively regulated by protein phosphatase 1 (PP1), phosphoinositide-dependent kinase-1 (PDK1), and src. PP1 and PDK1 directly interacted with HSP27, whereas src indirectly interacted with HSP27 via the tumor necrosis factor receptor-associated factor 6 complex. In conclusion, HSP27 modulated cell survival by its interactions with various binding partners, depending on the level of phosphorylated HSP27.

Abstract 694: Different pancreatic cancer cells have their own core signaling pathway

Gemcitabine as the standard chemotherapy agent to pancreatic cancer has proved effective; however, the response rate remains at 5.4% and the 5-year survival rate is extremely poor. Due to the shortcomings of gemcitabine and the presence of gemcitabine-resistant pancreatic cancer, the development of targeted agents into an anti-neoplastic agent remains a viable option. We have investigated the transforming growth factor-beta (TGF-β), which is known to play a central role in pancreas cell function. We have already observed that the suppression of TGF-β2 induced an unexpected down-regulation of both TGF-β1 and TGF-β3 with concomitant decreases of survival-related signals, such as N-caderin, β-catenin, pp65 in melanoma cell line. Here, two different pancreatic cancer cell lines (HPAC, BxPC3), which have opposite genetic backgrounds (K-ras, p53, Smad4, EGFR), have shown differing sensitivities to the down-regulation of TGF-β2. Through transcriptome analysis, HPAC showed a high expression of TGF-β2, MUC4, HSP27, BMP4 which are deeply related to tumor progression and metastasis and are composed to core signaling complex initiating from TGF-β2. This was confirmed by TGF-β2 down-regulation with gemcitabine to show a greater decrease in survival potential compared to BxPC3. On the contrary, BxPC3, which has more of the mutant type p53, exhibited greater down-regulation of MUC4 by downregulation of p53 compared to that of TGF-β2. Such observations suggest that BxPC3 is composed of a different core signaling pathway, p53-MUC4 rather than TGF-β2-related signalings.In conclusion, it is necessary that the core signaling pathways which are implicated in individual pancreatic carcinogenesis are more specifically considered so that the various gemcitabine-resistant pancreatic cancer cells on the basis of customized treatment can be controlled. Citation Format: Eunkyung Kim, Sujin Kang, So Y. Kim, Dongxu Kang, Seungha Lee, Hye J. Choi, Joo-Hang Kim, Jae Jin Song. Different pancreatic cancer cells have their own core signaling pathway. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 694. doi:10.1158/1538-7445.AM2014-694

Abstract C237: Overcoming gemcitabine resistance with TRAIL plus enhancer of anti-survival potential in pancreatic cancer.

Although the combination of gemcitabine with a variety of cytotoxic and targeted agents has generally shown no significant survival advantage as compared with gemcitabine alone, still, many targeted agents against various pathways remain to be tested in pancreatic cancer. Previously, we have observed that the combined treatment with TRAIL plus shRNA of HSP27 induced complete cell death in MiaPaCa-2 pancreatic cell. Here three different targets were invited with TRAIL for chemotherapy combination with gemcitabine using three pancreatic cancer cell lines. We observed that TRAIL treatment to all three pancreatic cells have relieved gemcitabine-induced resistance. More importantly, three different pancreatic cancer cell lines showed their own unique differential sensitivities to targeted agents examined. For example, in case of MiaPaCa-2, gemcitabine/TRAIL/shRNA of survivin was more effective to cell death than HPAC, whereas TRAIL plus downregulation of survival-related molecules (survivin, XIAP, Bcl-xL) effectively enhanced cell death of Panc-1 even without gemcitabine. Then, we extended the survival-related molecules to upstreams such as Src, PI3K-Akt, or NF-κB and inhibited their downstream signals using PP2, wortmannin, SN50, respectively. From their results, TRAIL plus SN50 decreased NF-κB activation and concomitant significant cell death in MiaPaCa-2 which has an acquired resistance of NF-κB after gemcitabine. Based on this result, it was found that combination of gemcitabine/TRAIL/SN50 dramatically increased cell death of MiaPaCa-2, whereas in case of Panc-1 or HPAC, gemcitabine/TRAIL/PP2 dramatically increased cell death. Taken all together, a better knowledge of the multiple molecular pathways implicated in pancreatic carcinogenesis should be much more specifically considered to control the various gemcitabine-resistant pancreatic cancer cells. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):C237. Citation Format: Seungha Lee, So Y. Kim, Dongxu Kang, Hye J. Choi, Joo-Hang Kim, Jae Jin Song. Overcoming gemcitabine resistance with TRAIL plus enhancer of anti-survival potential in pancreatic cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr C237.

Abstract 4104: TRAIL-induced p38 activation is regulated by MEKK4 with positive feedback manner

Proceedings: AACR 102nd Annual Meeting 2011‐‐ Apr 2‐6, 2011; Orlando, FL Previously, we demonstrated that TRAIL-induced p38 activation was responsible for the increases of Akt catalytic and invasive activity. Interestingly, TRAIL induced dramatically Akt catalytic activation in TRAIL-sensitive cells, whereas TRAIL did not induce Akt catalytic activation in TRAIL-resistant cells, suggesting Akt catalytic activation during TRAIL-induced apoptosis is likely to be a compensatory mechanism of cells. Here, we further provide that MEKK4 is the upstream molecule of TRAIL-induced p38 activation, and its catalytic activity is repressed by caspase-8 inhibitor. This result suggests that the caspase-8 activation induced by TRAIL is indispensible for MEKK4 activation. The CIN85, an adaptor molecule with three SH3 domains also interacted with MEKK4 and dissociated from MEKK4 in the presence of p38 inhibitor, providing the possibility that p38 induced the recruitment of CIN85 to MEKK4 for its activation by positive feedback manner. For the examination of p38 with CIN85, in vitro kinase assay, site-directed mutagenesis, and siRNA technique were used whether CIN85 was phosphorylated by p38, and the exact site of phosphorylation, and for the study of biological significance of CIN85 during TRAIL-induced p38 activation. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 4104. doi:10.1158/1538-7445.AM2011-4104

Abstract C42: Survival signals differentially turn on during TRAIL or curcumin/TRAIL treatment

Previously, we have shown that p38-HSP27 acted as modulators from survival to death depending on the apoptotic strength in prostate cancer cells. For example, if the apoptotic strength is in the moderate range, p38-HSP27 acted to increase Akt catalytic activity, whereas if in the stronger range, p38-HSP27 acted to decrease Akt catalytic activity as well as Akt dephosphorylation. Moreover, another survival signal, Src, a non-receptor tyrosine kinase was revealed to be highly phosphorylated (activated) during curcumin/TRAIL treatment, which leads to the conclusion of occurrence of more complex regulatory crosstalk between Akt and Src for the strict control of cell homeostasis. As a consequence, Akt dephosphorylation was regulated by protein phosphatase 1 (PP1), whose activity was subsequently controlled by dissociation from HSP27 (increase) and Src phosphorylation (deccrease). Src phosphorylation during curcumin/TRAIL was also positively regulated by TRAF6, whereas it was negatively regulated by HSP27. Taken together, the level of Akt dephosphorylation during curcumin/TRAIL could be decided by cumulative factors such as the levels of HSP27/TRAF6 and the activity of Src and PP1. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr C42.

Abstract 1277: Differential roles of TRAIL death receptors during TRAIL-induced signaling pathways

In this paper, we demonstrate that TRAIL death receptors (DR4, DR5) have their own distinctive function during TRAIL treatment. For the examination of each receptor9s role, FACS analysis, biotin labeling to plasma membrane and siRNA technique were used. Previously, we have shown that TRAIL has induced anti-apoptotic signals as well as apoptotic signals. Here, we investigated whether TRAIL death receptors (DR4 and DR5) have their own unique function. In TRAIL-sensitive cell line (DU-145), TRAIL induced much more rapid degradation of DR4 than that of DR5. On the other hand, in TRAIL-resistance cell line (SK-BR-3) with higher level of ErbB2/pAkt, DR5 was rarely expressed, while DR4 was distinctively expressed. We also have shown that relatively much more amount of c-Cbl in SK-BR-3 has played a role of supplying additional pAkt which can be also persistently provided by ErbB2. These phenomena are confirmed by knock down of c-Cbl expression using adenovirus expressing si-c-Cbl. Taken together, we suggest that DR4 functions to transmit apoptosis signals with a favor for anti-apoptosis signals, whereas DR5 functions to transmit apoptosis signals without a favor for anti-apoptosis signals. Correspondance: Jae J. Song (jjs109@yuhs.ac) and Joo-Hang Kim (kjhang@yuhs.ac) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1277.