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KSP inhibitor ARRY-520 as a substitute for Paclitaxel in Type I ovarian cancer cells.

Kim KH, Xie Y, Tytler EM, Woessner R, Mor G, Alvero AB - J Transl Med (2009)

Bottom Line: ARRY-520 and Paclitaxel exhibited the same cytotoxic effect on Type I and II cells.Unlike Paclitaxel, ARRY-520 did not induce NF-kappaB activation, did not enhance cytokine secretion, nor induce ERK phosphorylation in Type I EOC cells.However, unlike Paclitaxel, it does not induce these pro-tumor effects in Type I cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. ghkim@pusan.ac.kr

ABSTRACT

Background: We previously described a sub-population of epithelial ovarian cancer (EOC) cells with a functional TLR-4/MyD88/NF-kappaB pathway (Type I EOC cells), which confers the capacity to respond to Paclitaxel, a known TLR-4 ligand, by enhancing NF-kappaB activity and upregulating cytokine secretion - events that are known to promote tumor progression. It is therefore important to distinguish those patients that should not receive Paclitaxel; it is also important to identify alternative chemotherapy options that would benefit this sub-group of patients. The objective of this study is to determine if the KSP inhibitor, ARRY-520, can be a substitute for Paclitaxel in patients with Type I EOC.

Methods: EOC cells isolated from either ascites or tumor tissue were treated with increasing concentrations of ARRY-520 or Paclitaxel and cell viability determined. Activation of the apoptotic pathway was determined using Western blot analysis. Mitochondrial integrity was quantified using JC1 dye. Cytokine profiling was performed from supernatants using xMAP technology. NF-kappaB activity was measured using a Luciferase reporter system. In vivo activity was determined using a subcutaneous xenograft mouse model.

Results: ARRY-520 and Paclitaxel exhibited the same cytotoxic effect on Type I and II cells. The GI50 at 48 h for Type II EOC cells was 0.0015 microM and 0.2 microM for ARRY-520 and Paclitaxel, respectively. For Type I EOC cells, the GI50 at 48 h was > 3 microM and >20 microM for ARRY-520 and Paclitaxel, respectively. Decrease in the number of viable cells was accompanied by mitochondrial depolarization and caspase activation. Unlike Paclitaxel, ARRY-520 did not induce NF-kappaB activation, did not enhance cytokine secretion, nor induce ERK phosphorylation in Type I EOC cells.

Conclusion: Administration of Paclitaxel to patients with high percentage Type I cancer cells could have detrimental effects due to Paclitaxel-induced enhancement of NF-kappaB and ERK activities, and cytokine production (e.g. IL-6), which promote chemoresistance and tumor progression. ARRY-520 has similar anti-tumor activity in EOC cells as that of Paclitaxel. However, unlike Paclitaxel, it does not induce these pro-tumor effects in Type I cells. Therefore, the KSP inhibitor ARRY-520 may represent an alternative to Paclitaxel in this subgroup of EOC patients.

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ARRY-520 induces apoptosis in Type II EOC cells. Type II EOC cells were treated with 3 μM ARRY-520 for 6, 12, and 24 hours. "0" designation represents non-treated controls. (a) Activity of capase-3, -8, and -9 was measured using Caspase-Glo assay, and (b) effect on XIAP, Caspase-2, and Bid was determined using Western blot analysis. Results shown are for CP70. Similar results were observed with other cells tested.
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Figure 2: ARRY-520 induces apoptosis in Type II EOC cells. Type II EOC cells were treated with 3 μM ARRY-520 for 6, 12, and 24 hours. "0" designation represents non-treated controls. (a) Activity of capase-3, -8, and -9 was measured using Caspase-Glo assay, and (b) effect on XIAP, Caspase-2, and Bid was determined using Western blot analysis. Results shown are for CP70. Similar results were observed with other cells tested.

Mentions: To determine whether the decrease in cell viability is due to the induction of apoptosis, we measured caspase activity in ARRY-520-treated Type II EOC cells. Following ARRY-520 treatment, a significant increase in the activity of caspases- 8, 9, and 3 was observed in a time-dependent manner (Fig. 2a), with a corresponding decrease in the levels of XIAP (Fig. 2b). Moreover, we saw the appearance of the p30 XIAP fragment at 24 h post-treatment, which corresponded to the time point where the most significant increase in caspase-3 activity was observed.


KSP inhibitor ARRY-520 as a substitute for Paclitaxel in Type I ovarian cancer cells.

Kim KH, Xie Y, Tytler EM, Woessner R, Mor G, Alvero AB - J Transl Med (2009)

ARRY-520 induces apoptosis in Type II EOC cells. Type II EOC cells were treated with 3 μM ARRY-520 for 6, 12, and 24 hours. "0" designation represents non-treated controls. (a) Activity of capase-3, -8, and -9 was measured using Caspase-Glo assay, and (b) effect on XIAP, Caspase-2, and Bid was determined using Western blot analysis. Results shown are for CP70. Similar results were observed with other cells tested.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2719595&req=5

Figure 2: ARRY-520 induces apoptosis in Type II EOC cells. Type II EOC cells were treated with 3 μM ARRY-520 for 6, 12, and 24 hours. "0" designation represents non-treated controls. (a) Activity of capase-3, -8, and -9 was measured using Caspase-Glo assay, and (b) effect on XIAP, Caspase-2, and Bid was determined using Western blot analysis. Results shown are for CP70. Similar results were observed with other cells tested.
Mentions: To determine whether the decrease in cell viability is due to the induction of apoptosis, we measured caspase activity in ARRY-520-treated Type II EOC cells. Following ARRY-520 treatment, a significant increase in the activity of caspases- 8, 9, and 3 was observed in a time-dependent manner (Fig. 2a), with a corresponding decrease in the levels of XIAP (Fig. 2b). Moreover, we saw the appearance of the p30 XIAP fragment at 24 h post-treatment, which corresponded to the time point where the most significant increase in caspase-3 activity was observed.

Bottom Line: ARRY-520 and Paclitaxel exhibited the same cytotoxic effect on Type I and II cells.Unlike Paclitaxel, ARRY-520 did not induce NF-kappaB activation, did not enhance cytokine secretion, nor induce ERK phosphorylation in Type I EOC cells.However, unlike Paclitaxel, it does not induce these pro-tumor effects in Type I cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Obstetrics, Gynecology and Reproductive Sciences, Yale University School of Medicine, New Haven, CT, USA. ghkim@pusan.ac.kr

ABSTRACT

Background: We previously described a sub-population of epithelial ovarian cancer (EOC) cells with a functional TLR-4/MyD88/NF-kappaB pathway (Type I EOC cells), which confers the capacity to respond to Paclitaxel, a known TLR-4 ligand, by enhancing NF-kappaB activity and upregulating cytokine secretion - events that are known to promote tumor progression. It is therefore important to distinguish those patients that should not receive Paclitaxel; it is also important to identify alternative chemotherapy options that would benefit this sub-group of patients. The objective of this study is to determine if the KSP inhibitor, ARRY-520, can be a substitute for Paclitaxel in patients with Type I EOC.

Methods: EOC cells isolated from either ascites or tumor tissue were treated with increasing concentrations of ARRY-520 or Paclitaxel and cell viability determined. Activation of the apoptotic pathway was determined using Western blot analysis. Mitochondrial integrity was quantified using JC1 dye. Cytokine profiling was performed from supernatants using xMAP technology. NF-kappaB activity was measured using a Luciferase reporter system. In vivo activity was determined using a subcutaneous xenograft mouse model.

Results: ARRY-520 and Paclitaxel exhibited the same cytotoxic effect on Type I and II cells. The GI50 at 48 h for Type II EOC cells was 0.0015 microM and 0.2 microM for ARRY-520 and Paclitaxel, respectively. For Type I EOC cells, the GI50 at 48 h was > 3 microM and >20 microM for ARRY-520 and Paclitaxel, respectively. Decrease in the number of viable cells was accompanied by mitochondrial depolarization and caspase activation. Unlike Paclitaxel, ARRY-520 did not induce NF-kappaB activation, did not enhance cytokine secretion, nor induce ERK phosphorylation in Type I EOC cells.

Conclusion: Administration of Paclitaxel to patients with high percentage Type I cancer cells could have detrimental effects due to Paclitaxel-induced enhancement of NF-kappaB and ERK activities, and cytokine production (e.g. IL-6), which promote chemoresistance and tumor progression. ARRY-520 has similar anti-tumor activity in EOC cells as that of Paclitaxel. However, unlike Paclitaxel, it does not induce these pro-tumor effects in Type I cells. Therefore, the KSP inhibitor ARRY-520 may represent an alternative to Paclitaxel in this subgroup of EOC patients.

Show MeSH
Related in: MedlinePlus