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NF-kappaB activation enhances cell death by antimitotic drugs in human prostate cancer cells.

Parrondo R, de las Pozas A, Reiner T, Rai P, Perez-Stable C - Mol. Cancer (2010)

Bottom Line: In castration-resistant DU145 and PC3 cells, Doc and 2ME2 had little effect on the high basal NF-kappaB activity and addition of parthenolide did not enhance cell death.Our data suggest that the combination of antimitotic drugs with NF-kappaB inhibitors will have antagonistic effects in a common type of PC cell typical of LNCaP.However, combination strategies utilizing antimitotic drugs with BA, an activator of NF-kappaB, will universally enhance cell death in PC cells, notably in the aggressive, castration-resistant variety that does not respond to conventional therapies.

View Article: PubMed Central - HTML - PubMed

Affiliation: Geriatric Research, Education, and Clinical Center and Research Service, Bruce W, Carter Veterans Affairs Medical Center, Miami, FL 33125, USA.

ABSTRACT

Background: NF-kappaB is a transcription factor that promotes inhibition of apoptosis and resistance to chemotherapy. It is commonly believed that inhibition of NF-kappaB activity can increase sensitivity of cancer cells to chemotherapy. However, there is evidence that NF-kappaB activation can sensitize cells to apoptosis and that inhibition of NF-kappaB results in resistance to chemotherapy. In prostate cancer, it is not clear in the different cell types (androgen-dependent and castration-resistant) if activation or inhibition of NF-kappaB is required for stimulation of apoptosis by chemotherapy.

Results: Our data indicate that the response of prostate cancer (PC) cells to the antimitotic drugs docetaxel (Doc) and 2-methoxyestradiol (2ME2) is dependent on the levels of NF-kappaB activity. In androgen-dependent LNCaP cells, Doc and 2ME2 treatment increased the low basal NF-kappaB activity, as determined by Western blot analysis of phospho-IkappaBalpha/p65, NF-kappaB promoter reporter assays, and p65 localization. Treatment of LNCaP cells with parthenolide, a pharmacologic inhibitor of NF-kappaB, or introduction of dominant-negative IkappaBalpha, or an shRNA specific for p65, a component of the NF-kappaB heterodimer, blocked apoptosis induced by Doc and 2ME2. In castration-resistant DU145 and PC3 cells, Doc and 2ME2 had little effect on the high basal NF-kappaB activity and addition of parthenolide did not enhance cell death. However, the combination of Doc or 2ME2 with betulinic acid (BA), a triterpenoid that activates NF-kappaB, stimulated apoptosis in LNCaP and non-apoptotic cell death in DU145 and PC3 cells. Increased sensitivity to cell death mediated by the Doc or 2ME2 + BA combination is likely due to increased NF-kappaB activity.

Conclusions: Our data suggest that the combination of antimitotic drugs with NF-kappaB inhibitors will have antagonistic effects in a common type of PC cell typical of LNCaP. However, combination strategies utilizing antimitotic drugs with BA, an activator of NF-kappaB, will universally enhance cell death in PC cells, notably in the aggressive, castration-resistant variety that does not respond to conventional therapies.

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BA increases and parthenolide decreases expression of NF-κB target genes. RT-qPCR analysis showing that BA (B; 10 μM) increases expression of IκBα (top left), DR5 (top right), and A20 (bottom left) mRNAs in combination with 2ME2 (M; 5 μM) or Doc (D; 1 nM) (MB, DB) in LNCaP cells (48 h). Parthenolide (P; 10 μM) decreases the M and D increase in A20 (MP, DP) in LNCaP cells (72 h) (lower right panel). mRNA levels are normalized to control (C) = 1. n = 5, three independent experiments. *, P < 0.05.
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Figure 10: BA increases and parthenolide decreases expression of NF-κB target genes. RT-qPCR analysis showing that BA (B; 10 μM) increases expression of IκBα (top left), DR5 (top right), and A20 (bottom left) mRNAs in combination with 2ME2 (M; 5 μM) or Doc (D; 1 nM) (MB, DB) in LNCaP cells (48 h). Parthenolide (P; 10 μM) decreases the M and D increase in A20 (MP, DP) in LNCaP cells (72 h) (lower right panel). mRNA levels are normalized to control (C) = 1. n = 5, three independent experiments. *, P < 0.05.

Mentions: In addition, we investigated the expression of AIFshort (AIFsh) protein, an alternative transcription start site coding for AIF corresponding only to the C-terminal pro-death portion that translocates to the nucleus to induce cell death [32]. Western blot and RT-qPCR analysis showed that LNCaP cells treated with 2ME2/Doc + BA or BA alone increased AIFsh protein and mRNA (2-5-fold) compared to 2ME2 and Doc alone (Fig. 9B, D). IκBα, A20, and DR5 mRNAs, genes known to be regulated by NF-κB [1] also increased in LNCaP cells treated with the 2ME2/Doc + BA combination (Fig. 10). In DU145 and PC3 cells, there was also an increase in AIFsh protein after treatment with 2ME2/Doc + BA or BA alone (Fig. 9B).


NF-kappaB activation enhances cell death by antimitotic drugs in human prostate cancer cells.

Parrondo R, de las Pozas A, Reiner T, Rai P, Perez-Stable C - Mol. Cancer (2010)

BA increases and parthenolide decreases expression of NF-κB target genes. RT-qPCR analysis showing that BA (B; 10 μM) increases expression of IκBα (top left), DR5 (top right), and A20 (bottom left) mRNAs in combination with 2ME2 (M; 5 μM) or Doc (D; 1 nM) (MB, DB) in LNCaP cells (48 h). Parthenolide (P; 10 μM) decreases the M and D increase in A20 (MP, DP) in LNCaP cells (72 h) (lower right panel). mRNA levels are normalized to control (C) = 1. n = 5, three independent experiments. *, P < 0.05.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2908581&req=5

Figure 10: BA increases and parthenolide decreases expression of NF-κB target genes. RT-qPCR analysis showing that BA (B; 10 μM) increases expression of IκBα (top left), DR5 (top right), and A20 (bottom left) mRNAs in combination with 2ME2 (M; 5 μM) or Doc (D; 1 nM) (MB, DB) in LNCaP cells (48 h). Parthenolide (P; 10 μM) decreases the M and D increase in A20 (MP, DP) in LNCaP cells (72 h) (lower right panel). mRNA levels are normalized to control (C) = 1. n = 5, three independent experiments. *, P < 0.05.
Mentions: In addition, we investigated the expression of AIFshort (AIFsh) protein, an alternative transcription start site coding for AIF corresponding only to the C-terminal pro-death portion that translocates to the nucleus to induce cell death [32]. Western blot and RT-qPCR analysis showed that LNCaP cells treated with 2ME2/Doc + BA or BA alone increased AIFsh protein and mRNA (2-5-fold) compared to 2ME2 and Doc alone (Fig. 9B, D). IκBα, A20, and DR5 mRNAs, genes known to be regulated by NF-κB [1] also increased in LNCaP cells treated with the 2ME2/Doc + BA combination (Fig. 10). In DU145 and PC3 cells, there was also an increase in AIFsh protein after treatment with 2ME2/Doc + BA or BA alone (Fig. 9B).

Bottom Line: In castration-resistant DU145 and PC3 cells, Doc and 2ME2 had little effect on the high basal NF-kappaB activity and addition of parthenolide did not enhance cell death.Our data suggest that the combination of antimitotic drugs with NF-kappaB inhibitors will have antagonistic effects in a common type of PC cell typical of LNCaP.However, combination strategies utilizing antimitotic drugs with BA, an activator of NF-kappaB, will universally enhance cell death in PC cells, notably in the aggressive, castration-resistant variety that does not respond to conventional therapies.

View Article: PubMed Central - HTML - PubMed

Affiliation: Geriatric Research, Education, and Clinical Center and Research Service, Bruce W, Carter Veterans Affairs Medical Center, Miami, FL 33125, USA.

ABSTRACT

Background: NF-kappaB is a transcription factor that promotes inhibition of apoptosis and resistance to chemotherapy. It is commonly believed that inhibition of NF-kappaB activity can increase sensitivity of cancer cells to chemotherapy. However, there is evidence that NF-kappaB activation can sensitize cells to apoptosis and that inhibition of NF-kappaB results in resistance to chemotherapy. In prostate cancer, it is not clear in the different cell types (androgen-dependent and castration-resistant) if activation or inhibition of NF-kappaB is required for stimulation of apoptosis by chemotherapy.

Results: Our data indicate that the response of prostate cancer (PC) cells to the antimitotic drugs docetaxel (Doc) and 2-methoxyestradiol (2ME2) is dependent on the levels of NF-kappaB activity. In androgen-dependent LNCaP cells, Doc and 2ME2 treatment increased the low basal NF-kappaB activity, as determined by Western blot analysis of phospho-IkappaBalpha/p65, NF-kappaB promoter reporter assays, and p65 localization. Treatment of LNCaP cells with parthenolide, a pharmacologic inhibitor of NF-kappaB, or introduction of dominant-negative IkappaBalpha, or an shRNA specific for p65, a component of the NF-kappaB heterodimer, blocked apoptosis induced by Doc and 2ME2. In castration-resistant DU145 and PC3 cells, Doc and 2ME2 had little effect on the high basal NF-kappaB activity and addition of parthenolide did not enhance cell death. However, the combination of Doc or 2ME2 with betulinic acid (BA), a triterpenoid that activates NF-kappaB, stimulated apoptosis in LNCaP and non-apoptotic cell death in DU145 and PC3 cells. Increased sensitivity to cell death mediated by the Doc or 2ME2 + BA combination is likely due to increased NF-kappaB activity.

Conclusions: Our data suggest that the combination of antimitotic drugs with NF-kappaB inhibitors will have antagonistic effects in a common type of PC cell typical of LNCaP. However, combination strategies utilizing antimitotic drugs with BA, an activator of NF-kappaB, will universally enhance cell death in PC cells, notably in the aggressive, castration-resistant variety that does not respond to conventional therapies.

Show MeSH
Related in: MedlinePlus