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JNK is required for maintaining the tumor-initiating cell-like properties of acquired chemoresistant human cancer cells.

Liu Y, Zhang X, Wang J, Yang J, Tan WF - Acta Pharmacol. Sin. (2015)

Bottom Line: Additionally, inhibition of JNK activity significantly inhibited the in vitro and in vivo tumor-initiating abilities of KB/VCR cells.Furthermore, our data suggest that blocking JNK activity abundantly inhibited the Hedgehog (Hh) pathway activity, as reflected by reduction of Hedgehog (Hh) pathway target genes Gli1 and ptch1 at the mRNA level as well as Gli-luciferase activity.Thus, disruption of tumor-initiating cell-like properties by targeting JNK may be a new approach to combating acquired chemoresistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Pharmacy, Fudan University, Shanghai 201203, China.

ABSTRACT

Aim: Many studies reveal an association between the acquired chemoresistant phenotype of cancer cells and tumor-initiating cell-like properties. The aim of this study was to determine the impact of c-Jun N-terminal kinase (JNK) on the tumor-initiating cell-like properties of acquired chemoresistant human cancer cells.

Methods: Two well-established human acquired chemoresistant cancer cell lines K562/A02 and KB/VCR, as well as their respective parental counterparts K562 and KB were tested. The expression of relevant mRNAs and proteins was detected using qRT-PCR and Western blotting, respectively. Sphere formation and self-renewal assays were used to study the tumor-initiating cell-like properties. Soft agar and colony formation assays were used to investigate tumorigenic ability.

Results: We observed that suppressing JNK activity by its specific small molecule inhibitor SP600125 or by limiting JNK1/2 expression with JNK1/2 shRNA lentiviruses inhibited the expression of pluripotent stem cell markers such as Oct4, Sox2, and Nanog in KB/VCR cells and K562/A02 cells as well as sphere formation and self-renewal abilities of K562/A02 cells. Additionally, inhibition of JNK activity significantly inhibited the in vitro and in vivo tumor-initiating abilities of KB/VCR cells. Furthermore, our data suggest that blocking JNK activity abundantly inhibited the Hedgehog (Hh) pathway activity, as reflected by reduction of Hedgehog (Hh) pathway target genes Gli1 and ptch1 at the mRNA level as well as Gli-luciferase activity.

Conclusion: JNK maintains the tumor-initiating cell-like properties of acquired chemoresistant K562/A02 and KB/VCR cells potentially through activating the Hedgehog pathway. Thus, disruption of tumor-initiating cell-like properties by targeting JNK may be a new approach to combating acquired chemoresistance.

No MeSH data available.


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Inhibition of JNK activity suppressed Hh activity in acquired chemoresistant cancer cells. (A, B) After treatment with or without SP600125 (10 μmol/L) for 24 h, K562 and K562/A02 cells were collected for qRT-PCR analyses. SP600125 treatment reduced Gli1 (A) and Ptch1 (B) mRNA expression. (C, D) Knockdown of JNK1/2 expression using shRNA lentiviruses suppressed Gli1 (C) and Ptch1 (D) mRNA expression in KB/VCR cells. (E, F) Inhibition of JNK activity by SP600125 reduced Hh pathway activity in K562/A02 cells (E) KB/VCR cells (F) as reflected by Gli-luciferase reporter activity. Mean±SD. n=3. bP<0.05, cP<O.01.
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fig5: Inhibition of JNK activity suppressed Hh activity in acquired chemoresistant cancer cells. (A, B) After treatment with or without SP600125 (10 μmol/L) for 24 h, K562 and K562/A02 cells were collected for qRT-PCR analyses. SP600125 treatment reduced Gli1 (A) and Ptch1 (B) mRNA expression. (C, D) Knockdown of JNK1/2 expression using shRNA lentiviruses suppressed Gli1 (C) and Ptch1 (D) mRNA expression in KB/VCR cells. (E, F) Inhibition of JNK activity by SP600125 reduced Hh pathway activity in K562/A02 cells (E) KB/VCR cells (F) as reflected by Gli-luciferase reporter activity. Mean±SD. n=3. bP<0.05, cP<O.01.

Mentions: We have previously demonstrated that the Hh signaling pathway is required for maintaining tumor-initiating cell-like properties of acquired chemoresistant cancer cells6. In this context, we asked whether JNK maintained the tumor-initiating cell-like properties of acquired chemoresistant cancer cells by activating the Hh pathway. As anticipated, we observed that suppressing JNK activity in K562/A02 cells by SP600125 clearly inhibited the mRNA expression of Gli1 (Figure 5A) and Ptch1 (Figure 5B), which are two transcriptional target genes of Hh signaling pathway transcription factors and are often used as readouts of Hh pathway activity19. These observations were further confirmed by limiting JNK1/2 expression with JNK1/2 shRNA lentiviruses in KB/VCR cells (Figure 5C and 5D). We further examined Hh pathway activity using dual Gli-luciferase reporter assays in acquired chemoresistant cancer cells after SP600125 treatment. As demonstrated in Figure 5E and 5F, SP600125 treatment reduced Gli-luciferase activity in K562/A02 cells (Figure 5E) and KB/VCR cells (Figure 5F). Hence, our data suggest that JNK inhibition may suppress Hh pathway activity, thereby disrupting the tumor-initiating cell-like properties of acquired chemoresistant cancer cells.


JNK is required for maintaining the tumor-initiating cell-like properties of acquired chemoresistant human cancer cells.

Liu Y, Zhang X, Wang J, Yang J, Tan WF - Acta Pharmacol. Sin. (2015)

Inhibition of JNK activity suppressed Hh activity in acquired chemoresistant cancer cells. (A, B) After treatment with or without SP600125 (10 μmol/L) for 24 h, K562 and K562/A02 cells were collected for qRT-PCR analyses. SP600125 treatment reduced Gli1 (A) and Ptch1 (B) mRNA expression. (C, D) Knockdown of JNK1/2 expression using shRNA lentiviruses suppressed Gli1 (C) and Ptch1 (D) mRNA expression in KB/VCR cells. (E, F) Inhibition of JNK activity by SP600125 reduced Hh pathway activity in K562/A02 cells (E) KB/VCR cells (F) as reflected by Gli-luciferase reporter activity. Mean±SD. n=3. bP<0.05, cP<O.01.
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fig5: Inhibition of JNK activity suppressed Hh activity in acquired chemoresistant cancer cells. (A, B) After treatment with or without SP600125 (10 μmol/L) for 24 h, K562 and K562/A02 cells were collected for qRT-PCR analyses. SP600125 treatment reduced Gli1 (A) and Ptch1 (B) mRNA expression. (C, D) Knockdown of JNK1/2 expression using shRNA lentiviruses suppressed Gli1 (C) and Ptch1 (D) mRNA expression in KB/VCR cells. (E, F) Inhibition of JNK activity by SP600125 reduced Hh pathway activity in K562/A02 cells (E) KB/VCR cells (F) as reflected by Gli-luciferase reporter activity. Mean±SD. n=3. bP<0.05, cP<O.01.
Mentions: We have previously demonstrated that the Hh signaling pathway is required for maintaining tumor-initiating cell-like properties of acquired chemoresistant cancer cells6. In this context, we asked whether JNK maintained the tumor-initiating cell-like properties of acquired chemoresistant cancer cells by activating the Hh pathway. As anticipated, we observed that suppressing JNK activity in K562/A02 cells by SP600125 clearly inhibited the mRNA expression of Gli1 (Figure 5A) and Ptch1 (Figure 5B), which are two transcriptional target genes of Hh signaling pathway transcription factors and are often used as readouts of Hh pathway activity19. These observations were further confirmed by limiting JNK1/2 expression with JNK1/2 shRNA lentiviruses in KB/VCR cells (Figure 5C and 5D). We further examined Hh pathway activity using dual Gli-luciferase reporter assays in acquired chemoresistant cancer cells after SP600125 treatment. As demonstrated in Figure 5E and 5F, SP600125 treatment reduced Gli-luciferase activity in K562/A02 cells (Figure 5E) and KB/VCR cells (Figure 5F). Hence, our data suggest that JNK inhibition may suppress Hh pathway activity, thereby disrupting the tumor-initiating cell-like properties of acquired chemoresistant cancer cells.

Bottom Line: Additionally, inhibition of JNK activity significantly inhibited the in vitro and in vivo tumor-initiating abilities of KB/VCR cells.Furthermore, our data suggest that blocking JNK activity abundantly inhibited the Hedgehog (Hh) pathway activity, as reflected by reduction of Hedgehog (Hh) pathway target genes Gli1 and ptch1 at the mRNA level as well as Gli-luciferase activity.Thus, disruption of tumor-initiating cell-like properties by targeting JNK may be a new approach to combating acquired chemoresistance.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, School of Pharmacy, Fudan University, Shanghai 201203, China.

ABSTRACT

Aim: Many studies reveal an association between the acquired chemoresistant phenotype of cancer cells and tumor-initiating cell-like properties. The aim of this study was to determine the impact of c-Jun N-terminal kinase (JNK) on the tumor-initiating cell-like properties of acquired chemoresistant human cancer cells.

Methods: Two well-established human acquired chemoresistant cancer cell lines K562/A02 and KB/VCR, as well as their respective parental counterparts K562 and KB were tested. The expression of relevant mRNAs and proteins was detected using qRT-PCR and Western blotting, respectively. Sphere formation and self-renewal assays were used to study the tumor-initiating cell-like properties. Soft agar and colony formation assays were used to investigate tumorigenic ability.

Results: We observed that suppressing JNK activity by its specific small molecule inhibitor SP600125 or by limiting JNK1/2 expression with JNK1/2 shRNA lentiviruses inhibited the expression of pluripotent stem cell markers such as Oct4, Sox2, and Nanog in KB/VCR cells and K562/A02 cells as well as sphere formation and self-renewal abilities of K562/A02 cells. Additionally, inhibition of JNK activity significantly inhibited the in vitro and in vivo tumor-initiating abilities of KB/VCR cells. Furthermore, our data suggest that blocking JNK activity abundantly inhibited the Hedgehog (Hh) pathway activity, as reflected by reduction of Hedgehog (Hh) pathway target genes Gli1 and ptch1 at the mRNA level as well as Gli-luciferase activity.

Conclusion: JNK maintains the tumor-initiating cell-like properties of acquired chemoresistant K562/A02 and KB/VCR cells potentially through activating the Hedgehog pathway. Thus, disruption of tumor-initiating cell-like properties by targeting JNK may be a new approach to combating acquired chemoresistance.

No MeSH data available.


Related in: MedlinePlus