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Molecular signature of response and potential pathways related to resistance to the HSP90 inhibitor, 17AAG, in breast cancer.

Zajac M, Gomez G, Benitez J, Martínez-Delgado B - BMC Med Genomics (2010)

Bottom Line: In addition, different patterns of HSP90 client transcriptional changes after 17AAG were identified associated to the sensitive cell lines, which could be useful to evaluate drug effectiveness.Finally, we have found differentially expressed pathways associated to resistance to 17AAG.Our study shows that global mRNA expression analysis is a useful strategy to examine molecular effects of drugs, which allowed us the discovery of new biomarkers of 17AAG activity and provided more insights into the complex mechanism of 17AAG resistance.

View Article: PubMed Central - HTML - PubMed

Affiliation: Human Genetics Group, Spanish National Cancer Centre, Madrid, Spain.

ABSTRACT

Background: HSP90 may be a favorable target for investigational therapy in breast cancer. In fact, the HSP90 inhibitor, 17AAG, currently has entered in phase II clinical trials as an anticancer agent in breast and other tumors. Since HSP90 inhibition leads to global depletion of oncogenic proteins involved in multiple pathways we applied global analysis using gene array technology to study new genes and pathways involved in the drug response in breast cancer.

Methods: Gene expression profiling using Whole Human Genome Agilent array technology was applied to a total of six sensitive and two resistant breast cancer cell lines pre-treatment and treated with the 17AAG for 24 and 48 hours.

Results: We have identified a common molecular signature of response to 17AAG composed of 35 genes which include novel pharmacodynamic markers of this drug. In addition, different patterns of HSP90 client transcriptional changes after 17AAG were identified associated to the sensitive cell lines, which could be useful to evaluate drug effectiveness. Finally, we have found differentially expressed pathways associated to resistance to 17AAG. We observed significant activation of NF-κB and MAPK pathways in resistant cells upon treatment, indicating that these pathways could be potentially targeted to overcome resistance.

Conclusions: Our study shows that global mRNA expression analysis is a useful strategy to examine molecular effects of drugs, which allowed us the discovery of new biomarkers of 17AAG activity and provided more insights into the complex mechanism of 17AAG resistance.

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Related in: MedlinePlus

Gene signature of response to 17AAG in breast cancer cell lines. A) Heat-map represents the expression of significant differentially expressed 35 genes included in the molecular signature of response to 17AAG. Untreated samples were compared to treated samples from all sensitive cell lines to obtain common variations. B) Comparison between the changes in expression in the genes included in the signature in sensitive and resistant cells. Fold changes after 17AAG for sensitive cell lines, taken the averaged expression of all of them, compared to fold changes in resistant MDA-MB-231 cells or T47 D.Error bars represent standard deviation. C) Western blot analysis of different HSP70 isoforms before and after 17AAG in resistant MDA-MB-231 and sensitive MCF-7 cell line showed differential HSP70 induction after 17AAG. Lack of induction of the HSPA2 and HSPA1L in resistant cells, and increased expression of HSP72 and HSC70 in both resistant and sensitive cell lines upon treatment is shown. GAPDH was used as loading control.
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Figure 2: Gene signature of response to 17AAG in breast cancer cell lines. A) Heat-map represents the expression of significant differentially expressed 35 genes included in the molecular signature of response to 17AAG. Untreated samples were compared to treated samples from all sensitive cell lines to obtain common variations. B) Comparison between the changes in expression in the genes included in the signature in sensitive and resistant cells. Fold changes after 17AAG for sensitive cell lines, taken the averaged expression of all of them, compared to fold changes in resistant MDA-MB-231 cells or T47 D.Error bars represent standard deviation. C) Western blot analysis of different HSP70 isoforms before and after 17AAG in resistant MDA-MB-231 and sensitive MCF-7 cell line showed differential HSP70 induction after 17AAG. Lack of induction of the HSPA2 and HSPA1L in resistant cells, and increased expression of HSP72 and HSC70 in both resistant and sensitive cell lines upon treatment is shown. GAPDH was used as loading control.

Mentions: Regardless of the specific changes found in each cell line, we searched for genes commonly associated to sensitivity to 17AAG. Analysis of differentially expressed genes between all treated compared with all untreated 17AAG sensitive breast cancer cell lines was performed. A list of 35 genes with a significant FDR value <0.05 was obtained, which constituted a molecular signature of response to 17AAG in breast cancer cells (Figure 2A). Among the 35 genes, 15 were down-regulated and 20 up-regulated genes following treatment (Table 1). The up-regulated genes following treatment included heat shock family proteins such as HSP90AA1, HSPA8, DNAJB12, HSPA1L, DNAJA1, HSPA4L, consistently with other array studies in different tumor cell types [24,32,33]. Among other genes showing increased expression after treatment were ubiquitin-conjugating enzyme E2C (UBE2C), death effector domain containing 2 (DEDD2), zinc finger protein 587 (ZNF587), zinc finger protein 473 (ZNF473), Rac GTPase activating protein 1 (RACGAP1), MHC class I polypeptide-related sequence B (MICB), regulator of G-protein signalling 2 (RGS2), cysteine and histidine-rich domain (CHORD)-containing 1 (CHORDC1), PPAR binding protein. There were also down-regulated genes in response to 17AAG which included e.g. JUNB, CYCLIN D1, NFKBIA, immediate early response 3 (IER3), transmembrane protein 129 (TMEM129) or replication factor C (activator 1) 4 (RFC4).


Molecular signature of response and potential pathways related to resistance to the HSP90 inhibitor, 17AAG, in breast cancer.

Zajac M, Gomez G, Benitez J, Martínez-Delgado B - BMC Med Genomics (2010)

Gene signature of response to 17AAG in breast cancer cell lines. A) Heat-map represents the expression of significant differentially expressed 35 genes included in the molecular signature of response to 17AAG. Untreated samples were compared to treated samples from all sensitive cell lines to obtain common variations. B) Comparison between the changes in expression in the genes included in the signature in sensitive and resistant cells. Fold changes after 17AAG for sensitive cell lines, taken the averaged expression of all of them, compared to fold changes in resistant MDA-MB-231 cells or T47 D.Error bars represent standard deviation. C) Western blot analysis of different HSP70 isoforms before and after 17AAG in resistant MDA-MB-231 and sensitive MCF-7 cell line showed differential HSP70 induction after 17AAG. Lack of induction of the HSPA2 and HSPA1L in resistant cells, and increased expression of HSP72 and HSC70 in both resistant and sensitive cell lines upon treatment is shown. GAPDH was used as loading control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2959047&req=5

Figure 2: Gene signature of response to 17AAG in breast cancer cell lines. A) Heat-map represents the expression of significant differentially expressed 35 genes included in the molecular signature of response to 17AAG. Untreated samples were compared to treated samples from all sensitive cell lines to obtain common variations. B) Comparison between the changes in expression in the genes included in the signature in sensitive and resistant cells. Fold changes after 17AAG for sensitive cell lines, taken the averaged expression of all of them, compared to fold changes in resistant MDA-MB-231 cells or T47 D.Error bars represent standard deviation. C) Western blot analysis of different HSP70 isoforms before and after 17AAG in resistant MDA-MB-231 and sensitive MCF-7 cell line showed differential HSP70 induction after 17AAG. Lack of induction of the HSPA2 and HSPA1L in resistant cells, and increased expression of HSP72 and HSC70 in both resistant and sensitive cell lines upon treatment is shown. GAPDH was used as loading control.
Mentions: Regardless of the specific changes found in each cell line, we searched for genes commonly associated to sensitivity to 17AAG. Analysis of differentially expressed genes between all treated compared with all untreated 17AAG sensitive breast cancer cell lines was performed. A list of 35 genes with a significant FDR value <0.05 was obtained, which constituted a molecular signature of response to 17AAG in breast cancer cells (Figure 2A). Among the 35 genes, 15 were down-regulated and 20 up-regulated genes following treatment (Table 1). The up-regulated genes following treatment included heat shock family proteins such as HSP90AA1, HSPA8, DNAJB12, HSPA1L, DNAJA1, HSPA4L, consistently with other array studies in different tumor cell types [24,32,33]. Among other genes showing increased expression after treatment were ubiquitin-conjugating enzyme E2C (UBE2C), death effector domain containing 2 (DEDD2), zinc finger protein 587 (ZNF587), zinc finger protein 473 (ZNF473), Rac GTPase activating protein 1 (RACGAP1), MHC class I polypeptide-related sequence B (MICB), regulator of G-protein signalling 2 (RGS2), cysteine and histidine-rich domain (CHORD)-containing 1 (CHORDC1), PPAR binding protein. There were also down-regulated genes in response to 17AAG which included e.g. JUNB, CYCLIN D1, NFKBIA, immediate early response 3 (IER3), transmembrane protein 129 (TMEM129) or replication factor C (activator 1) 4 (RFC4).

Bottom Line: In addition, different patterns of HSP90 client transcriptional changes after 17AAG were identified associated to the sensitive cell lines, which could be useful to evaluate drug effectiveness.Finally, we have found differentially expressed pathways associated to resistance to 17AAG.Our study shows that global mRNA expression analysis is a useful strategy to examine molecular effects of drugs, which allowed us the discovery of new biomarkers of 17AAG activity and provided more insights into the complex mechanism of 17AAG resistance.

View Article: PubMed Central - HTML - PubMed

Affiliation: Human Genetics Group, Spanish National Cancer Centre, Madrid, Spain.

ABSTRACT

Background: HSP90 may be a favorable target for investigational therapy in breast cancer. In fact, the HSP90 inhibitor, 17AAG, currently has entered in phase II clinical trials as an anticancer agent in breast and other tumors. Since HSP90 inhibition leads to global depletion of oncogenic proteins involved in multiple pathways we applied global analysis using gene array technology to study new genes and pathways involved in the drug response in breast cancer.

Methods: Gene expression profiling using Whole Human Genome Agilent array technology was applied to a total of six sensitive and two resistant breast cancer cell lines pre-treatment and treated with the 17AAG for 24 and 48 hours.

Results: We have identified a common molecular signature of response to 17AAG composed of 35 genes which include novel pharmacodynamic markers of this drug. In addition, different patterns of HSP90 client transcriptional changes after 17AAG were identified associated to the sensitive cell lines, which could be useful to evaluate drug effectiveness. Finally, we have found differentially expressed pathways associated to resistance to 17AAG. We observed significant activation of NF-κB and MAPK pathways in resistant cells upon treatment, indicating that these pathways could be potentially targeted to overcome resistance.

Conclusions: Our study shows that global mRNA expression analysis is a useful strategy to examine molecular effects of drugs, which allowed us the discovery of new biomarkers of 17AAG activity and provided more insights into the complex mechanism of 17AAG resistance.

Show MeSH
Related in: MedlinePlus