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Inhibition of mutant IDH1 decreases D-2-HG levels without affecting tumorigenic properties of chondrosarcoma cell lines.

Suijker J, Oosting J, Koornneef A, Struys EA, Salomons GS, Schaap FG, Waaijer CJ, Wijers-Koster PM, Briaire-de Bruijn IH, Haazen L, Riester SM, Dudakovic A, Danen E, Cleton-Jansen AM, van Wijnen AJ, Bovée JV - Oncotarget (2015)

Bottom Line: Specific inhibition of mutant IDH1 using AGI-5198 decreased levels of D-2-HG in a dose dependent manner.After 72 hours of treatment one out of three mutant IDH1 cell lines showed a moderate decrease in viability , while D-2-HG levels decreased >90%.Likewise, prolonged treatment (up to 20 passages) did not affect proliferation and migration.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Leiden University Medical Center, Leiden, The Netherlands.

ABSTRACT
Mutations in isocitrate dehydrogenase 1 (IDH1) and IDH2 are found in a subset of benign and malignant cartilage tumors, gliomas and leukaemias. The mutant enzyme causes the production of D-2-hydroxyglutarate (D-2-HG), affecting CpG island and histone methylation. While mutations in IDH1/2 are early events in benign cartilage tumors, we evaluated whether these mutations play a role in malignant chondrosarcomas. Compared to IDH1/2 wildtype cell lines, chondrosarcoma cell lines harboring an endogenous IDH1 (n=3) or IDH2 mutation (n=2) showed up to a 100-fold increase in intracellular and extracellular D-2-HG levels. Specific inhibition of mutant IDH1 using AGI-5198 decreased levels of D-2-HG in a dose dependent manner. After 72 hours of treatment one out of three mutant IDH1 cell lines showed a moderate decrease in viability , while D-2-HG levels decreased >90%. Likewise, prolonged treatment (up to 20 passages) did not affect proliferation and migration. Furthermore, global gene expression, CpG island methylation as well as histone H3K4, -9, and -27 trimethylation levels remained unchanged. Thus, while IDH1/2 mutations cause enchondroma, malignant progression towards central chondrosarcoma renders chondrosarcoma growth independent of these mutations. Thus, monotherapy based on inhibition of mutant IDH1 appears insufficient for treatment of inoperable or metastasized chondrosarcoma patients.

No MeSH data available.


Related in: MedlinePlus

Unsupervised hierarchical clustering of the top 2000 most differentially methylated CpG sites in CpG islands(A) Unsupervised hierarchical clustering for the top 2000 most differentially methylated CpG sites resulted in a wildtype cluster (blue samples), a mutation specific cluster (red samples) and a cluster with a mixture of wildtype and mutant IDH1/2 cell lines. (B) CpG islands are known to be located in promoter regions. As expected, methylation in the CpG islands was higher in the mutant IDH1/2 cell lines compared to the wildtype counterparts.
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Figure 2: Unsupervised hierarchical clustering of the top 2000 most differentially methylated CpG sites in CpG islands(A) Unsupervised hierarchical clustering for the top 2000 most differentially methylated CpG sites resulted in a wildtype cluster (blue samples), a mutation specific cluster (red samples) and a cluster with a mixture of wildtype and mutant IDH1/2 cell lines. (B) CpG islands are known to be located in promoter regions. As expected, methylation in the CpG islands was higher in the mutant IDH1/2 cell lines compared to the wildtype counterparts.

Mentions: Since mutations in IDH1 were shown to cause global hypermethylation in tumors [9, 26, 32], we assessed the methylome in the chondrosarcoma cell lines. Unsupervised hierarchical clustering for the top 2000 most differentially methylated CpG sites resulted in a cluster containing IDH1/2 wildtype and a cluster containing mutant IDH1/2 cell lines (Supplementary Figure 1A). Unexpectedly, when all 480 000 probes are included, comparison of the mean β-values between mutant IDH1/2 and IDH1/2 wildtype cell lines suggested global hypermethylation of the IDH1/2 wildtype cell lines (Supplementary Figure 1B). When analyzing the different probes separately, only CpG islands probes showed hypermethylation in the chondrosarcoma cell lines with an IDH1/2 mutation (Figure 2B), whereas the shores (regions flanking the CpG islands) and shelves (regions flanking the shores) show hypermethylation in the IDH1/2 wildtype cell lines (Supplementary Figure 1B). Since CpG islands are known to be located in promoter regions and because IDH1/2 mutations are associated with DNA hypermethylation at CpG islands but not at other genomic regions [26], we focused on the probes that were CpG island specific. Unsupervised hierarchical clustering of CpG islands specific probes resulted in three clusters. The first containing mutant IDH1/2 cell lines only, the second containing IDH1/2 wildtype cell lines and the third cluster contained a mixture of wildtype and mutant IDH1/2 cell lines (Figure 2A).


Inhibition of mutant IDH1 decreases D-2-HG levels without affecting tumorigenic properties of chondrosarcoma cell lines.

Suijker J, Oosting J, Koornneef A, Struys EA, Salomons GS, Schaap FG, Waaijer CJ, Wijers-Koster PM, Briaire-de Bruijn IH, Haazen L, Riester SM, Dudakovic A, Danen E, Cleton-Jansen AM, van Wijnen AJ, Bovée JV - Oncotarget (2015)

Unsupervised hierarchical clustering of the top 2000 most differentially methylated CpG sites in CpG islands(A) Unsupervised hierarchical clustering for the top 2000 most differentially methylated CpG sites resulted in a wildtype cluster (blue samples), a mutation specific cluster (red samples) and a cluster with a mixture of wildtype and mutant IDH1/2 cell lines. (B) CpG islands are known to be located in promoter regions. As expected, methylation in the CpG islands was higher in the mutant IDH1/2 cell lines compared to the wildtype counterparts.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4494954&req=5

Figure 2: Unsupervised hierarchical clustering of the top 2000 most differentially methylated CpG sites in CpG islands(A) Unsupervised hierarchical clustering for the top 2000 most differentially methylated CpG sites resulted in a wildtype cluster (blue samples), a mutation specific cluster (red samples) and a cluster with a mixture of wildtype and mutant IDH1/2 cell lines. (B) CpG islands are known to be located in promoter regions. As expected, methylation in the CpG islands was higher in the mutant IDH1/2 cell lines compared to the wildtype counterparts.
Mentions: Since mutations in IDH1 were shown to cause global hypermethylation in tumors [9, 26, 32], we assessed the methylome in the chondrosarcoma cell lines. Unsupervised hierarchical clustering for the top 2000 most differentially methylated CpG sites resulted in a cluster containing IDH1/2 wildtype and a cluster containing mutant IDH1/2 cell lines (Supplementary Figure 1A). Unexpectedly, when all 480 000 probes are included, comparison of the mean β-values between mutant IDH1/2 and IDH1/2 wildtype cell lines suggested global hypermethylation of the IDH1/2 wildtype cell lines (Supplementary Figure 1B). When analyzing the different probes separately, only CpG islands probes showed hypermethylation in the chondrosarcoma cell lines with an IDH1/2 mutation (Figure 2B), whereas the shores (regions flanking the CpG islands) and shelves (regions flanking the shores) show hypermethylation in the IDH1/2 wildtype cell lines (Supplementary Figure 1B). Since CpG islands are known to be located in promoter regions and because IDH1/2 mutations are associated with DNA hypermethylation at CpG islands but not at other genomic regions [26], we focused on the probes that were CpG island specific. Unsupervised hierarchical clustering of CpG islands specific probes resulted in three clusters. The first containing mutant IDH1/2 cell lines only, the second containing IDH1/2 wildtype cell lines and the third cluster contained a mixture of wildtype and mutant IDH1/2 cell lines (Figure 2A).

Bottom Line: Specific inhibition of mutant IDH1 using AGI-5198 decreased levels of D-2-HG in a dose dependent manner.After 72 hours of treatment one out of three mutant IDH1 cell lines showed a moderate decrease in viability , while D-2-HG levels decreased >90%.Likewise, prolonged treatment (up to 20 passages) did not affect proliferation and migration.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Leiden University Medical Center, Leiden, The Netherlands.

ABSTRACT
Mutations in isocitrate dehydrogenase 1 (IDH1) and IDH2 are found in a subset of benign and malignant cartilage tumors, gliomas and leukaemias. The mutant enzyme causes the production of D-2-hydroxyglutarate (D-2-HG), affecting CpG island and histone methylation. While mutations in IDH1/2 are early events in benign cartilage tumors, we evaluated whether these mutations play a role in malignant chondrosarcomas. Compared to IDH1/2 wildtype cell lines, chondrosarcoma cell lines harboring an endogenous IDH1 (n=3) or IDH2 mutation (n=2) showed up to a 100-fold increase in intracellular and extracellular D-2-HG levels. Specific inhibition of mutant IDH1 using AGI-5198 decreased levels of D-2-HG in a dose dependent manner. After 72 hours of treatment one out of three mutant IDH1 cell lines showed a moderate decrease in viability , while D-2-HG levels decreased >90%. Likewise, prolonged treatment (up to 20 passages) did not affect proliferation and migration. Furthermore, global gene expression, CpG island methylation as well as histone H3K4, -9, and -27 trimethylation levels remained unchanged. Thus, while IDH1/2 mutations cause enchondroma, malignant progression towards central chondrosarcoma renders chondrosarcoma growth independent of these mutations. Thus, monotherapy based on inhibition of mutant IDH1 appears insufficient for treatment of inoperable or metastasized chondrosarcoma patients.

No MeSH data available.


Related in: MedlinePlus