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Whole genome sequence analysis links chromothripsis to EGFR, MDM2, MDM4, and CDK4 amplification in glioblastoma.

Furgason JM, Koncar RF, Michelhaugh SK, Sarkar FH, Mittal S, Sloan AE, Barnholtz-Sloan JS, Bahassi el M - Oncoscience (2015)

Bottom Line: The data derived from this study was further expanded on using fluorescence in situ hybridization (FISH) analysis and susceptibility studies with colony formation assays.We show that primary GBMs are associated with higher chromothripsis scores and establish a link between chromothripsis and gene amplification of receptor tyrosine kinases (RTKs), as well as modulators of the TP53 and RB1 pathways.Utilizing a newly introduced bioinformatic tool, we provide evidence that chromothripsis is associated with the formation of amplicons containing several oncogenes involved in key pathways that are likely essential for post-chromothriptic cell survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Division of Hematology/Oncology and UC Brain Tumor Center, University of Cincinnati, Cincinnati OH, USA.

ABSTRACT

Background: Findings based on recent advances in next-generation sequence analysis suggest that, in some tumors, a single catastrophic event, termed chromothripsis, results in several simultaneous tumorigenic alterations. Previous studies have suggested that glioblastoma (GBM) may exhibit chromothripsis at a higher rate (39%) than other tumors (9%). Primary glioblastoma is an aggressive form of brain cancer that typically appears suddenly in older adults. With aggressive treatment, the median survival time is only 15 months. Their acute onset and widespread genomic instability indicates that chromothripsis may play a key role in their initiation and progression. GBMs are often characterized by EGFR amplification, CDKN2A and PTEN deletion, although approximately 20% of GBMs harbor additional amplifications in MDM2 or MDM4 with CDK4.

Methods: We used the chromothripsis prediction tool, Shatterproof, in conjunction with a custom whole genome sequence analysis pipeline in order to generate putative regions of chromothripsis. The data derived from this study was further expanded on using fluorescence in situ hybridization (FISH) analysis and susceptibility studies with colony formation assays.

Results: We show that primary GBMs are associated with higher chromothripsis scores and establish a link between chromothripsis and gene amplification of receptor tyrosine kinases (RTKs), as well as modulators of the TP53 and RB1 pathways.

Conclusions: Utilizing a newly introduced bioinformatic tool, we provide evidence that chromothripsis is associated with the formation of amplicons containing several oncogenes involved in key pathways that are likely essential for post-chromothriptic cell survival.

No MeSH data available.


Related in: MedlinePlus

TCGA-19-2624 appears to contain a population of chromothripsis-derived regions that facilitate the amplification of EGFR, MDM2, and CDK4(A) Circos diagram depicting all suspect chromothriptic regions resulting from Shatterproof analysis. (B) Enhancement of the chromosome 7 and 12 recombination seen in 3a supporting the assertion that EGFR, along with MDM2 and CDK4, have been translocated onto the same amplicon. (C) FISH performed on paraffin embedded tissue from TCGA-19-2624 reveals that both EGFR and MDM2 are present at numerous loci within the nucleus, consistent with the formation of double minutes.
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Figure 3: TCGA-19-2624 appears to contain a population of chromothripsis-derived regions that facilitate the amplification of EGFR, MDM2, and CDK4(A) Circos diagram depicting all suspect chromothriptic regions resulting from Shatterproof analysis. (B) Enhancement of the chromosome 7 and 12 recombination seen in 3a supporting the assertion that EGFR, along with MDM2 and CDK4, have been translocated onto the same amplicon. (C) FISH performed on paraffin embedded tissue from TCGA-19-2624 reveals that both EGFR and MDM2 are present at numerous loci within the nucleus, consistent with the formation of double minutes.

Mentions: Chromothripsis results in shattering a genome into many fragments and then stitching them back together, in a seemingly random process, often resulting in the formation of extra-chromosomal double minutes. To test whether this was the case in our patients, tissue slides were obtained from TCGA-19-2624, which shows numerous translocations and massive amplifications involving EGFR, MDM2 and CDK4 (Figure 3a-b). This is consistent with the findings of Garsed et al. which linked 12q amplification to double minutes in several tumors. Fluorescent in-situ hybridization on tissue slides from this patient revealed that many copies of both EGFR and MDM2 are spread throughout the nuclear regions, which is consistent with extra-chromosomal double minutes (Figure 3c). The presence of EGFR in the double minutes would suggest that the shattering affected chromosome 7 as well, although we have no evidence beyond the translocations in our bioinformatic data that suggests that MDM2 and EGFR are within the same double minutes. Unfortunately, it was impossible to confirm the presence of double minutes using metaphase FISH, as tumor-derived cells were not available. Consistent with previous reports, we found that the stitching of the broken fragments involves small homologous sequences indicative of micro-homology mediated repair (Figure S4).


Whole genome sequence analysis links chromothripsis to EGFR, MDM2, MDM4, and CDK4 amplification in glioblastoma.

Furgason JM, Koncar RF, Michelhaugh SK, Sarkar FH, Mittal S, Sloan AE, Barnholtz-Sloan JS, Bahassi el M - Oncoscience (2015)

TCGA-19-2624 appears to contain a population of chromothripsis-derived regions that facilitate the amplification of EGFR, MDM2, and CDK4(A) Circos diagram depicting all suspect chromothriptic regions resulting from Shatterproof analysis. (B) Enhancement of the chromosome 7 and 12 recombination seen in 3a supporting the assertion that EGFR, along with MDM2 and CDK4, have been translocated onto the same amplicon. (C) FISH performed on paraffin embedded tissue from TCGA-19-2624 reveals that both EGFR and MDM2 are present at numerous loci within the nucleus, consistent with the formation of double minutes.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4549359&req=5

Figure 3: TCGA-19-2624 appears to contain a population of chromothripsis-derived regions that facilitate the amplification of EGFR, MDM2, and CDK4(A) Circos diagram depicting all suspect chromothriptic regions resulting from Shatterproof analysis. (B) Enhancement of the chromosome 7 and 12 recombination seen in 3a supporting the assertion that EGFR, along with MDM2 and CDK4, have been translocated onto the same amplicon. (C) FISH performed on paraffin embedded tissue from TCGA-19-2624 reveals that both EGFR and MDM2 are present at numerous loci within the nucleus, consistent with the formation of double minutes.
Mentions: Chromothripsis results in shattering a genome into many fragments and then stitching them back together, in a seemingly random process, often resulting in the formation of extra-chromosomal double minutes. To test whether this was the case in our patients, tissue slides were obtained from TCGA-19-2624, which shows numerous translocations and massive amplifications involving EGFR, MDM2 and CDK4 (Figure 3a-b). This is consistent with the findings of Garsed et al. which linked 12q amplification to double minutes in several tumors. Fluorescent in-situ hybridization on tissue slides from this patient revealed that many copies of both EGFR and MDM2 are spread throughout the nuclear regions, which is consistent with extra-chromosomal double minutes (Figure 3c). The presence of EGFR in the double minutes would suggest that the shattering affected chromosome 7 as well, although we have no evidence beyond the translocations in our bioinformatic data that suggests that MDM2 and EGFR are within the same double minutes. Unfortunately, it was impossible to confirm the presence of double minutes using metaphase FISH, as tumor-derived cells were not available. Consistent with previous reports, we found that the stitching of the broken fragments involves small homologous sequences indicative of micro-homology mediated repair (Figure S4).

Bottom Line: The data derived from this study was further expanded on using fluorescence in situ hybridization (FISH) analysis and susceptibility studies with colony formation assays.We show that primary GBMs are associated with higher chromothripsis scores and establish a link between chromothripsis and gene amplification of receptor tyrosine kinases (RTKs), as well as modulators of the TP53 and RB1 pathways.Utilizing a newly introduced bioinformatic tool, we provide evidence that chromothripsis is associated with the formation of amplicons containing several oncogenes involved in key pathways that are likely essential for post-chromothriptic cell survival.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine, Division of Hematology/Oncology and UC Brain Tumor Center, University of Cincinnati, Cincinnati OH, USA.

ABSTRACT

Background: Findings based on recent advances in next-generation sequence analysis suggest that, in some tumors, a single catastrophic event, termed chromothripsis, results in several simultaneous tumorigenic alterations. Previous studies have suggested that glioblastoma (GBM) may exhibit chromothripsis at a higher rate (39%) than other tumors (9%). Primary glioblastoma is an aggressive form of brain cancer that typically appears suddenly in older adults. With aggressive treatment, the median survival time is only 15 months. Their acute onset and widespread genomic instability indicates that chromothripsis may play a key role in their initiation and progression. GBMs are often characterized by EGFR amplification, CDKN2A and PTEN deletion, although approximately 20% of GBMs harbor additional amplifications in MDM2 or MDM4 with CDK4.

Methods: We used the chromothripsis prediction tool, Shatterproof, in conjunction with a custom whole genome sequence analysis pipeline in order to generate putative regions of chromothripsis. The data derived from this study was further expanded on using fluorescence in situ hybridization (FISH) analysis and susceptibility studies with colony formation assays.

Results: We show that primary GBMs are associated with higher chromothripsis scores and establish a link between chromothripsis and gene amplification of receptor tyrosine kinases (RTKs), as well as modulators of the TP53 and RB1 pathways.

Conclusions: Utilizing a newly introduced bioinformatic tool, we provide evidence that chromothripsis is associated with the formation of amplicons containing several oncogenes involved in key pathways that are likely essential for post-chromothriptic cell survival.

No MeSH data available.


Related in: MedlinePlus