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Recurrent chromosomal copy number alterations in sporadic chordomas.

Le LP, Nielsen GP, Rosenberg AE, Thomas D, Batten JM, Deshpande V, Schwab J, Duan Z, Xavier RJ, Hornicek FJ, Iafrate AJ - PLoS ONE (2011)

Bottom Line: One copy loss of the 10q23.31 region which encodes PTEN was found in 16/20 (80%) cases.Our sporadic chordoma cases did not show hotspot point mutations in some common cancer gene targets.Moreover, most of these sporadic tumors are not associated with T (brachyury) duplication or amplification.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America. lple@partners.org

ABSTRACT
The molecular events in chordoma pathogenesis have not been fully delineated, particularly with respect to copy number changes. Understanding copy number alterations in chordoma may reveal critical disease mechanisms that could be exploited for tumor classification and therapy. We report the copy number analysis of 21 sporadic chordomas using array comparative genomic hybridization (CGH). Recurrent copy changes were further evaluated with immunohistochemistry, methylation specific PCR, and quantitative real-time PCR. Similar to previous findings, large copy number losses, involving chromosomes 1p, 3, 4, 9, 10, 13, 14, and 18, were more common than copy number gains. Loss of CDKN2A with or without loss of CDKN2B on 9p21.3 was observed in 16/20 (80%) unique cases of which six (30%) showed homozygous deletions ranging from 76 kilobases to 4.7 megabases. One copy loss of the 10q23.31 region which encodes PTEN was found in 16/20 (80%) cases. Loss of CDKN2A and PTEN expression in the majority of cases was not attributed to promoter methylation. Our sporadic chordoma cases did not show hotspot point mutations in some common cancer gene targets. Moreover, most of these sporadic tumors are not associated with T (brachyury) duplication or amplification. Deficiency of CDKN2A and PTEN expression, although shared across many other different types of tumors, likely represents a key aspect of chordoma pathogenesis. Sporadic chordomas may rely on mechanisms other than copy number gain if they indeed exploit T/brachyury for proliferation.

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CDKN2A and PTEN methylation                            specific PCR.Bisulfite-treated chordoma DNA samples were tested with methylation                            specific PCR to evaluate for hypermethylation of the                                CDKN2A and PTEN promoter regions.                            Two sets of unmethylated (U) and methylated (M) PCR primers were used                            for each target gene (bottom labels, MSP1 and MSP2). Unmethylated and                            methylated controls are shown along with results for case CH33. Results                            for other tested cases are summarized in Tables 3 and 4 under the CDKN2A                            and PTEN MSP1 and MSP2 columns. Tick marks on the left                            and right of each panel indicate 100, 200, 300, and 400 base pair sizes                            (bottom to top).
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pone-0018846-g005: CDKN2A and PTEN methylation specific PCR.Bisulfite-treated chordoma DNA samples were tested with methylation specific PCR to evaluate for hypermethylation of the CDKN2A and PTEN promoter regions. Two sets of unmethylated (U) and methylated (M) PCR primers were used for each target gene (bottom labels, MSP1 and MSP2). Unmethylated and methylated controls are shown along with results for case CH33. Results for other tested cases are summarized in Tables 3 and 4 under the CDKN2A and PTEN MSP1 and MSP2 columns. Tick marks on the left and right of each panel indicate 100, 200, 300, and 400 base pair sizes (bottom to top).

Mentions: CDKN2A and PTEN promoter hypermethylation was evaluated by using methylation specific PCR (MSP). Two sets of previously reported unmethylated and methylated specific primers were used in a touchdown PCR protocol to amplify CpG islands in the promoter regions after bisulfite treatment of the genomic DNA [18], [19]. Only one tested chordoma case (CH33) showed definitive evidence of CDKN2A promoter methylation with positive PCR amplification products using both sets of methylation specific primers (Table 3 and Figure 5). Equivocal PTEN promoter methylation specific PCR results (positive amplification with only one out of two primer sets) were obtained for five tested chordoma cases (CH34, CH36, CH37, P527, and P984, Table 4). No tumors showed definitive PTEN promoter methylation.


Recurrent chromosomal copy number alterations in sporadic chordomas.

Le LP, Nielsen GP, Rosenberg AE, Thomas D, Batten JM, Deshpande V, Schwab J, Duan Z, Xavier RJ, Hornicek FJ, Iafrate AJ - PLoS ONE (2011)

CDKN2A and PTEN methylation                            specific PCR.Bisulfite-treated chordoma DNA samples were tested with methylation                            specific PCR to evaluate for hypermethylation of the                                CDKN2A and PTEN promoter regions.                            Two sets of unmethylated (U) and methylated (M) PCR primers were used                            for each target gene (bottom labels, MSP1 and MSP2). Unmethylated and                            methylated controls are shown along with results for case CH33. Results                            for other tested cases are summarized in Tables 3 and 4 under the CDKN2A                            and PTEN MSP1 and MSP2 columns. Tick marks on the left                            and right of each panel indicate 100, 200, 300, and 400 base pair sizes                            (bottom to top).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3094331&req=5

pone-0018846-g005: CDKN2A and PTEN methylation specific PCR.Bisulfite-treated chordoma DNA samples were tested with methylation specific PCR to evaluate for hypermethylation of the CDKN2A and PTEN promoter regions. Two sets of unmethylated (U) and methylated (M) PCR primers were used for each target gene (bottom labels, MSP1 and MSP2). Unmethylated and methylated controls are shown along with results for case CH33. Results for other tested cases are summarized in Tables 3 and 4 under the CDKN2A and PTEN MSP1 and MSP2 columns. Tick marks on the left and right of each panel indicate 100, 200, 300, and 400 base pair sizes (bottom to top).
Mentions: CDKN2A and PTEN promoter hypermethylation was evaluated by using methylation specific PCR (MSP). Two sets of previously reported unmethylated and methylated specific primers were used in a touchdown PCR protocol to amplify CpG islands in the promoter regions after bisulfite treatment of the genomic DNA [18], [19]. Only one tested chordoma case (CH33) showed definitive evidence of CDKN2A promoter methylation with positive PCR amplification products using both sets of methylation specific primers (Table 3 and Figure 5). Equivocal PTEN promoter methylation specific PCR results (positive amplification with only one out of two primer sets) were obtained for five tested chordoma cases (CH34, CH36, CH37, P527, and P984, Table 4). No tumors showed definitive PTEN promoter methylation.

Bottom Line: One copy loss of the 10q23.31 region which encodes PTEN was found in 16/20 (80%) cases.Our sporadic chordoma cases did not show hotspot point mutations in some common cancer gene targets.Moreover, most of these sporadic tumors are not associated with T (brachyury) duplication or amplification.

View Article: PubMed Central - PubMed

Affiliation: Department of Pathology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts, United States of America. lple@partners.org

ABSTRACT
The molecular events in chordoma pathogenesis have not been fully delineated, particularly with respect to copy number changes. Understanding copy number alterations in chordoma may reveal critical disease mechanisms that could be exploited for tumor classification and therapy. We report the copy number analysis of 21 sporadic chordomas using array comparative genomic hybridization (CGH). Recurrent copy changes were further evaluated with immunohistochemistry, methylation specific PCR, and quantitative real-time PCR. Similar to previous findings, large copy number losses, involving chromosomes 1p, 3, 4, 9, 10, 13, 14, and 18, were more common than copy number gains. Loss of CDKN2A with or without loss of CDKN2B on 9p21.3 was observed in 16/20 (80%) unique cases of which six (30%) showed homozygous deletions ranging from 76 kilobases to 4.7 megabases. One copy loss of the 10q23.31 region which encodes PTEN was found in 16/20 (80%) cases. Loss of CDKN2A and PTEN expression in the majority of cases was not attributed to promoter methylation. Our sporadic chordoma cases did not show hotspot point mutations in some common cancer gene targets. Moreover, most of these sporadic tumors are not associated with T (brachyury) duplication or amplification. Deficiency of CDKN2A and PTEN expression, although shared across many other different types of tumors, likely represents a key aspect of chordoma pathogenesis. Sporadic chordomas may rely on mechanisms other than copy number gain if they indeed exploit T/brachyury for proliferation.

Show MeSH
Related in: MedlinePlus