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Geographical mapping of a multifocal thyroid tumour using genetic alteration analysis & miRNA profiling.

Aherne ST, Smyth PC, Flavin RJ, Russell SM, Denning KM, Li JH, Guenther SM, O'Leary JJ, Sheils OM - Mol. Cancer (2008)

Bottom Line: Several studies have investigated the genetic alteration status of multifocal thyroid tumours, with discordant results.Our data corroborated miRNAs previously discovered in this carcinoma, and additional miRNAs linked to various processes involved in tumour growth and proliferation.The initial genetic alteration analysis indicated that pluriform PTC did not necessarily evolve from classic PTC progenitor foci.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Histopathology, Trinity College, Dublin, Ireland. ahernesi@tcd.ie

ABSTRACT

Background: Papillary thyroid carcinoma (PTC) frequently presents as multiple tumour-foci within a single thyroid gland or pluriform, with synchronous tumours comprising different histological variants, raising questions regarding its clonality. Among the genetic aberrations described in PTC, the BRAF V600E mutation and ret/PTC activation occur most commonly. Several studies have investigated the genetic alteration status of multifocal thyroid tumours, with discordant results. To address the question of clonality this study examined disparate geographical and morphological areas from a single PTC (classic PTC, insular and anaplastic foci, and tumour cells adjacent to vascular invasion and lymphocytic infiltrate) for the presence of ret/PTC 1 or BRAF mutations. Moreover, we wanted to investigate the consistency of miRNA signatures within disparate areas of a tumour, and geographical data was further correlated with expression profiles of 330 different miRNAs. Putative miRNA gene targets were predicted for differentially regulated miRNAs and immunohistochemistry was performed on tissue sections in an effort to investigate phenotypic variations in microvascular density (MVD), and cytokeratin and p53 protein expression levels.

Results: All of the morphological areas proved negative for ret/PTC 1 rearrangement. Two distinct foci with classic morphology harboured the BRAF mutation. All other regions, including the insular and anaplastic areas were negative for the mutation. MiRNA profiles were found to distinguish tumours containing the BRAF mutation from the other tumour types, and to differentiate between the more aggressive insular & anaplastic tumours, and the classic variant. Our data corroborated miRNAs previously discovered in this carcinoma, and additional miRNAs linked to various processes involved in tumour growth and proliferation.

Conclusion: The initial genetic alteration analysis indicated that pluriform PTC did not necessarily evolve from classic PTC progenitor foci. Analysis of miRNA profiles however provided an interesting variation on the clonality question. While hierarchical clustering analysis of miRNA expression supported the hypothesis that discrete areas did not evolve from clonal expansion of tumour cells, it did not exclude the possibility of independent mutational events suggesting both phenomena might occur simultaneously within a tumour to enhance cancer progression in geographical micro-environments within a tumour.

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Related in: MedlinePlus

TaqMan® SNP assays for BRAF V600E mutation detection. DNA from the TPC1, BC-PAP, and K-2 cell lines was used for positive control purposes. TPC1 contains the wild type BRAF allele, K-2 is heterozygous for the mutant allele, and BC-PAP is homozygous for the mutant allele. The figure shows clustering of the samples into 3 distinct groups depending on their respective levels of VIC/FAM fluorescence: homozygous T1799A mutation (●), homozygous wild-type/normal (●) and heterozygous T1799 mutation (●). Negative controls and undetermined samples are also displayed (×). The two classic PTC tumour areas are the only ones that exhibit the BRAF mutation.
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Figure 1: TaqMan® SNP assays for BRAF V600E mutation detection. DNA from the TPC1, BC-PAP, and K-2 cell lines was used for positive control purposes. TPC1 contains the wild type BRAF allele, K-2 is heterozygous for the mutant allele, and BC-PAP is homozygous for the mutant allele. The figure shows clustering of the samples into 3 distinct groups depending on their respective levels of VIC/FAM fluorescence: homozygous T1799A mutation (●), homozygous wild-type/normal (●) and heterozygous T1799 mutation (●). Negative controls and undetermined samples are also displayed (×). The two classic PTC tumour areas are the only ones that exhibit the BRAF mutation.

Mentions: TaqMan® SNP detection was used to detect the presence of the BRAF mutation. This assay acts as its own endogenous control as the wild type allele will be detected in the absence of the mutant. The two foci with classic morphology harboured the BRAF mutation. All other tumour areas, including the insular and anaplastic areas were negative for the mutation (Figure 1). This result indicates that the anaplastic lesions may not have evolved from progenitor classic PTC foci.


Geographical mapping of a multifocal thyroid tumour using genetic alteration analysis & miRNA profiling.

Aherne ST, Smyth PC, Flavin RJ, Russell SM, Denning KM, Li JH, Guenther SM, O'Leary JJ, Sheils OM - Mol. Cancer (2008)

TaqMan® SNP assays for BRAF V600E mutation detection. DNA from the TPC1, BC-PAP, and K-2 cell lines was used for positive control purposes. TPC1 contains the wild type BRAF allele, K-2 is heterozygous for the mutant allele, and BC-PAP is homozygous for the mutant allele. The figure shows clustering of the samples into 3 distinct groups depending on their respective levels of VIC/FAM fluorescence: homozygous T1799A mutation (●), homozygous wild-type/normal (●) and heterozygous T1799 mutation (●). Negative controls and undetermined samples are also displayed (×). The two classic PTC tumour areas are the only ones that exhibit the BRAF mutation.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2612696&req=5

Figure 1: TaqMan® SNP assays for BRAF V600E mutation detection. DNA from the TPC1, BC-PAP, and K-2 cell lines was used for positive control purposes. TPC1 contains the wild type BRAF allele, K-2 is heterozygous for the mutant allele, and BC-PAP is homozygous for the mutant allele. The figure shows clustering of the samples into 3 distinct groups depending on their respective levels of VIC/FAM fluorescence: homozygous T1799A mutation (●), homozygous wild-type/normal (●) and heterozygous T1799 mutation (●). Negative controls and undetermined samples are also displayed (×). The two classic PTC tumour areas are the only ones that exhibit the BRAF mutation.
Mentions: TaqMan® SNP detection was used to detect the presence of the BRAF mutation. This assay acts as its own endogenous control as the wild type allele will be detected in the absence of the mutant. The two foci with classic morphology harboured the BRAF mutation. All other tumour areas, including the insular and anaplastic areas were negative for the mutation (Figure 1). This result indicates that the anaplastic lesions may not have evolved from progenitor classic PTC foci.

Bottom Line: Several studies have investigated the genetic alteration status of multifocal thyroid tumours, with discordant results.Our data corroborated miRNAs previously discovered in this carcinoma, and additional miRNAs linked to various processes involved in tumour growth and proliferation.The initial genetic alteration analysis indicated that pluriform PTC did not necessarily evolve from classic PTC progenitor foci.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Histopathology, Trinity College, Dublin, Ireland. ahernesi@tcd.ie

ABSTRACT

Background: Papillary thyroid carcinoma (PTC) frequently presents as multiple tumour-foci within a single thyroid gland or pluriform, with synchronous tumours comprising different histological variants, raising questions regarding its clonality. Among the genetic aberrations described in PTC, the BRAF V600E mutation and ret/PTC activation occur most commonly. Several studies have investigated the genetic alteration status of multifocal thyroid tumours, with discordant results. To address the question of clonality this study examined disparate geographical and morphological areas from a single PTC (classic PTC, insular and anaplastic foci, and tumour cells adjacent to vascular invasion and lymphocytic infiltrate) for the presence of ret/PTC 1 or BRAF mutations. Moreover, we wanted to investigate the consistency of miRNA signatures within disparate areas of a tumour, and geographical data was further correlated with expression profiles of 330 different miRNAs. Putative miRNA gene targets were predicted for differentially regulated miRNAs and immunohistochemistry was performed on tissue sections in an effort to investigate phenotypic variations in microvascular density (MVD), and cytokeratin and p53 protein expression levels.

Results: All of the morphological areas proved negative for ret/PTC 1 rearrangement. Two distinct foci with classic morphology harboured the BRAF mutation. All other regions, including the insular and anaplastic areas were negative for the mutation. MiRNA profiles were found to distinguish tumours containing the BRAF mutation from the other tumour types, and to differentiate between the more aggressive insular & anaplastic tumours, and the classic variant. Our data corroborated miRNAs previously discovered in this carcinoma, and additional miRNAs linked to various processes involved in tumour growth and proliferation.

Conclusion: The initial genetic alteration analysis indicated that pluriform PTC did not necessarily evolve from classic PTC progenitor foci. Analysis of miRNA profiles however provided an interesting variation on the clonality question. While hierarchical clustering analysis of miRNA expression supported the hypothesis that discrete areas did not evolve from clonal expansion of tumour cells, it did not exclude the possibility of independent mutational events suggesting both phenomena might occur simultaneously within a tumour to enhance cancer progression in geographical micro-environments within a tumour.

Show MeSH
Related in: MedlinePlus