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Intratumoral genetic heterogeneity in metastatic melanoma is accompanied by variation in malignant behaviors.

Anaka M, Hudson C, Lo PH, Do H, Caballero OL, Davis ID, Dobrovic A, Cebon J, Behren A - BMC Med Genomics (2013)

Bottom Line: Minor clones were identified based on dissimilarity to the parental cell line, and these clones were the most clonogenic and least sensitive to drugs.Finally, treatment of a polyclonal cell line with paclitaxel to enrich for drug-resistant cells resulted in the adoption of a gene expression profile with features of one of the minor clones, supporting the idea that these populations can mediate disease relapse.Our results support the hypothesis that minor clones might have major consequences for patient outcomes in melanoma.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cancer Immuno-biology Lab, Ludwig Institute for Cancer Research Melbourne, Austin Branch, Melbourne, Victoria 3084, Australia. jonathan.cebon@ludwig.edu.au.

ABSTRACT

Background: Intratumoral heterogeneity is a major obstacle for the treatment of cancer, as the presence of even minor populations that are insensitive to therapy can lead to disease relapse. Increased clonal diversity has been correlated with a poor prognosis for cancer patients, and we therefore examined genetic, transcriptional, and functional diversity in metastatic melanoma.

Methods: Amplicon sequencing and SNP microarrays were used to profile somatic mutations and DNA copy number changes in multiple regions from metastatic lesions. Clonal genetic and transcriptional heterogeneity was also assessed in single cell clones from early passage cell lines, which were then subjected to clonogenicity and drug sensitivity assays.

Results: MAPK pathway and tumor suppressor mutations were identified in all regions of the melanoma metastases analyzed. In contrast, we identified copy number abnormalities present in only some regions in addition to homogeneously present changes, suggesting ongoing genetic evolution following metastatic spread. Copy number heterogeneity from a tumor was represented in matched cell line clones, which also varied in their clonogenicity and drug sensitivity. Minor clones were identified based on dissimilarity to the parental cell line, and these clones were the most clonogenic and least sensitive to drugs. Finally, treatment of a polyclonal cell line with paclitaxel to enrich for drug-resistant cells resulted in the adoption of a gene expression profile with features of one of the minor clones, supporting the idea that these populations can mediate disease relapse.

Conclusion: Our results support the hypothesis that minor clones might have major consequences for patient outcomes in melanoma.

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Related in: MedlinePlus

LM-MEL-62 features clonal copy number heterogeneity. A) Hierarchical clustering and heatmap of copy number data from single cell clones of LM-MEL-62, the parental cell line, and a fresh frozen fragment of Tumor 1. Black regions represent normal copy number (2), green represents amplifications, red deletions. B) number of copy number abnormalities found in from single cell clones of LM-MEL-62, the parental cell line, and a fresh frozen fragment of Tumor 1. C) Proportion of clones in which specific copy number changes were identified.
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Figure 4: LM-MEL-62 features clonal copy number heterogeneity. A) Hierarchical clustering and heatmap of copy number data from single cell clones of LM-MEL-62, the parental cell line, and a fresh frozen fragment of Tumor 1. Black regions represent normal copy number (2), green represents amplifications, red deletions. B) number of copy number abnormalities found in from single cell clones of LM-MEL-62, the parental cell line, and a fresh frozen fragment of Tumor 1. C) Proportion of clones in which specific copy number changes were identified.

Mentions: The copy number profiles of a cell line derived from Tumor 1 (LM-MEL-62), ten single cell clones derived from LM-MEL-62, and a fresh frozen tumor fragment of Tumor 1 were assessed using Illumina SNP microarrays. All cell line-derived samples clearly shared many common copy number changes with the original tumor, such as gain of 6p (Figure 4A). The clones harbored between 55 and 69 copy number changes, which represents significantly less variation than was observed in the different regions of Tumor 1 (Figure 4B). Nevertheless, the cell clones were also heterogeneous for many of the detected aberrations (Figure 4C), and the LM-MEL-62 clones displayed heterogeneity at chromosome regions similarly affected in the archival FFPE tumor material (Figure 5A and B). This suggests that early passage melanoma cell lines are polyclonal, and that they can retain or recapitulate genetic heterogeneity representative of that found in the patient’s tumor.


Intratumoral genetic heterogeneity in metastatic melanoma is accompanied by variation in malignant behaviors.

Anaka M, Hudson C, Lo PH, Do H, Caballero OL, Davis ID, Dobrovic A, Cebon J, Behren A - BMC Med Genomics (2013)

LM-MEL-62 features clonal copy number heterogeneity. A) Hierarchical clustering and heatmap of copy number data from single cell clones of LM-MEL-62, the parental cell line, and a fresh frozen fragment of Tumor 1. Black regions represent normal copy number (2), green represents amplifications, red deletions. B) number of copy number abnormalities found in from single cell clones of LM-MEL-62, the parental cell line, and a fresh frozen fragment of Tumor 1. C) Proportion of clones in which specific copy number changes were identified.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3852494&req=5

Figure 4: LM-MEL-62 features clonal copy number heterogeneity. A) Hierarchical clustering and heatmap of copy number data from single cell clones of LM-MEL-62, the parental cell line, and a fresh frozen fragment of Tumor 1. Black regions represent normal copy number (2), green represents amplifications, red deletions. B) number of copy number abnormalities found in from single cell clones of LM-MEL-62, the parental cell line, and a fresh frozen fragment of Tumor 1. C) Proportion of clones in which specific copy number changes were identified.
Mentions: The copy number profiles of a cell line derived from Tumor 1 (LM-MEL-62), ten single cell clones derived from LM-MEL-62, and a fresh frozen tumor fragment of Tumor 1 were assessed using Illumina SNP microarrays. All cell line-derived samples clearly shared many common copy number changes with the original tumor, such as gain of 6p (Figure 4A). The clones harbored between 55 and 69 copy number changes, which represents significantly less variation than was observed in the different regions of Tumor 1 (Figure 4B). Nevertheless, the cell clones were also heterogeneous for many of the detected aberrations (Figure 4C), and the LM-MEL-62 clones displayed heterogeneity at chromosome regions similarly affected in the archival FFPE tumor material (Figure 5A and B). This suggests that early passage melanoma cell lines are polyclonal, and that they can retain or recapitulate genetic heterogeneity representative of that found in the patient’s tumor.

Bottom Line: Minor clones were identified based on dissimilarity to the parental cell line, and these clones were the most clonogenic and least sensitive to drugs.Finally, treatment of a polyclonal cell line with paclitaxel to enrich for drug-resistant cells resulted in the adoption of a gene expression profile with features of one of the minor clones, supporting the idea that these populations can mediate disease relapse.Our results support the hypothesis that minor clones might have major consequences for patient outcomes in melanoma.

View Article: PubMed Central - HTML - PubMed

Affiliation: Cancer Immuno-biology Lab, Ludwig Institute for Cancer Research Melbourne, Austin Branch, Melbourne, Victoria 3084, Australia. jonathan.cebon@ludwig.edu.au.

ABSTRACT

Background: Intratumoral heterogeneity is a major obstacle for the treatment of cancer, as the presence of even minor populations that are insensitive to therapy can lead to disease relapse. Increased clonal diversity has been correlated with a poor prognosis for cancer patients, and we therefore examined genetic, transcriptional, and functional diversity in metastatic melanoma.

Methods: Amplicon sequencing and SNP microarrays were used to profile somatic mutations and DNA copy number changes in multiple regions from metastatic lesions. Clonal genetic and transcriptional heterogeneity was also assessed in single cell clones from early passage cell lines, which were then subjected to clonogenicity and drug sensitivity assays.

Results: MAPK pathway and tumor suppressor mutations were identified in all regions of the melanoma metastases analyzed. In contrast, we identified copy number abnormalities present in only some regions in addition to homogeneously present changes, suggesting ongoing genetic evolution following metastatic spread. Copy number heterogeneity from a tumor was represented in matched cell line clones, which also varied in their clonogenicity and drug sensitivity. Minor clones were identified based on dissimilarity to the parental cell line, and these clones were the most clonogenic and least sensitive to drugs. Finally, treatment of a polyclonal cell line with paclitaxel to enrich for drug-resistant cells resulted in the adoption of a gene expression profile with features of one of the minor clones, supporting the idea that these populations can mediate disease relapse.

Conclusion: Our results support the hypothesis that minor clones might have major consequences for patient outcomes in melanoma.

Show MeSH
Related in: MedlinePlus