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Genetic profiling of tumours using both circulating free DNA and circulating tumour cells isolated from the same preserved whole blood sample.

Rothwell DG, Smith N, Morris D, Leong HS, Li Y, Hollebecque A, Ayub M, Carter L, Antonello J, Franklin L, Miller C, Blackhall F, Dive C, Brady G - Mol Oncol (2015)

Bottom Line: However, accurate assessment of both CTCs and ctDNA requires all blood cells to be maintained intact until samples are processed.We demonstrate that yields of circulating free DNA (cfDNA) obtained from whole blood CellSave samples are equivalent to those obtained from conventional EDTA plasma processed within 4 h of blood draw.Targeted and genome-wide NGS revealed comparable DNA quality and resultant sequence information from cfDNA within CellSave and EDTA samples.

View Article: PubMed Central - PubMed

Affiliation: Nucleic Acid Biomarker Laboratory, Clinical Experimental Pharmacology Group, CR-UK Manchester Institute, University of Manchester, M20 4BX, UK. Electronic address: dominic.rothwell@cruk.mancehster.ac.uk.

No MeSH data available.


Related in: MedlinePlus

A. Schematic showing procedure for processing of a single blood sample to give NGS analysis of cfDNA and CTCs. B & C. Unsupervised, hierarchical clustering of CNA profiles in two SCLC patients. CNA profiles were generated from isolated CTCs, EDTA cfDNA, CellSave cfDNA, two CDX tumours (A only), germline gDNA and isolated WBC. Matching patterns of gain (regions of red) and loss (regions of blue) were seen across all tumour material and were absent from germline controls. Arrows indicate location of common copy number aberrations found in SCLC with red indicating gain and blue loss. Dark filled arrows indicate loci altered in the patient sample.
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fig4: A. Schematic showing procedure for processing of a single blood sample to give NGS analysis of cfDNA and CTCs. B & C. Unsupervised, hierarchical clustering of CNA profiles in two SCLC patients. CNA profiles were generated from isolated CTCs, EDTA cfDNA, CellSave cfDNA, two CDX tumours (A only), germline gDNA and isolated WBC. Matching patterns of gain (regions of red) and loss (regions of blue) were seen across all tumour material and were absent from germline controls. Arrows indicate location of common copy number aberrations found in SCLC with red indicating gain and blue loss. Dark filled arrows indicate loci altered in the patient sample.

Mentions: As well as the identification of tumour associated mutations, low depth whole genome sequencing (WGS) of cfDNA can be used to characterise CNA patterns arising from the circulating tumour DNA (ctDNA) present in the total cfDNA (Leary et al., 2012, Mohan et al., 2014). Since we and others have shown CNA analysis can be readily applied to CTCs isolated following CellSearch enrichment (Hodgkinson et al., 2014, Gasch et al., 2013, Heitzer et al., 2013), use of CellSave for cfDNA isolation would enable combined CTC and cfDNA analysis from the same collection tube (Figure 4A), thereby maximizing the potential clinical information that can be elucidated.


Genetic profiling of tumours using both circulating free DNA and circulating tumour cells isolated from the same preserved whole blood sample.

Rothwell DG, Smith N, Morris D, Leong HS, Li Y, Hollebecque A, Ayub M, Carter L, Antonello J, Franklin L, Miller C, Blackhall F, Dive C, Brady G - Mol Oncol (2015)

A. Schematic showing procedure for processing of a single blood sample to give NGS analysis of cfDNA and CTCs. B & C. Unsupervised, hierarchical clustering of CNA profiles in two SCLC patients. CNA profiles were generated from isolated CTCs, EDTA cfDNA, CellSave cfDNA, two CDX tumours (A only), germline gDNA and isolated WBC. Matching patterns of gain (regions of red) and loss (regions of blue) were seen across all tumour material and were absent from germline controls. Arrows indicate location of common copy number aberrations found in SCLC with red indicating gain and blue loss. Dark filled arrows indicate loci altered in the patient sample.
© Copyright Policy - CC BY-NC-ND
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4834815&req=5

fig4: A. Schematic showing procedure for processing of a single blood sample to give NGS analysis of cfDNA and CTCs. B & C. Unsupervised, hierarchical clustering of CNA profiles in two SCLC patients. CNA profiles were generated from isolated CTCs, EDTA cfDNA, CellSave cfDNA, two CDX tumours (A only), germline gDNA and isolated WBC. Matching patterns of gain (regions of red) and loss (regions of blue) were seen across all tumour material and were absent from germline controls. Arrows indicate location of common copy number aberrations found in SCLC with red indicating gain and blue loss. Dark filled arrows indicate loci altered in the patient sample.
Mentions: As well as the identification of tumour associated mutations, low depth whole genome sequencing (WGS) of cfDNA can be used to characterise CNA patterns arising from the circulating tumour DNA (ctDNA) present in the total cfDNA (Leary et al., 2012, Mohan et al., 2014). Since we and others have shown CNA analysis can be readily applied to CTCs isolated following CellSearch enrichment (Hodgkinson et al., 2014, Gasch et al., 2013, Heitzer et al., 2013), use of CellSave for cfDNA isolation would enable combined CTC and cfDNA analysis from the same collection tube (Figure 4A), thereby maximizing the potential clinical information that can be elucidated.

Bottom Line: However, accurate assessment of both CTCs and ctDNA requires all blood cells to be maintained intact until samples are processed.We demonstrate that yields of circulating free DNA (cfDNA) obtained from whole blood CellSave samples are equivalent to those obtained from conventional EDTA plasma processed within 4 h of blood draw.Targeted and genome-wide NGS revealed comparable DNA quality and resultant sequence information from cfDNA within CellSave and EDTA samples.

View Article: PubMed Central - PubMed

Affiliation: Nucleic Acid Biomarker Laboratory, Clinical Experimental Pharmacology Group, CR-UK Manchester Institute, University of Manchester, M20 4BX, UK. Electronic address: dominic.rothwell@cruk.mancehster.ac.uk.

No MeSH data available.


Related in: MedlinePlus