nach oben

International Journal of Clinical Oncology

Erschienen in:

04.04.2016 | Original Article

Quantitative analysis of the BRAF ^V600E mutation in circulating tumor-derived DNA in melanoma patients using competitive allele-specific TaqMan PCR

verfasst von: Atsuko Ashida, Kaori Sakaizawa, Asuka Mikoshiba, Hisashi Uhara, Ryuhei Okuyama

Erschienen in: International Journal of Clinical Oncology | Ausgabe 5/2016

Einloggen, um Zugang zu erhalten

Abstract

Background

BRAF ^V600E is a common mutation in melanoma, and BRAF inhibitors are effective in treating of BRAF mutation-positive melanoma. DNA carrying this mutation is released from melanoma cells into the circulation. As such, circulating tumor-derived DNA (ctDNA) in peripheral blood represents a novel biomarker for evaluating tumor features in cancer patients. However, ctDNA is present in the peripheral blood at very low levels, which makes the detection of specific mutations in this DNA a challenge. Competitive allele-specific TaqMan PCR (castPCR), a straightforward commercially available assay, is a sensitive technique for quantitating a small amount of DNA.

Methods

The level of BRAF ^V600E ctDNA was quantified by castPCR in 26 consecutive plasma samples from six melanoma patients.

Results

The castPCR assay was performed using a mixture of BRAF ^V600E DNA and BRAF ^wild DNA and found to be able to detect BRAF ^V600E at a fractional abundance of ≥0.5 % in 2- to 10-ng samples of genomic DNA. Cell-free DNA was then extracted from peripheral blood samples collected from six patients with melanoma harboring the BRAF ^V600E mutation. BRAF ^V600E ctDNA was detected in three patients, at a fractional abundance of between 1.28 and 58.0 % of total BRAF cell-free DNA. The abundance of BRAF ^V600E ctDNA correlated with tumor burden, as determined by computed tomography imaging. In two cases, an increase in the level of BRAF ^V600E ctDNA preceded exacerbation of clinical symptoms.

Conclusion

The castPCR assay can detect and quantitate small amounts of BRAF ^V600E ctDNA in samples containing large amounts of BRAF ^wild cell-free DNA. Thus, we suggest that the castPCR assay is suitable for monitoring ctDNA in the plasma of melanoma patients.

Menzies AM, Long GV (2014) Systemic treatment for BRAF-mutant melanoma: where do we go next? Lancet Oncol 15:e371–e381CrossRefPubMed

Diaz-Lagares A, Alegre E, Arroyo A et al (2011) Evaluation of multiple serum markers in advanced melanoma. Tumour Biol 32:1155–1161CrossRefPubMed

Sanmamed MF, Fernandez-Landazuri S, Rodriguez C et al (2014) Relevance of MIA and S100 serum tumor markers to monitor BRAF inhibitor therapy in metastatic melanoma patients. Clin Chim Acta 429:168–174CrossRefPubMed

Schwarzenbach H, Hoon DS, Pantel K (2011) Cell-free nucleic acids as biomarkers in cancer patients. Nat Rev Cancer 11:426–437CrossRefPubMed

Dawson SJ, Tsui DW, Murtaza M et al (2013) Analysis of circulating tumor DNA to monitor metastatic breast cancer. N Engl J Med 368:1199–1209CrossRefPubMed

Oxnard GR, Paweletz CP, Kuang Y et al (2014) Noninvasive detection of response and resistance in EGFR-mutant lung cancer using quantitative next-generation genotyping of cell-free plasma DNA. Clin Cancer Res 20:1698–1705CrossRefPubMedPubMedCentral

Taly V, Pekin D, Benhaim L et al (2013) Multiplex picodroplet digital PCR to detect KRAS mutations in circulating DNA from the plasma of colorectal cancer patients. Clin Chem 59:1722–1731CrossRefPubMed

Thierry AR, Mouliere F, El Messaoudi S et al (2014) Clinical validation of the detection of KRAS and BRAF mutations from circulating tumor DNA. Nat Med 20:430–435CrossRefPubMed

Huggett JF, Whale A (2013) Digital PCR as a novel technology and its potential implications for molecular diagnostics. Clin Chem 59:1691–1693CrossRefPubMed

10.

Siravegna G, Bardelli A (2014) Minimal residual disease in breast cancer: in blood veritas. Clin Cancer Res 20:2505–2507CrossRefPubMed

11.

Ashida A, Uhara H, Kiniwa Y et al (2012) Assessment of BRAF and KIT mutations in Japanese melanoma patients. J Dermatol Sci 66:240–242CrossRefPubMed

12.

Cheng SP, Hsu YC, Liu CL et al (2014) Significance of allelic percentage of BRAF c.1799T > A (V600E) mutation in papillary thyroid carcinoma. Ann Surg Oncol 21[Suppl 4]:S619–S626CrossRefPubMed

13.

Didelot A, Le Corre D, Luscan A et al (2012) Competitive allele specific TaqMan PCR for KRAS, BRAF and EGFR mutation detection in clinical formalin fixed paraffin embedded samples. Exp Mol Pathol 92:275–280CrossRefPubMed

14.

Ashida A, Uhara H, Mikoshiba A et al (2015) Melanoma with BRAF mutation in circulating cell-free DNA despite no mutation in the primary lesion: a case report. Acta Derm Venereol 96:128–129CrossRef

15.

Schreuer M, Meersseman G, van Den Herrewegen S et al (2016) Applications for quantitative measurement of BRAF V600 mutant cell-free tumor DNA in the plasma of patients with metastatic melanoma. Melanoma Res 26:157–163CrossRefPubMed

16.

Wakamatsu K, Ito S, Horikoshi T (1991) Normal values of urinary excretion and serum concentration of 5-S-cysteinyldopa and 6-hydroxy-5-methoxyindole-2-carboxylic acid, biochemical markers of melanoma progression. Melanoma Res 1:141–147CrossRefPubMed

17.

Sakaizawa K, Goto Y, Kiniwa Y et al (2012) Mutation analysis of BRAF and KIT in circulating melanoma cells at the single cell level. Br J Cancer 106:939–946CrossRefPubMedPubMedCentral

18.

Stoecklein NH, Hosch SB, Bezler M et al (2008) Direct genetic analysis of single disseminated cancer cells for prediction of outcome and therapy selection in esophageal cancer. Cancer Cell 13:441–453CrossRefPubMed

19.

Diehl F, Schmidt K, Choti MA et al (2008) Circulating mutant DNA to assess tumor dynamics. Nat Med 14:985–990CrossRefPubMed

20.

Taniguchi K, Uchida J, Nishino K et al (2011) Quantitative detection of EGFR mutations in circulating tumor DNA derived from lung adenocarcinomas. Clin Cancer Res 17:7808–7815CrossRefPubMed

21.

Sanmamed MF, Fernandez-Landazuri S, Rodriguez C et al (2015) Quantitative cell-free circulating BRAFV600E mutation analysis by use of droplet digital PCR in the follow-up of patients with melanoma being treated with BRAF inhibitors. Clin Chem 61:297–304CrossRefPubMed

22.

Kukita Y, Uchida J, Oba S et al (2013) Quantitative identification of mutant alleles derived from lung cancer in plasma cell-free DNA via anomaly detection using deep sequencing data. PLoS One 8:e81468CrossRefPubMedPubMedCentral

23.

Wakamatsu K, Kageshita T, Furue M et al (2002) Evaluation of 5-S-cysteinyldopa as a marker of melanoma progression: 10 years’ experience. Melanoma Res 12:245–253CrossRefPubMed

24.

Colombino M, Capone M, Lissia A et al (2012) BRAF/NRAS mutation frequencies among primary tumors and metastases in patients with melanoma. J Clin Oncol 30:2522–2529CrossRefPubMed

25.

Saint-Jean M, Quereux G, Nguyen JM et al (2014) Is a single BRAF wild-type test sufficient to exclude melanoma patients from vemurafenib therapy? J Invest Dermatol 134:1468–1470CrossRefPubMed

Titel: Quantitative analysis of the BRAF V600E mutation in circulating tumor-derived DNA in melanoma patients using competitive allele-specific TaqMan PCR
verfasst von: Atsuko Ashida
Kaori Sakaizawa
Asuka Mikoshiba
Hisashi Uhara
Ryuhei Okuyama
Publikationsdatum: 04.04.2016
Verlag: Springer Japan
Erschienen in: International Journal of Clinical Oncology / Ausgabe 5/2016
Print ISSN: 1341-9625
Elektronische ISSN: 1437-7772
DOI: https://doi.org/10.1007/s10147-016-0976-y

Neu im Fachgebiet Onkologie

19.04.2024 | EAU 2024 | Kongressbericht | Nachrichten

Live-Webinar: Aktuelle Leitlinien bei Herz-Kreislauf-Erkrankungen

Springer Medizin

Quantitative analysis of the BRAF ^V600E mutation in circulating tumor-derived DNA in melanoma patients using competitive allele-specific TaqMan PCR

Abstract

Background

Methods

Results

Conclusion

Neu im Fachgebiet Onkologie

Prostatakarzinom: EU initiiert neues Screeningkonzept

Blasenkarzinom – Biomarker statt Zytologie?

Stereotaktische Radiatio schlägt konventionelle Bestrahlung

Adjuvante Brustkrebstherapie: Doppelt hält besser

Update Onkologie

Live-Webinar: Aktuelle Leitlinien bei Herz-Kreislauf-Erkrankungen

Springer Medizin

Abstract

Background

Methods

Results

Conclusion

Bitte loggen Sie sich ein, um Zugang zu diesem Inhalt zu erhalten

Weitere Artikel der Ausgabe 5/2016

Spectrum of cancer risk among Taiwanese with chronic obstructive pulmonary disease

Identification of adverse events that have a negative impact on quality of life in a clinical trial comparing docetaxel versus S-1 with cisplatin in lung cancer

MAGE-A1–6 expression in patients with head and neck squamous cell carcinoma: impact on clinical patterns and oncologic outcomes

Treatment results and prognostic factors for advanced squamous cell carcinoma of the head and neck treated with salvage surgery after concurrent chemoradiotherapy

Approving molecularly targeted drugs: different approval processes for cytotoxic agents

Comment on Ueno et al.: Prevalence of laparoscopic surgical treatment and its clinical outcomes in patients with familial adenomatous polyposis in Japan

Neu im Fachgebiet Onkologie

Prostatakarzinom: EU initiiert neues Screeningkonzept

Blasenkarzinom – Biomarker statt Zytologie?

Stereotaktische Radiatio schlägt konventionelle Bestrahlung

Adjuvante Brustkrebstherapie: Doppelt hält besser

Update Onkologie