BACKGROUND: Exosomes arise from viable cancer cells and may reflect a different biology than circulating cell-free DNA (cfDNA) shed from dying tissues. We compare exosome-derived DNA (exoDNA) to cfDNA in liquid biopsies of patients with pancreatic ductal adenocarcinoma (PDAC). PATIENTS AND METHODS: Patient samples were obtained between 2003 and 2010, with clinically annotated follow up to 2015. Droplet digital PCR was performed on exoDNA and cfDNA for sensitive detection of KRAS mutants at codons 12/13. A cumulative series of 263 individuals were studied, including a discovery cohort of 142 individuals: 68 PDAC patients of all stages; 20 PDAC patients initially staged with localized disease, with blood drawn after resection for curative intent; and 54 age-matched healthy controls. A validation cohort of 121 individuals (39 cancer patients and 82 healthy controls) was studied to validate KRAS detection rates in early-stage PDAC patients. Primary outcome was circulating KRAS status as detected by droplet digital PCR. Secondary outcomes were disease-free and overall survival. RESULTS: KRAS mutations in exoDNA, were identified in 7.4%, 66.7%, 80%, and 85% of age-matched controls, localized, locally advanced, and metastatic PDAC patients, respectively. Comparatively, mutant KRAS cfDNA was detected in 14.8%, 45.5%, 30.8%, and 57.9% of these individuals. Higher exoKRAS MAFs were associated with decreased disease-free survival in patients with localized disease. In the validation cohort, mutant KRAS exoDNA was detected in 43.6% of early-stage PDAC patients and 20% of healthy controls. CONCLUSIONS: Exosomes are a distinct source of tumor DNA that may be complementary to other liquid biopsy DNA sources. A higher percentage of patients with localized PDAC exhibited detectable KRAS mutations in exoDNA than previously reported for cfDNA. A substantial minority of healthy samples demonstrated mutant KRAS in circulation, dictating careful consideration and application of liquid biopsy findings, which may limit its utility as a broad cancer-screening method.

Department of Cancer Epidemiology and Genetics Masaryk Memorial Cancer Institute Brno Czech Republic

Department of Clinical Cancer Prevention The University of Texas MD Anderson Cancer Center Houston USA

Department of Epidemiology and Public Health Faculty of Medicine University of Ostrava Ostrava Czech Republic

Department of GI Medical Oncology The University of Texas MD Anderson Cancer Center Houston USA

Department of Investigational Cancer Therapeutics and the Institute for Personalized Cancer Therapy Houston USA

Department of Pathology The University of Texas MD Anderson Cancer Center Houston USA

Department of Preventive Medicine Palacky University of Medicine Olomouc Czech Republic

Department of Surgical Oncology The University of Texas MD Anderson Cancer Center Houston USA

Department of Translational Molecular Pathology The University of Texas MD Anderson Cancer Center Houston USA

Genetic Epidemiology Group International Agency for Research on Cancer Lyon France

Institute of Public Health and Preventive Medicine 2nd Faculty of Medicine Charles University Prague Czech Republic

Regional Authority of Public Health in Banska Bystrica Banska Bystrica Slovakia

Section of Experimental Pathology University of Texas M D Anderson Cancer Center Houston USA

Sheikh Ahmed Pancreatic Cancer Research Center The University of Texas MD Anderson Cancer Center Houston USA

Ann Oncol. 2017 Apr 1;28(4):677-678. doi: 10.1093/annonc/mdx056. PubMed

Zobrazit více v PubMed

Ryan DP, Hong TS, Bardeesy N.. Pancreatic adenocarcinoma. N Engl J Med 2014; 371: 1039–1049. PubMed

Yeo TP. Demographics, epidemiology, and inheritance of pancreatic ductal adenocarcinoma. Semin Oncol 2015; 42: 8–18. PubMed

Warner E. Clinical practice. Breast-cancer screening. N Engl J Med 2011; 365: 1025–1032. PubMed

Arteaga CL. Progress in breast cancer: overview. Clin Cancer Res 2013; 19: 6353–6359. PubMed

Lieberman DA. Clinical practice. Screening for colorectal cancer. N Engl J Med 2009; 361: 1179–1187. PubMed

Lee MS, Kopetz S.. Current Future Approaches to target the epidermal growth factor receptor and its downstream signaling in metastatic colorectal cancer. Clin Colorectal Cancer 2015; 14: 203–218. PubMed

Zhou C, Wu YL, Chen G. et al. Erlotinib versus chemotherapy as first-line treatment for patients with advanced EGFR mutation-positive non-small-cell lung cancer (OPTIMAL, CTONG-0802): a multicentre, open-label, randomised, phase 3 study. Lancet Oncol 2011; 12: 735–742. PubMed

Smith RA, Manassaram-Baptiste D, Brooks D. et al. Cancer screening in the United States, 2015: a review of current American cancer society guidelines and current issues in cancer screening. CA Cancer J Clin 2015; 65: 30–54. PubMed

Katz MH, Wang H, Fleming JB. et al. Long-term survival after multidisciplinary management of resected pancreatic adenocarcinoma. Ann Surg Oncol 2009; 16: 836–847. PubMed PMC

Howlader N NA, Krapcho M, Miller D. et al. (eds). SEER Cancer Statistics Review, 1975–2013. Bethesda, MD: National Cancer Institute; 2016.

Siegel RL, Miller KD, Jemal A.. Cancer statistics, 2016. CA Cancer J Clin 2016; 66: 7–30. PubMed

Chakraborty S, Baine MJ, Sasson AR, Batra SK.. Current status of molecular markers for early detection of sporadic pancreatic cancer. Biochim Biophys Acta 2011; 1815: 44–64. PubMed PMC

Vietsch EE, van Eijck CH, Wellstein A.. Circulating DNA and micro-RNA in patients with pancreatic cancer. Pancreat Disord Ther 2015; 5(2): 156. PubMed PMC

Mulcahy HE, Lyautey J, Lederrey C. et al. A prospective study of K-ras mutations in the plasma of pancreatic cancer patients. Clin Cancer Res 1998; 4: 271–275. PubMed

Dianxu F, Shengdao Z, Tianquan H. et al. A prospective study of detection of pancreatic carcinoma by combined plasma K-ras mutations and serum CA19-9 analysis. Pancreas 2002; 25: 336–341. PubMed

Maire F, Micard S, Hammel P. et al. Differential diagnosis between chronic pancreatitis and pancreatic cancer: value of the detection of KRAS2 mutations in circulating DNA. Br J Cancer 2002; 87: 551–554. PubMed PMC

Uemura T, Hibi K, Kaneko T. et al. Detection of K-ras mutations in the plasma DNA of pancreatic cancer patients. J Gastroenterol 2004; 39: 56–60. PubMed

Dabritz J, Preston R, Hanfler J, Oettle H.. Follow-up study of K-ras mutations in the plasma of patients with pancreatic cancer: correlation with clinical features and carbohydrate antigen 19-9. Pancreas 2009; 38: 534–541. PubMed

Chen H, Tu H, Meng ZQ. et al. K-ras mutational status predicts poor prognosis in unresectable pancreatic cancer. Eur J Surg Oncol 2010; 36: 657–662. PubMed

Castells A, Puig P, Mora J. et al. K-ras mutations in DNA extracted from the plasma of patients with pancreatic carcinoma: diagnostic utility and prognostic significance. J Clin Oncol 1999; 17: 578–584. PubMed

Diaz LA Jr, Bardelli A.. Liquid biopsies: genotyping circulating tumor DNA. J Clin Oncol 2014; 32: 579–586. PubMed PMC

Bettegowda C, Sausen M, Leary RJ. et al. Detection of circulating tumor DNA in early- and late-stage human malignancies. Sci Transl Med 2014; 6: 224ra224. PubMed PMC

Lu L, Risch HA.. Exosomes: potential for early detection in pancreatic cancer. Future Oncol 2016; 12: 1081–1090. PubMed

Melo SA, Luecke LB, Kahlert C. et al. Glypican-1 identifies cancer exosomes and detects early pancreatic cancer. Nature 2015; 523: 177–182. PubMed PMC

San Lucas FA, Allenson K, Bernard V. et al. Minimally invasive genomic and transcriptomic profiling of visceral cancers by next-generation sequencing of circulating exosomes. Ann Oncol 2016; 27: 635–641. PubMed PMC

Sluijter JP, Verhage V, Deddens JC. et al. Microvesicles and exosomes for intracardiac communication. Cardiovasc Res 2014; 102: 302–311. PubMed

Skog J, Wurdinger T, van Rijn S. et al. Glioblastoma microvesicles transport RNA and proteins that promote tumour growth and provide diagnostic biomarkers. Nat Cell Biol 2008; 10: 1470–1476. PubMed PMC

Demory Beckler M, Higginbotham JN, Franklin JL. et al. Proteomic analysis of exosomes from mutant KRAS colon cancer cells identifies intercellular transfer of mutant KRAS. Mol Cell Proteomics 2013; 12: 343–355. PubMed PMC

Dictionaries NCI. U.S. Department of Health and Human Services. National Institutes of Health. National Cancer Institute 2015.

Kahlert C, Melo SA, Protopopov A. et al. Identification of double-stranded genomic DNA spanning all chromosomes with mutated KRAS and p53 DNA in the serum exosomes of patients with pancreatic cancer. J Biol Chem 2014; 289: 3869–3875. PubMed PMC

Jahr S, Hentze H, Englisch S. et al. DNA fragments in the blood plasma of cancer patients: quantitations and evidence for their origin from apoptotic and necrotic cells. Cancer Res 2001; 61: 1659–1665. PubMed

Diamandis EP. A word of caution on new and revolutionary diagnostic tests. Cancer Cell 2016; 29: 141–142. PubMed

Krimmel JD, Schmitt MW, Harrell MI. et al. Ultra-deep sequencing detects ovarian cancer cells in peritoneal fluid and reveals somatic TP53 mutations in noncancerous tissues. Proc Natl Acad Sci USA 2016; 113: 6005–6010. PubMed PMC

Poruk KE, Gay DZ, Brown K. et al. The clinical utility of CA 19-9 in pancreatic adenocarcinoma: diagnostic and prognostic updates. Curr Mol Med 2013; 13: 340–351. PubMed PMC

Katz MH, Pisters PW, Evans DB. et al. Borderline resectable pancreatic cancer: the importance of this emerging stage of disease. J Am Coll Surg 2008; 206: 833–846. Discussion 846–838. PubMed PMC

Extracellular vesicles in cancer´s communication: messages we can read and how to answer