Detection of mutant KRAS and TP53 DNA in circulating exosomes from healthy individuals and patients with pancreatic cancer

Abstract

Pancreatic cancer presents with a dismal mortality rate and is in urgent need of methods for early detection with potential for timely intervention. All living cells, including cancer cells, generate exosomes. We previously discovered double stranded genomic DNA in exosomes derived from the circulation of pancreatic cancer patients, which enabled the detection of prevalent mutations associated with the disease. Here, we report a proof-of-concept study that demonstrates the potential clinical utility of circulating exosomal DNA for identification of KRAS and TP53 mutations in patients with pancreas-associated pathologies, including pancreatic ductal adenocarcinoma (PDAC), chronic pancreatitis (CP) and intraductal papillary mucinous neoplasm (IPMN), and in healthy human subjects. In 48 clinically annotated serum samples from PDAC patients, digital PCR analyses of exosomal DNA identified KRAS mutation in 39.6% of cases, and TP53 mutation in 4.2% of cases. KRAS and TP53 mutations were also detected in exosomal DNA from IPMN patients (2 out of 7 with KRAS, one of which also co-presented with TP53 mutation). Circulating exosomal DNA in 5 out of 9 CP patients enabled the detection of KRAS mutation. In 114 healthy subject-derived circulating exosomal DNA, 2.6% presented with KRAS mutation and none with TP53 mutation. This study highlights the value of circulating exosomal DNA for a rapid, low-cost identification of cancer driving mutations. The identification of mutations in IPMN patients and healthy subjects suggests that liquid biopsies may allow potential assessment of cancer risk but with a cautionary note that detection of clinical cancer cannot be assumed.

Liquid biopsy - emergence of a new era in personalized cancer care

“…Both KRAS G12D and TP53 R273H mutations have been detected in exosomal DNA from patients with pancreas-associated pathologies, including pancreatic ductal adenocarcinoma, chronic pancreatitis and intraductal papillary mucinous neoplasm compared to healthy human subjects [145]. A combined isolation and analysis of exosomal RNA and cfDNA (together referred to as "exoNA") seems to improve blood-based liquid biopsy for EGFR mutation detection in NSCLC patients.…”

miRNA Profiling of Magnetic Nanopore–Isolated Extracellular Vesicles for the Diagnosis of Pancreatic Cancer

“…Pancreatic cancer is the third leading cause of cancer-related death in the United States with a median overall survival of less than one year and five-year survival of only 7.7% (1,2). At the time of diagnosis, most patients with pancreatic cancer have metastatic disease and are at a disease stage that is no longer curable (3,4).…”

A comparative study of extracellular vesicle-associated and cell-free DNA and RNA for HPV detection in oropharyngeal squamous cell carcinoma

“…Cell-free (cf-DNA) is one of the fastest-growing liquid biopsy biomarkers that has been extensively validated and is currently used routinely in the clinical setting to test for specific tumoral mutations in oncogene driven cancers [7][8][9][10][11] . A number of studies have also demonstrated the emerging role of extracellular vesicles (EV) as another complementary liquid biopsy source [12][13][14][15][16] . EV are bioactive vesicles secreted by all cell types, including vesicles of various sizes and biogenetic origin (e.g.…”

“…exosomes, microvesicles, oncosomes and microparticles) and are implicated in carcinogenesis and in the facilitation of metastatic potential 12 . EV have also been found to contain molecular content reflective of the characteristics of the tumor and thus, EV and their contents have the potential to represent a novel liquid biopsy biomarker [12][13][14][15][16] . As EV are hypothesized to contain more intact molecular contents, EV-based liquid biopsy approaches provide an alternative to cell-free based approaches which rely on the detection of degraded, apoptotic cellular material 17 .…”