title: DREAMing: a simple and ultrasensitive method for assessing intratumor epigenetic heterogeneity directly from liquid biopsies.
abstract: Many cancers comprise heterogeneous populations of cells at primary and metastatic sites throughout the body. The presence or emergence of distinct subclones with drug-resistant genetic and epigenetic phenotypes within these populations can greatly complicate therapeutic intervention. Liquid biopsies of peripheral blood from cancer patients have been suggested as an ideal means of sampling intratumor genetic and epigenetic heterogeneity for diagnostics, monitoring and therapeutic guidance. However, current molecular diagnostic and sequencing methods are not well suited to the routine assessment of epigenetic heterogeneity in difficult samples such as liquid biopsies that contain intrinsically low fractional concentrations of circulating tumor DNA (ctDNA) and rare epigenetic subclonal populations. Here we report an alternative approach, deemed DREAMing (Discrimination of Rare EpiAlleles by Melt), which uses semi-limiting dilution and precise melt curve analysis to distinguish and enumerate individual copies of epiallelic species at single-CpG-site resolution in fractions as low as 0.005%, providing facile and inexpensive ultrasensitive assessment of locus-specific epigenetic heterogeneity directly from liquid biopsies. The technique is demonstrated here for the evaluation of epigenetic heterogeneity at p14(ARF) and BRCA1 gene-promoter loci in liquid biopsies obtained from patients in association with non-small cell lung cancer (NSCLC) and myelodysplastic/myeloproliferative neoplasms (MDS/MPN), respectively.
authors: Johns Hopkins Institute for NanoBioTechnology, Baltimore, MD 21218, USA.
authors: Thomas R
authors: TR
authors: Pisanic
authors: Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD 21218, USA.
authors: Pornpat
authors: P
authors: Athamanolap
authors: Cancer Biology Program, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA.
authors: Weijie
authors: W
authors: Poh
authors: Cancer Biology Program, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA Department of Thoracic Surgery, The Second Xiangya Hospital of Central South University, Changsha, Hunan 410011, China.
authors: Chen
authors: C
authors: Chen
authors: Cancer Biology Program, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA.
authors: Alicia
authors: A
authors: Hulbert
authors: Cancer Biology Program, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA.
authors: Malcolm V
authors: MV
authors: Brock
authors: Cancer Biology Program, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA hermanj3@upmc.edu.
authors: James G
authors: JG
authors: Herman
authors: Johns Hopkins Institute for NanoBioTechnology, Baltimore, MD 21218, USA Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD 21218, USA Cancer Biology Program, The Sidney Kimmel Comprehensive Cancer Center at Johns Hopkins, Baltimore, MD 21287, USA Department of Mechanical Engineering, Johns Hopkins University, Baltimore, MD 21218, USA thwang@jhu.edu.
authors: Tza-Huei
authors: TH
authors: Wang
mesh_terms: D000483:Alleles; D001706:Biopsy; D002289:Carcinoma, Non-Small-Cell Lung / Q000235:genetics; D018899:CpG Islands; D019175:DNA Methylation; D017931:DNA Primers; D004273:DNA, Neoplasm / Q000097:blood* / Q000737:chemistry; D003627:Data Interpretation, Statistical; D044127:Epigenesis, Genetic; D057890:Epigenomics / Q000379:methods; D014644:Genetic Variation; D006801:Humans; D008175:Lung Neoplasms / Q000235:genetics; D008297:Male; D009190:Myelodysplastic Syndromes / Q000235:genetics; D009369:Neoplasms / Q000235:genetics* / Q000473:pathology; D009691:Nucleic Acid Denaturation; D017422:Sequence Analysis, DNA; D025803:Tumor Suppressor Protein p14ARF / Q000235:genetics
chemical_list: D017931:DNA Primers; D004273:DNA, Neoplasm; D025803:Tumor Suppressor Protein p14ARF