title: Functional characterization of circulating tumor cells with a prostate-cancer-specific microfluidic device.
abstract: Cancer metastasis accounts for the majority of cancer-related deaths owing to poor response to anticancer therapies. Molecular understanding of metastasis-associated drug resistance remains elusive due to the scarcity of available tumor tissue. Isolation of circulating tumor cells (CTCs) from the peripheral blood of patients has emerged as a valid alternative source of tumor tissue that can be subjected to molecular characterization. However, issues with low purity and sensitivity have impeded adoption to clinical practice. Here we report a novel method to capture and molecularly characterize CTCs isolated from castrate-resistant prostate cancer patients (CRPC) receiving taxane chemotherapy. We have developed a geometrically enhanced differential immunocapture (GEDI) microfluidic device that combines an anti-prostate specific membrane antigen (PSMA) antibody with a 3D geometry that captures CTCs while minimizing nonspecific leukocyte adhesion. Enumeration of GEDI-captured CTCs (defined as intact, nucleated PSMA+/CD45- cells) revealed a median of 54 cells per ml identified in CRPC patients versus 3 in healthy donors. Direct comparison with the commercially available CellSearch® revealed a 2-400 fold higher sensitivity achieved with the GEDI device. Confocal microscopy of patient-derived GEDI-captured CTCs identified the TMPRSS2:ERG fusion protein, while sequencing identified specific androgen receptor point mutation (T868A) in blood samples spiked with only 50 PC C4-2 cells. On-chip treatment of patient-derived CTCs with docetaxel and paclitaxel allowed monitoring of drug-target engagement by means of microtubule bundling. CTCs isolated from docetaxel-resistant CRPC patients did not show any evidence of drug activity. These measurements constitute the first functional assays of drug-target engagement in living circulating tumor cells and therefore have the potential to enable longitudinal monitoring of target response and inform the development of new anticancer agents.
authors.affiliation: Sibley School of Mechanical and Aerospace Engineering, Cornell University, Ithaca, New York, USA. bk88@cornell.edu
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Kirby
authors.forename: Mona
authors.initials: M
authors.lastname: Jodari
authors.forename: Matthew S
authors.initials: MS
authors.lastname: Loftus
authors.forename: Gunjan
authors.initials: G
authors.lastname: Gakhar
authors.forename: Erica D
authors.initials: ED
authors.lastname: Pratt
authors.forename: Chantal
authors.initials: C
authors.lastname: Chanel-Vos
authors.forename: Jason P
authors.initials: JP
authors.lastname: Gleghorn
authors.forename: Steven M
authors.initials: SM
authors.lastname: Santana
authors.forename: He
authors.initials: H
authors.lastname: Liu
authors.forename: James P
authors.initials: JP
authors.lastname: Smith
authors.forename: Vicente N
authors.initials: VN
authors.lastname: Navarro
authors.forename: Scott T
authors.initials: ST
authors.lastname: Tagawa
authors.forename: Neil H
authors.initials: NH
authors.lastname: Bander
authors.forename: David M
authors.initials: DM
authors.lastname: Nanus
authors.forename: Paraskevi
authors.initials: P
authors.lastname: Giannakakou
mesh_terms: D001483:Base Sequence; D001681:Biological Assay; D045744:Cell Line, Tumor; D003198:Computer Simulation; D004867:Equipment Design; D006801:Humans; D008297:Male; D046210:Microfluidic Analytical Techniques / Q000379:methods*; D057054:Molecular Imaging; D008969:Molecular Sequence Data; D009362:Neoplasm Metastasis; D009360:Neoplastic Cells, Circulating / Q000187:drug effects / Q000378:metabolism / Q000473:pathology*; D015514:Oncogene Proteins, Fusion / Q000378:metabolism; D009928:Organ Specificity / Q000187:drug effects; D017354:Point Mutation / Q000235:genetics; D011471:Prostatic Neoplasms / Q000378:metabolism / Q000473:pathology*; D011485:Protein Binding / Q000187:drug effects; D011944:Receptors, Androgen / Q000235:genetics; D043823:Taxoids / Q000494:pharmacology; D014404:Tubulin / Q000378:metabolism
chemical_list: C410798:AR protein, human; D015514:Oncogene Proteins, Fusion; D011944:Receptors, Androgen; C515553:TMPRSS2-ERG fusion protein, human; D043823:Taxoids; D014404:Tubulin