title: ROCK inhibitor and feeder cells induce the conditional reprogramming of epithelial cells.
abstract: We demonstrate that a Rho kinase inhibitor (Y-27632), in combination with fibroblast feeder cells, induces normal and tumor epithelial cells from many tissues to proliferate indefinitely in vitro, without transduction of exogenous viral or cellular genes. Primary prostate and mammary cells, for example, are reprogrammed toward a basaloid, stem-like phenotype and form well-organized prostaspheres and mammospheres in Matrigel. However, in contrast to the selection of rare stem-like cells, the described growth conditions can generate 2 × 10(6) cells in 5 to 6 days from needle biopsies, and can generate cultures from cryopreserved tissue and from fewer than four viable cells. Continued cell proliferation is dependent on both feeder cells and Y-27632, and the conditionally reprogrammed cells (CRCs) retain a normal karyotype and remain nontumorigenic. This technique also efficiently establishes cell cultures from human and rodent tumors. For example, CRCs established from human prostate adenocarcinoma displayed instability of chromosome 13, proliferated abnormally in Matrigel, and formed tumors in mice with severe combined immunodeficiency. The ability to rapidly generate many tumor cells from small biopsy specimens and frozen tissue provides significant opportunities for cell-based diagnostics and therapeutics (including chemosensitivity testing) and greatly expands the value of biobanking. In addition, the CRC method allows for the genetic manipulation of epithelial cells ex vivo and their subsequent evaluation in vivo in the same host.
authors.affiliation: Department of Pathology, Lombardi Comprehensive Cancer Center, Georgetown University Medical School, Washington, DC 20057, USA.
authors.forename: Xuefeng
authors.initials: X
authors.lastname: Liu
authors.forename: Virginie
authors.initials: V
authors.lastname: Ory
authors.forename: Sandra
authors.initials: S
authors.lastname: Chapman
authors.forename: Hang
authors.initials: H
authors.lastname: Yuan
authors.forename: Chris
authors.initials: C
authors.lastname: Albanese
authors.forename: Bhaskar
authors.initials: B
authors.lastname: Kallakury
authors.forename: Olga A
authors.initials: OA
authors.lastname: Timofeeva
authors.forename: Caitlin
authors.initials: C
authors.lastname: Nealon
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Dakic
authors.forename: Vera
authors.initials: V
authors.lastname: Simic
authors.forename: Bassem R
authors.initials: BR
authors.lastname: Haddad
authors.forename: Johng S
authors.initials: JS
authors.lastname: Rhim
authors.forename: Anatoly
authors.initials: A
authors.lastname: Dritschilo
authors.forename: Anna
authors.initials: A
authors.lastname: Riegel
authors.forename: Alison
authors.initials: A
authors.lastname: McBride
authors.forename: Richard
authors.initials: R
authors.lastname: Schlegel
mesh_terms: D000577:Amides / Q000494:pharmacology*; D000818:Animals; D001940:Breast / Q000166:cytology; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects*; D065150:Cellular Reprogramming / Q000187:drug effects / Q000502:physiology*; D003094:Collagen; D004338:Drug Combinations; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects*; D061252:Feeder Cells / Q000166:cytology / Q000502:physiology*; D005260:Female; D006801:Humans; D007797:Laminin; D008297:Male; D051379:Mice; D016513:Mice, SCID; D009368:Neoplasm Transplantation; D011467:Prostate / Q000166:cytology; D011471:Prostatic Neoplasms / Q000473:pathology; D011509:Proteoglycans; D011725:Pyridines / Q000494:pharmacology*; D014183:Transplantation, Heterologous; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors*
chemical_list: D000577:Amides; D004338:Drug Combinations; D004791:Enzyme Inhibitors; D007797:Laminin; D011509:Proteoglycans; D011725:Pyridines; C059485:matrigel; C108830:Y 27632; D003094:Collagen; D054460:rho-Associated Kinases