title: Conditional reprogramming and long-term expansion of normal and tumor cells from human biospecimens.
abstract: Historically, it has been difficult to propagate cells in vitro that are derived directly from human tumors or healthy tissue. However, in vitro preclinical models are essential tools for both the study of basic cancer biology and the promotion of translational research, including drug discovery and drug target identification. This protocol describes conditional reprogramming (CR), which involves coculture of irradiated mouse fibroblast feeder cells with normal and tumor human epithelial cells in the presence of a Rho kinase inhibitor (Y-27632). CR cells can be used for various applications, including regenerative medicine, drug sensitivity testing, gene expression profiling and xenograft studies. The method requires a pathologist to differentiate healthy tissue from tumor tissue, and basic tissue culture skills. The protocol can be used with cells derived from both fresh and cryopreserved tissue samples. As approximately 1 million cells can be generated in 7 d, the technique is directly applicable to diagnostic and predictive medicine. Moreover, the epithelial cells can be propagated indefinitely in vitro, yet retain the capacity to become fully differentiated when placed into conditions that mimic their natural environment.
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Xuefeng
authors.identifier: 0000-0002-9922-9627
authors.initials: X
authors.lastname: Liu
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Ewa
authors.initials: E
authors.lastname: Krawczyk
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Frank A
authors.initials: FA
authors.lastname: Suprynowicz
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Nancy
authors.initials: N
authors.lastname: Palechor-Ceron
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Hang
authors.initials: H
authors.lastname: Yuan
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Aleksandra
authors.initials: A
authors.lastname: Dakic
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Vera
authors.initials: V
authors.lastname: Simic
authors.affiliation: Department of Oncology, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Yun-Ling
authors.initials: YL
authors.lastname: Zheng
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Praathibha
authors.initials: P
authors.lastname: Sripadhan
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Chen
authors.initials: C
authors.lastname: Chen
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Jie
authors.initials: J
authors.lastname: Lu
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Tung-Wei
authors.initials: TW
authors.lastname: Hou
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Sujata
authors.initials: S
authors.lastname: Choudhury
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Bhaskar
authors.initials: B
authors.lastname: Kallakury
authors.affiliation: Department of Epigenetics and Molecular Carcinogenesis, University of Texas MD Anderson Cancer Center, Smithville, Texas, USA.
authors.forename: Dean G
authors.initials: DG
authors.lastname: Tang
authors.affiliation: Department of Dermatology, Uniformed Services University of the Health Sciences, Bethesda, Maryland, USA.
authors.forename: Thomas
authors.initials: T
authors.lastname: Darling
authors.affiliation: Department of Dermatology, Uniformed Services University of the Health Sciences, Bethesda, Maryland, USA.
authors.forename: Rajesh
authors.initials: R
authors.lastname: Thangapazham
authors.affiliation: Department of Oncology, Georgetown University Medical Center, Washington, DC, USA.|Department of Radiation Medicine, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Olga
authors.initials: O
authors.lastname: Timofeeva
authors.affiliation: Department of Radiation Medicine, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Anatoly
authors.initials: A
authors.lastname: Dritschilo
authors.affiliation: Department of Cell Biology and Physiology, The University of North Carolina School of Medicine, Chapel Hill, North Carolina, USA.
authors.forename: Scott H
authors.initials: SH
authors.lastname: Randell
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.|Department of Oncology, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Christopher
authors.initials: C
authors.lastname: Albanese
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Seema
authors.initials: S
authors.lastname: Agarwal
authors.affiliation: Department of Pathology, Georgetown University Medical Center, Washington, DC, USA.|Center for Cell Reprogramming, Georgetown University Medical Center, Washington, DC, USA.
authors.forename: Richard
authors.initials: R
authors.lastname: Schlegel
mesh_terms: D000577:Amides / Q000494:pharmacology; D000818:Animals; D002471:Cell Transformation, Neoplastic; D065150:Cellular Reprogramming; D018920:Coculture Techniques / Q000379:methods*; D061252:Feeder Cells / Q000166:cytology / Q000187:drug effects; D005347:Fibroblasts / Q000166:cytology / Q000187:drug effects; D006801:Humans; D051379:Mice; D009369:Neoplasms / Q000473:pathology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D054460:rho-Associated Kinases / Q000037:antagonists & inhibitors
chemical_list: D000577:Amides; D047428:Protein Kinase Inhibitors; D011725:Pyridines; C108830:Y 27632; D054460:rho-Associated Kinases