Abstract Details

Title Rapid Generation of Induced Pluripotent Stem Cells from Lymphoblastoid Cell Lines Using an Episomal Plasmid Containing Multiple Reprogramming Factors in a Single Cassette

Topic Movement Disorders

Presentation(s) P05 - (-)

Poster/Presentation
Number 046

Objective

Background BACKGROUND: B cells represent a larger fraction of the peripheral blood mononuclear cell population ([sim]20%) and can be transformed in vitro by EBV to generate lymphoblastoid cell lines (LB), creating an unlimited proliferative source of cells for reprogramming trials. LB cells are a precious resource for immunologic, epidemiologic, and rare disease studies. Generating iPSCs from LB cells offers the advantage of working with minimal amounts of blood from living donors as well as frozen LB collections banked worldwide. Recently, iPSCs have been generated by delivering the reprogramming factors (RFs) via oriP/EBNA-1-based plasmids in fibroblasts and peripheral blood CD34+cells.

Design/Methods DESIGN/METHODS: We have generated a novel episomal plasmid where RFs (SOX2, OCT3/4, c-MYC and KLF4) are controlled by individual CMV promoters in a single cassette, designated pAd-GFP-SOcMK, which allows coordinated expression of the four RFs. EBV-B cells were electroporated with plasmid, pAd-GFP-SOcMK and plated on inactivated MEF cells with ES cell medium containing basic fibroblast growth factor (bFGF).

Results RESULTS: After 5-6 days of culture of electroporated EBV-B cells with ES cell medium, cells tend to attach and start to form colonies. EBV-B cell line-derived iPSCs showed ES cell morphology, stained with alkaline phosphatase (AP), and expressed pluripotent cell-specific genes.

Conclusions CONCLUSIONS: We developed a rapid method to generate iPSCs from EBV-immortalized B-lymphocyte cell lines using a novel 4-factor vector. The ability to reprogram banked patient EBV-transformed cell lines efficiently will offer an unprecedented opportunity to rapidly generate genetic disease models and also provide a translational platform for therapeutic drug development.

Authors/Disclosures
PRESENTER	No disclosure on file
	No disclosure on file
Karla P. Figueroa (University of Utah)	Ms. Figueroa has nothing to disclose.
Anthony E. Lang, MD, FAAN (Toronto Western Hospital)	Dr. Lang has received personal compensation in the range of $5,000-$9,999 for serving as a Consultant for AbbVie, Amylyx, Aprinoia, Biogen, BioAdvance, Biohaven, BioVie, BlueRock, BMS, Denali, Janssen, Lilly, Pharma 2B, Sun Pharma, and UCB. Dr. Lang has received personal compensation in the range of $50,000-$99,999 for serving as an Expert Witness for medicolegal cases related to paraquat. The institution of Dr. Lang has received research support from AbbVie. Dr. Lang has received intellectual property interests from a discovery or technology relating to health care. Dr. Lang has received publishing royalties from a publication relating to health care.
Stefan M. Pulst, MD, FAAN (University of Utah)	Dr. Pulst has received personal compensation in the range of $500-$4,999 for serving as a Consultant for venrock. Dr. Pulst has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Arrowhead. Dr. Pulst has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Leverna. Dr. Pulst has received personal compensation in the range of $10,000-$49,999 for serving as an Editor, Associate Editor, or Editorial Advisory Board Member for AAN. Dr. Pulst has received personal compensation in the range of $500-$4,999 for serving as an Expert Witness for Leninthal LLC. The institution of Dr. Pulst has received research support from NINDS. Dr. Pulst has received intellectual property interests from a discovery or technology relating to health care.

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