iCell Cardiomyocytes

Induced Pluripotent Stem (iPS) Cells

How does CDI's technology work?

  1. A human biological sample, for example blood or skin, is obtained, and the cells within the sample are grown under appropriate cell culture conditions.
  2. In the episomal reprogramming method, vectors containing multiple reprogramming genes are introduced into the cells.

    While the vectors turn genes in the cell on and off, reprogramming them to a stem cell state, they do not integrate into the genome itself. This method alleviates concerns arising over the potential risks associated with the insertion of foreign DNA to induce reprogramming, which other prior iPS methods use (bottom row in illustration above).
  3. After approximately four weeks pluripotent stem cells can be observed, isolated, and expanded under cell culture conditions. The ability to successfully create and grow iPS cells requires significant stem cell expertise. The iPS cell colonies are very fragile and sensitive to manipulation.
  4. CDI uses patented, proprietary methods to direct the cell differentiation process into specific cell lineages, including cardiomyocytes, neurons, hepatocytes, and more.

CDI’s Technology

What are induced pluripotent stem (iPS) cells?

iPS cells are somatic cells (e.g., skin or blood) that have been genetically reprogrammed to a pluripotent stem cell state through forced expression of pluripotency genes. By definition, iPS cells replicate indefinitely and have the potential to differentiate into any cell type in the human body.

What are reprogramming factors?

Reprogramming factors are the genes introduced into somatic cells that induce a pluripotent stem cell state.  Initial reports describing the creation of human iPS cells utilized four reprogramming factors: OCT4, SOX2, KLF4 and MYC (OSKM) (Takahashi, et al. 2007) or OCT4, SOX2, NANOG and LIN28 (OSNL) (Yu, et al. 2007).  Subsequent studies revealed that reprogramming using a specific combination of all 6 of these factors combined with SV40LT and a cocktail of small molecules yields iPS cells at much higher efficiency (Yu, et al. 2009; Yu, et al. 2011). 

What is episomal reprogramming?

iPS cells are genetically reprogrammed through forced expression of pluripotency genes into somatic cells. The expression of these genes can be accomplished using a variety of different methods. The episomal reprogramming method introduces pluripotency genes into a target cell using circular DNA plasmid vectors (i.e. episomes) that replicate autonomously within the cell cytoplasm and do not integrate into the host cell genome.

What are the benefits of episomal reprogramming?

Initial methods of iPS cell reprogramming utilized retroviral and lentiviral vectors to introduce pluripotency genes into somatic cells.  While these methods generally work well, the viral DNA integrates into the genome of the target cell, and the resulting iPS cells (and cells differentiated from them) will contain foreign DNA, which may result in defects and errors. 
By contrast, episomal vectors replicate autonomously within the cell cytoplasm and do not integrate into the host genome.  In addition, the episomal vectors are released from the target cell at a rate of ~5% per cell cycle resulting in transgene-free or “footprint-free” iPS cells. These features, combined with recent advancements in episomal reprogramming efficiency, have led to a strong preference for this method to alleviate concerns about genome integrity for drug discovery and cell therapy applications.   

What cell types have been successfully reprogrammed using the episomal method?

Episomal reprogramming has been reported successful from a variety of somatic cells, including fibroblasts, lymphoblastoid cells, and peripheral blood mononuclear cells.  Importantly, CDI has optimized its episomal reprogramming method to achieve high efficiency iPS cell generation from small amounts of human peripheral blood.  Not only does this enable more streamlined and less invasive collection of donor samples, but ensures increased sterility and lower cost production of iPS cells.  In addition, efficient iPS cell production from peripheral blood enables access to large banks of normal and disease-associated clinical samples for disease research and drug screening. 

How can I access CDI’s episomal reprogramming technology?

CDI’s suite of MyCell Products includes episomal reprogramming of customer-provided donor samples and subsequent genetic engineering and/or differentiation of the iPS cells. 
In addition, for researchers who would like to generate their own iPS cells, CDI’s episomal reprogramming technology is available as a kit from Life Technologies, including Episomal iPSC Reprogramming Vectors, Vitronectin, and Essential 8™ Medium.  Customer-generated iPS cells using this kit may then be transferred to CDI for genetic engineering and/or differentiation through MyCell Products

How does episomal reprogramming compare to other integration-free or
“footprint-free” methods?

Integration-free iPS cells have been generated using a variety of methods including adenovirus, Sendai virus, piggyBac, minicircle vectors, and direct introduction of protein or synthesized mRNA.  The efficiency and success rate of these methods varies depending on the source of somatic cells and experimental conditions, but in general these approaches are limited by impractically low reprogramming efficiency, requirement for higher biosafety containment, and/or labor- and cost-intensive protocols that require repeated transfection/infection. Compared to these methods, episomal reprogramming is virus-free, safe to use, stable, and inexpensive.

Can iPS cells be generated using small molecules?

A variety of small molecules have been identified that can functionally substitute for one or more reprogramming factors and/or improve the efficiency of iPS cell reprogramming.  However, no combination of small molecules has been shown to functionally substitute for all four reprogramming factors. 
The use of small molecules in iPS cell reprogramming offers some practical advantages including the ability to optimize the chemical structure, fine-tune dose and concentration, and simplify handling and application protocols. 
However, the use of small molecules presents a number of scientific challenges.  Most notably, small molecules may have more than one target, which may or may not be known.  In addition, unexpected toxicity and other side effects in vivo may interfere with the clinical application of small molecules. 

Intellectual Property Questions

What does the CDI episomal reprogramming patent cover?

The issued CDI patent No. 8,546,140 covers all reprogramming by vectors that include the genetic elements EBNA-1 and OriP, which are the elements that enable the episomal vectors to replicate when cells divide.  All known episomal reprogramming methods use these elements.

Does the CDI episomal reprogramming patent also cover the iPS cells made by this method?

Yes.  The US patent statute provides at 35 USC 271g that whoever “offers to sell, sells or uses within the United States a product which is made by a process patented in the United States shall be liable as an infringer” if the activity occurs during term of the patent on the process.  All commercial use of iPS cell lines made by episomal reprogramming is thus covered by the No. 8,546,140 patent. 

What is the term of the CDI episomal reprogramming patent?

The No. 8,546,140 patent will have a term extending until at least May 2030.

Who needs a license?

Parties interested in licensing this episomal reprogramming technology should contact CDI at

Is this the only patent on episomal reprogramming?

No. Already issued is US Patent No. 8,268,260 to Drs. James Thomson and Junying Yu. This patent is owned by Wisconsin Alumni Research Foundation (WARF) and exclusively licensed to CDI. Also, episomal reprogramming requires the use of reprogramming factors, some of which may be licensed from CDI and others of which may be licensed from iPS Academia Japan, Kyoto, Japan.

Forward-looking Statements
To the extent that statements contained in these FAQs are not descriptions of historical facts regarding Cellular Dynamics International, Inc., including the impact of the issuance of U.S. Patent No. 8,546,140 and episomal reprogramming, they are forward-looking statements reflecting the current beliefs and expectations of management made pursuant to the safe harbor provisions of the Private Securities Litigation Reform Act of 1995. Words such as "may," "will," “believe,” "expect," "anticipate," "estimate," "intend," and similar expressions (as well as other words or expressions referencing future events, conditions or circumstances) are intended to identify forward-looking statements.  Forward-looking statements in this release involve substantial risks and uncertainties that could cause our product development efforts, actual results, performance or achievements to differ materially from those expressed or implied by the forward-looking statements. Cellular Dynamics undertakes no obligation to update or revise any forward-looking statements. For a further description of the risks and uncertainties that could cause actual results to differ from those expressed in these forward-looking statements, as well as risks relating to the business of the Company in general, see Cellular Dynamic’s quarterly report on Form 10-Q filed with the Securities and Exchange Commission on August 29, 2013.