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  Project Overview


Chemically Induced Pluripotent Stem Cells (CiPSCTM) .

Ongoing studies in our laboratories, initiated in 2007, have developed a robust iPS methodology using a cocktail of small molecule inducers, termed “Chemically induced Pluripotent Stem Cells” (CiPSC, patent pending) eliminating the need for any exogenous gene transduction. The CiPSC cocktail of chemical compounds was identified using cell based high throughput screens of biomaterial microarrays (www.dnamicroarray.com/r&d_projects_cma.html). CiPSC have been generated from Human Dermal Fibroblasts (HDF). We are currently working on derivation of CiPSC from other human cell types. The CiPSC methodology has multiple practical advantages:

1) CiPSC does not interfere with the natural genome of the resulting cells as drugs are washed off after induction.

2) CiPSC derivation is practical and cost effective.

3) CiPSC lines are safe and most translatable into clinical settings.

4) CiPSC Generates fully reprogrammed cells with high level of efficiency. Analysis of expression of pluripotency markers, methylation at the OCT4 and NANOG promoters, and differentiation into teratomas (Ectoderm, Mesoderm, and Endoderm), has shown that CiPSCs are true iPSCs (fully reprogrammed), as opposed to reprogramming intermediates using the criteria of Chan et al.

5) CiPSCs are stable, have normal karyotype, and maintain their iPS characteristics in more than 20 passages post derivation.

In comparison with other iPS methodologies, CiPSC offers high efficiency, low cost, low “hands-on” time, high genomic/ epigenomic integrity, high clinical feasibility, and is very amenable to kit development:

  Sendai Virus Lenti/ Retro Adeno

Episomal/ Minicircle

Protein Modified RNA Partial Chemical CiPSC
Efficiency 0.01-1% 0.001-0.01% 0.0001% 0.0001% 0.00001% 0.01-0.1% 0.01-0.1% >0.1%
Genomic Integration No Yes No No No No Yes No
Multiple Transfections No No No No/Yes Yes Yes No No
Overal Assay Cost Medium High High Medium/High High High Medium Low
Kit Possibility Yes Yes Yes Yes Low Possib. Low Possib. High Possib. High Possib.
"Hands On" Time Low Low Low Low/Medium High High Low/Medium Low
Clinical Feasibility Medium Low Medium Medium Medium Medium Low High
Most Prone to Optimization Low Low Low Low Low Low Low High
Genomic/ Epigenomic Integrity Medium Low Medium Low Medium Medium Low High

CiPSC procedure is a breakthrough technology for developing reliable and host-compatible pluripotent cells that offer practical “paradigm shifting” opportunities for studying human disease models in a dish, creating a personalized renewable source of cells for patient-specific cell therapies and enabling various regenerative medicine applications without immune rejection.

For updates on progress please refer to our R&D Projects page. For collaborations email us at TAP@dnamicroarray.com.

Click here for “5 Things to Know Before Jumping on the iPS Bandwagon” (Cyranoski D., 2008, Nature, 452:406-408). Click here for “Growing Pains of Pluripotency” (Check Hayden E., 2011, Nature, 473:272-274). Click here for “Reprogramming: faithful reporters” (Baker, M., 2012, Nature Methods, 9:231-234).

CiPSC cocktail of inducers rescue the phenotype of control genetically (Lentiviral “Yamanaka factors”) induced iPS cells in a 32 days CiPSC reprogramming protocol. One of our initial findings showed that less than all of the transduced genes were necessary for induction of pluripotency in HDF cells and in fact a select combination of small molecules could be used to induce the pluripotent state in the absence of ectopic gene expression (columns I, J, K, L, R, S, T, V, X vs. column A lentiviral control). Note change in the efficiency of reprogramming (# of CiPSC colonies obtained per 100K host cells after 32 days of derivation, y axis on the graph). Through multiple rounds of screening and optimizations we observed that different cocktails of small molecule combinations increase efficiency of CiPSC induction to values approaching 0.1%. The preliminary cocktail (CS4356, Column J above) has an efficiency of 0.005% similar to Lentiviral induced iPS (Column A). Modifications to this cocktail resulted in CS56754 with 0.04%, CS58876 (same as column X above) with 0.05%, and CS11543 with greater than 0.1% efficiencies (not shown in figure). Data presented as number of CiPSC/ iPSC per 100K HDF cells used at the beginning of the reprogramming experiment

References

  1. Takahashi, K. et. al, (2007), Cell 131:1-12.
  2. Esmaeli-Azad B. MICROENVIRONMENT NICHE ASSAY FOR CIPS SCREENING is subject to world-wide protection under PCT International Patent Application No. PCT/US09/67950 and in the U.S. under Patent Application Serial number 12/637,738; 2008.
  3. Junying Yu et al. (2007) Science 318: 1917-1920.
  4. Meissner, A et al. (2007) Nature Biotechnology 25 (10): 1177-1181.
  5. Chan E.M., et al. (2009) Nature Biotechnology 27:1033-1037.

 
 
 
 

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