New Allele Biotech Publication on Stem Cells

Feeder-Free Reprogramming of Human Fibroblasts with Messenger RNA
Current Protocols in Stem Cell Biology • November 13, 2013
DOI: 10.1002/9780470151808.sc04a06s27

Authors: Luigi Warren, Jiwu Wang

This unit describes a feeder-free protocol for deriving induced pluripotent stem cells (iPSCs) from human fibroblasts by transfection of synthetic mRNA. The reprogramming of somatic cells requires transient expression of a set of transcription factors that collectively activate an endogenous gene regulatory network specifying the pluripotent phenotype. The necessary ectopic factor expression was first effected using retroviruses; however, as viral integration into the genome is problematic for cell therapy applications, the use of footprint-free vectors such as mRNA is increasingly preferred. Strong points of the mRNA approach include high efficiency, rapid kinetics, and obviation of a clean-up phase to purge the vector. Still, the method is relatively laborious and has, up to now, involved the use of feeder cells, which brings drawbacks including poor applicability to clinically oriented iPSC derivation. Using the methods described here, mRNA reprogramming can be performed without feeders at much-reduced labor and material costs relative to established protocols.

Allele iPSC Service and Technology Licensing Contact: http://www.allelebiotech.com/cell-line-and-culture-services/#ips-line

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Tags: feeder-free, footprint free, high efficiency reprogramming, iPSC, iPSCs, Jiwu Wang, Luigi Warren, mRNA iPSC, mRNA reprogrammin, pluripotent, stem cell protocol, stem cells, xeno-free

Allele Biotech Receives $200,000 Grant to Update Its mRNA Reprogramming Commercial Products and Services

On June 10, 2013 Allele received an SBIR award from the National Institute of Drug Abuse (NIDA/NIH) entitled “Revolutionary Technology for Efficient Derivation of Human iPSCs with Messenger RNA”. The goal of the proposed project is to provide to the biomedical research market an advanced reagent kit and services for highly efficient reprogramming of high quality human induced pluripotent stem cells (iPSCs). At the core of this kit is the Allele team’s recent development transcribed messenger RNA (mRNA). Compared to other reprogramming methods, such as lentivirus, Sendai virus, protein, small molecules or any combinations of these reagents, our new generation of the mRNA method often requires less than half the time while sometimes achieving “bulk conversion” efficiency.

While the Allele reprogramming technology was designed for clinical use as the process is feeder-free, xeno-free, chromosome integration-free, as well as without the need for cell splitting, PI, Dr. Jiwu Wang states, “Our purpose of executing the NIH-funded research it to make our method so easy that any researcher can integrate iPSC into his or her projects.” In addition to the extremely high efficiency, mRNA-generated iPSCs should also be more stable because there are no genetic alterations, more uniform among all clones as there is no clonal event, and ultimately suitable for future autologous cell therapy now that creating iPSCs from patient tissue cells should no longer be the rate-limiting steps.

Allele’s business model is to provide cGMP-grade iPSCs to pharmaceutical companies and perform large scale reprogramming by partnering first with university-affiliated hospitals. Great progress has been made in both directions, which has prompted the initiation of a cGMP unit within Allele’s newly acquired building in San Diego.

Tags: cell therapy, feeder-free, footprint free, iPSC, mRNA iPSC, pluripotent, stem cells, tissue regeneration, xeno-free

Reprogramming Life

President Obama is expected to lift the ban on federal fund for embryonic stem cell research soon. However, that does not seem to be the hottest topic these days concerning stem cell research. In 2006, Shinya Yamanaka showed that mouse skin cells could be reprogrammed back into something called induced pluripotent stem (iPS) cells by introducing a handful of cDNAs using retroviral vectors. The process was later repeated in human cells and by other groups including those of Thomson and Melton, sometimes with a slightly different set of inducing cDNAs, or with chemicals or shRNA repressing the repressors of the inducer genes.

The iPS cells are not exactly the same as ES cells, and no animals have been created using iPS cells, but they are close enough to be of great interest to lots of people, particularly for basic research purposes. The method to create iPS by reversing chromosomal changes along differentiation pathways appears to be surprisingly simple, like erasing an old audio tape, there may still be acoustic information left if analyzed by the right equipment, but to most people it is as clean as new. You’d wish a few things in life could be reversed that easily!

For labs that are not already in the stem cell field but feel a need to get their feet wet, then they want reagents that are pre-assembled and pre-tested. Such reagents may include: iPS cultures, iPS inducing viral particles, antibodies to stem cell specific markers, cell assays, and even PCR primer sets (synthesizing hundreds of oligos used in the Yamanaka papers alone will take a lot time and unnecessary costs). That’s where a fast-moving, research-oriented company like Allele comes in. We will bring what we think as starter sets for you, and listen to what you think as needed as we along. The new iPS product line will be launched within weeks, hopefully coinciding with our brand new webpages for all our current product lines!

Tags: c-myc, desease tissue regeneration, differetiation, iPS, Klf4, Lin28, Nanog, Oct3/4, pathways, pluripotent, re-programming, Sox2, stem cells, therapy