Generation of GMP-grade human induced pluripotent stem cells

Generation of GMP-grade human induced pluripotent stem cells
seyedeh shima mohammadi,¹ zohreh saltanat pour,² amir ali hamidieh,^3,*
1. 1/ Stem Cell and Regenerative Medicine Center of Excellence, Tehran University of Medical Sciences, Iran 2/Department of Stem Cells and Regenerative Medicine, Institute of Medical Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran
2. 1)Pediatric cell therapy research center 2)Stem Cell and Regenerative Medicine Center of Excellence, Tehran University of Medical Sciences, Iran
3. 1)Pediatric cell therapy research center 2)Stem Cell and Regenerative Medicine Center of Excellence, Tehran University of Medical Sciences, Iran
Introduction: Stem cells (SCs) classified into two essential groups based on their plasticity and sources. Adult stem cells and embryonic stem cells (ESCs) (Inner cell mass of mammalian blastocysts) are two primary sources that naturally existing in body of human. Moreover, in 2006 the third SC sources announced by Yamanaka and Takashi via genetic manipulation. Induced pluripotent stem (iPS) is an artificial stem cell that has been discovered by overexpression of three or four factors (Oct4, Klf4, Sox2, and [c-Myc or l-Myc]) that highly expressed in ESC or Thomson 4 factors (Sox2, Oct3/4, Nanog, and Ln28) and reprogrammed mouse and human somatic cells. However, iPS cells have essential characteristics of ESCs such as normal karyotypes, telomerase activity, maintain the ability to differentiate into all three germ layers, moreover they have same genes and cell surface markers. After that, some groups had investigated the procedure of Yamanaka with other mouse tissues for verification and validation. Even, human differentiated cells could have successfully reprogrammed toward iPS cells. But the most important reason for its expanded studies and usage is personalized medicine means generating iPS cells from the patients' own cells that remove immune rejection by recipient and ethical difficulties regarding the use of the human embryo. This unlimited source of cells has been exploiting for many disease modeling, toxicology assays, drug screening and discovery in the comprehensive field of regenerative medicine. Presenting proto-oncogenes (C-MYC and KLF4) into the Yamanaka method and better understanding of molecular mechanism of pluripotency have been persuading scientists to consider different strategies that have more safety and efficiency. Although, c-Myc has been identified as a dispensable factor for reprogramming, absence in this procedure intensive affects efficiency. Retro and lentiviruses are the first vehicles that have been used in reprogramming, but because of oncogenic activation by reason of integration into host genome, subsequently low safety, their clinical application have been abolished, however have still the highest reprogramming efficiency. Recently, In order to safe, clinical-grade and GMP (good manufacturing practice ) manufacturing, integration-free and non-viral methods have been preferring for generation of iPS, such as using episomal plasmid vector, synthesized mRNA, microRNA, Sendai virus, recombinant proteins, small molecules. Also, several studies have accomplished in investigating a safe and effective method in clinical application. But these strategies need to be continued for further scrutiny.
Methods: Reprogramming factors such as DNA, mRNA, miRNA, protein, and small molecules have been delivering by integration or non-integration methods. Integration methods are including Retro and lentiviruses, that have the possibility of tumorigenicity. Various types of non-integration methods classified into two groups based on virus usage: 1/viral approaches (adenovirus, sendai virus (SeV)), 2/Non-viral approaches (plasmid transfection, piggy Bac transposon, minicircle vector, episomal plasmid, modified mRNA, self-replication RNA (srRNA), micro RNAs, xeno-free condition, recombinant proteins, and small molecules. Also, chemical and physical methods have established for reprogramming process.
Results: The integration method that at first step had used for the generation of iPS cells is not a safety method, although high efficiency. Thus, this method is currently used for drug discovery and disease modeling. Scientists are leading to a new non-integration strategy with high efficiency and safety for clinical application.
Conclusion: Biophysical and biochemical factors are prompting for cell reprogramming. For example, extracellular matrix forces variation, 3D microenvironment, nanoparticles, and 3D scaffold coated with growth factors and small molecules. Studies are continued and need to consider in GMP grade iPS generation.
Keywords: iPS, cell reprogramming, nanotechnology, biomaterial