Mumbai, Sept 16, 2008: Over the past 20 months, reports claiming the generation of Induced Pluripotent Stem (iPS) cell from somatic cells and having embryonic stem (ES) cell-like characteristics, has captured great attention of scientific community and general public. A lot of enthusiasm was generated particularly in the West  since iPS cell line was proving to be a viable alternative to the controversial Embryonic Stem Cells. This led to the rapid and premature acceptance of iPS cells for regenerative therapy. Evolution of iPS- Induced Pluripotent Stem Cells are Stem Cells established from a mouse and human somatic cells by inducing a “forced” expression of defined transcription factors. Nuclear reprogramming studies were earlier carried out using the SCNT where it was seen that trans-activating factors present in the mammalian oocyte are required to reprogram a somatic cell nuclei to an undifferentiated state. These experiments showed that the developmental state of a nucleus from an adult somatic cell could be reprogrammed upon its transfer into an unfertilized oocyte. Such a strategy generated the first cloned embryo such as Dolly, the world’s first cloned animal. Such experiments provided a definitive proof that pluripotency can be restored to the nucleus of a terminally differentiated cell. Subsequently, cell fusion experiments where adult somatic cells are fused with ES cells have shown that these pluripotent cells also harbor reprogramming activities.

 

An astonishing breakthrough in Stem Cells occurred in 2006 when Japanese Stem Cell researcher Shinya Yamanaka demonstrated that iPS cells could be generated from mouse embryonic or adult fibroblasts by retrovirus-mediated introduction of four factors Oct3/4, Sox2, c-Myc and Klf4 (Figure 1). These iPS cells were similar to ES cells in morphology and teratoma formation. The race then began in establishing a similar result in humans and incidentally the race ended in a tie when Yamanaka’s group from the Kyoto University along with Thomson’s group from the University of Wisconsin-Madison both reported that they reprogrammed a Human Somatic cell into an embryonic stem-like cell. Shi explored alternative strategy combining genetic and chemical approach wherein small molecules were capable of replacing viral integration of certain transcription factors and promote reprogramming process. Use of specific small molecule inhibitors suggests that loss of function of certain genes may be an effective mechanism for generating iPS cells.

 

This study has generated hope that the use of additional small molecules or other non genetic methods could improve reprogramming that could ultimately allow the generation of iPS cells or multipotent tissue-specific stem cells in completely chemically defined conditions without any genetic modification. Shinya Yamanaka announced development in terms of characterization, directed differentiation of iPS cells into neurons and beating cardiomyocytes (Figure 2). Besides mouse/human fibroblasts, some groups have also worked on reprogramming of adult mouse hepatocytes, gastric epithelial cells, bronchial epithelial cells, keratinocytes, adrenal gland cells, immature B-lymphocytes and muscle cells. However, the conditions of reprogramming do differ depending on organ source, and whether cells are from fetal or adult source. iPS cells have high clonogenic potential. They express ES cell-specific antigen profile. These cells have growth potential, gene expression pattern, telomerase activities and epigenetic status similar to ES cells.

 

Applications of iPS cell technology- iPS cells promise several practical applications. Procedures already validated for ES cells will investigate iPS cells' potential to differentiate into functioning, specialized tissues. Working in sickle-cell anaemia, Rudolf Jaenisch and co-workers have already shown, in an elegant proof-of-concept study in the mouse, how reprogramming, tissue-specific differentiation and gene therapy can be used to cure inherited disorders.

 

The potential advantage of using this technology will be for generation of patient-specific cells and evaluation of diagnostic or pharmacological tests in an individualized medicine approach. In contrast to the term 'therapeutic cloning' coined for SCNT-derived applications, this would be 'therapeutic reprogramming'. As with other biomedical research discoveries, the fields that will benefit most from these recent discoveries are that of Tissue engineering and regenerative medicine where customized tissue grafts could be made from a persons’ own adult fibroblastic cell. In addition, the technology may help avoid immune rejection of replacement tissues, because an adult patient's cell could be the source of stem cells that are a genetic match to that individual.

 

Indeed, the potential applications for iPS technology are endless. An overlooked application of iPS cells would be using them to bypass the difficulty of working with species for which establishing ES cells is difficult or impossible. The ability to perform genetic manipulations would help to engineer traits such as disease resistance or greater muscle mass in domestic or threatened animals. Freezing batches of iPS cells from endangered species may also help to preserve them.

 

Ethical concern - The technology sidesteps the ethical objections raised against research in human embryonic stem cells, which are derived from early stage human embryos that are often destroyed in the process. The new technique is not without its own set of limitations, although some of those have already been resolved. One of the original genes used for reprogramming (c-MYC) has been shown to produce tumors and cancers. With current level of knowledge, it would not be a preferred choice for patient therapy.

 

Commercialization: As a step towards development and commercialization, Japan Education, Science & Technology Ministry has established CiRA, Center  for iPS research & Application and allocated approximately 1 billion yen ($ 10 million). CiRA will promote basic research and clinical application of iPS cells in understanding disease mechanisms, drug screening, toxicology and regenerative medicine.

 

Collaborative work between iZumi Foundation and San Francisco based J. David of Gladstone Institutes are undertaking research in iPS cells that can be coaxed to regenerate injured spinal cords or damaged hearts.

 

PrimeGen Biotech from Irvine, CA, USA has claimed to have successfully used nonviral technologies to reprogram adult human cells into stem cells that the Company refers to as intermediate iPS cells.  They employed high efficiency particle delivery system to transport proteins and DNA molecules directly into cells from human skin, retina & kidney. They are the first to use methods that do not involve potentially tumor-causing viruses or genetic manipulations. They report that this method is faster and efficient than other known methods. However, they have not yet disclosed the details which need to be validated by various laboratories.

 

Conclusions- Induction of pluripotency to produce embryonic-like stem cells is the hot field in stem cell research. The fact is that iPS cells have been produced in at least six different laboratories within a few months after the initial animal studies shows that the technique is robust and easily reproducible. In contrast, the competing technique, human SCNT (cloning), has never been transferred from animal studies to human application, despite years of attempts. At this point, it seems pretty certain that the iPS technique will soon replace ES cells as the preferred means of generating human stem cell lines. However, this study is still in infancy of basic science, the practical aspect of a therapy using autologous iPS is not yet clear. The safety and efficacy of these cells need to be worked out before testing in human Clinical trials.