EGF enhances HSC regeneration following myelosuppressive injury

Maintenance of the hematopoietic system requires constant replenishment of mature blood cells from HSCs.  For patients with malignant and non-malignant disorders of the blood and immune system, myeloablation and subsequent HSC transplantation is often necessary. describe the imageHowever, exposure to ionizing radiation to induce myeloablation also causes DNA damage that can induce cell-cycle arrest or apoptosis of HSCs and their progenitor cells 1.  Studies have shown that treatment with cytokines can prevent cell-cycle arrest.  For example, administration of stem cell factor (SCF) before radiation exposure protected mice from radiation-induced lethality by inducing HSCs into late S phase 2, which is the most radioresistant phase of the cell cycle.  Further studies to identify additional cytokines that mediate HSC regeneration following radiation exposure are critical for the development of therapies to minimize myelosuppression in patients receiving chemotherapy.

Recently, in Nature Medicine, Doan et al discovered a new function of epidermal growth factor (EGF) signaling in regulation of HSC regeneration following myelosuppressive injury 3.  The authors previously generated a mouse model in which pro-apoptotic proteins, BAK and BAX, were deleted in Tie2+ bone marrow endothelial cells 4.  Mice lacking BAK and BAX expression demonstrated significantly increased numbers of HSCs and progenitor cells and increased survival following total body irradiation (TBI) compared to wild-type mice expressing the pro-apoptotic proteins.  This was the first indication that bone marrow endothelial cells might have therapeutic potential in enhancing hematopoietic reconstitution following myelosuppression.  However, the mechanism through which these cells regulate hematopoietic regeneration was unknown.

In their most recent study, Doan et al performed a cytokine array on bone marrow serum from mice lacking BAK and BAX expression and found a significant enrichment of EGF compared to wild-type mice 3.  Using multiparametric flow cytometry, they demonstrated that ~9% of c-Kit+Sca-1+LinSLAM+ HSCs express functional EGF receptor (EGFR), and expression increased by 6-fold following irradiation.  Systemic administration of EGF augmented HSC recovery in vivo and improved the survival of mice following TBI compared to saline-treated control mice.  In contrast, administration of erlotinib, an EGFR antagonist, suppressed HSC regeneration and significantly decreased the survival of mice following TBI, further suggesting that EGFR signaling is critical for radioprotection of bone marrow HSCs and progenitor cells.  They found that EGFR signaling promotes HSC proliferation by activation of the PI3K-AKT pathway.  In addition, EGF treatment inhibited expression of the p53 upregulated modulator of apoptosis (PUMA), an essential mediator of radiation-induced HSC apoptosis.

describe the imageIn summary, EGF promotes HSC cycling and survival following radiation-induced myelosuppression.  The study by Doan et al was the first demonstration that bone marrow HSCs express functional EGFR, and that EGFR signaling plays a role in HSC self-renewal.  The results of this study suggest that EGF may have therapeutic potential to enhance hematopoietic regeneration in patients receiving myelosuppressive chemotherapy or undergoing HSC transplantation.




1. Liu, Y. et al. p53 regulates hematopoietic stem cell quiescence. Cell Stem Cell 4, 37-48, doi:10.1016/j.stem.2008.11.006 (2009).

2. Zsebo, K. M. et al. Radioprotection of mice by recombinant rat stem cell factor. Proc Natl Acad Sci U S A 89, 9464-9468 (1992).

3. Doan, P. L. et al. Epidermal growth factor regulates hematopoietic regeneration after radiation injury. Nat Med, doi:10.1038/nm.3070 (2013).

4. Doan, P. L. et al. Tie2(+) Bone Marrow Endothelial Cells Regulate Hematopoietic Stem Cell Regeneration Following Radiation Injury. Stem Cells, doi:10.1002/stem.1275 (2012).

A novel role of TPO in the regulation of HSC DNA repair

HSC transplantation is routinely used to treat patients with malignant and non-malignant disorders of the blood and immune system, but its therapeutic application is often restricted by difficulties in in vitro maintenance and expansion of HSCs.  Studies aimed at understanding the mechanisms governing self-renewal of HSCs within hematopoietic tissues identified a set of growth factors and cytokines that can augment in vitro expansion of HSCs, including stem cell factor (SCF), fms-like tryrosine kinase 3 ligand (FLT3l) and thrombopoietin (TPO) 1.  TPO and its receptor, Mpl, are primarily known for their role in megakaryopoiesis, but TPO has also been shown to support HSC quiescence during adult hematopoiesis, with the loss of signaling associated with bone marrow failure and thrombocytopenia 2.

Recently, in Cell Stem Cell, de Lavel et al. identified a novel role of TPO in the regulation of DNA repair in HSCs 3.  Exposure to genotoxic agents, such as ionizing radiation (IR), induces DNA damage comprised of double-strand breaks (DSBs).  DNA damage is repaired through two main pathways: homologous recombination (HR) and nonhomologous end-joining (NHEJ).  DNA repair is essential for cell survival, and studies have shown that NHEJ is necessary for HSC maintenance 4,5.  In their study, de Lavel et al. found that γH2AX foci, a marker of DSB formation, were significantly increased in Mpl-deficient HSCs and in their progenitors following IR exposure.  Moreover, a TPO injection into mice prior to IR reduced the number of γH2AX foci in HSCs in vivo, while HSCs exposed to IR in the absence of TPO demonstrated an increased number of γH2AX foci.  Other experiments showed that TPO modulates the efficiency of the NHEJ pathway by increasing the phosphorylation of the DNA-PK catalytic subunit, a major enzyme involved in NHEJ.  Pharmacological or genetic inhibition of DNA-PK abrogated TPO-mediated DNA repair.  Interestingly, the other cytokines involved in HSC maintenance and expansion, SCF and FLT3l, did not have the same effects as TPO, suggesting that DNA repair activity is a specific function of TPO.

clinical trialIn short, TPO regulates NHEJ-mediated DNA repair of DSBs by stimulating DNA-PK activity in HSCs.  This is the first demonstration that a cytokine involved in HSC maintenance may also regulate DSB repair machinery.  Since TPO treatment prior to IR exposure reduces DNA damage, TPO agonists could potentially be given to patients prior to receiving chemotherapy to reduce the risk of developing oncogenic mutations and defects in HSC function.  Romiplostim, a TPO peptide mimetic, and eltrombopag, a non-peptide TPO mimetic, have been successfully used for the treatment of immune thrombocytopenic purpura (ITP) and are approved by the FDA 6.  de Lavel et al showed that injection of romiplostim prior to IR exposure also completely abolished persistent DNA damage in HSCs, similar to TPO.  Thus, these TPO agonists might also be suited for clinical applications involving protection of normal HSC from DNA-damaging agents.



1          Ohmizono, Y. et al. Thrombopoietin augments ex vivo expansion of human cord blood-derived hematopoietic progenitors in combination with stem cell factor and flt3 ligand. Leukemia 11, 524-530 (1997).

2          Ballmaier, M., Germeshausen, M., Krukemeier, S. & Welte, K. Thrombopoietin is essential for the maintenance of normal hematopoiesis in humans: development of aplastic anemia in patients with congenital amegakaryocytic thrombocytopenia. Ann N Y Acad Sci 996, 17-25 (2003).

3          de Laval, B. et al. Thrombopoietin-Increased DNA-PK-Dependent DNA Repair Limits Hematopoietic Stem and Progenitor Cell Mutagenesis in Response to DNA Damage. Cell Stem Cell 12, 37-48, doi:10.1016/j.stem.2012.10.012 (2013).

4         Rossi, D. J. et al. Deficiencies in DNA damage repair limit the function of haematopoietic stem cells with age. Nature 447, 725-729, doi:10.1038/nature05862 (2007).

5          Nijnik, A. et al. DNA repair is limiting for haematopoietic stem cells during ageing. Nature 447, 686-690, doi:10.1038/nature05875 (2007).

6          Kuter, D. J. New thrombopoietic growth factors. Clin Lymphoma Myeloma 9 Suppl 3, S347-356, doi:10.3816/CLM.2009.s.034 (2009).

R&D 3.0: The Return of the Patient

For years, patients and their caregivers were mostly uninformed outsiders as pharmaceutical companies and research institutes developed new drugs and strategies for future growth.

Now, however, in order to increase brand awareness or accelerate predictive medicine development, more and more pharmaceutical and biotechnology companies realize the importance of patient engagement, starting at the pre-clinical R&D process all the way through clinical trials and the commercialization phase.

The Problem:

Despite all the ongoing efforts to engage patients, Big Pharma ranks near bottom in patient attitudes”. The FiercePharma article from January 18, 2013 briefly describes the outcome of a survey of 500 international, national, and regional patient groups, and their views of the different stages of the drug/device development as well as the paying and distribution process. As expected, high drug pricing was ranked on top of the list as the main cause for negative attitudes towards Big Pharma. The survey also indicates that patients found drug companies “too secretive, doing a poor job of letting patients know about adverse drug event news”.  Consequently, many patients have issues with the industry’s integrity.

describe the imageOverall, I think companies are slowly getting better in bridging the gap between industry and patients. As we all know, change – whether it needs to occur within a company or the mind of a patient – takes time. Gaining patients’ trust will be a long-term task, an infinite circle of strategizing, implementing, observing, and analyzing. Generally speaking, most companies could still need a healthy portion of empathy. If they really care about patients, they would need to develop a real understanding of patients’ or caregivers’ needs, fears, and reasoning of current attitudes.

The good news is: There are several ways to connect with patients on a deeper level.

Depending on a) purpose, b) stage of drug or biomarker development, and c) patient or, eventually, caregiver demographics, one has to develop appropriate recruitment/engagement strategies.

Below, I am outlining some of my thoughts that are based on my own experiences as well as reading many articles and blog posts. It is not a complete list of current status/issues/solutions, and is written to rather start a discussion.

Nevertheless, here are my 2 cents about current developments:


A) Pre-clinical Drug Development

The status quo:

For drug target/validation studies or biomarker development researchers oftentimes needdescribe the image larger quantities of patient samples (often, from hundreds of patients) in order to obtain statistically significant data. Hospital-based biobanks and, eventually, independent biobanks, are organizations that usually have direct access to patients. Although some US-based hospitals have established patient-friendly/engaging websites, those are not used to source samples “on demand”. As a result, it becomes quite time-consuming to source needed samples to start or continue certain research projects.


The Problem:

As Matt Jones, staff reporter for GenomeWeb Daily News, pointed out in his most recent article:

  • Two-thirds of the nation’s biobanks were established over the past decade – it is estimated that there are about 800 biobanks (with 90% embedded within institutions) within the US.
  • The increase of genomics and related large-scale studies have led biobanks to play an increasingly central part in biomedical R&D – genomic research appears as if it is a key driver in the “biobank explosion”.

In short, based on increasing demand, competition is increasing, and given the fact that sequencing costs are rapidly decreasing, one can imagine that this trend will continue. But as biobanks increase and specific patient populations stay the same (especially rare diseases), sample/data sourcing becomes a big issue.

Here is another issue to think about:

Matt Jones also wrote: Several issues that have been flagged by the National Human Genome Research Institute’s Ethical, Legal, and Social Implications Research program are stirred up by the expansion of biobanks, such as questions about policies governing data sharing and security, privacy and the identifiability of genomic information, how and when to return research results and incidental findings, how governance structures function at genomic repositories, and informed consent issues caused by the multiple uses for samples by genome researchers.”

In short, with all the data being generated, certain new regulations might “kick in” within the next few years. Privacy protection always was, and increasingly will be, a topic of many future discussions – especially after the Yaniv Ehrlich’s recent Science article: Identifying Personal Genomes by Surname Inference”. No matter what your attitude (as a researcher) towards these findings is, patients that become aware of these potential “threats” to privacy will increasingly fear breach of “protected” private data and therefore probably be less willing to donate and share their valuable biosamples and medical information. (If interested, further comments can be read in a New York Times article from January 17, 2013: Web Hunt for DNA Sequences Leaves Privacy Compromised”).


Why engage with patients?

describe the imageIt is necessary to connect and build trust. Real trust, however, can only be established through an open, genuine 2-way communication. Since there are already many articles written about this topic, I will not go too far into any details. However, patients/caregivers could potentially be engaged through social media (despite IRB regulations and/or oftentimes long turnaround times) and/or patient portals: for instance, at the biobank Sanguine Biosciences’ patient blog, patients are writing about their experiences, their fears and how they overcame those. At Sanguine, we realize that it is not only important to educate (potential) customers (as Sanguine does too: here is our researcher blog) but even more important to actually work with patients to learn about and educate them – learning, listening and educating needs to go both ways.

Furthermore, increased transparency is another way to develop trust. In the near future, Sanguine Biosciences will notify patients about the impact their samples made by informing them about where they have been sent to and/or for what purpose they were used for.  For instance: “Today, we sent some of your serum and CSF to a pharmaceutical company located on the East Coast. This company is seeking to develop a new biomarker that will help detect Alzheimer’s disease at a very early stage. We thank you for helping us accelerate research for Alzheimer’s!”

Sure, some companies or research institutes will not allow us to communicate any information at all; however, others are very interested in working with us to help them bridge the company-patient-gap.

Additional engagement strategies include partnerships/collaborations with non-profit organizations to provide patients the right education and to learn first-hand about the difficulties patients are dealing with on a daily basis.  Engaged patients are more likely to share their experiences with others.  This leads to a word-of-mouth campaign, which can help increase participation in research studies.

Ultimately, if there is already an existing hospital/biobank-patients-bond, sourcing samples and data will be faster, and less costly, and longitudinal collection studies will become easier.  Recruiting donors for clinical trials (i.e. based on their genetic profile) will become easier too.


B) Clinical Trials Phase

describe the imageOftentimes, pharmas, CROs, clinics/hospitals are faced with relatively high patient recruitment costs (about $2000/patient) and/or lengthy recruitment periods. Having engaged patients already “on-hand” might therefore lead to reduced recruitment cost and time. Besides, and as mentioned above, a 2-way dialogue (as much as possible) will help companies build better reputations.

The Pfizer-way:

As a former Pfizer R&D executive LaMattina suggested, big pharma should consider providing more transparency regarding payments to physicians and data derived from clinical trials. In addition, LaMattina advised “to stop pushing drugs for unapproved uses and give up television advertising”. Despite some strong criticism, several pharmaceutical companies already started following these recommendations.

However, regulations will always be a barrier for an open 2-way dialogue. As Todd Kolm, Director, Emerging Channel Strategy at Pfizer pointed out: “Because of the regulatory environment, we’re not able to engage in a true dialogue”.

Furthermore, what makes patient engagement even more difficult is the relatively poor understanding of new media channels, a lack of FDA guidance, a lack of pharmaceutical executives promoting it, the scarcity of demonstrated success stories, the often inadequate resources allocated to new patient engagement strategies, and the simple fact that it is often not really perceived as a must-have, value-added strategy.

Nonetheless, Pfizer’s REMOTE (Research on Electronic Monitoring of OAB (overactive bladder) Treatment Experience) trial proved that through innovative thinking, courage and implementation of completely new strategies, hurdles can slowly be overcome.

REMOTE, which ended in mid-2012, was the first randomized virtual clinical trial conducted under an IND application.

The main problem, however, was recruiting the right patients needed for the study. This was mainly caused by being very regulatory-cautious so no wrong patients would enter the virtual trial. Ultimately, it became very difficult for patients to actually get into the study.

To quote Craig Lipset, Worldwide Head of Clinical Innovation at Pfizer: “As an industry, we will continue to fail to recruit patients in our studies if we cannot create an ecosystem of patients already engaged and aware about research studies and research participation”.

The good news: Having a web-based recruiting site simplified patient recruitment (including consenting) and allowed patients to participate from their own home – web-based and mobile platforms were used to capture self-reported data from those patients. In addition, primary care physicians engaged with patients by screening and caring for them during the trial. Furthermore, patients received their data at the end of the trial period and were able to even share those with their physician.


C) Post-approval/Commercialization Phase

Pharmaceutical biotechnology companies are slowly becoming more patient-centric. Belgium-based UCB, for example, developed a very patient-focused website, engaging patients through education and disease-specific patient-support programs.

Other companies are also trying to bridge the gap to their patients. Major firms such as Biogen Idec , Pfizer, and French pharmaceutical giant Sanofi, the parent of Genzyme, are partnering on a number of initiatives with patient groups such as The Michael J. Fox Foundation for Parkinson’s Research.


Taken together, it is key to provide top-quality, objective, reliable, unbiased education, training and information to patient organizations and patients at large on all aspects of R&D, in order for patients to get involved and become empowered players in the medical drug development process.

Companies that stay on top of this and develop new strategies will ultimately create the necessary paradigm shift, making sure that patients’ needs and insights are at the center in all relevant areas of medical R&D.


Further Reading:

Big Pharma ranks near bottom in patient attitudes – FiercePharma

US Sees Boom in Diverse Range of Biobanks, But Regulations are Lacking – GEN

Identifying Personal Genomes by Surname Inference – Science

Web Hunt for DNA Sequences Leaves Privacy Compromised – New York Times

Between patients and pharmas online, a disconnect – Medical Marketing & Media

Pfizer Perseveres In Pioneering Virtual Clinical Trials – Life Science Leader

In vitro modeling of hematopoiesis: from pluripotency to blood

Pluripotent stem cells (PSCs) derived from the inner part of a blastocyst (embryonic stem cells, ESCs) or through reprogramming of terminally differentiated adult cells (induced pluripotent stem cells, iPSCs) are capable of self-renewal and differentiation into almost all cell types in the human body. Their differentiation capacities and proliferation potential make pluripotent stem cells a promising source of cells for various clinical applications including regenerative medicine.

fetal red blood cells resized 600Blood is considered to be a connective tissue both functionally and embryologically. It originates from the mesodermal layer, the same germ layer that gives rise to the other connective tissues such as bone, cartilage and muscle. Blood cells and blood vessels develop in parallel and form a functional circulatory system. Various studies have shown that hematopoietic differentiation of PSCs in vitro closely resembles early steps of blood development in the embryo and induces blood forming cell populations with mesodermal and hemato-endothelial properties [1]. Different types of mature blood cells were successfully generated from murine, primate and human pluripotent stem cells. Here, we will briefly review the major in vitro systems of hematopoietic differentiation from PSCs.

Embryoid Body formation

Hematopoietic differentiation of PSCs can be carried out in either a two-dimensional system (2D), where cells are attached to the plate during differentiation, or in a three-dimensional system (3D), where isolated cells are dispersed into a liquid or a semisolid medium to form embryoid bodies (EBs).

Embryoid bodies are spherical structures that are formed by embryonic bodies resized 600pluripotent stem cells grown in non-adherent culture conditions (3D system). Differentiation of PSCs in aggregates mimics three-dimensional embryonic development and yields the establishment of cell adhesion, paracrine signaling and a microenvironment similar to native tissue structures. Thus, EB formation is often used as a method for initiating spontaneous differentiation of PSCs towards all three germ lines.

Differentiation in the presence of growth factors specific for mesoderm (BMP4, FGF, activin A) and blood formation (VEGF, SCF, Flt3, IL-3, IL-6, G-SCF, TPO) promotes hematopoiesis within embryoid aggregates and may result in the appearance of tissue-like structures such as blood islands and early blood vessels. The combination of BMP4 with hematopoietic cytokines yields up to 20% of CD34+CD45+ cells that will give rise to erythroid, macrophage, granulocytic and megakaryocytic colonies [2].

To produce EBs of equal size and standardize differentiation, a certain number of cells can be used to form aggregates by a spin technique (centrifugation) or in a hanging drops method. Hanging drops are single 10-20μl droplets with known cell densities that are placed on a glass surface or into hanging drop plates. Several studies have shown improvements of this method that would allow it in practical application.

Coculture with stromal cells

This two-dimensional differentiation system is based on induction of hematopoiesis upon exposure to extrinsic signals from the feeder cells that underlie the PSCs in coculture. Stromal cells with the capacity to induce and support hematopoiesis can be isolated from a variety of anatomical sites associated with the hematopoietic development in vivo. A number of cell lines were established from mouse bone marrow (OP9, MS5 and S17), yolk sac endothelium (C166), fetal liver (mFLSC, EL08) and other sources. The genetically modified stromal cells, immortalized or expressing specific growth factors and signaling molecules, are widely used in hematopoietic coculture.

The standard coculture conditions comprise prolonged, up to 4 weeks, incubation of undifferentiated pluripotent stem cells on top of the stromal cells in the presence of fetal bovine serum (FBS) and/or hematopoietic cytokines. Both mouse and human pluripotent cells can be successfully differentiated into CD34+ multi-lineage blood progenitors in a coculture, though the efficiency of hematopoietic differentiation significantly varies between different stromal cell lines and compositions of differentiation media [3].

Defined feeder-free, serum-free systems

These systems are designed to avoid the use of undefined, animal-origin components such as FBS and stroma cells to achieve highly reproducible and efficient outputs. Thus, PSCs can be plated on matrix protein collagen IV and differentiated into primitive CD34+CD43+ hematopoietic progenitors by exposure to BMP4, bFGF and VEGF. This initial differentiation is more efficient when accompanied with the hypoxic conditions (5% oxygen tension) that resemble the environment of a developing embryo. A further incubation of blood progenitors with the various combinations of cytokines yields maturation of CD71+CD235a+ erythroid cells, CD41a+ CD42b+ megakaryocytes, HLA-DR+CD1a+ dendritic cells, CD14+CD68+ macrophages, CD45+CD117+ mast cells and CD15+CD66+ neutrophils [4].

Despite the great progress achieved in the in vitro modeling of hematopoiesis, blood production from PSCs is still a variable process.  The final goal of intensive research in this area – a consistent production of engraftable cells, capable of reconstituting all blood lineages in the body, remains a major challenge. Finding critical intrinsic and extrinsic factors that can recreate the unique properties of a hematopoietic stem cell niche in vitro could advance the generation and expansion of PSCs-derived hematopoietic stem cells in the future.



1. Moreno-Gimeno I, Ledran MH, Lako M. Hematopoietic differentiation from human ESCs as a model for developmental studies and future clinical translations. FEBS J. 2010 Dec;277(24):5014-25.Review.

2. Chadwick K, Wang L, Li L, Menendez P, Murdoch B, Rouleau A, Bhatia M.Cytokines and BMP-4 promote hematopoietic differentiation of human embryonic stem cells. Blood. 2003 Aug 1;102(3):906-15.

3. Vodyanik MA, Bork JA, Thomson JA, Slukvin II. Human embryonic stem cell-derived CD34+ cells: efficient production in the coculture with OP9 stromal cells and analysis of lymphohematopoietic potential. Blood. 2005 Jan 15;105(2):617-26.

4. Salvagiotto G, Burton S, Daigh CA, Rajesh D, Slukvin II, Seay NJ. A defined, feeder-free, serum-free system to generate in vitro hematopoietic progenitors and differentiated blood cells from hESCs and hiPSCs. PLoS One. 2011 Mar 18;6(3):e17829.

Direct conversion or iPSCs: Do all roads lead to Rome?

During development, when cells are programmed to perform specific tasks via terminal differentiation, they don’t undergo fate changes and perform their specific tasks throughout life, a process referred to as phenotypic stability. The idea of reprogramming challenged this concept and introduced the notion that the genomic material from any given cell can be reprogrammed to move back in the developmental cascade and become more unrestricted in terms of what other cell types it could generate in the process. It was initially shown that through Somatic Cell Nuclear Transfer (SCNT), one can reprogram the nucleus of any somatic cell by injecting it into oocytes that were experimentally devoid of their own nucleus. Once the oocyte divides and begins the process of developing, pluripotent cells can be isolated to be used for many purposes (e.g., further differentiation into a specific cell type, such as neurons, expansion, etc.).

stem cells

Over the last decade, induced pluripotent stem cell (iPSC) technology revolutionized the way we now create pluripotent cells from differentiated, somatic cells. By ectopically expressing handful of genes -as low as 2 of them in some cases- differentiated cells can be pushed to express genes that are associated with a more undifferentiated state and eventually transform into “embryonic stem cell like” colonies. Numerous laboratories around the world is utilizing this technology to generate iPSCs for future cell-replacement therapy, studying disease mechanisms, testing drug toxicity and more. However, depending on the cell type that’s being differentiated from the iPSCs for these particular studies, one may not need to generate iPSCs and go straight to the last step of generating a particular cell type (e.g., neurons, beta endocrine cells, etc.) by using direct conversion.

describe the image

In direct conversion (also referred to as transdifferentiation), the ectopic expression of transcription factors converts somatic cells from one lineage to another without reverting the cell all the way back to a fully undifferentiated state. Direct conversion has been around a lot longer than the iPSC technology. For instance, conversion of fibroblasts into muscle cells by overexpression of MyoD [1] or conversion of lymphoid cells into macrophages by expressing Pu-1 [2] were among the first demonstrations that a single transcription factor could directly convert one cell type into another. Note that these initial studies achieved conversion from one cell type to another within the same lineage. More recently, using the iPSC approach, there have been studies that achieved direct conversion from one germ layer to that of another, such as fibroblasts to neurons or liver cells. Furthermore, there have been studies that utilized direct conversion in vivo, such as in vivo conversion of exocrine to endocrine pancreas cells [3], or in vivo conversion of fibroblasts into in functional myocytes in an infarct area [4]. Without a doubt, the success of recent experiments using direct conversion was inspired by Yamanaka’s discovery of iPSC technology, further indicating that the approach has importance beyond just the reprogramming of somatic cells into pluripotent cells.

In direct conversion, one cell type is transformed into another without going through the intermediate pluripotent state (i.e., iPSCs). In specific applications where the pluripotent state is not necessary, this is a welcome change, since the process of creating and maintaining iPSCs are labor intensive and costly. Furthermore, for cell-based replacement therapies, differentiation of cells from iPSCs and transplanting them into recipients carry additional risk factors, such as increased chance of tumor formation, while dealing with pluripotent cells. In the two examples stated above, in which the pancreatic exocrine cells were converted directly into endocrine cells or the resident fibroblasts in the injured heart that were converted into functional cardiomyocytes, there was no need for an intermediate pluripotent state to achieve the goal of transdifferentiation, and the process of conversion took place in vivo. By utilizing the right set of transcription factors, the cells transformed from one lineage into another and achieved the final result of repair and/or restoration of function following disease/injury. Even though, iPSCs would be necessary for many applications, for certain tasks, we can reach the same final aim by using direct conversion without the need for iPSCs. Furthermore, direct lineage conversion could provide important new sources of human cells for creating in vitro disease models and cell-based replacement therapies. While the use of direct conversion gains more popularity, it will be significant to carefully determine the fidelity of reprogramming and to develop methods for robustly and efficiently generating one specific human cell type from another, directly.


[1] Expression of a single transfected cDNA converts fibroblasts to myoblasts. Davis et al., (1987) Cell, 51, 987-1000

[2] Stepwise Reprogramming of B Cells into Macrophages. Xie et al., (2004) Cell, 117, 5, 663-676

[3] In vivo reprogramming of adult pancreatic exocrine cells to beta cells. Zhou et al., (2008) Nature 455, 627-632.

[4] Heart repair by reprogramming non-myocytes with cardiac transcription factors. Song et al., (2012) Nature, 485, 599–604.

The Promise of Induced Pluripotent Stem Cells

Human embryonic stem cells (hESCs) hold great potential for cell replacement therapies, where cells are lost due to disease and/or injury. During the last decade, we have witnessed the development of the induced pluripotent stem cell (iPSC) technology, which revolutionized the stem cell field, as well as the regenerative medicine. Since the source of the reprogrammed cells (e.g., fibroblasts, keratinocytes, etc.) as part of the generation of iPSCs is readily accessible, this opened up an entirely new chapter in regenerative medicine by significantly reducing immune rejection problems after transplantation. Furthermore, by taking somatic cells directly from patients, we now have the ability to create disease models (also know as “disease in a dish”) for many conditions that were not possible before. As of today, numerous diseases have been modeled using iPSCs and many researchers are exploring the possibility of utilizing iPSC-derived cells for cell replacement therapy.

One of the pioneers of the iPSC work, Shinya Yamanaka, was awarded the Nobel Prize in Physiology or Medicine (together with John B. Gurdon) this year. Considering that Dr. Yamanaka’s first study related to iPSCs was published in 2006, it’s easy to appreciate the enormous impact that this technology has created as substantiated by the decision of the Nobel Prize committee. If we look back at the chronological events, the initial report from the Yamanaka group using mouse fibroblast and 4 transcription factors (i.e., Oct4, Sox2, Klf4 and c-Myc) to create iPSCs was published in 2006 (Takahashi and Yamanaka, 2006), followed by the demonstration in 2007 (Takahashi et al., 2007) that a similar approach was applicable for human fibroblasts, and by introducing a defined set of transcription factors, human iPSCs can be generated. On the same day Yamanaka published his human iPSC paper, Jamie Thomson’s group also demonstrated the generation of iPSCs from human cells using a different set of factors (Yu et al., 2007).

describe the imageEven though, iPSCs can be generated reproducibly, the efficiency of the reprograming remains low (i.e., around 1% using the original method). Furthermore, the initial generation of iPSCs utilized either retroviruses or lentiviruses, which both integrate into the host genome and might cause insertional mutagenesis, collectively creating a risk for translational applications. However, the beauty of the iPSC technology is its simplicity and reproducibility. Since 2007, iPSCs have been in the central focus of stem cell research and regenerative medicine, and researchers throughout the world have made a number of improvements to Yamanaka’s original protocol. Today, we have safer and more efficient methods for generation human iPSCs for disease modeling or cell replacement therapies, and by the look of how much has been achieved so far, the technology is only going to get better and more proficient. Considering that human embryonic stem cell derived approaches have been successfully used at the clinical level for cell replacement therapies, there is no doubt that iPSCs will eventually prove to be the ultimate source for regenerative medicine in the future.