This week in stem cell news: Fibronectin in a clinical trials success story, STAP cells not so remarkable anymore
Every week, discoveries in the regenerative medicine field span the gamut from remarkable to groundbreaking to surprising, not to mention they add up to, in numbers, more than the space we have to mention them. This week, we highlight a couple of interesting developments that have generated a significant amount of discussion in the regenerative medicine field over the past week.
- STAP cells: Not so remarkable? A few weeks ago, we wrote about STAP cells, after Haruko Obokata and colleagues at Harvard Medical School made a big splash in the stem cell field by publishing a paper in Nature claiming to have discovered a new type of stem cells, called STAP, which are adult skin cells that acquire pluripotency after a simple exposure to an acidic environment. Since the publication, these findings have been under increasing scrutiny. First came the good news: Nature made history when it agreed to make the paper in question open access, i.e. available online for free. Then, the not-so-good news followed: While much praise was lavished on the study at the time of publication, the past week the stem cell community has grown increasingly skeptical of the study after findings surfaced about the paper that paint a much bleaker picture. Serious allegations of data irregularities accuse the authors of plagiarism both in the data and the figures, bringing into question Obokata’s entire study. The RIKEN Center for Developmental Biology in Kobe announced that is has begun a formal investigation into the studies, while independent groups have been attempting to reproduce the results of the study, so far unsuccessfully. The lively discussion that has surfaced in the stem cell community is exciting to follow, even though it’s anybody’s guess how this will all play out. We will be watching.
- Fibronectin: A clinical trials success story. We know that fibronectin is good at its job. A remarkable extracellular matrix protein, fibronectin has found wide-ranging application in tissue engineering constructs such as Dermagraft that have been successfully commercialized. Another success story of the remarkable biological activity of fibronectin comes from Kyoto Prefectures University of Medicine in Japan. The results of a Phase 1 clinical trial on T cell therapy to treat patients with advanced cancer have just been published, and they signal good news. The study, Phase I Clinical Trial of Fibronectin CH296-Stimulated T Cell Therapy in Patients with Advanced Cancer was published in PLoS One by Takeshi Ishikawa and colleagues. The authors report that fibronectin (fragment CH296)-stimulated T cell transfer therapy showed very good tolerance and efficacy over a course of 3 months. Fibronectin acts together with anti-CD3 to induce T cell proliferation. In this way, the authors used fibronectin to generate “fit T cells”, and showed that such an approach could be extended to various T cell based therapies.
For more on human fibronectin, including custom bioassay design, see our fibronectin product page.
This week, we extend our conversation on scaffolds used in tissue engineering by featuring an exclusive interview with Dr. Michael Hiles, the Vice President for Research and Clinical Affairs at Cook Biotech Incorporated. Cook Biotech is a company that develops extracellular matrix (ECM) technologies for implantable and topical medical devices.
Dr. Hiles is an expert on biomaterials and biological scaffolds as they are used in tissue engineering applications. Dr. Hiles received his BS and MS degrees in Electrical Engineering from Purdue and his Ph.D. in Veterinary Physiology and Pharmacology from the Veterinary Medical School at Purdue and holds over 30 patents. In his extensive career, he has published articles that span a broad array of tissue engineering topics, including catheter-based medical instrumentation, pharmacological intervention in acute animal disease, composition and structure of biomaterials, and biomechanics of soft tissues. Mike has provided his expert opinion on the current state of biological scaffolds.
Cook Biotech is a major developer of ECM-based biological tissue grafts – or scaffolds. Can you tell us a little bit about your company and the main sources of tissue graft materials that you work with?
ECMs can come from many species, including humans, and are used as tissue grafts in animals and people, but the processing and safety of each are not equivalent. For example, cadaveric grafts can convey fatal diseases and animal tissues need to be proven free of viruses and TSEs. Fortunately, the acellular animal tissues have proven to be quite compatible with humans and definitely safer and more ethical than cadaveric tissue. We currently work with several tissue ECMs from a porcine source. Most of our work is with the strong, growth factor rich, plentiful submucosa of the small intestine.
How have your tissue grafts been used? Can you tell us a little about some of your main products and applications?
Our grafts based on small intestinal submucosa (SIS) have been used to repair nearly every soft tissue in the body, from the dura mater directly on the brain to stimulating closure of venous ulcers on the foot. Our main products are used to repair large ventral hernias, common inguinal hernias, colorectal fistulas, chronic wounds, and for dental, plastics, and nerve repair procedures.
What are the most important requirements for a successful biological scaffold?
A successful biologic scaffold must interact with the body on macroscopic and cellular levels to be not only truly biocompatible but also be cell-friendly, tissue conductive, non-immunogenic, sterile, and possess the necessary handling characteristics to give surgeons confidence in using it. Unsuccessful biologic scaffolds behave more like synthetics, harbor infections, or are outright rejected by the body. We take a great deal of care to preserve the natural ECM because we have seen that this environment is important for healing.
Obviously you have had a lot of clinical success with your scaffold technology. Is this a trend you are seeing – in other words, are biological scaffolds finding a more straightforward path to approval?
Although we have seen literally millions of patient uses of SIS without rejection in tissues all over the body and in countries all over the world, the pathways to approval and regulations in general for new uses are murkier and more unnecessarily burdensome than ever. Most regulatory agencies are finding ways to exclude biologic scaffolds because their compositions cannot be fully defined, because their secondary or tertiary mechanisms of action are complex interactions with the body, or simply because they erroneously fear animal tissues are somehow inherently unsafe. Many aspects of regulation are driving up costs at a time when price pressures are also knocking biologic grafts out of many procedures.
Recurrence is often mentioned as a major issue with implanted tissue grafts. What are the main causes of elevated recurrence with certain grafts and what are strategies that companies are employing to reduce recurrence?
A successful outcome with a biologic graft can only result when there is a proper balance between patient, procedure, and product. Most recurrences, adverse events, or other complications are often blamed on the product when improper technique or patient selection may have been the root cause. This is not to say that recurrence is never a fault of the product. Overtly infected surgical fields and hyperactive metabolic states can cause premature graft degradation and recurrence, and improper material selection or graft design can cause graft failures. For example, highly elastic materials like dermis are not suited for applications that require long term strength with dimensional stability because they tend to stretch over time and form new tissue that is also stretchy. Thus, two strategies to reduce recurrence are to start with more structural ECMs and then teach practitioners how best to implant them.
The complexity of tissues is obviously a major challenge to developing functional grafts. Can you name a few tissue-specific challenges that you have encountered?
As a first example, although SIS isn’t as highly thrombogenic as pure collagen, it doesn’t seem to work well by itself for small diameter vascular grafts because the grafts thrombose before they can fully heal. We spent a lot of time, money, and effort trying to make this work and couldn’t. Secondly, because our source material is intestine, levels of bacterial endotoxin in the raw material can be very high. Dura mater replacement and contact with the CNS require a very low endotoxin load, and although difficult, we managed to get there and have a very successful dura product.
What are, in your opinion, the main general hurdles that accompany the development of new tissue graft products?
We’ve already touched on a few of the hurdles, such as increasing regulatory burden and tissue-specific designs, and other hurdles include viral validations and best practices determinations. If you don’t understand the chemical, biological, and mechanical needs of your intended use, then you won’t be able to make an effective product design. If you can’t prove that your source materials are safe, you won’t be helping anyone, and if you can’t help surgeons learn the best ways to use your products, you’ll end up with many more unhappy customers and poor patient outcomes than are acceptable.
Biological grafts have come a very long way. What is, in your opinion, the yet untapped potential of biological scaffolds looking ahead?
Scaffolds are perhaps the most important part of tissue engineering, and as we move more and more into the realms of cellular therapies, I think we will find that co-delivering cells with a matrix will make a much better product. Thus, biologic grafts in the form of combination products have a very large and untapped potential in virtually all tissues that can be accessed with a needle or a catheter. Further, we have learned that ECM can convert cancer cells into less pathologic phenotypes, that ECM itself can be used as vaccine adjuvants, and that drugs and cytokines can be preferentially bound and protected from degradation by ECMs. The limits of ECM utility are truly limited only by imagination!
What happens after you injure yourself? Wound healing is a complex process involving a number of controlled steps, each involving the appearance of a distinct cell type the function of which is regulated by a number of extracellular matrix proteins. Traditionally, the wound healing process has been divided into three main steps: inflammation, proliferation, and maturation and remodeling (Yolanda et al., Stem Cell Research and Therapy, 2014, 4:1).
One the cellular side, after wounding, hematopoietic stem cells and, particularly, mesenchymal stem cells from the bone marrow migrate to the wound site, where they regulate cell proliferation by recruiting other cells and releasing growth factors and matrix proteins. The beneficial properties of MSCs on wound healing was observed in a number of animal models and in clinical cases and they are considered one of the most important components of the wound healing cascade. On the extracellular matrix protein side, after wounding fibronectin is part of the fibrin clot and distributed along fibrin strands.
Fibronectin’s role in this process is remarkable and critically important. During the proliferation phase, the crosstalk between fibronectin and platelet-derived growth factor receptor (PDGFR)-β controls the migration of mesenchymal stem cells. This is a fundamental determinant of cell migration and wound repair. This interaction between fibronectin and MSCs has been reported extensively in the literature (Veevers-Lowe, 2014, J Cell Sci, 124:1288-1300).
But it doesn’t stop here. Fibronectin is a key driver not only of the wound healing process in stem cell-driven tissue repair.
Fibronectin-gold nanoparticle-coated composite scaffolds in cardiovascular devices have also shown to favorably interact with mesenchymal stem cells and enhance MSC migration as well as protein expression (Hung et al, 2013, PLOS One). This importantly suggests that the FN-Au nanocomposites may significantly improve blood-contacting devices such as vascular grafts.
Fibronectin expression has also been linked to increased MSC lung adherence (Nystedt, 2013, Stem Cells, 31(2):317-326), which has implications in the development of infusion-driven delivery routes of MSCs for cell therapy.
These are just some examples – and this is by no means a comprehensive list – of the critical role fibronectin plays in the myriad interactions that regulate tissue repair.
Akron is a leading supplier of human plasma-derived fibronectin, and can assist with custom bioassay design and investigating fibronectin function in complex 3D scaffolds. Order your sample today. Contact us for information.
In a seminal paper published in 2009, Place and colleagues described the then-current state of the art of the field of biopolymers used in tissue engineering. The paper is still a comprehensive go-to reference for anyone working in the field and looking to broaden their understanding of both the history and the challenges of this growing area. When the paper was published, there were over 170 companies investing in tissue engineering research and development. This number has since increased, as has the number of discoveries that have turned into commercial products. For instance, Osiris was in 2009 a young but promising company developing a mesenchymal stem cell-based therapy for acute myocardial infarction called Prochymal. Four months ago, Osiris announced the sale of their entire MSC business to Mesoblast, a big player in the tissue engineering field.
Such transitions are not uncommon in the tissue engineering (TE) field, but they highlight the growing impact that scaffold-based TE therapy is having on the development of biotechnology businesses.
A few entries ago, you may remember us writing about synthetic and biological scaffolds. Synthetic biodegradable polymers used in tissue engineering scaffold design include polyesters, polyanhydride, polyorthoester, polycarbonate and polycaprolactone. Among these, those that are most commonly used includes poly(lactic acid) (PLA), poly(glycolic acid) (PGA), poly(lactic acid-co-glycolic acid) (PLGA), and poly(ɛ-caprolactone) (PCL).
One of the biggest areas of change since Place et al.’s Nature Materials paper has been the development of new combinations of synthetic biopolymer scaffolds that are both simple to manufacture and amenable to being used by others, and the synergystic application of molecular biology and chemical methods to the engineering design of novel biomaterials. An example of this is a new technology described in the Journal of Biological Chemistry last year, wherein authors developed a new electrospun 3D matrix composed of PLGA and polymerized allylamine with heparan sulfate non-covalently integrated on the surface of the fibres. Heparan sulfate triggered the neural differentiation capacity of mouse embryonic stem cells which in turn supported neural activity across the scaffold surface. The technique is both simple, straightforward, scalable and is amenable to testing by others.
PLGA has also been used in conjunction with lipid nanoparticles to generate polymer-lipid hybrids which support the delivery of drugs. The paper, by Willem Mulder’s lab at Mount Sinai School of Medicine, then used these constructs to successfully deliver the drugs doxorubicin and sorafenib into tumor-bearing mice.
These are just some of the examples – and they only involve PLGA – of how technological advancements, and the synergy between engineering, chemistry and biology – is poised to usher in new, simple and application-driven scaffold-based solutions for the treatment of a widening range of diseases. As a result, we expect the business arena to restructur itself and adapt to these new developments in the lab.
Akron is part of that process. Our expertise in scaffold design enables us to be at the cutting edge of scaffold-based TE developments and readily adapt to market needs. Akron can accommodate small scale development and high throughput electrospinning. Please call us for more information.
This week, the stem cell field was shaken by a remarkable discovery that has generated waves across the world. The journal Nature published two studies, a letter and a research article, by Haruko Obokata and collaborators at Charles Vacanti’s lab at Harvard Medical School, that are being heralded as a “medical breakthrough” that could “revolutionize personalized medicine“. The studies report on the controlled reprogramming of differentiated cells into pluripotent cells by simply exposing the cells to an acidic environment.
The authors call this phenomenon stimulus-triggered aquisition of pluripotency (STAP) which, in essence, generates iPS-like cells from any differentiated cell by simply placing the cells in an acid bath for 30 minutes. In the past, a related achievement, that of reprogamming adult cells into induced plutipotent stem cells (iPSC) won Shinya Yamanaka and John Gurdon, the two scientists who came up with the discovery, a Nobel Prize in 2012. Is Obokata’s discovery just as remarkable?
In the Nature study, Obokata showed that three different stresses - membrane pore formation by a bacterial toxin, exposure to acidic pH and physical squeezing – all triggered cells to a pluripotent state. To prove their point, Obokata isolated lymphocytes from mice that were engineered to carry a gene that glows in the presence of Oct-4, a protein found in pluripotent cells. The cells that grew after being exposed to a pH of 5.7 for 30 minutes grew bright green, indicating their pluripotency. After injecting these STAP cells into blastocysts, the cells incorporated into every tissue in the body.
But with the praise came the questions - the most ubiquitous of all being whether this technique will work with real adult cells, since the authors used cells from one week old infant mice. Others are liking the findings to the controversial discovery of very small embryonic-like stem cells (VSEL) 2006, which were recently brought into question by reports that these cells are nothing more than an “aberrant and inactive population” of cell debris and fragments of dying cells. Other opinions verge on calling STAP cells cancer-like, and adding that the stress induced on the STAP cells might remain and manifest itself later.
Whatever the truth, it will hopefully come out as more labs attempt to reproduce Obokata’s results, and in doing so, shed more light on this fascinating new phenomenon.
For many laboratory researchers, the transition from pre-clinical development to clinical application is either an afterthought or a convoluted, documentation-riddled process. Yet cGMP-compliant manufacturing processes should be at the core of any product development campaign, as good planning can make an otherwise extensive process significantly more straightforward. Good planning starts from the beginning: the selection of raw materials. Because researchers are ultimately responsible for the quality of the final product, traceability and quality of raw materials are key. Since the FDA does not regulate raw materials, but rather final products, researchers are responsible for ensuring final product quality standards are achieved through a combination of raw material quality and process compliance. As Akron Biotech CEO Claudia Zylberberg, Ph.D, put it in a recent interview with Stem Cell Assays, “By selecting raw materials early and verifying the availability, consistency, traceability, toxicity and contaminant studies, you control the risks of variability in effectiveness downstream. A qualification process and on-site audit brings confidence and closeness in the relationship with your materials and reagents manufacturer that supports systems, process modifications, and regulatory compliance.”
But that’s not all. cGMP manufacturing compliance is part of a larger Quality Assurance program that also requires compliance of other aspects of the manufacturing process:
- intermediate and end-products
- in process controls
- extensive process documentation and validation
- product identity, packaging and transport
- personnel training
For tissue-engineered products, GMP regulations apply to all phases of product development, including tissue collection, cell and tissue processing and expansion, and storage.
A helpful overview of such guidelines and the current status of the ongoing efforts on the regulatory space are given in “Raw Materials for production of cell-based and gene therapy products” a report by the European Directorate for the Quality of Medicines & HealthCare (EDQM), Council of Europe and the European Medicines Agency (EMA) last April in France, and initiatives from USP such as “Cell and Gene Therapy Products” on Chapter <1046> and “Ancillary Materials for Cell, Gene and Tissue-Engineered Products” on < Chapter 1043>
A lot is known about extracellular matrix protein vitronectin. Its structure, composed of three main domains, and its direct implication in vitronectin’s cellular function, has been studied in extensive detail for a long time. We know vitronectin, like fibronectin, is a key protein of the ECM with important roles in wound healing (Koivisto et al.; Advances in Wound Care; 2013). Which is why new studies implicating vitronectin in novel functions are fascinating to come across.
One such study in question, published by the lab of Nicolai Sidenius at the European Institute of Oncology’s IFOM-IEO research institute in the journal Blood (Pirazzoli et al.; Blood; 2013, 121:2316), presents evidence of vitronectin’s direct influence on tumor growth and recession and confirms vitronectin as an interesting drug target.
It has been known for some time that vitronectin’s Somatomedin B domain interacts with the urokinase plasminogen activator receptor (uPAR) (Gardsvoll and Ploug; J Biol Chem.; 2007 282(18):13561-72). By preparing mutants of uPAR incapable of binding vitronectin and both vitronectin and uPA, the authors demonstrated that cells with these mutants, which lack wild type uPAR function, lacked cell adhesion, spreading, migration, and proliferation function. The authors then followed findings with in vivo studies in severe combined immunodeficiency mice. And the findings were fascinating: In vivo, they showed that the uPAR mutant incapable of binding vitronectin resulted in strongly reduced tumor formation. This is a remarkable finding that directly relates the interaction between uPAR and vitronectin to tumor growth.
Previously, the vitronectin family of integrins, key components of the extracellular matrix, had been identified as interesting drug targets, due to their direct involvement in cell survival upon tumor invasion (Murphy and Stupack; Cell-Extracellular Matrix Interactions in Cancer; 2010, 137-170). This is beyond its well-known function in wound healing and viral infection.
This is interesting to us, because, as a supplier of vitronectin, we work with researchers to provide pure, biologically active human plasma-derived vitronectin to aid biomedical research and further such discoveries. Contact us to enquire about our vitronectin grades and our bioassay development capabilities, and find out how we can help move your research.