Stem Cell-based Therapeutics Laboratory

Stem Cell Therapeutics

Stem cells form the basis of the field of regenerative medicine and are thought to have tremendous promise for treatment of a wide range of diseases. To maintain cutting edge knowledge in this field and provide information for effective decision making, the Centre for Biologics Evaluation has a developed a research program focused on stem cell-based therapeutics, which include stem/progenitor cells and their derivatives (i.e., extracellular vesicles).

What are Stem Cell based Therapeutics?

Stem cells are the building blocks of all organs and tissues in the human body. As such, stem cells have a vast potential in medical applications. Stem cell-based therapeutics are being investigated at the preclinical and clinical levels for their therapeutic effects to treat a wide variety of diseases and disorders with unmet clinical needs. Much of the therapeutic promise of stem cell therapies are based on their abilities to repair and regenerate tissues and organs, as well as a capacity to regulate the immune system. This latter property of stem cells is mediated either through direct interactions between the stem cell and immune cells or through the secretion of substances into their surrounding environment. One of the most therapeutically important of these secreted substances are extracellular vesicles.

Stem cells have two features that make them distinct from other cells of the body:

  • First, stem cells are capable of long-term self-renewal
    • that is, they are able to give rise to identical copies of themselves over and over again
  • Second, stem cells are defined by their unique ability to develop into different functional cell types
    • for example, stem cells in the brain can develop into all of the different cell types of the brain such as neurons, astroglia and oligodendrocytes, each of which has a separate function

Stem cells can be grouped into two broad categories: pluripotent stem cells (e.g. embryonic stem cells and induced pluripotent stem cells made by reprogramming somatic cells) and somatic stem cells (e.g. adult stem cells). The distinguishing properties of these cells are as follows:

Pluripotent stem cells hold the potential to become any cell type of the human body. The embryonic stem cells are found under this category. They are derived from embryos at a specific stage in early development termed the blastocyst stage and can be used to derive an entire organism. A “man-made” version of embryonic stem cells were recently engineered using terminally differentiated cells or somatic cells such as fibroblasts (i.e. skin cells). These cells are called induced pluripotent stem cells and behave similarly to embryonic stem cells, without holding the ethical issues that come with using embryonic tissues to derive them. Induced pluripotent stem cells have shown an increased interest over the years for making disease models and the use of their differentiated counterparts for transplantation purposes.

Somatic or adult stem cells are found in fully developed organs and tissues of the body. They can replace dying cells within their organ of residence and can repair minor tissue damage. Adult stem cells are usually restricted to forming only the type of cells found in their tissue of origin; however, some have demonstrated pluripotency.

Therapeutics derived from stem cells are currently being developed for treating conditions such as muscle injuries, neuronal disorders, loss of blood flow, heart disease, liver failure and cartilage deterioration. To evaluate these upcoming biological products fully, a better understanding of their properties is needed in order to promote the development of standardized tools and assays for examining their safety and effectiveness.

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What are Extracellular Vesicles?

Extracellular vesicles derived from stem/progenitor cells are increasingly known for their therapeutics benefits, which can be comparable to the parental cells from which they are derived. Since recent pre-clinical evidence is showing that stem/progenitor cell therapies may work through the transfer of extracellular vesicle released from cells rather than by the transplanted cells themselves, a great deal of attention has turned towards the study and the use of the extracellular vesicles as potential new therapeutics.

Extracellular vesicles are nanometric (50-200 nm in diameter) membrane vesicles that are released by all cell types, including stem/progenitor cells. Their nanosized nature means that they can only be measured and visualised by instruments able to detect nanoparticles such as electron microscopes and nano tracking analysis instruments. They are composed of a cargo of bio active molecules (proteins, DNA, RNA, lipids) that can be transferred to other cells to mediate their beneficial (therapeutic) or detrimental (tumor promoting) effect. The importance of extracellular vesicles as therapeutics lies in their potential to mediate and regulate immune cells, as well as by promoting tissue repair and regeneration.

The Stem Cell-based Therapeutics Laboratory is looking at standardizing the manufacturing steps and characterisation procedures for extracellular vesicle-based therapeutics for obtaining scalable, reproducible and GMP-compliant protocols that fit the regulatory framework. When focusing on regulatory challenges, issues regarding safety, quality, efficacy and manufacturing requirements still remain to be addressed. Indeed, there is a strong need for standardization of production/enrichment procedures, as well as establishment for accurate methods of quantification (concentration, yield and purity) and qualification (identity and cargo content). Therefore, research aimed at better defining stem cell-derived extracellular vesicle products, such as MSC-derived, in terms of mode of production/enrichment as well as cargo content (proteins, RNA, DNA, lipids), and the effect of long term storage and handling on quantity/quality attributes is needed to assess their clinical potential as therapeutics.

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Challenges Associated with Safety and Efficacy with Stem Cell-based Therapeutics

Stem cell based therapeutics are often based on or from cells that have been extensively expanded or grown in the laboratory in order to obtain sufficient material to treat patients in the clinic. As cells are expanded, they can be subjected to increased genetic instability, which can lead to increased risk of tumorigenicity. Also, as cells are expanded they tend to lose their efficacy in performing their normal functions. Moreover, since stem cells are derived from human tissues, they can present variability in terms of functionality from donor-to-donor tissue. Assessment of the functionality of the cells and variation in the amount and types of extracellular vesicles they generate is therefore a key aspect for ensuring their therapeutic quality. The laboratory’s research focuses on those safety and efficacy issues that can arise when preparing stem cells and extracellular vesicles for therapeutic use in humans.

Defects at the stem cell level can also form the basis of a disease. Mutations in stem cells, for example are thought to be the root cause for several types of blood cancers and may be the principal reason behind sarcoma formation. Understanding how to differentiate between normal healthy stem cells and stem cells that contribute to the disease state is also and important aspect of ensuring that stem cell based therapeutics are safe and effective. Our research program focuses on the comparison of stem cells derived from a healthy environment with those derived from various disease states in order to tease out molecular and behavioural differences between these cells. The idea of this work is to develop methods to identify the tipping point between when a stem cell moves from a healthy cell to a diseased cell.

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Concepts and Tools We Use to Study Stem Cell-based Therapeutics

At the Centre for Biologics Evaluation, we want to develop new methods and models for testing the safety and efficacy of therapeutics that are based on stem cells. The centre's research team does this by growing adult stem cells and generating extracellular vesicles in the laboratory and analysing them with sophisticated instruments. They identify characteristics of stem cells associated with therapeutic effectiveness. Examples of these characteristics include particular patterns of protein and gene expression to identify molecular markers that are linked to the safety and efficacy of the products. We examine these properties in the laboratory through the combined use of fluorescence microscopy, polymerase chain reaction (PCR), mass spectrometry (MS), RNA sequencing and flow cytometric assays.

This research will be important for:

  • Providing scientific information on how to evaluate therapeutic products that are based on stem cells
  • Understanding the positive therapeutic potential of stem cells and extracellular vesicles
  • Understanding how to identify potential safety issues associated with stem cells and extracellular vesicles
  • Developing new assays and tools to monitor the quality and safety of stem cell–based therapeutics
  • Applying and developing in vitro models to better evaluate the effectiveness of stem cell-basedtherapeutics for treating various medical conditions

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Research Highlight 1: Molecular markers to characterize stem cell-based therapeutics regarding safety and efficacy.  

The term molecular marker is used to refer to something that may be an indicator for a specific biological substance, process or function. For example, specific DNA or RNA sequences can provide a genetic marker for the presence of cancer and may provide useful information for treating the disease. A marker may be a valuable tool to confirm the identity of a cell-based therapeutic. For stem cells, the expression of specific genes and the presence of certain proteins and enzymatic functions may serve as suitable markers. Our studies aim to identify markers that will help in the understanding of the efficacy and safety of adult stem cell based therapeutics (e.g. mesenchymal stromal/stem cells; MSCs) or induced pluripotent stem cells, and their derivatives (e.g. extracellular vesicles).

The centre is working to

  • Understand the various properties and mechanisms of growth and differentiation of adult mesenchymal stem cells and induced pluripotent stem cells and their derivatives that will provide insight into possible medical applications
  • Characterize the properties of mesenchymal stem cell and induced pluripotent stem cells and their derivatives, to provide guidance on how to evaluate the safety, efficacy and quality of therapeutic products that are based on stem cells
  • Use a detection assay, based on markers to help develop standardized methods for demonstrating a product’s quality, efficacy and safety aspects before it is used in therapeutic applications

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Research Highlight 2: Development of strategies and assays to improve understanding of stem cell-based therapeutics.

Stem cells have the potential to help repair damaged tissues and organs, as well as regulate the immune cells. The Centre for Biologics Evaluation is interested in developing testing methods to evaluate the application of stem cells and extracellular vesicles to perform their normal functions which range from tissue repair to modulation of the immune system.

The establishment of in vitro assays and testing methods for the measurement of the immune modulatory and tissue regenerative properties of stem cells and extracellular vesicles are of great importance for assessment of their quality, potency and safety attributes. Being able to measure the activity of stem cells and extracellular vesicles using in vitro assays also aim at reducing animal use, thereby providing other means of assessment which do not rely on animal models.

For example, the effects of stem cells can be assessed using an in vitro assay, herein called T cell proliferation (TCP) assay, which involves the co-culture of MSCs with activated human T cells, where the proliferation of T cells is reduced over time after incubation with the stem cells. This assay is currently utilized for quality control testing of ProchymalTM and other cell-based products in the early stages of development. Protection and regeneration of ischemic tissue damage is a second key function of stem cells such as MSCs. In light of this, the Laboratory is looking at testing the efficacy of stem cell therapies and extracellular vesicles in an in vitro assay that measures the ability of MSCs to protect neurons from an oxygen/glucose deprivation (OGD)-like insult, as occurs in stroke.

As multiple stem cell types are being investigated in clinical trials for neurodegenerative diseases and autoimmune disorders with unmet clinical needs, these assays will serve to measure the activity of the stem cells and their extracellular vesicles with the aim to put in place assays for quality and efficacy control assessments.

The centre is working to

  • Develop assay systems that provide information to predict the therapeutic potential as well as safety aspects of stem cell-based products and their derivatives
  • Provide increased understanding of the function of human stem cells within human tissue

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