- Massachusetts Institute of Technology, Cambridge, MA, U.S., Research Fellow in Bioengineering and Environmental Health, 1997-1998.
- University of British Columbia, PhD in Chemical Engineering and Biotechnology, 1997.
- McGill University, BEng in Chemical Engineering and Biotechnology, 1992.
- Institute of Biomaterials and Biomedical Engineering, University of Toronto.
- Department of Chemical Engineering and Applied Chemistry, University of Toronto.
- McLaughlin Centre for Molecular Medicine, University of Toronto.
- McEwen Centre for Regenerative Medicine, Toronto.
- Interim Executive Director, Medicine by Design.
- Chief Scientific Officer, Centre for the Commercialization for Regenerative Medicine, Toronto.
MY RESEARCH OVERVIEW (GO TO SCIENTIFIC OVERVIEW)
Stem Cell Bioengineering Approaches to Generate Specialized Cells for Therapeutic Applications
There are two main motivations driving our research on stem cells. First and foremost, we have a desire to contribute to the health and welfare of Canadians and people all over the world. Second, we are interested in tackling complex health problems using bioengineering approaches.
Stem cells hold eno rmous potential as a renewable source of cells for use in a variety of regenerative medicine applications. However significant challenges remain in translating their demonstrated biological properties into robust and effective therapies. Living cells present an opportunity to develop therapeutics such as tissue repair and regeneration, metabolic support, immunomodulation, and gene replacement affecting every organ system to tackle medical problems that cannot be treated with small or biological molecule drugs.
The use of live human cells as a therapeutic modality is by no means a new concept; blood transfusions have been in clinical practice for almost 200 years, the first successful solid-organ transplant (kidney) was performed over 50 years ago, and bone marrow reconstitution following myeloablative conditioning (the only currently available stem cell therapy) was first performed (albeit clinically unsuccessfully) in 1959. However, despite the huge expectations, only a handful of cell therapy products currently exist on the market (fibroblasts and keratinocyte-based products for skin burns and ulcerations) because the vast majority of products are in pre-clinical and clinical development.
What differentiates the new generation of cellular therapeutics currently under development is that investigators are no longer simply processing cells or tissues, but rather designing, engineering and manufacturing cell-based products. Despite these advances, the demand continues to exceed the supply of cell-based products, thereby requiring scalable manufacturing processes. Bioprocesses that reproducibly and robustly control cell fate would address both the quality requirement for these cell-based products, as well as the critical gap between discovery and commercialization. By utilizing engineering-based approaches such as mathematical modeling, molecular engineering and bioreactor design, our research aims to enable stem cell based therapies and technologies, and thus positively impact health care.
In my lab, our research focuses on the generation of functional tissue from adult, embryonic, and induced pluripotent stem cells. Our quantitative, technology-driven approach strives to gain new insights into the fundamental mechanisms that control the fate of stem cells, as well as to develop robust systems for the controlled generation of clinically relevant numbers of functional stem cells and their derivatives. Our work is concentrated specifically on the growth of human blood stem cells and the generation of blood and cardiac cells from pluripotent stem cells. The long-term goal of our work is to generate transplantable blood stem cells and repair damaged tissues such as hearts with stem or progenitor cells.
My lab shares space with the van der Kooy and Audet labs, specializing in stem cell biology and bioengineering respectively. Members of our labs routinely collaborate, communicate and synergize to solve problems. Recently we have interacted with a number of labs in the Donnelly Centre including the Bader, Emili, Blencowe and Chan labs. Being surrounded by other like-minded researchers interested in exploring high-impact applications is very motivating. It is in these multi-disciplinary environments that excellence and innovation can be fostered.
SCIENTIFIC RESEARCH OVERVIEW
Research in my lab is focused on the regeneration of functional tissues from stem cells and the development and utilization of tools to modulate the responses of stem cells in vitro and in vivo. Interest in the generation of primary cells and tissues ex vivo is driven both by their potential use as a direct clinical modality and as human tissue analogues for the development of novel therapeutic agents. Our group is particularly interested in understanding the role that the extracellular environment, the stem cell niche, plays in controlling stem cell fate decisions. Using quantitative bioengineering approaches including micro-fabrication, bioreactors, mathematical modeling and protein and cell engineering we are discovering how stem cells integrate their complex microenvironment to make self-renewal or differentiation decisions. The insights gained from our research enable new therapeutically and technologically relevant strategies in regenerative medicine. We specifically focus on understanding how to grow human blood stem cells, and how to differentiate pluripotent stem cells into functional blood, cardiac and pancreatic tissue.
Areas of focus in the lab are:
1. Increasing the yield of hematopoietic stem cells (HSCs) isolated from umbilical cord blood (UCB)
The clinical utility of HSCs arises from their ability to engraft and sustain multilineage hematopoiesis in compromised hosts. Umbilical cord blood is a very attractive source of HSCs. The major limitation of the use of UCB for hematopoietic transplantation is the limited number of stem and progenitor cells obtained in each UCB collection. We have examined the role of cell population dynamics on HSC expansion of UCB-derived cells, and have developed automated systems to manipulate subpopulations of cells and to control the delivery of soluble proteins to the culture. Our predominant observation from this work is that blood stem cell growth in vitro, and perhaps also in vivo, is limited by feedback networks from differentiated cells. Ongoing development for an ex vivo bioprocess for HSC expansion capable of generating clinically relevant cell numbers is underway. Currently, such processes have large media and cytokine requirements, limiting their use. Therefore, we aim to develop a bioprocess that is robust, economical and automated. To that end we have developed an alternative fed-batch media dilution bioreactor and are working towards implementing automated feedback control to reduce media requirements and improve the reliability of this bioprocess.
2. Stirred-suspension based pluripotent stem cell (PSC) expansion and differentiation
PSCs are of significant interest as a source of therapeutically useful cells. Enabling PSC culture in stirred suspension bioreactors would lead to the production of scalable quantities of PSC-derived blood, endothelial, and cardiac cells under strictly controlled operating conditions. An advantage of this system is that it allows for the investigation of physiochemical parameters (oxygen, glucose) on PSC development, as well as the further engineering of PSC differentiation. A design criterion in these studies is to make PSC propagation and differentiation robust, developmentally relevant and amenable to scalable cell production. To this end, we are exploring methods to use novel biomimetic strategies (gradients and scaffold-mediated signaling) to influence PSC differentiation trajectories, and combining microfabrication technologies and bioreactors to generate PSC aggregates and cells in the quantities necessary for biotechnological applications. To be clinically useful such a technology should be capable of routinely producing large cell numbers (as many as 5x108 to 5x109 functional cells). We have been developing novel bioprocess engineering strategies to expand PSCs (based on aggregate size control, pH, dissolved oxygen concentration, media analysis and other parameters). This research will not only define approaches that dramatically increase the availability of specific cell types for use in transplantation and drug discovery, but will also inform our understanding of the cellular and molecular mechanisms by which PSCs commit to specific lineages.
3. Investigating microenvironmental control of PSC fate using cell niche engineering approaches
Current technologies for investigating the PSC response to exogenous signals are commonly performed in 6-well plates or with PSC aggregates, take several days, and are typically variable and undefined, limiting mechanistic insight. Additionally, these systems typically do not account for the complex microenvironments (i.e. niches), in which the PSCs reside. These niches, like in the embryo, often consist of heterogeneous spatial organization and multiple populations of factor-secreting cells. These microenvironmental heterogeneities confound results, reduce assay robustness, and limit our ability to gain mechanistic insight into stem cell fate processes. We have previously demonstrated that micro-contact printing of PSCs into colonies of specified shape, size, and colony-to-colony separation distance can be used to engineer the microenvironment and mitigate heterogeneous response in PSC culture. This design principal has been applied to develop a robust and scalable micro-contact printing based high-throughput (µCP-HTP) platform, consisting of PSC colonies arrayed in 96-well plates, appropriate for high-content (automated microscopy, single-cell based) screening. We are currently utilizing the µCP-HTP platform to perform biochemical assays that will aid in optimization of stem cell expansion and differentiation, and we are also investigating kinase activation in PSCs in response to stimuli with the goal of generating stem cell-specific systems-level models to link agonists/antagonists, kinase activation, and PSC fate decisions. We have further used this niche engineering approach to drive the differentiation of PSC-derived HSCs towards mature definitive blood lineages such as T cells.
4. Predictive computational and synthetic biology approaches to regenerative medicine
In each of these areas, we routinely use computational approaches to enhance our abilities to explore and understand stem cell biology and regenerative medicine. Therefore, this has become a relatively new, yet vital area of research on which we focus. This work combines approaches from computer science and bioinformatics, numerical simulation and visualization, and theoretical science to study basic and applied problems in stem cell biology. For instance, In collaboration with John Dick’s lab, we are currently using bioinformatic analyses in combination with data from high throughput (HT) gene expression screens to identify novel biomarkers that correlate with clinical outcome in patients with acute myeloid leukemia (AML) (Ng et al., manuscript in preparation). By stratifying patients based on their gene expression profiles, we hope to allow clinicians to better determine the optimal treatment regime for each individual patient.
We are also attempting to better understand the development of the blood system with the ultimate goal of being able to more efficiently produce particular blood cell types for clinical use. To this end, we have developed a novel computational approach to determine the diffusible ligand-based cell interactions between cells in the blood developmental hierarchy (Qiao et al., Mol.Syst.Biol. 10:741, 2014) and how those interactions govern how blood stem cells produce all the cell types in the adult hematopoietic system. We also use HT expression data from stem cells and their differentiated progeny to generate simplified Boolean logic models of the intracellular genetic regulatory networks (GRNs) required for both the maintenance of stem cell pluripotency and for their transition to differentiated cell fates (Yachie et al., submitted). Combined with cell position data, we also develop and analyze spatially-explicit multiscale models of how diffusible factors secreted from individual cells either help maintain the fate of surrounding cells, or initiate changes in their GRNs that induce subsequent fate transitions. Finally, we are beginning to develop simplified, abstract models based on dynamical and complex systems theory in an effort to understand these biological systems without having to incorporate the high degree of biophysical realism required by the more detailed simulation approaches mentioned above. Using these interdisciplinary approaches, we aim to quantitatively understand stem cell biology so as to advance the health of Canadians and people around the world.
- CD24 tracks divergent pluripotent states in mouse and human cells. Shakiba N, White CA, Lipsitz YY, Yachie-Kinoshita A, Tonge PD, Hussein SM, Puri MC, Elbaz J, Morrissey-Scoot J, Li M, Munoz J, Benevento M, Rogers IM, Hanna JH, Heck AJ, Wollscheid B, Nagy A, Zandstra PW. Nat Commun. 2015 Jun 16;6:7329.
Rapid expansion of human hematopoietic stem cells by automated control of inhibitory feedback signaling. Csaszar E, Kirouac DC, Yu M, Wang W, Qiao W, Cooke MP, Boitano AE, Ito C, Zandstra PW. Cell Stem Cell. 2012 Feb 3;10(2):218-29.
Growth factors, matrices, and forces combine and control stem cells. Discher DE, Mooney DJ, Zandstra PW. Science. 2009 Jun 26;324(5935):1673-7.
View Pubmed search of Dr. Zandstra's full list of publications.