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‫תל השומר מחקרים תשתיות ושירותים רפואיים בע"מ‬
Tel Hashomer Medical Research, Infrastructure and Services Ltd.
Tel: +972-3-5305998 Fax: +972-3-5305944
Tel Hashomer Medical Research
Infrastructure and Services Ltd.
Human Stem Cell Research
& Regenerative Medicine
Contact :
Sylvie Luria PhD.
Technology Transfer and Business & Development Manager
Tel Hashomer Medical Research, Infrastructure and Services Ltd.
Tel: +972-3-5305998 Fax: +972-3-5305944 Cell: 052-6667277
[email protected]
http://research.sheba.co.il/e/
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‫תל השומר מחקרים תשתיות ושירותים רפואיים בע"מ‬
Tel Hashomer Medical Research, Infrastructure and Services Ltd.
Tel: +972-3-5305998 Fax: +972-3-5305944
Enrichment and Expansion of Human Stem Cells from Peripheral Blood,
and Umbilical vein Cord Blood (UCB)
Although stem cells from UCB can be used for therapeutic transplantations, their number in
donor UCB units is limited. Therefore, there is a need to expand these stem cells under
growth in cell culture conditions for several weeks. The current preexisting expansion
technologies are laborious, expensive and only partially effective, as many of the expanded
stem cell differentiate and loose their stemness (self renewal and multi potency). The
research carried at the Laboratory of Molecular Immunobiology concentrate on the
development of a unique and novel technology to enrich and expand stem cells form human
UCB. This technology is based on relatively high expression levels of ABC (MDR)
transporter genes in the stem cells, as compared to more differentiated progenitor and
mature cells. As these ABC transporters confer resistance (MDR) to cytostatic drugs and
predominantly expressed by the stem cells, the undifferentiated stem cells are enriched by
selective killing (or growth arrest) of non-progenitor cells using treatment with a
combination of a cytostatic agents together with a cocktail of growth factors. This invented
procedure (Patent pending) for the preparation of enriched populations of stem/progenitor
cells for various cell therapy applications exclude the need for laborious and expensive
alternative methods that are based on continuous use of xenogeneic antibodies directed
against cell surface molecules, affinity columns or density gradients. Preliminary studies
indicated that this technology supply relatively large quantities of enriched, high quality
stem cells that have multi potential ability to differentiate into all various hematopoietic
cells.
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Other Non-Hematopoietic Stem Cells from UCB Mesenchymal and
Neuronal Regenerative Medicine
Human UCB is also a source for non-hematopoietic stem cells as mesenchymal stem cells
(MSCs) and neuronal progenitors (UCBNPs). These stem cells selectively adhere to gelatin
and collagen matrixes, respectively. However, these progenitor cells are also separated from
differentiated non-progenitors, based on higher expression levels of the abovementioned
ABC-transporters.
The isolated population of MSCs is successfully expanded in culture and can be enforced to
differentiate into bone or endothelial cells under appropriate growth conditions. These cells
are putatively useful for bone- and blood vessels regenerative medicine.
The isolated neuronal progenitors (UCBNPs) differentiate into cells expressing specific
neuronal markers with extensive neurite outgrowth under culturing at a special conditioning
medium developed in collaboration with scientists at the School of Pharmacy of the Hebrew
University. These cells were found very potent in regenerating neuron cells in ischemia
(neuronal-degenerative) model in vitro, and their efficacy is currently studied in vivo.
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Stem Cell Therapy for Renal Failure
An increasing number of patients suffer from chronic kidney disease (CKD). The
progression of CKD is often relentless with the majority of patients reaching end-stage renal
disease (ESRD) requiring dialysis or organ transplantation, where demand of organs exceeds
availability. Dialysis and kidney transplantation are the only current therapies for patients
suffering chronic renal failure. A demand for kidney progenitor cells is increasing because
of the shortage of donor organs for orthotopic transplantation. Renal progenitor cells allow
gene therapy, cell transplantation, and tissue engineering of bioartificial organs.
Regenerative medicine is focused on the development of cells, tissues and organs for the
purpose of restoring function through transplantation. The use of stem cells as starting
material offers new and powerful strategies for future tissue development and engineering.
When considering cell replacement in diseased kidneys via stem cell transfer, one should
carefully dissect the timing of such a therapy, as end-stage kidneys are already small and
fibrotic and would therefore not allow for the integration of cells. In contrast, earlier stages
of CKD, when residual kidney function and kidney histology are still preserved, are
expected to be applicable for stem cell therapy. Stem cells have been shown to exert
beneficial effects on acute renal injury. We hypothesize that the progression of CKD into
ESRD can be halted with their use.
The invention consists of a method for derivation of kidney stem cells based on
surface marker expression. Such markers provide a feasible approach for
experimental cell sorting of human renal progenitors as well as a framework for
developing cell selection strategies for renal cell-based therapies.
We have identified unique cellular markers and developed a method for derivation of kidney
stem cells based on surface marker expression. Such markers provide a feasible approach for
experimental cell sorting of human renal progenitors as well as a framework for developing
cell selection strategies for renal cell-based therapies. Currently we are establishing preclinical models of CKD in immunodeficient hosts to evaluate the capabilities of the
aforementioned cells in comparison with well-characterized bone marrow - derived stem
cells in the CKD animal models.
We anticipate that the finding of human stem cell populations that contribute to chronic
kidney repair would have tremendous impact on the practice of renal medicine. Halting or
preferably reversal of CKD would reduce the need for organ transplantation and, in turn,
affect the chronic problem of organ shortage.
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Methods of Monitoring Differentiation of - and Reprogramming Renal Cells
During mammalian development, stem cells become progressively restricted in the tissue
types, to which they can give rise, eventually differentiating into a single mature cell type.
Classically, this process was considered irreversible. However, recent evidence suggests that
differentiated cells can be reprogrammed back to pluripotency to support the re-expression
of all developmental programs. These breakthroughs highlight the importance of
transcription factors as well as epigenetic modifiers in the establishment, maintenance, and
rewiring of cell identity.
The early development of the mammalian metanephros, the direct precursor tissue of the
adult kidney, is a complex process that involves highly regulated interactions between two
derivatives of the intermediate mesoderm, the wolffian duct and the metanephric /
nephrogenic mesenchyme. Reciprocal signaling between the neohrogenic/metanephric
mesenchyme and a derivative of the nephric duct known as the ureteric bud results in
branching of the ureteric bud (UB) and condensation of metanephric mesenchyme (MM) at
its tips. The condensed mesenchyme (CM) is thought to form a precursor cell population,
which both maintains itself at the tips of the UB (via proliferation and/or addition from the
surrounding non-condensed mesenchyme) and gives off cells that differentiate into
nephrons, the functional filtration unit of the kidney. Recent experiments have established
that the progenitor cell in the MM fulfils the criteria of a true committed stem cell in that is
capable of self-renewing and of differentiating towards different types of nephron epithelia.
Prior to ureteric bud induction, the condensing metanephric mesenchyme expresses a unique
combination of markers, including the Hox11 paralogs, Osr1, Pax2, Eya1, Wt1, Six2, cited1
and Gdnf . These genes are considered early markers of kidney progenitor cells. Of all, the
continued expression of the transcription factor Six2 in CM is required for maintenance of
this stem cell population during kidney development. The human metanephros appears at
the 5th of gestation and renal stem/progenitor cells in the MM are induced to form nephrons
until 34 weeks of gestation. Thus, endowment of new nephrons is restricted to prenatal
development in humans, while in rodents it persists only until the immediate postnatal
period (up to 2 weeks postnatal).
In a previous study, we observed that the cessation of nephrogenesis was coincident with
dramatic down-regulation of the renal progenitor genes. Given the lack of definitive
evidence for an endogenous renal stem cell in the adult kidney and the apparent absence of
embryonic progenitors after the cessation of nephrogenesis, re-activation of renal progenitor
genes, especially SIX2, might represent a crucial step in reprogramming of somatic kidney
cells towards a progenitor phenotype required for repair or regeneration in the kidney.
Interestingly, during ischemia-reperfusion renal injury and consequent regeneration there
seems to be re-expression of renal developmental genes and pathways, although to a limited
extent.
Our observation that renal progenitor gene silencing in the adult human kidney was
associated with epigenetic changes in the form of DNA hypermethylation, led to
hypothesize that epigenetic reprogramming via VPA or TSA/5-Aza treatment can alter the
expression of these genes in concert with 'stemness' pluripotency genes, which in turn may
have an impact on the developmental competence of renal somatic cells.
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Stem Cell Therapy for Cardiac Aapplications
Tissue engineering offers a potential of developing new treatment options for the repair of
injured organs, using functional tissues grown in vitro. The field of cardiology is poised for
a revolution in which cell-based methods will be used to re-grow healthy heart muscle after
myocardial infarction or in chronic heart failure. Cardiac stem cells (CSCs), are resident
stem cells within the heart with regenerative potential, show great promise for regenerative
therapy.
The present invention enables the potential repair of damaged heart muscle and/or valves.
We have discovered and developed a unique technique for the isolation of stem/progenitor
cells from human adult myocardial tissue. The isolated cells have the potential to regenerate
and repair human cardiac tissue and possibly other tissues. A novel "cocktail" of enzymes
and a unique isolation protocol which yields significant numbers of progenitor cells was
developed. The isolated human cells are characterized with the unique markers representing
cardiac cells with regenerative potential. The invention enables the freezing and banking of
cells for future needs. The significance of the findings enables for the first time the creation
of a human "cardiac stem/progenitor cell bank" which, in the future, will serve as an
autologous pool of stem/progenitor cells for transplantation and tissue repair without the risk
of rejection or autoimmune reaction, and without the need for lifelong immunosuppressive
therapy.
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Tel Hashomer Medical Research, Infrastructure and Services Ltd.
Tel: +972-3-5305998 Fax: +972-3-5305944
Diabetes Therapy:
Adult Human Cells Transformed into Insulin-Producing Cells
Both type 1 and type 2 diabetes are characterized by the loss and dysfunction of insulinproducing cells, also known as beta cells. Beta cells reside in the pancreatic islets, which
comprise less than 2% of the pancreas.
Type 1 diabetes is caused by the patient's own immune system destroying insulin-producing
cells in the pancreas. The ideal therapy for diabetes would mimic the two essential features
of normal beta cells: the ability to sense glucose continually coupled with intelligent and
appropriate release of insulin in response to changes in blood glucose. One solution could be
an implantable, glucose-sensing insulin delivery system. Our approach provides a method
for generating a robust number of pluripotent cells by inducing adult cells dedifferentiation.
Our recent studies demonstrate that pancreatic-specific transcription factors, previously
known to control organ differentiation in the embryo, also possess instructive roles in
diverting the developmental fate of adult liver cells along the pancreatic lineage. We have
demonstrated that transducing liver adult cells with a single gene in a replication-defective
adenovirus vector delivered ex-vivo, induced them to transdifferentiate into human beta
cells. The delivered gene showed that in mice it leads to failure of development of the entire
pancreas.
We have uncovered the mechanism of the pancreatic lineage activation in liver. We have
developed a unique methods inducing regulated pancreatic hormone production in nonpancreatic islet tissues.
We have previously shown that PDX-1 transcription factor, activates pancreatic lineage
development and insulin production following ectopic expression in liver. This findings
suggests a surprising degree of residual plasticity in adult liver cells. The activation of the
pancreatic lineage and function in liver (and additional extra-pancreatic adult tissues) is
protected by previous patents for cell therapy for the treatment of diabetes patients.
The potential of the current finding is that PDX-1 transcription fuctor induces hepatic dedifferentiation in each liver cell, but activates the pancreatic lineage only in a restricted
subpopulation of these cells. The other “de-differentiated cells” may have acquired
increased plasticity and the capacity of being converted into other tissues, such as muscle.
The present technology and conceptenable the use of PDX-1 as a novel inducer of adult
cells plasticity, as such can promote developmental redirection of adult tissues. The results
demonstrate that Pdx-1 facilitates MyoD dependent activation of the myogenic marker Mif5,
for muscle tissue differentiation.
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Tel: +972-3-5305998 Fax: +972-3-5305944
Regenerative potential of cells recovered form
Peripheral Blood of Adult people to treat Human Diseases
A novel proprietary technology to generate therapeutic quantities of stem and progenitor
cells from one unit of adult peripheral blood (BRGTM cells) was developed. This unique
regenerative cell composition, which contains all lineages of stem and progenitor cells, has
significant advantages for regenerative medicine applications and is produced by a simple
procedure, from an easily accessible tissue.
The technology mainly addresses the need for isolation and enrichment of cells with
regenerative potential from one unit of non-mobilized peripheral blood of healthy adult
people. The cell composition obtained contains endothelial, mesenchymal and
hematopoietic progenitor cells as well as monocytic cells which can differentiate into
various lineages. The production process is associated with significant increase in the
number of colonies with differentiation potential to these lineages, as compared to the
number of colonies in fresh peripheral blood, before the process. The resulting cell
population is termed Bioregenerative (BRg) cells according to their potential functional
capacity.
The BRg cells were also able to increase the blood flow in ischemic limbs of rats, in an
animal disease model of limb ischemia. The enrichment process is simple and includes
stages of gradient separation, adherence and culture with cytokine mixtures. It is also
possible to selectively enrich for defined cell populations according to the clinical indication
required. For example, certain conditions will enrich for endothelial progenitors while others
will enrich for hematopoietic progenitors. Moreover, we demonstrated the extraction of BRg
cells from peripheral blood of cardiac ischemic patients, but in smaller numbers than
extracted from healthy individuals. Such composition of several cell populations with
regenerative potential could be a better candidate for therapeutic applications than purified
single stem or progenitor cell population, which is isolated by different methods.
Currently we aim for clinical indication of treatment of cardiovascular ischemic diseases. In
addition, based on the therapeutic potential of the BRg cells, the company will soon offer to
healthy people a service for long term storage of cells from peripheral blood. The cells will
be extracted from one unit of peripheral blood with no prior mobilization or any other
treatment of the donor. The cells will be cryopreserved for future clinical needs of the
individual, in which cell therapy could be applicable. This storage service will be provided
in collaboration with "Taburit", which currently operates together with Sheba the family
cord blood banking program.
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