31.10.
2023
Types of stem cells:
3)Induced pluripotent (iPS)
stem cells
Induced pluripotent (iPS) stem cells
• The «deprogrammed «» cells are known as induced pluripotent stem
(iPS) cells because, in using this fairly simple laboratory technique to
return them to their undifferentiasted state (pluripotency) and could
do everything ES cells can do.
1
31.10.2023
Why IPS Cells?
ES cells do not match
the donor’s genetic makeup for therapeutic purposes
Historical Timeline Showing Events That Led To The Development Of IPS Cells
2
31.10.2023
Somatic-derived stem cells via nuclear transfer
Nucleus
directed differentiation
Fibroblasts from Enucleated
ES cell Progenitor Neuron
patients oocyte
• Create ES cells that match the donor’s genetic makeup for therapeutic purposes.
• ES cells derived from patients can be directed to differentiate into specific lineages (e.g.
dopaminergic neurons) to study a particular disease (e.g. Parkinson’s disease).
• This method may be used for cell-based therapies that would circumvent immune rejection.
Somatic cells can turn into embryonic state
(Prof Shinya Yamaka)
3
31.10.2023
Dr. Shinya Yamanaka, PhD
Dr. Kazutoshi Takahashi, PhD
Human iPSCs
Production of iPS cells
directed
iPS reprogramming differentiation
factors
ectodermal cell brain
iPS cell mesodermal cell heart
fibroblasts from
patients
endodermal cell
pancreas
8
4
31.10.2023
Induced pluripotent stem cells (iPS cells)
‘genetic reprogramming’
= add certain genes to the cell
cell from the body induced pluripotent stem (iPS) cell
behaves like an embryonic stem cell
differentiation
culture iPS cells in the lab
all possible types of
Advantage: no need for embryos! specialized cells
Induced pluripotent stem cells (iPS cells)
genetic reprogramming
cell from the body (skin) pluripotent stem cell
(iPS)
differentiation
5
31.10.2023
iPS cells - a recent advance
• IPS cells were first produced in 2006 from mouse tissue, and in 2007 from human.
• This is an groundbreaking advance in stem cell research, because it allows researchers
to obtain pluripotent stem cells, which are important in research and potentially
have therapeutic uses, without the controversial use of embryos.
• Reprogramming adult cells to obtain iPS cells may pose significant risks that could limit
their use in humans. If viruses are used to alter the cells’ genome, the expression of
cancer-causing genes or oncogenes may potentially be triggered after these cells are
introduced into animals.
11
Induced pluripotent stem cells (iPS cells)
• A type of pluripotent stem cell artificially derived from an adult somatic
cell by "forcing" expression of specific genes.
• iPS cells are believed to be similar to ES cells with respect to:
• stem cell gene and protein expression
• ability to differentiate into all lineages in vitro
• forming viable chimeras after injection into blastocysts or tumors
when transplanted into adult tissues
• potential to form an entire organism, such as a mouse
12
6
31.10.2023
Alternatives to hES Cells
Induced Pluripotent Stem Cells (IPS Cells)
• Induced Pluripotent Stem Cells (iPS)
• Uses Oct4, Sox2, Klf4 and c-Myc genes
• Show higher potency than cord cells
• Low efficiency (recent improvements)
• Other problems
• Abnormal aging
• Tumor production
• Incomplete reprogramming
7
31.10.2023
First Generation
• First generated by Shinya Yamanaka’s team at Kyoto University, Japan
in 2006.
• Four key pluripotency genes essential for the production of
pluripotent stem cells were used; Oct-3/4, Sox2, c-Myc and Klf4
• Retroviruses were used to transfect Mouse fibrblasts.
Pluripotency Factors
• C-Myc, Oct ¾ and SOX2 promote self-renewal and drive proliferation
in pluripotent cells.
• Sox2: Essential for embryonic development. Down regulation of this
factor leads to differentiation of ES and IPS cells.
8
31.10.2023
Limitations
• This IPSCs showed methylation errors compared to original patterns
in ESC lines and failed to produce viable chimeras if injected into
developing embryoes.
Second Generation
• In June 2007, by the same group.
• These cell lines were also derived from mouse fibroblast by retroviral mediated
reactivation of the four endogenous pluripotent factors, but instead of Fbx15, they
used Nanog which is an important gene in ESCs.
• DNA methylation patterns and producing viable chimeras (and thereby contributing
to subsequent germ-line production) indicated that Nanog is a major determinant of
cellular pluripotency.
• Nanog: In embronic stem cells, Nanog, along with Oct-3/4 and Sox2, is necessary in
promoting pluripotency.
9
31.10.2023
Limitations
• One of the four genes used (namely, c-Myc) is oncogenic, and 20% of
the chimeric mice developed cancer.
• (In a later study, Yamanaka reported that one can create iPSCs even
without c-Myc, although process takes londer and is not as efficient,
but he resulting chimeras did not develop cancer).
Complications
• Because of viral transfection systems, the created cells might be prone to
form tumors.
• However Konrad Hochedlinger and his Harward University research team
successfully used an adenovirus to transport the requisite four genes into
the DNA of skin and liver cells of mice.
• Since the adenovirus does not combine any of its own genes with the
targeted host, the danger of creating tumors is eliminated.
10
31.10.2023
Delivery Methods of iPSC Induction
11
31.10.2023
Alternative to “Therapeutic” cloning
transplantation
differentiated donor cells
embryonic stem cells
early embryo
egg
? skin cells
nucleus
Induced pluripotent stem (iPS) cells
transplantation
differentiated donor cells
iPS cells
blastocyst Oct4
Klf4 Sox2
c-Myc
enucleated oocyte
? adult cells
nucleus
12
31.10.2023
Gene repair in iPS cells
transplantation
differentiated donor cells
repaired iPS cells
by CHRISPER
gene targeting
Oct4
iPS cells
Klf4 Sox2
c-Myc
adult cells
Applications of IPS cells
13
31.10.2023
Regenerative Medicine Using IPS Cells
fffffffffffffffffffffffffffffffbbbbbbbbf
Regenerative Medicine Using IPS Cells
• Neurons - Parkinson’s Disease
• Retinal/Corneal Cells - Eye Diseases
• Heart Muscle Cells - Heart Diseases
• Neural Stem Cells - Spinal Cord Injury
• Platelets - Blood Transfusion
• Immune Cells - Cancer
• Cartilage - Arthritic Disorder
14
31.10.2023
Drug Development Using IPS cells
• Disease Modeling with Organoids
• Drug Discovery Applications
15
31.10.2023
Clinical Trails of IPS Cells
RPE: Retinal pigment epithelium
AMD: Age-related Macular degeneration
(World’s first operation with IPS cells)
http://msemporda.blogspot.com/p/rik-ip.html
16
31.10.2023
Clinical Trails of IPS Cells
RPE: Retinal pigment epithelium
AMD: Age-related Macular degeneration
(World’s first operation with IPS cells)
17
31.10.2023
•Print
Clinical Trails of IPS Cells
• A clinical trial to treat Parkinson’s disease patients with human induced pluripotent stem (iPS) cells.
• The studies will involve transplanting nerve cells produced by human iPS cells into the brains of
Parkinson’s disease patients. The trial is Japan’s third case of transplanting iPS cells into humans in a
clinical setting, following treatments for eye and heart disease patients.
July 30, 2018
http://www.asahi.com/ajw/articles/AJ201807300029.html
18
31.10.2023
Genome analysis for one patient was half million US dollars (for first clinical trial)
19
31.10.2023
Allogenic Cell Therapy with IPS cells
• Instead of each individual, healthy volunteers
• Allogenic transplantation – immune rejection problem
• Match HLA
• Super donors – IPS cell stocks
20
31.10.2023
21
31.10.2023
22
31.10.2023
Limitations of IPS cells
➢ iPS cells are not identical to human embryonic stem (hES)
iPS cell morphology, proliferation, surface antigens, gene expression, telomerase
activity, and the epigenetic status of pluripotent cell-specific genes are similar to
the same characteristics in embryonic stem cells but not identical.
➢There are even differences between IPS cells derived from the same patient.
human cells as DNA microarray analysis detected differences between the two
pluripotent stem cell lines (Takahashi et al. 2007); and (iii) iPS cells are derived
from somatic cells such as fibroblasts and peripheral blood cells.
23
31.10.2023
Limitations of IPS cells
❖Uncertainty about the functional equivalence to hES
Due to the lack of long-term follow-up studies;
❖the risk of tumourigenesis — as iPS cell teratoma formation can lead to tumour
formation
❖the risk of immunogenicity.
Technology innovations have reduced the risk of tumourigenesis (Yu et al. 2009;
Chen et al. 2011; Okita et al. 2011; Okita et al. 2013) and it was reported limited
immunogenicity of the transplanted cells differentiated from iPSCs and ES cells
(Araki et al. 2013)
24
