Gene and Stem Cell Therapy

Week 13-14 - Bioengineering and Biotechnology 1

Second Semester, SY 2023-2024
Intended Learning At the end of this lessons, students should be able

Outcomes to:

1 DIFFERENT FORMS Describe gene therapy and stem cell and its forms.

Understand the different approaches and types of both stem

2 TYPES & APPROACHES
cells therapy and gene therapy.

Assess the issues potential benefits and detriments to global

3 PROS & CONS
health.

4 CURRENT To discover recent studies and breakthrough on gene therapy and

BREAKTHROUGHS stem cells.

5 MORAL & FUTURE

Discuss the moral and ethical implications, identify the effects,
ISSUES particularly in the context of science, health and economy of
gene- & stem cell therapy and potential future impacts on
society, health and economy.
 1
Definition & History
❑ DNA Replication Mistake

GENE
❑ Gene Therapy
❑ How it Started?
❑ First Case

THERAPY
DNA REPLICATION MISTAKES

In humans, genes vary in size from a few

hundred DNA bases to more than 2
million bases.
If genes don't produce the right proteins
or don't produce them correctly, a child
can have a genetic disorder.
DNA replication is a highly accurate
process, but mistakes can occasionally
occur as when a DNA polymerase inserts a
wrong base. Uncorrected mistakes may
sometimes lead to serious consequences,
such as cancer.
WHAT IS GENE THERAPY?

Gene therapy is an experimental technique

that uses genes to treat or prevent disease.
It seeks to alter genes to correct genetic
defects in order to prevent or cure genetic
diseases.

It is a medical approach that treats or prevents disease by correcting the

underlying genetic problem.

Gene therapy techniques allow doctors to treat a disorder by altering a

person's genetic makeup instead of using drugs or surgery.
HISTORY

In the 1980s, Scientists began to investigate gene therapy.

• Researchers would insert human genes into a bacteria cell.

• Then the bacteria cell would transcribe and translate the information into a protein.

• Then researchers would introduce the protein into human cells.

On January 19, 1989, the director of the National Institutes of Health (NIH), Dr. James A.

Wyngaarden, approved the first clinical protocol to insert a foreign gene into the immune

cells of persons with cancer (Roberts, 1989). On September 14, 1990, W. French Anderson

and his colleagues at the NIH performed the first approved gene therapy procedure on a

four-year-old girl born with severe combined immunodeficiency (SCID) (Anderson, 1990).
FIRST CASE

Despite the viral horror stories written by the popular media, this initial trial was largely a success,

and the most recent report on this individual in 2004 noted that she is thriving as an 18-year-old

teenager in suburban Cleveland (Springen, 2004). Over the next ten years, 300 clinical gene

therapy trials were performed on about 3000 individuals (McKie, 2000). The field was then

blackened with the death of an 18-year-old male four days after the introduction of 38 trillion

particles of recombinant adenovirus into his liver (Somia and Verma, 2000).

Despite this tragedy, researchers continue to move forward because of the great promise of novel

genetic treatments that, when perfected, will likely outshine current methods, such as protein

therapy or pharmacotherapeutics, for treatment of many diseases and defects.

HISTORICAL OVERVIEW
 2
Types & Approaches
❑ Types of Gene Therapy

GENE
❑ What are the Approaches
of Gene Therapy
❑ How does Ex Vivo Works?
❑ How does In Vivo Works?
❑ Types of vectors

THERAPY
TYPES OF GENE THERAPY

SOMATIC CELL GENE THERAPY (SCGT)

the transfer of a section of DNA to any cell of the body that doesn’t produce sperm or
eggs therapeutic genes transferred into the somatic cells of the patient:
such as, bone marrow cells, blood cells, skin cells, etc. and hence the new DNA doesn’t
enter the eggs or sperm any such modifications affects the individual patient only…and
are not inherited by the offspring (are not passed on to the patient’s children).

GERMLINE GENE THERAPY (GGT)

The transfer of a section of DNA to cells that produce eggs or sperm (therapeutic genes
transferred into the germ cells) involves the modification of germ cells (gametes) that will
pass the change on to the next generation (inheritable by offspring) for safety, ethical and
technical reasons, IT IS NOT attempted at present Countries prohibiting the application of
GGT in human beings:
AUSTRALIA, CANADA, GERMANY, ISRAEL, NETHERLANDS, SWITZERLAND
 WHAT ARE THE APPROACHES OF GENE THERAPY
1. EX VIVO 2. IN VIVO
❖transfer of genes in cultured ❖introduction of therapeutic gene
into the vector injected directly to
cells and will be reinjected to the body.
the body of the patient. ❖Involves direct delivery of the
Performed with the genetic therapeutic gene into the target
alterations of patient’s target cells of a particular tissue of a
patient
cells happening outside of the
body in a culture. Target cells ❖Potential tissue candidates
from the patient are infected include liver, muscle, skin, spleen,
with a recombinant virus lung, brain and blood cells.
containing the desired ❖Can be carried out using (1) viral
therapeutic gene. These or (2) non-viral vectors
modified cells are then
reintroduced into the patient's ❖Success depends on:
body, where they produce the ❖efficiency of the uptake of the
therapeutic gene by the target cells
needed proteins that ❖intracellular degradation of the
correspond to the inserted gene and its uptake by nucleus
gene. ❖the expression capability of the
gene.

The ff. are steps of EX VIVO Gene Therapy

1. Treatment begins by isolating cells with genetic defect from a patient.
2. These cells are then grown in culture in the laboratory.
3. Therapeutic gene will be introduced into the cultured cells using a virus called Vector. The virus’s DNA is taken out and
replaced with the one that treats the patient.
4. Patient’s sample target cells are now genetically altered with therapeutic genes and reinjected into the patient’s body.
 TYPES OF VECTORS
To transfer the desired gene into a target cell, a carrier is required.
VECTORS are responsible for gene delivery.
2 main classes :
Non-Viral vectors
Viral Vectors
Naked DNA or DNA complexes
Recombinant viruses (sometimes called
Non-viral vectors are currently being
biological nanoparticles)
evaluated for long-term expression of the
therapeutic genetic material.
Viral vector gene delivery has been
The most actively researched non-viral
utilized in a number of gene therapies due
vectors include
to the virus’ natural ability to access the
chemical disruption
cells of the body.
electroporation, and
polymer-based vectors
a) Retro Viruses
a) Naturally Occurring compounds
b) Adeno Viruses b) Enhance delivery by physical methods
c) Adeno Associated Viruses c) Enhance delivery by chemical methods
d) Herpes Simplex Virus
WHAT ARE VIRUSES?

Replicate by inserting their DNA into a host

cell

Gene therapy can use this to insert genes

that encode for a desired protein to create
the desired trait

4 different types of VIRAL VECTORS

• Retroviruses
• Adenoviruses
• Adeno-associated viruses
• Herpes Simplex viruses
 TYPES OF VIRUSES USED FOR VIRAL VERCTOR

Virus Name Uses/ Examples

RETROVIRUS It is a kind of virus that inserts a copy of its RNA genome into the DNA of a host cell
that it invades, thus changing the genome of that cell
It is a virus composed of RNA and not of DNA
Have an enzyme, called REVERSE TRANSCRIPTASE, that gives them the unique
property of transcribing their RNA into DNA after entering a cell.
The retroviral DNA can then integrate into the chromosomal DNA of the host cell, to
be expressed there.
HIV is a RETROVIRUS.
ADENOVIRUS These are a group of common viruses that infect the lining of your eyes, airways and
lungs, intestines, urinary tract, and nervous system
They're common causes of fever, coughs, sore throats, diarrhea, and pink eye.
Infections happen in children more often than in adults, but anyone can get them.
Target cell’s Immune response is HIGH.
The common cold is an ADENOVIRUS.
 TYPES OF VIRUSES USED FOR VIRAL VERCTOR

Virus Name Uses/ Examples

ADENO- These are small (25 nm) viruses that infect humans and some other primate species.
ASSOCIATED Small, single stranded DNA that insert genetic material at a specific point on
VIRUSES
chromosome 19.
(AAV)
From parvovirus family- causes no known disease and doesn't trigger patient immune
response.
Low information capacity: gene is always "on" so the protein is always being
expressed, possibly even in instances when it isn't needed.
Target cell’s immune response is VERY LOW
HERPES Double stranded DNA viruses that infect neurons; contagious and can be transmitted
SIMPLEX from person to person through direct contact
VIRUSES Example:
human herpes viruses; herpes simplex virus type 1 (HSV-1) – highly contagious;
herpes simplex virus type 2; varicella-zoster virus; cytomegalovirus; Epstein-Barr
virus; human herpesvirus 6; human herpesvirus 7; Kaposi’s Sarcoma herpesvirus
 VIRAL VECTOR USED IN COVID19 VACCINES
Using DNA

For COVID-19 viral vector vaccines, the vector (not the virus that causes COVID-19, but a different, harmless virus) will enter a cell in our
body and then use the cell’s machinery to produce a harmless piece of the virus that causes COVID-19. This piece is known as a spike
protein, and it is only found on the surface of the virus that causes COVID-19.
The cell displays the spike protein on its surface, and our immune system recognizes it doesn’t belong there. This triggers our immune
system to begin producing antibodies and activating other immune cells to fight off what it thinks is an infection.
At the end of the process, our bodies have learned how to protect us against future infection with the virus that causes COVID-19. The
benefit is that we get this protection from a vaccine, without ever having to risk the serious consequences of getting sick with COVID-19.
Any temporary discomfort experienced after getting the vaccine is a natural part of the process and an indication that the vaccine is
working.
 OTHER VECTORS IN COVID-19 VACCINES
Other vaccines for Covid-19 uses nanotechnology, and
traditional approach (Inactivated virus of SARS-CoV-2)

Using Inactivated Virus

Using mRNA (Traditional Vaccine)
NON-VIRAL VECTORS
Direct introduction of therapeutic DNA
But only with certain tissue
Requires a lot of DNA
Creation of artificial lipid sphere with aqueous core, liposome
Carries therapeutic DNA through membrane
Chemically linking DNA to molecule that will bind to special cell receptors
DNA is engulfed by cell membrane
Less effective
Trying to introduce a 47th chromosome
Exist alongside with the 46 others
Could carry a lot of information
But how to get the big molecule through membranes?
 3
Status & Issues
❑ Current Status

GENE
❑ Issues on Gene Therapy
❑ First Gene Edited Babies

THERAPY
CURRENT STATUS OF GENE THERAPY
FDA hasn’t approved any human gene therapy product for sale
Reasons:

In 1999, 18-year-old Jesse Gelsinger died from multiple organ failure 4 days after
treatment for omithine transcarboxylase deficiency.

Death was triggered by severe immune response to adenovirus carrier January

2003, halt to using retrovirus vectors in blood stem cells because children
developed leukemia-like condition after successful treatment for X-linked severe
combined immunodeficiency disease.
CURRENT STATUS OF GENE THERAPY

Published in 2021.
Gene therapy can be done in either somatic or germline cells. In somatic cells, gene
therapy only the modified tissues will be affected, but in germline cell gene therapy,
genetic changes transmit to the offspring. So, there is no clinical trial on human germline
gene therapy. Currently, somatic gene therapy is safe for the management of several
disorders in human beings.
Gene therapy effectively treats several diseases due to increased understanding of disease
pathogenesis and improved gene delivery technologies.
ISSUES ON GENE THERAPY

Short Lived
Hard to rapidly integrate therapeutic DNA into genome and rapidly dividing
nature of cells prevent gene therapy from long time
Would have to have multiple rounds of therapy
Immune Response
new things introduced leads to immune response
increased response when a repeat offender enters
Viral Vectors
patient could have toxic, immune, inflammatory response
also may cause disease once inside
Multigene Disorders
Heart disease, high blood pressure, Alzheimer’s, arthritis and diabetes are hard
to treat because you need to introduce more than one gene.
May induce a tumor if integrated in a tumor suppressor gene because
insertional mutagenesis
 THE FIRST GENE EDITED BABIES

He Jiankui and his team had used the CRISPR-Cas9 gene-editing tool on twin girls when they were just embryos, resulting in the birth
of the world’s first genetically modified babies. A third gene-edited child was born a year later. However, Chinese existing regulation,
thought not very detailed, does not provide legal basis for the experiment carried out by He Jiankui and his team (Nie, 2018; Nie &
Cheung, 2019). In particular, the 2003 “Ethical Guiding Principles for Research on Embryonic Stem Cell” issued by China's Ministry of
Science and Technology and then Ministry of Health (now National Health Commission), very clearly bans research to be performed on
human in vitro embryos after the 14th day of existence, and its subsequent implantation into a human uterus. In 2021, China also
made human germline editing for clinical use a crime. He’s original goal was to use gene editing to attempt—many call this a live
human experiment—to rewrite the CCR5 gene to create resistance to HIV. He says the genes were edited successfully and believed it
gave the babies either complete or partial HIV resistance because of the mutation.
The new rules, formally called the Measures for Ethical Review of Life Sciences and Medical Research Involving Humans, aim to
strengthen a host of existing guidelines and rules, says Ruipeng Lei, a bioethicist at Huazhong University of Science and Technology.
THANK YOU