1 - BSMLS

CYTOGENETICS
TERM
MLS 102

BREAK
CLASS
LECTURE & LABORATORY NOTES
School: Lyceum of the Philippines University – Batangas Academic Year: 2020 - 2021
Program: Bachelor of Science in Medical Laboratory Science Professor: Deignielle Bert Arellano

S(Source: Powerpoint Presentation)

LESSON 16: GENE THERAPY

THE BEGINNING…

 In the 1980s, scientists began to look into

gene therapy.
 They would insert human genes
into a bacterial cell.
GENES  Then the bacterial cell would
transcribe and translate the
 Are carried on a chromosome information into a protein.
 The basic units of heredity  Then they would introduce the
 When altered, it causes dysfunction of a protein into human cells.
protein
 When there is mutation in the geen, then it THE FIRST CASE
will change the codon, which will change the
amino acid called for, changing the  The first gene therapy was performed on
conformation of the protein and its function September 14, 1990.
 Genetic disorder result from mutations in  Ashanti DeSilva was treated for
the genome SCID. (Severe combined
immunodeficiency)
WHAT IS GENE THERAPY?  Doctors removed her white blood
cells, inserted the missing gene
 It is a technique for correcting defective into the WBC, and then put them
genes that are responsible for disease back into her blood stream.
development.  This strengthened her immune
 There are four approaches: system.
1. A normal gene is inserted to  Only worked for a few months
compensate for a nonfunctional
gene. HOW IT WORKS
2. An abnormal gene is traded for a
normal gene.  A vector delivers the therapeutic gene
3. An abnormal gene is repaired into a patient’s target cell.
through selective reverse mutation.  The target cells become infected with the
4. Change the regulation of gene pairs. viral vector.
 The vector’s genetic material is inserted
into the target cell.
 Functional proteins are created from
the therapeutic gene causing the cell
to return to a normal state.

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VIRUSES ADENO-ASSOCIATED VIRUSES

 Replicate by inserting their DNA into a host  Small, single-stranded DNA that inserts
cell genetic material at a specific point on
 Gene therapy can use this to insert chromosome 19
genes that encode for a desired  From Parvovirus family- causes no known
protein to create the desired trait. disease and does not trigger patient
 Four different types: immune response.
1. Retrovirus  Low information capacity
2. Adenovirus  Gene is always “on” so the protein is
3. Adeno-associated virus always being expressed, possibly even in
4. Herpes simplex virus instances when it isn’t needed.
 Example: Hemophilia treatments - a gene-
RETROVIRUSES carrying vector could be injected into a
muscle, prompting the muscle cells to
 Creates double stranded DNA copies from produce Factor IX and thus, prevent
RNA bleeding.
genome  Study by Wilson and Kathy High
 The retrovirus goes through reverse  (University of Pennsylvania)
transcription using reverse transcriptase  Patients have not needed Factor IX
and RNA. injections for more than a year.
 The double stranded viral genome
integrates into the human genome using HERPES SIMPLEX VIRUSES
integrase.
 Integrase inserts the gene anywhere  Double-stranded DNA viruses infect
because it has no specific site. neurons
 May cause insertional mutagenesis  Examples: Herpes simplex virus type 1
 One gene disrupts
another gene’s code
(disrupted cell division
causes cancer from
uncontrolled cell division).
 Vectors used are derived from the
human immunodeficiency virus (HIV) and
are being evaluated for safety.

ADENOVIRUSES

 Double – stranded DNA genome that NON-VIRAL OPTIONS

causes respiratory, intestinal, and eye
infections in hums  Direct introduction of therapeutic DNA
 The inserted DNA is NOT incorporated into o Only with certain tissues
the genome. o Required a lot of DNA
 Not replicated; has to be inserted when  Creation of artificial lipid sphere with
more cells divide aqueous core, liposome
 Example: Common Cold o Carries therapeutic DNA through
membrane

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 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.

NON-VIRAL OPTIONS

 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 the 46 others
 Could carry a lot of information

Alipogene tiparvovec, sold under the brand

name Glybera, is a gene therapy treatment
designed to reverse lipoprotein lipase
deficiency (LPLD), a rare inherited disorder which
can cause severe pancreatitis. It was
recommended for approval by the European
Medicines Agency in July 2012 and approved by
the European Commission in November of the
same year. It was the first marketing
authorization for a gene therapy treatment in
either Europe or the United States.

PROBLEMS WITH GENE THERAPY

 Short Lived
 Hard to rapidly integrate therapeutic
DNA into genome and rapid
CURRENT STATUS cell division prevents gene
therapy from long time
 Slow approval  Multiple rounds of therapy is
Reasons: needed.
 In 1999, 18 – year – old, Jesse Gelsinger  Immune Response
died from multiple organ failure 4 days after  New things introduced leads
treatment for ornithine transcarboxylase to immune response.
deficiency.  Increased response when a repeat
offender enters

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 Viral Vectors  In these patients, peripheral T cells
 Patient could have toxic, immune, were transduced with a vector bearing
inflammatory response. the gene for adenosine deaminase.
 May cause disease once inside  The experiment was extremely
 Multigene Disorders labor intensive, because mature
 Heart disease, peripheral-blood T cells were modified
hypertension, Alzheimer’s, arthritis rather than stem cells, and the procedure
and diabetes are hard to treat therefore had to be repeated many
because you need to introduce times to achieve success.
more than one gene.
 May induce a tumor if integrated in a SUCCESSFUL ONE YEAR GENE THERAPY
tumor suppressor gene because of TRIAL FOR PARKINSON'S DISEASE
insertional mutagenesis

UNSUCCESSFUL GENE THERAPIES  Neurologix, a biotech company

announced that they have successfully
 Jesse Gelsinger, a gene therapy patient completed its landmark Phase I trial of
who lacked ornithine transcarbamylase gene therapy for Parkinson's Disease.
(OTC) activity, died in 1999.  This was a 12 patient study with four
 Within hours after doctors shot the normal patients in each of three dose
OTC gene attached to a therapeutic escalating cohorts. All procedures were
virus into his liver, Jesse developed a high performed under local anesthesia and
fever. all 12 patients were discharged from
 His immune system began raging out of the hospital within 48 hours of the
control, his blood began clotting, procedure, and followed for 12 months.
ammonia levels climbed, his liver  Primary outcomes of the study design,
hemorrhaged and a flood of white blood safety and tolerability, were successfully
cells shut down his lungs. met. There were no adverse events
 One problem with gene therapy is that one reported relating to the treatment.
does not have control over where the gene
will be inserted into the genome. PARKINSON'S DISEASE
 The location of a gene in the genome is of
importance for the degree of  The gene transfer procedure utilized the
expression of the gene and for the AAV (adeno-associated virus) vector,
regulation of the gene (the so-called a virus that has been used safely in
"position effect"), and thus the gene a variety of clinical gene therapy trials, and
regulatory aspects are always the vehicle that will be used in all of the
uncertain after gene therapy. company’s first generation products,
including epilepsy and Huntington’s
SUCCESSFUL GENE THERAPY FOR SEVERE disease.
COMBINE IMMUNODEFICIENCY  In its Parkinson’s disease trial, Neurologix
used its gene transfer technology.
 Infants with SCID are unable to mount
an adaptive immune response, because RECENT DEVELOPMENTS
they have a profound deficiency of
lymphocytes.  Genes get into brain using liposomes
 Severe combined immunodeficiency coated in polymer called polyethylene
is inherited as an X-linked recessive glycol.
disease, which for all practical purposes o Potential treatment for Parkinson’s
affects only boys. disease
 In the other half of the patients with
severe combined immunodeficiency, the  RNA interference or gene silencing to treat
inheritance is autosomal recessive — Huntington’s
and there are several abnormalities  siRNAs used to degrade RNA of particular
in the immune system when the sequence.
defective gene is encoded on an  Abnormal protein will not be produced.
autosome.  Creation of tiny liposomes that can carry
therapeutic DNA through pores of nuclear
SEVERE COMBINE IMMUNODEFICIENCY membrane
 Sickle cell disease successfully treated in
 A previous attempt at gene therapy mice
for immunodeficiency was successful in
children with SCID due to a deficiency of
adenosine deaminase.

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GENE EDITING

Would you want to have designer babies?

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