3. The beginning…
• The first link between genetic inheritance
and a human condition was made in 1902
when alkaptonuria was identified as an
inherited disorder. (51 years prior to
Watson & Crick discovering DNA)
• Alkaptonuria is a condition in which urine
turns black on exposure to air.
• Sir Archibald E. Garrod (1857–1936) related
this and similar diseases to a lack of
particular enzymes in the body.
4. Causes
Genetic defects can be due to problems with
an entire chromosome, multiple genes, or
even a single gene (section of a
chromosome).
The human –Globin gene is 626 base pairs
long. If a single base substitution mutation
occurs in a particular place, the person will
develop Sickle Cell Anaemia.
Eg: small section of gene
ACT CCT GAG GAG AAG (unaffected person)
ACT CCT GTG GAG AAG (sickle-cell individual)
6. The following conditions can be tested by conductiong a
simple heel-prick test on a baby 2-3 days after birth,
1. Phenylketonuria (PKU) is a disorder characterized by an inability to
produce the enzyme phenylalanine hydroxylase, resulting in a
potentially fatal or damaging build up of the amino acid
phenalalanine, and occurs in one in every 10000 babies.
1. Hypothyroidism is a disorder caused by a small or improperly
functioning thyroid gland, or even its complete absence, and occurs in
one in every 3500 babies.
2. Galactosaemia is an inherited disorder and occurs in one in every
40,000 babies. Lactose is digested into galactose and glucose. A baby
with galactosaemia lacks the enzyme that metabolises galactose and
will die if untreated because of the build-up of galactose in the blood.
3. Cystic fibrosis (CF) is an inherited disorder and occurs in one in every
2500 babies. A person with CF produces abnormal secretions that
have a serious adverse effect on the function of lungs and digestion.
Recent advances in treatment have greatly improved the prognosis for
these babies so early diagnosis and treatment are important
8. PKU – a case study
• The aforementioned conditions are tested for at birth as early
detection is essential in successful treatment.
• Individuals born with PKU can’t produce phenylalanine hydroxylase
(an enzyme).
• Many products contain the amino acid phenylalanine, which can’t be
broken down, the build-up starts to destroy brain tissue, thereby
having a detrimental effect on development.
• Initial detection technique was the “wet nappy” test using ferric
chloride but this often occurred too late
9. PKU – a case study
• An alternative technique was the Guthrie
plate. Blood was collected via the heel prick
test.
• Foetal blood was tested for high levels of
phenalalanine. Using bacteria that would
only grow under certain concentrations
• Blood is still collected via a heel prick test
but can be tested far more efficiently via
mass spectrometry (a biochemical analysis
of the make-up of the babie’s blood)
11. Gene therapy
• Many genetic diseases are caused by a single gene.
• Eg: PKU
• this single gene defect, could be „treated‟ by consuming a special diet.
• This approach is not possible with many conditions.
• Why not try to insert a normal functional gene into cells
that contain a defect?
• Such a procedure is called gene therapy and is a medical procedure
that modifies the genetic material of living cells of an individual so that a
genetic defect is corrected.
13. Transferring a piece of DNA into a cell
• The functional piece of DNA inserted into a cell is specially
prepared and is called a cloned gene.
• How does the cloned gene, get into a patient‟s cells?
• This action is most successful when a vector is used to
carry the gene into a cell.
• The most commonly used vectors for gene therapy are
modified viruses. (Retroviruses and adenoviruses are
the main types used.)
14. Gene therapy using a
retrovirus vector:
• Some viruses have DNA, some have RNA,
Retroviruses have RNA that codes for DNA
once it enters a cell
• 1 – remove harmful RNA but leave bits that
code for infection and transformation to DNA
• 2 – insert RNA version of normal gene in to
virus
• 3 – introduce virus to cells ex-vivo and allow
cells to replicate
• 4 – inject infected cells in to patient
15. Gene therapy using an adenovirus
• Adenoviruses contain DNA, in to which a normal copy of a
gene can be integrated
• When the adenovirus enters the host cell, its DNA enters the
nucleus and starts coding for the required proteins.
• Adenovirus DNA does not integrate with host DNA, so when
mitosis occurs, it does not replicate.
17. Success of gene therapy
• Many successful experiments have been conducted with animals such as
mice…
• Still very experimental in humans and can pose risks to patients‟ safety.
• Non-viral gene therapy would be very difficult as this is one of the few ways
of bypassing our immune system
18. Screening during Pregnancy
Ultrasound is commonly used to view the
uterus and fetus during pregnancy.
Scans are often performed at 18 to 20 weeks into a
pregnancy.
Ultrasound maybe performed earlier if there are signs
or indications that the fetus is not growing normally.
Ultrasound is also used
to diagnose:
multiple pregnancies
(twins, triplets)
gross fetal abnormalities
such as trisomy
(e.g. Down, Klinefelter,
and Turner syndromes)
19. Other types of pre-natal testing
CSV (Chorionic Villus Sampling)
Performed at 6-8 weeks
Material gathered can be used for
both metabolic and genetic testing
Amniocentesis
Amniotic fluid contains both skin cells
and urine
Thin tube extracts fetal tissue
Material gathered can be used for
both metabolic and genetic testing
Miscarriages are often not random
occurrences but actually the result of the
foetus dying doue to a genetic
abnormality
Amniotic fluid
21. Electrophoresis
• DNA collected during screening procedures can be cut in specific
locations using REs (restriction enzymes).
• The DNA can be run through an agarose gel. A current is put through
the gel.
• DNA is negatively charged and will be drawn towards the positive
terminal.
• Smaller fragments will move faster while larger fragment will take more
time to move through the gel
• Every person has 2 copies of every gene and depending on the
abnormality, these can be observed on the gel
22. • In this case the disease form of the gene is smaller than the normal
version
• The gel shows that both parents were carriers for the condition and that
their first child received each parent’s diseased gene and therefor
developed the condition.
• Fortunately their unborn child received both normal versions of the gene
24. Genetic disease vs predisposition
DISEASE
• If you have two affected copies the Huntingtons gene you are
guaranteed to get the disease.
• Onset is usually around the age of 40 and manifests in fairly rapid
brain degeneration. There is no currently available treatment
PREDISPOSITION
• If you have affected copies of the BRCA1 or BRCA 2 genes for breast
cancer, there is no guarantee that you will get the disease.
• You have a much higher chance of it developing it, but the right
environmental conditions are required to trigger its onset
25. Genetics and Disease
Down syndrome is often
characterized by
impairment of cognitive
ability and physical
growth, as well as some
very recognizable facial 21
features
It is caused by an individual
being born with a 3rd copy
of chromosome 21, due to a
separation error during
meiosis.
Photo: Art Today
Occurrence is estimated at
approximately 1:750 births.
26. Who should be screened
• Anyone with a history of genetic disease
• Two unaffected individuals who have had an affected child are
both carriers of a recessive disease.
• Their chances of having another affected child are 1 in 4
• An affected individual with a dominant disease has one or
both of their alleles carrying the disease
• They have a 1 in 1 or at best a 1 in 2 chance of having an
affected child
29. Rational drug design
• When our immune system fights off the flu it develops a memory of
it, the next time we encounter the same strain we may not even
develop symptoms before it is fought off.
• The problem is that new strains are constantly appearing, so the
process must be repeated time and time again.
• Two surface proteins on influenza virus:
• Haemagglutinin is active in gaining entry to a cell.
• Neuraminidase allows the exit of new virus particles from a
cell, freeing them to infect other cells.
30. Rational drug design
• Neuraminidase is an enzyme that varies in structure from
strain to strain.
• Examination of strains of the virus from past years demonstrated
that although most of the molecule changed
dramatically, one small part remained constant.
• Fortunately, this non-variable part is the active site of the
enzyme.
31. Using the active site
• If a drug was to be designed to inhibit
the active site of neuraminidase,
• the molecular structure of the site
had to be worked out.
• This was done by computer modelling so
that the active site‟s exact shape and the Computer
representation of the
spatial arrangements of the atoms anti-flu drug in the active
surrounding it became known. site of neuraminidase.
32. Designing the drug
• Once the detailed structure of the active site was
known
• a molecule could be designed to fit and bind to
the active site,
Photo: CDC
• Then an anti-flu drug was created!
• This technique, in which the active site of a
molecule is determined and a second molecule
(the drug) is constructed to fit into that active site
to inhibit the activity of the first molecule, is
called rational drug design.
35. Types of Vaccine
There are two basic types of vaccine:
subunit vaccines and whole-agent vaccines.
Recombinant
vaccines
Subunit Vaccine
Contains some part or Toxoids
product of micro-
organisms that can
Conjugated
produce an immune
response
vaccines
Acellular
vaccines
Attenuated
Whole-Agent (weakened)
Vaccine
Contains
whole, nonvirulent
microorganisms Inactivated
(killed) ie. just antigen
37. Malaria
There are a number of strains, all carried by the Anopheles mosquito
1/3 of the world‟s population is at risk. Every year there are 4 million new cases
and 2 million deaths
Involves a very complex life cycle
Illness mainly due to:
Capillaries becoming blocked by “sticky protein” on red blood cells
Release of the toxin GPL
Healthy blood cells being overtaken by the parasite
Only Plasmodium Faliciparum curable as in other strains parasites lay
dormant in gut
Resistance
Mosquitoes becoming resistant to DDT
Plasmodium becoming resistant to Chloroquine
38. Developing Vaccines
MALARIA
Developing a vaccine is difficult due to Malarial DNA constantly
changing and therefore its recognizable protein coat aswell.
A potential vaccine could pre-expose people to a microscopic
quantity of the GPL toxin
Another option is to create an inhibitor to block the receptor site
on the “sticky protein” so that it is unable to bind to tissue and
block capillaries
MEASELS
Still a major cause of death in developing countries due to
unavailability of vaccine and difficulty in storing it
Current trials have spliced Measles antigen into a strain of lettuce
Lettuce is freeze-dried in to powder and then packaged in to
tablets
Cheap and does not require refrigeration
40. Manufacturing biological molecules
Insulin:
• produced by the pancreas, a hormone that controls
the level of glucose in the blood by controlling its
uptake from the blood by cells…
• A deficiency of this hormone results in an
abnormally high level of glucose in the blood -
diabetes.
• Because the amino acid sequence of the active
molecule of insulin is known, a piece of DNA
carrying the instructions (code) for insulin can be
built and inserted into a plasmid vector.
42. Manufacturing biological molecules
Application for manufactured molecules
Insulin (for diabetics)
Growth hormone
Factor VIII (blood clotting agent missing in hemophiliacs)
Advantages
Can be produced in very large quantities
If viral antigen can be copied, can be produced using no materials
of human biological origin – minimises risk of associated disease
Eg. in the past haemophiliacs receiving transfusions of Factor VIII
from blood risked HIV
Eg. patients receiving growth hormone from cadavers risked the
brain disease CJD
44. Manufacturing biological molecules
Very small human-made particle (0.1-100 nM in diameter)
Being used to deliver drugs directly to the cells requiring them
In cancer trials with mice, survival chances increased by 30%
Made up of 3 parts
Fluorescent stain – to follow progress
Methotrexate – a drug that destroys cancerous cells
Folic Acid – a vitamin required by rapidly reproducing cells
