Emerging Treatments

Question 1

What are inborn errors of metabolism?

Answer 1

Largest group of genetic disease

Affect basically all pathways, carbohydrate, fatty acid and protein metabolism

Usually a result of a lack of an enzyme

Eg. PKE, MCAD deficiency, maple syrup urine disease

Decreases the levels of a useful product. Also gives an increase in the substrate. This may be pushed down a different pathway

Question 2

What is PKU?

Answer 2

Phenylketonuria

Lack of enzyme phenylalanine hydroxylass

Means no phenylalanine —> tyrosine

Phenylalanine is instead pushed down another route to turn into phenylketones

Symptoms: cognitive imparement, behavioural difficulties, fairer skin hair and eyes, vomiting

Question 3

How is PKU treated?

Answer 3

Treated with a low protein diet

Also tyrosine supplements

Both ideally started before symptoms develop

Screening is now available

Question 4

What is haemophilia?

Answer 4

Blood clotting disorder

Symptoms: uncontrolled bleeding, pain from bleeding into joints, bleeding into brain

Fatal if untreated

Question 5

What are treatments for haemophilia?

Answer 5

First diluted snake venom was used, then whole blood transfusion but this required very large volumes

Infusions of factor VIII in plasma (also needs very large volumes)

Factor VIII concentrate became available and was turned into a freeze dried plasma derived factor concentrate

Now we use recombinant factor VIII

However this led to the blood scandal where up to 30000 people were given HIV infected blood

Question 6

What are some diseases treated by gene replacement?

Answer 6

Growth hormone deficiency - treated with recombinant growth hormone

Haemophilia - recombinant FVIII

Lysosomal storage diseases: fabry disease- injection of recombinant alpha galactosiadass A , pompe disease - injection of alpha glucosidase

Question 7

What do pharmacological therapies targeting proteins do?

Answer 7

Treat the underlying condition not the symptoms

Are treatments not cures

They normalise the function of the mutant protein

Question 8

What are pharmacological chaperones?

Answer 8

Protein folding is complex and sometimes fails

System in ER degrades misfolded proteins

Some mutations prevent proteins folding properly, these are subject to the degradation pathway

Pharmacological chaperones treat these

Question 9

What is Fabry disease?

Answer 9

Deficiency in alpha galactosidase A

Causes a build up of globotriaosylceramide

Some mutations cause misfolding

Migalastat is a small molecule chaperone that stabilises the enzyme in its correct shape

Migalastat is mutation specific

Question 10

What are pharmacological modulators?

Answer 10

Most commonly used drugs are these

They are receptor agonists or antagonists

Or ion channel activators or blockers

Eg used in some cystic fibrosis people

Question 11

How are pharmacological modulators used to treat cystic fibrosis?

Answer 11

CF may be caused by a defective chloride ion channel

Some mutations (33) cause this channel not to open

So a drug was designed that caused this channel to open - ivacaftor

Ivacaftor is mutation specific

Question 12

How is combination therapy used to treat some CF cases?

Answer 12

some people have a defective ion channel and a misfolding protein caused by the same mutation (f508del)

This is treated with a combination of a chaperone and an activator

Eg. Orkambi (ivacaftor/lumacaftor)

Not a cure but does improve lung function

Question 13

What are drugs that treat stop codon read through?

Answer 13

Some diseases are caused by a non sense mutation. This introduces a premature stop codon

This stops protein production and gives unusually short proteins

Drugs can be used to prevent this

Question 14

What is an example of a drug that treats stop codon read through?

Answer 14

Aminoglycoside anyobiotics bind to ribosome

This causes a mis translation

But drugs can be based on these so that non sense mutations are actually read through

This produces a full length protein as opposed to a truncated one

Question 15

How are stop codon read through drugs used to treat duchenne muscular dystrophy?

Answer 15

DMD is caused by a premature stop codon that produces truncated dystrophin

Becker muscular dystrophy has dystrophin with a missing section

If a drug can cause readthrough of a premature stop codon, it can give a phenotype for DMD that is more like BMD

Ataluren does this

A treatment not a cure

Non sense mutation specific

Question 16

What are some types of drugs, broad catagories?

Answer 16

Pharmacological chaperones

Pharmacological modulators

Prevent stop codon read through

Question 17

What is gene therapy?

Answer 17

Conceptually:

Recessive - replace the defective gene

Dominant - dlte the defective gene

But in practice very difficult to achieve :
Specificity, getting the therapy to the right place, maintain expression

Much easier in vitro than in Vivo

Question 18

How is gene therapy used to treat mitochondrial my inherited disease?

Answer 18

Ex Vivo

Requires IVF

Thank dna (spindle) from an unfertilised patient egg. Transfer dna to a donor egg that had normal mitochondria. Then fertilise and implant the new egg

Or take pro nucleus from fertilised patient egg, transfer this to a fertilised donor egg

Question 19

What is viral gene therapy?

Answer 19

Ex Vivo

Can engineer a virus to carry the therapeutic gene

Wide variety of virus used: AAV, adenovirus, lentivirus - HIV, vaccinia

Virus choice depends on the target tissue (viral tropism) and the amount of DNA

Question 20

What is SCID?

Answer 20

Severe combined immune deficiency

Patients lack both B and T cell responses

Most common is X linked (75%) also adenosine deaminase deficiency (15%)

Can now be treated with a bone marrow transplant but this isn’t always possible and comes with its own complications

Question 21

How is gene therapy used to treat SCID?

Answer 21

In vitro

Treats ADA-SCID

Called strimvelis

Autologous transplant:

Take bone marrow from patient

Isolate haemapoietic stem cells

Isolate and expand CD34+ cells

Transfected with ADA-lentivirus and grow these cells

Treat patient with Bulfusan (kills HSCs)

Reinfuse transformed cells into patient

Question 22

What is in Vivo therapy supplement?

Answer 22

Lack of functional gene

A virus can be used to carry in a working copy

Can be injected systemically but more effective if injected into eye or spine or brain

Question 23

What is the only in Vivo therapy supplement in use?

Answer 23

Leber congenital amaurosis type 2

Recessive disease caused by mutation in RPE65

Causes progressive blindness due to loss of retinal cells

Treatment is called Luxturna, uses rAAV2 virus expressing RPE65 gene

Not a cure but greatly improves vision

Question 24

What are anti sense oligonucleotides?

Answer 24

Short modified nucleic acids complementary to the target

Modification allows entry to cell and prevents degradation

They bind to target and block translation

Can also alter splicing

Cheap to make

Aka knockdown approach

Question 25

What is an example of anti sense oligo nucleotide therapy?

Answer 25

In Vivo

Useful in treating diseases caused by gain of function

Inotersen:

Treat transthyretin-related hereditary amyloidosis

Caused by mutation in transthyretin (TTH)

TTH can’t form tetramers, forms aggregates

Mutation specific

Question 26

How can anti sense oligonucleotides be used for exon skipping?

Answer 26

During pre rna processing

Oligonucleotides can cause exon to be skipped ie. a disease causing exon can be skipped

However is useful in limited circumstances: the exon skipped must not be vital and it generally only work in large proteins

May be used in Duchenne muscular dystrophy - eteplirsen but not approved in Europe

Question 27

What is a possible future gene therapy?

Answer 27

CRISPR-Cas9

Clustered regularly interspaced short palindromic repeats

Derived from bacteriophages (viruses that infect bacteria)

Cas9 - crispr associated protein 9 - endonuclease

Recognised and clever DNA CRISPR hybrids

May be able to correct relatively small errors. May have off target effects

Not currently used in humans

Question 28

What is the one case in which CRISPR-Cas9 is used?

Answer 28

He Jiankui used CRISPR-Cas to alter genome in IVF embryos

He deleted CCR5 In twins, this is involved in the entry of HIV into cells

However this is controversial and illegal in most parts of the world