Gene therapy and other novel therapies for haemophilia

Question 1

What are the current limitations of haemophilia replacement therapy?

Answer 1

Poor pharmacokinetics
Peaks and troughs
Antibody inhibitor development
IV administration
Short half life

Question 2

What are the aims behind new techniques?

Answer 2

Extend half lives
rebalance coagulation
substitute of factors
gene therapy

Question 3

What is pegylation?

Answer 3

Addition of a long inorganic molecule which prevents clearance by the kidney due to size

Question 4

WHat are the problems with pegylation?

Answer 4

They may interfere with the factor

There could be unwanted effects

Question 5

What is the effect of pegylation on half life?

Answer 5

~1.5x control

Question 6

What are the different PEGs available?

Answer 6

Peg: Bay 94
Bax855
N8 - glycan linked
Glycopegylated FIX N9-GP

Question 7

Is pegging safe?

Answer 7

Not sure yet as still relativelynew could have bad effects later
there has been some evidence which suggests that foamy macrophages form at high doses in toxicity models but lots of pegs are approved and have no problems.

Question 8

What is albumin fusion for FIX?

Answer 8

Addition of an activation peptide to the factor which is cleaved when FIX is activated
reduced doses from twice per week to once weekly

(Santagostino E et al 2012)

Question 9

What is Fc Fusion?

Answer 9

Linking of a factor to an Fc so it is covalently linked to IgG
It binds to the Fc receptor on the endothelial cells and is recycled and avoids lysosomal degradation which prolongs its half life

Question 10

What effect does Fc Fusion have?

Answer 10

Increase of half life by 1.7 fold
no adverse effects and no antibodies

(powell Js et al, 2012)

Question 11

What are the pros and cons for prologing half life?

Answer 11

Higher trough levels which means there are less spontaneous bleeds and more freedom with activity
It extends the dosing interval so greater quality of life for patient

Question 12

Why is prolonging halflifes more effective in FIX?

Answer 12

In FVIII the half life is already prolonged because it is bound by vwf, this determines its half life

Question 13

What is a bispecific antibody and how may it treat haemophilia?

Answer 13

It has two molecular targets
FVIIIa has the role of bringing FIX and FX together, a bispecific antibody can mimic this
Works even in those with inhibition

Question 14

What is the study that investigated the bleeding rate of ACE910

Answer 14

Oldenburg J et al. N Engl J Med 2017

Question 15

What is the pharmacokinetics of ACE910?

Answer 15

There are peaks and troughs but this is much less dynamic

ACE910 can be administered monthly

Question 16

What is gene therapy?

Answer 16

Delivery of nucleic acid polymers in cells to treat disease

Question 17

why is haemophilia an attractive case for gene therapy?

Answer 17

There is one abnormal protein, knock this out and switch with the good one

• single gene disorder
• simple loss of expression
• level of expression is unlikely to be critical
• specific tissue/cells is unimportant as it is simply secreted into the plasma
• the effect can be easily quantified and monitored

Question 18

Why are the following less attractive for gene therapy?

Answer 18

Beta thal - the level of expression is important
Antitrypsin/hereditary amyloidosis - dominant effect toxic gene product
Huntingtons/marfans - cells inaccesible, also dominant
Hereditary cancer syndromes - gain of function multiple tissue and all cellular regulation is bad

Question 19

What is the mechanism of gene therapy?

Answer 19

Cells are taken from the patients, they insert a virus containing the genome which depending on the vector can integrate into the genome, this is then transfered back to the patient who produces cells now containing the corrected gene

Question 20

What is the effectiveness of this data for gene therapy?

Answer 20

There is only small increases of production of factor however this is massive in terms of haemophilia as they are no longer characterised as “severe” - increased up to 6%

Question 21

What was the pauda gene?

Answer 21

This is FIX pauda gene therapy for Haemophilia B and gave a higher level of FIX but had thrombotic tendency in haemophiliacs

Question 22

What are the issues with gene therapy for haemophilia A?

Answer 22

The FVIII gene is much bigger and more immunogenic
But they can remove the B domain as this isnt functionally important in FVIII and this can just about fit into an adenovirus once removed

(biomarin trial 2017)

Question 23

What is the evidence for the use of gene therapy in haemophilia A

Answer 23

Slow start of trend for FVIII production but once gets going can overshoot the normal levels
significantly reduced bleeding
(Rangarajan 2017 )

Question 24

What happens to the immune response with gene therapy?

Answer 24

May have viral activation of MHC to cause cytotoxicity leading to reduced expression - steroids to treat this
Immunity to AAV- this is rare
Malignancy - AAV can integrate into the genome which causes DNA instability - although rare
Inhibition formation - also unliekly but lack of clinicla trial data

Question 25

What is gene editing?

Answer 25

Alters the existing allele however this is preferable when there is heterozygosity - the abnormal gene is deleted doesn’t really work in haemophilia as they are x linked
You can cut out the defective gene and insert a functional one

Question 26

How is dna cut and what is the effect?

Answer 26

CRISPR mechanism can target and cut the sequence
Effects only seen if there is partial hepetomy as this means when liver regrows it can regenerate normal gene carrying cells

Question 27

What are the issues that remain with gene therapy for haemophilia?

Answer 27

The duration of effect
Immunity - some patients may be immune to AAV
Mutagenesis - AAV integration and off target editing
early hepatitis?