Engineering Antibodies

Question 1

What characteristics of antibodies make them such useful therapeutics?

Answer 1

1. They are specific
2. They are proteins - hopefully no immune response

Question 2

What regions do ximab antibodies have?

Answer 2

Consist of the mouse Fab regions fused with human Fc (they are 65% human)

Question 3

What do zumab antibodies have on them?

Answer 3

Mouse CDRs are engrafted into a human antibody (which is 95% human)

Question 4

What variations of an antibody can expression of natural monoclonal anitbodies give rise to?

Answer 4

Glycosylation
Charge
Structural stability

Question 5

What can glyco engineering do?

Answer 5

Ensure you only have certain sugar groups on glycosylated protein

Question 6

How can you go glyco engineering?

Answer 6

Engineering the cell that you are going to express the antibody in so it doesn’t have the enzyme to put the sugar on the glycine change or make sure you have an excess of the enzyme so everything is pushed in the other direction

Question 7

How can you link something to an anitbody through engineering?

Answer 7

Engineer additional cysteine residues - these normally make disulphide bonds

Question 8

Why is IgG4 unusual?

Answer 8

Has a weak heavy chain to heavy chain linkage, when it gets taken into a cell they can come apart

Question 9

What is pyroglutamation?

Answer 9

A common charge variant - from a point of purity or quality control, you don’t want this

Question 10

How can you prevent aggregation?

Answer 10

If you get charge changes, may not want to stay in solution or it may attract other molecules, so can change amino acids to bring it to its most stable structure

Question 11

Why are antibodies engineered?

Answer 11

Humanisation
Glycosylation
Half-life alteration
Conjugation capabilities
Hinge region disulphides
Complement fixation

Question 12

What can moving a sugar group off an anitbody change its ability to do?

Answer 12

Change its ability to trigger antibody dependent cellular cytotoxicity

Question 13

What happens when you add mannose groups to antibodies?

Answer 13

More complement mediated cytotoxicity

Question 14

What can changing CD-R regions do?

Answer 14

Change affinity of antibody
Changes ability to interact with complement especially with C1q and ability to interact with Fc receptors

Question 15

What is KIH engineering used for?

Answer 15

Used to generate bispecific antibodies

Question 16

What is KIH engineering?

Answer 16

Interactions between tyrosine (knob) and threonine (hole) residues, improves heterodimer formation

Question 17

How are CH2 domains brought together?

Answer 17

Through glycan chain interactions

Question 18

Why are bispecific antibodies useful>

Answer 18

You can link two thing together

Question 19

What are improved version of the same termed?

Answer 19

Bio better / me better antibodies

Question 20

How can you get biosimilars to market with 1/20th of the cost?

Answer 20

Once patent runs out, have it prove it has the same specificity as you have already done the safety

Question 21

What could happen if you increase drug to antibody ratio?

Answer 21

Improve potency, but not as good clearance , exposure, aggregation and toxicity

Question 22

What happens if you make different attachment sites?

Answer 22

Different binding
Different drug release rate etc.

Question 23

What are antibody—conjugate uses?

Answer 23

Radio-imaging/radiotherapy
Delivery of a cytokine to a specific cell type
Delivery of a drug or pro-drug to specific cells
Longevity of anitbody in serum

Question 24

What can you do with cytokines and antibodies?

Answer 24

Can have an anti-cytokine specific antibody

Question 25

What is the link between human serum albumin and antibodies?

Answer 25

Can link something that binds to HSA, gives it a bigger size so isn’t filtered out so quickly