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cysteines used in antibody conjugation

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nkhanbham

Master

Posts: 53

Joined: Wed Feb 25, 2015 3:03 pm

Post Tue Nov 07, 2017 6:37 pm

cysteines used in antibody conjugation

Does anybody know whether the cysteine groups used to bind the polymer in MaxPar conjugation are restricted to the heavy chain or also include ones on the light chain? There are internal disulfide bonds on both heavy and light chains and potentially both could be used. On all the diagrams I have seen the polymer is always bound to the heavy chain.
Is it correct that these are more likely to be reduced by TCEP while the ones in the hinge region connecting the two heavy chains are both (or at least one is) intact?
Thanks,
Naeem

(This may be a topiuc previously discussed - if so please redirect me to the thread)
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mleipold

Guru

Posts: 5792

Joined: Fri Nov 01, 2013 5:30 pm

Location: Stanford HIMC, CA, USA

Post Tue Nov 07, 2017 7:02 pm

Re: cysteines used in antibody conjugation

Hi Naeem,

Short answer: I'm not aware of anyone doing mass spec (digested peptide) to formally determine any and all points of attachment of the MAXPAR polymer to a given antibody, or even given antibody class. So, as far as I know, it's formally unknown which Cys are involved on a given antibody. And, formally, reacting with Cys close to the antigen-binding site would be one way of messing up an antibody.


Longer answer: there's a wealth of literature on antibody-drug conjugates (ADC), many of which are attached via maleimides. To my knowledge, pharma generally *does* have to figure this out during drug development (number of attachments, points of attachment, etc). This is for reproducibility manufacturing claims: they may not be able to claim, say, exactly 4 conjugates every single time, but they have to give the range of modifications and their points of attachment.

In all cases I'm aware of, even for small molecule attachments, the Fc region Cys are the ones modified for ADCs.


Here's some literature to get you started:
http://dx.doi.org/10.1021/ac3032355
http://dx.doi.org/10.1021/ac404003q
http://dx.doi.org/10.1021/bc5005262
http://dx.doi/10.1002/cbic.201500667 (see especially comment about Genentech's THIOMAB approach on p. 5)


Mike

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