GTC Biotherapeutics (fka GTCB)

[Pre_Clinical]

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Re: Two isoforms of antithrombin

>…so let me guess this straight, there are two glycoforms of AT produced in transgenic goats, one form that is essentially bio-equivalent to pAT, and another form that is more rapidly cleared.<

This is inaccurate on two counts:

1. Both ATryn and endogenous AT have two isoforms: AT-alpha and AT-beta. The beta isoform has a shorter half-life. In endogenous AT (and plasma-derived AT), the alpha isoform comprises 90-95% of total AT and the beta isoform comprises 5-10%:

http://www.ncbi.nlm.nih.gov/sites/entrez?cmd=Retrieve&db=PubMed&list_uids=12372930&dopt=....

In ATryn, the beta isoform comprises considerably more than 10% of total AT (but GTC has not publicly disclosed the actual number). This is what gives ATryn a shorter half-life than plasma-derived AT.

2. To my knowledge, the two isoforms of AT are not variant glycoforms, as you assumed. Thus, the PK difference between ATryn and plasma-derived AT is not a glycosylation issue.

Regards, Dew
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Hi Dew, to my knowledge, the two isoforms of AT, alpha, and beta, are different glycoforms. According to pubmed ID 12651125 (Kalsson G et al 2003), "AT consists of a single polypeptide chain containing 432 amino acid residues and 3 disulfide bridges. The total molecular weight for ATalpha, which is glycosylated at four amino acid residue positions (asparagines 96, 135, 155, and 172), is approximately 58 kDa, while ATbeta, which does not have the Asn135 glycosylation, has a molecular weight of about 56 kDa."

So it seems that there are two glycoforms of AT, alpha and beta, in plasma AND produced in transgenic goats. In plasma, the alpha glycoform, the fully glycosylated form, comprises about 90-95% of the total protein. So the real question is, what % of the material produced in goats is the beta glycoform, the under-glycosylated form (ie lacking glycosylation at Asn135)?

On top of that, what are the differences in the goat mammary gland secretory pathway that lead to this heterogeneity and is it a universal aspect of this expression platform?