Site-selective protein conjugation at histidine.

Site-selective conjugation generally requires both (i) molecular engineering of the protein of interest to introduce a conjugation site at a defined location and (ii) a site-specific conjugation technology. Three N-terminal interferon α2-a (IFN) variants with truncated histidine tags were prepared and conjugation was examined using a bis-alkylation reagent, PEG-mono-sulfone . A histidine tag comprised of two histidines separated by a glycine (His-tag) underwent PEGylation.

Development of a high yield expression and purification system for Domain I of Beta-2-glycoprotein I for the treatment of APS.

Background: In this paper we describe a novel method to achieve high yield bacterial expression of a small protein domain with considerable therapeutic potential; Domain I of Beta-2-glycoprotein I (β2GPI). β2GPI is intrinsic to the pathological progression of the Antiphospholipid Syndrome (APS). Patients develop autoantibodies targeting an epitope located on the N-terminal Domain I of β2GPI rendering this domain of interest as a possible therapeutic.

In Vitro and In Vivo Evaluation of Cysteine Rebridged Trastuzumab-MMAE Antibody Drug Conjugates with Defined Drug-to-Antibody Ratios.

The conjugation of monomethyl auristatin E (MMAE) to trastuzumab using a reduction bis-alkylation approach that is capable of rebridging reduced (native) antibody interchain disulfide bonds has been previously shown to produce a homogeneous and stable conjugate with a drug-to-antibody ratio (DAR) of 4 as the major product. Here, we further investigate the potency of the DAR 4 conjugates prepared by bis-alkylation by comparing to lower drug loaded variants to maleimide linker based conjugates possessing typical mixed DAR profiles. Serum stability, HER2 receptor binding, internalization, in vitro potency, and in vivo efficacy were all evaluated.

Expression of soluble and active interferon consensus in SUMO fusion expression system in E. coli.

Protein production can be improved if methods for soluble protein expression are developed. Interferon consensus (IFN-con) is used to treat hepatitis C. IFN-con has superior activity compared to other clinically used interferon α subtypes.

Site-specific PEGylation at histidine tags.

The efficacy of protein-based medicines can be compromised by their rapid clearance from the blood circulatory system. Achieving optimal pharmacokinetics is a key requirement for the successful development of safe protein-based medicines. Protein PEGylation is a clinically proven strategy to increase the circulation half-life of protein-based medicines.