Phenotype-specific recombinant haptoglobin polymers co-expressed with C1r-like protein as optimized hemoglobin-binding therapeutics

Autor: Paul W. Buehler, Jeremy W. Deuel, Catherine M. Owczarek, Stefan Schauer, Jin Hyen Baek, Matthew P. Hardy, Dominik J. Schaer, Peter Schmidt, Peter Soupourmas, Ayla Yalamanoglu, Pierre Scotney, Christian A. Schaer, Matthias Pelzing
Přispěvatelé: University of Zurich, Schaer, Dominik J
Jazyk: angličtina
Rok vydání: 2018
Předmět:
Male
0301 basic medicine
Hemoglobin binding
Swine
10216 Institute of Anesthesiology
lcsh:Biotechnology
Guinea Pigs
610 Medicine & health
10071 Functional Genomics Center Zurich
Heme
030204 cardiovascular system & hematology
Biology
Nitric Oxide
Protein Engineering
law.invention
Hemoglobins
03 medical and health sciences
0302 clinical medicine
law
lcsh:TP248.13-248.65
Human Umbilical Vein Endothelial Cells
medicine
Animals
Humans
Haptoglobins
Serine Endopeptidases
Haptoglobin
medicine.disease
Coronary Vessels
Phenotype
Recombinant Proteins
Hemolysis
In vitro
030104 developmental biology
Biochemistry
biology.protein
Recombinant DNA
1305 Biotechnology
570 Life sciences
biology
Lipid Peroxidation
Hemoglobin
10029 Clinic and Policlinic for Internal Medicine
Research Article
Biotechnology
Cysteine
Zdroj: BMC Biotechnology
BMC Biotechnology, Vol 18, Iss 1, Pp 1-13 (2018)
BMC Biotechnology, 18 (1)
ISSN: 1472-6750
Popis: Background Preclinical studies have evaluated haptoglobin (Hp) polymers from pooled human plasma as a therapeutic protein to attenuate toxic effects of cell-free hemoglobin (Hb). Proof of concept studies have demonstrated efficacy of Hp in hemolysis associated with transfusion and sickle cell anemia. However, phenotype-specific Hp products might be desirable to exploit phenotype specific activities of Hp 1–1 versus Hp 2–2, offering opportunities for recombinant therapeutics. Prohaptoglobin (proHp) is the primary translation product of the Hp mRNA. ProHp is proteolytically cleaved by complement C1r subcomponent-like protein (C1r-LP) in the endoplasmic reticulum. Two main allelic Hp variants, HP1 and HP2 exist. The larger HP2 is considered to be the ancestor variant of all human Hp alleles and is characterized by an α2-chain, which contains an extra cysteine residue that pairs with additional α-chains generating multimers with molecular weights of 200–900 kDa. The two human HP1 alleles (HP1F and HP1S) differ by a two-amino-acid substitution polymorphism within the α-chain and are derived from HP2 by recurring exon deletions. Results In the present study, we describe a process for the production of recombinant phenotype specific Hp polymers in mammalian FS293F cells. This approach demonstrates that efficient expression of mature and fully functional protein products requires co-expression of active C1r-LP. The functional characterization of our proteins, which included monomer/polymer distribution, binding affinities as well as NO-sparing and antioxidant functions, demonstrated that C1r-LP-processed recombinant Hp demonstrates equal protective functions as plasma derived Hp in vitro as well as in animal studies. Conclusions We present a recombinant production process for fully functional phenotype-specific Hp therapeutics. The proposed process could accelerate the development of Hb scavengers to treat patients with cell-free Hb associated disease states, such as sickle cell disease and other hemolytic conditions.
BMC Biotechnology, 18 (1)
ISSN:1472-6750
Databáze: OpenAIRE
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