Vasoconstriction, Hypertension and Oxidative Toxicity are Regulated by Polymerized Hemoglobin Size

Belcik, Brian M.; Palmer, Andre F.

doi:10.1007/978-3-642-40717-8_37

Cited by 2 publications

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“…Of these, only Oxyglobin® was approved by the FDA for veterinary use (Driessen et al, 2001). Unfortunately, Hemopure® and PolyHeme® elicited vasoconstriction, systemic hypertension, and oxidative tissue injury during Phase III clinical trials, which hindered their commercial development (Belcik & Palmer, 2013; Jahr et al, 2008, 2010). Previous work has attributed these side‐effects to the presence of low‐molecular weight (MW) Hb species in formulations of Hemopure® and PolyHeme®, which can extravasate through the blood vessel wall into the tissue space, where the Hb species can scavenge nitric oxide (NO) and deposit redox‐active iron, which induces vasoconstriction, systemic hypertension and oxidative tissue injury (Baek et al, 2012; Cabrales et al, 2009).…”

Section: Introductionmentioning

confidence: 99%

Tangential flow filtration facilitated fractionation and PEGylation of low and high‐molecular weight polymerized hemoglobins and their biophysical properties

Savla

Palmer

2021

Biotech & Bioengineering

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Various types of hemoglobin (Hb)‐based oxygen carriers (HBOCs) have been developed as red blood cell substitutes for treating blood loss when blood is not available. Among those HBOCs, glutaraldehyde polymerized Hbs have attracted significant attention due to their facile synthetic route, and ability to expand the blood volume and deliver oxygen. Hemopure®, Oxyglobin®, and PolyHeme® are the most well‐known commercially developed glutaraldehyde polymerized Hbs. Unfortunately, only Oxyglobin® was approved by the FDA for veterinary use in the United States, while Hemopure® and PolyHeme® failed phase III clinical trials due to their ability to extravasate from the blood volume into the tissue space which facilitated nitric oxide scavenging and tissue deposition of iron, which elicited vasoconstriction, hypertension and oxidative tissue injury. Fortunately, conjugation of poly (ethylene glycol) (PEG) on the surface of Hb is capable of reducing the vasoactivity of Hb by creating a hydration layer surrounding the Hb molecule, which increases its hydrodynamic diameter and reduces tissue extravasation. Several commercial PEGylated Hbs (MP4®, Sanguinate®, Euro‐PEG‐Hb) have been developed for clinical use with a longer circulatory half‐life and improved safety compared to Hb. However, all of these commercial products exhibited relatively high oxygen affinity compared to Hb, which limited their clinical use. To dually address the limitations of prior generations of polymerized and PEGylated Hbs, this current study describes the PEGylation of polymerized bovine Hb (PEG‐PolybHb) in both the tense (T) and relaxed (R) quaternary state via thiol‐maleimide chemistry to produce an HBOC with low or high oxygen affinity. The biophysical properties of PEG‐PolybHb were measured and compared with those of commercial polymerized and PEGylated HBOCs. T‐state PEG‐PolybHb possessed higher hydrodynamic volume and P50 than previous generations of commercial PEGylated Hbs. Both T‐ and R‐state PEG‐PolybHb exhibited significantly lower haptoglobin binding rates than the precursor PolybHb, indicating potentially reduced clearance by CD163 + monocytes and macrophages. Thus, T‐state PEG‐PolybHb is expected to function as a promising HBOC due to its low oxygen affinity and enhanced stealth properties afforded by the PEG hydration shell.

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Section: Introductionmentioning

confidence: 99%

Tangential flow filtration facilitated fractionation and PEGylation of low and high‐molecular weight polymerized hemoglobins and their biophysical properties

Savla

Palmer

2021

Biotech & Bioengineering

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“…19 Presumably, the increased size prevents these species from penetrating the endothelia. 20 However, the PEGylation imparts high oxygen affinity along with low cooperativity. 21 Those materials have been utilized in specific applications 22 and are a potential therapeutic directed to very hypoxic regions of circulation.…”

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confidence: 99%

Subunit-directed click coupling via doubly cross-linked hemoglobin efficiently produces readily purified functional bis-tetrameric oxygen carriers

et al. 2015

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While cross-linked hemoglobin (Hb) tetramers can deliver oxygen as a supplement to red cells, they also cause unacceptable increases in blood pressure, presumably from their penetration of the linings of blood vessels (endothelia) where the internal hemes bind endogenous nitric oxide (NO). This penetration would lower the local concentration of NO that normally induces vasodilation. Enlarging the effective size of the oxygen-carrying protein by coupling two Hbs can prevent their extravasation. Efficient and selective protein-protein coupling to produce those species has been a significant challenge. Introduction of an azide within a protein provides a directionally-oriented reaction site for utilization of the Cu(i)-catalyzed azide-alkyne cycloaddition (CuAAC) in the protein-protein-coupling process based on solubility-directed sequential addition to a bis-alkyne. However, it is known that Hb with an azide-containing cross-link between α-subunits is unreactive in CuAAC. To direct reaction away from the α-subunits of Hb, a specific fumaryl cross-link is installed exclusively between the most reactive sites on those subunits, thereby blocking the α-99 lysyl groups and preventing any further reaction. This modification allows installation of an azide-containing cross-link exclusively between lysine-82 ε-amino groups of the β-subunits of Hb. The multiply interconnected sites establish a geometry that permits initial interfacial interaction of the cross-linked Hb-azide with Cu(i) and a bis-alkyne. After coupling, the protein-linked azide product undergoes CuAAC at the remaining alkyne with a second cross-linked Hb-azide, producing a fully functional cross-linked Hb bis-tetramer whose oxygenation and structural properties include cooperativity and oxygen affinity that should be suitable for testing as an alternative to red cells in transfusions.

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Vasoconstriction, Hypertension and Oxidative Toxicity are Regulated by Polymerized Hemoglobin Size

Cited by 2 publications

References 83 publications

Tangential flow filtration facilitated fractionation and PEGylation of low and high‐molecular weight polymerized hemoglobins and their biophysical properties

Tangential flow filtration facilitated fractionation and PEGylation of low and high‐molecular weight polymerized hemoglobins and their biophysical properties

Subunit-directed click coupling via doubly cross-linked hemoglobin efficiently produces readily purified functional bis-tetrameric oxygen carriers

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