The carboxyl-terminal region of human coagulation factor X as a natural linker for fusion strategies

Ferrarese, Mattia; Pignani, Silvia; Lombardi, Silvia; Balestra, Dario; Bernardi, Francesco; Pinotti, Mirko; Branchini, Alessio

doi:10.1016/j.thromres.2018.11.007

Cited by 8 publications

(7 citation statements)

References 44 publications

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“…Data file S1. Raw data References (64)(65)(66)(67)(68)(69)(70)(71)(72)(73)(74)(75)(76)(77)(78) View/request a protocol for this paper from Bio-protocol.…”

Section: Discussionmentioning

confidence: 99%

An engineered human albumin enhances half-life and transmucosal delivery when fused to protein-based biologics

et al. 2020

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Needle-free uptake across mucosal barriers is a preferred route for delivery of biologics, but the efficiency of unassisted transmucosal transport is poor. To make administration and therapy efficient and convenient, strategies for the delivery of biologics must enhance both transcellular delivery and plasma half-life. We found that human albumin was transcytosed efficiently across polarized human epithelial cells by a mechanism that depends on the neonatal Fc receptor (FcRn). FcRn also transported immunoglobulin G, but twofold less than albumin. We therefore designed a human albumin variant, E505Q/T527M/K573P (QMP), with improved FcRn binding, resulting in enhanced transcellular transport upon intranasal delivery and extended plasma half-life of albumin in transgenic mice expressing human FcRn. When QMP was fused to recombinant activated coagulation factor VII, the half-life of the fusion molecule increased 3.6-fold compared with the wild-type human albumin fusion, without compromising the therapeutic properties of activated factor VII. Our findings highlight QMP as a suitable carrier of protein-based biologics that may enhance plasma half-life and delivery across mucosal barriers.

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“…Data file S1. Raw data References (64)(65)(66)(67)(68)(69)(70)(71)(72)(73)(74)(75)(76)(77)(78) View/request a protocol for this paper from Bio-protocol.…”

Section: Discussionmentioning

confidence: 99%

An engineered human albumin enhances half-life and transmucosal delivery when fused to protein-based biologics

et al. 2020

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“…Expression studies in HEK293 cells and stable clone selection were performed as described previously. 12 , 39 …”

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

“…9,10 Idelvion is designed with a cleavable linker connecting the two fusion partners that, upon FIX activation, drives the release of albumin and activated FIX. 11,12 Hence, this fusion strategy improves FIX halflife without compromising its functional properties. 9,10 The fact that albumin is biodegradable, non-immunogenic and effector negative, which minimises the risk of side effects, makes it attractive in this context.…”

Section: Introductionmentioning

confidence: 99%

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Fusion of engineered albumin with factor IX Padua extends half‐life and improves coagulant activity

et al. 2021

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The short half-life of coagulation factor IX (FIX) for haemophilia B (HB) therapy has been prolonged through fusion with human serum albumin (HSA), which drives the neonatal Fc receptor (FcRn)-mediated recycling of the chimera. However, patients would greatly benefit from further FIX-HSA half-life extension. In the present study, we designed a FIX-HSA variant through the engineering of both fusion partners. First, we developed a novel cleavable linker combining the two FIX activation sites, which resulted in improved HSA release. Second, insertion of the FIX R338L (Padua) substitution conferred hyperactive features (sevenfold higher specific activity) as for FIX Padua alone. Furthermore, we exploited an engineered HSA (QMP), which conferred enhanced human (h)FcRn binding [dissociation constant (K D ) 0Á5 nM] over wild-type FIX-HSA (K D 164Á4 nM). In hFcRn transgenic mice, Padua-QMP displayed a significantly prolonged half-life (2Á7 days, P < 0Á0001) versus FIX-HSA (1 day). Overall, we developed a novel FIX-HSA protein with improved activity and extended half-life. These combined properties may result in a prolonged functional profile above the therapeutic threshold, and thus in a potentially widened therapeutic window able to improve HB therapy. This rational engineering of both partners may pave the way for new fusion strategies for the design of engineered biotherapeutics.

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“…Fusion strategies such as those exploiting albumin, in which half-life of the target albumin-fused coagulation factor is prolonged by the albumin recycling pathway mediated by the neonatal Fc receptor, will not be the focus of this review. However, three key coagulation factors, namely, factor FVII [32], FIX [33] and FX [34], with different requirements in terms of fusion strategy, have been produced and tested in different models and trials, with FIX having approached the market.…”

Section: Overview Of Correction Approaches At Dna Mrna and Proteimentioning

confidence: 99%

Molecular Mechanisms and Determinants of Innovative Correction Approaches in Coagulation Factor Deficiencies

Balestra

Branchini

2019

IJMS

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Molecular strategies tailored to promote/correct the expression and/or processing of defective coagulation factors would represent innovative therapeutic approaches beyond standard substitutive therapy. Here, we focus on the molecular mechanisms and determinants underlying innovative approaches acting at DNA, mRNA and protein levels in inherited coagulation factor deficiencies, and in particular on: (i) gene editing approaches, which have permitted intervention at the DNA level through the specific recognition, cleavage, repair/correction or activation of target sequences, even in mutated gene contexts; (ii) the rescue of altered pre-mRNA processing through the engineering of key spliceosome components able to promote correct exon recognition and, in turn, the synthesis and secretion of functional factors, as well as the effects on the splicing of missense changes affecting exonic splicing elements; this section includes antisense oligonucleotide- or siRNA-mediated approaches to down-regulate target genes; (iii) the rescue of protein synthesis/function through the induction of ribosome readthrough targeting nonsense variants or the correction of folding defects caused by amino acid substitutions. Overall, these approaches have shown the ability to rescue the expression and/or function of potentially therapeutic levels of coagulation factors in different disease models, thus supporting further studies in the future aimed at evaluating the clinical translatability of these new strategies.

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The carboxyl-terminal region of human coagulation factor X as a natural linker for fusion strategies

Cited by 8 publications

References 44 publications

An engineered human albumin enhances half-life and transmucosal delivery when fused to protein-based biologics

An engineered human albumin enhances half-life and transmucosal delivery when fused to protein-based biologics

Fusion of engineered albumin with factor IX Padua extends half‐life and improves coagulant activity

Molecular Mechanisms and Determinants of Innovative Correction Approaches in Coagulation Factor Deficiencies

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