PEGylation of Recombinant Human Deoxyribonuclease I Provides a Long‐Acting Version of the Mucolytic for Patients with Cystic Fibrosis

Abstract

Inhalation of recombinant human deoxyribonuclease I (rhDNase) is a gold-standard therapy in the management of cystic fibrosis (CF). Yet, the rapid elimination of the mucolytic from the lungs requires its daily administration and rhDNase contributes to the high therapy burden of patients. Here, a long-acting version of rhDNase that is delivered once weekly instead of once daily is presented. Conjugation of rhDNase to a polyethylene glycol (PEG) chain sustains its presence and mucolytic activity within murine lungs for more than 15 days. One single dose of PEGylated rhDNase is as effective as 1 daily dose of unconjugated rhDNase during five days to decrease the DNA content in the lungs of-ENaC mice, a model of the CF lung disease. Moreover, once weekly administration of PEGylated rhDNase over four weeks decreases both the DNA content and the neutrophil counts in the lungs of-ENaC mice. PEGylated rhDNase is stable to jet nebulization. Finally, multiple high-dose administrations of PEGylated rhDNase for up to 3 months do not cause any significant pulmonary or systemic toxicity, nor accumulation of the rhDNase or PEG moieties in biological fluids. PEGylation of rhDNase may offer a convenient long-acting mucolytic to patients with CF.

Biodistribution and elimination pathways of PEGylated recombinant human deoxyribonuclease I after pulmonary delivery in mice

“…Macrophage uptake was also reported for PEG40-Fab', PEG30-rhDNase, and PEGs of up to 40 kDa after pulmonary delivery [19,25,27,33]. While no quantitative data were collected in the present work, another study from our laboratory demonstrated that the uptake of PEGylated anti-IL13 Fab' by alveolar macrophages was slightly less efficient than its non-PEGylated counterpart in mice [19].…”

“…Previously published works have shown that conjugation of proteins to PEGs of MWs ≥ 20 kDa has a high potential for increasing their residence time in the lungs [13,[18][19][20]27]. More recently, Guichard et al demonstrated the prolonged residence time of 30 and 40 kDa PEG conjugates of rhDNase in the lungs of mice for more than 15 days compared to merely 24 h for the unconjugated rhDNase [25]. They also demonstrated the improved therapeutic efficacy of PEG30-rhDNase in β-ENaC mice, a model of the CF lung disease, as well as its safety after single and repeated administration to healthy mice.…”

“…The PEGylation of rhDNase was adapted from the method described by Guichard et al [23,25]. PEG20-rhDNase, PEG30-rhDNase, and PEG40-rhDNase were produced by conjugation of the N-terminal amino acid of rhDNase to PEG20, PEG30, and PEG40, respectively.…”

“…Unconjugated rhDNase, on the other hand, had a retention of less than 24 hours. Furthermore, a single dose of PEG30-rhDNase or PEG40-rhDNase exhibited an equivalent DNA hydrolysis activity after 5 days as one daily dose of rhDNase during 5 days in β-transepithelial Na+ channel-overexpressing mice (β-ENaC) [25].…”

Protein Engineering Strategies for Improved Pharmacokinetics

“…[19] Conjugation of PEG to recombinant human deoxyribonuclease I (rhDNase) resulted in an impressive extension of its residence time (≥15 days) in the murine lungs. [20] Moreover, one single dose of PEGylated rhDNase was as effective as 1 daily dose of unconjugated rhDNase during 5 days in decreasing the DNA content in the lungs of β-ENaC mice, a model of the cystic fibrosis lung disease. The lack of marketed PEGylated proteins for pulmonary delivery reflects the paucity of approved proteins for inhalation in the first place.…”