Species differences in plasma protein binding of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) main protease inhibitor nirmatrelvir

Abstract

1. Plasma protein binding (PPB) studies on the SARS-CoV-2 main protease inhibitor nirmatrelvir revealed considerable species differences primarily in dog and rabbit, which prompted further investigations into the biochemical basis for these differences.

2. The unbound fraction (f_u) of nirmatrelvir in dog and rabbit plasma was concentration (2–200 µM)-dependent (dog f_u,p 0.024–0.69, rabbit f_u,p 0.010–0.82). Concentration (0.1–100 µM)-dependent binding in serum albumin (SA) (f_u,SA 0.040–0.82) and alpha-1-acid glycoprotein (AAG) (f_u,AAG 0.050–0.64) was observed in dogs. Nirmatrelvir showed minimal binding to rabbit SA (1–100 µM: f_u,SA 0.70–0.79), while binding to rabbit AAG was concentration-dependent (0.1–100 µM: f_u,AAG 0.024–0.66). In contrast, nirmatrelvir (2 µM) revealed minimal binding (f_u,AAG 0.79–0.88) to AAG from rat and monkeys. Nirmatrelvir showed minimal-to-moderate binding to SA (1–100 µM; f_u,SA 0.70–1.0) and AAG (0.1–100 µM; f_u,AAG 0.48–0.58) from humans across tested concentrations.

3. Nirmatrelvir molecular docking studies using published crystal structures and homology models of human and preclinical species SA and AAG were used to rationalise the species differences to plasma proteins. This suggested that species differences in PPB are primarily driven by molecular differences in albumin and AAG resulting in differences in binding affinity.

Keywords:

• Plasma protein binding
• albumin
• alpha-1-acid glycoprotein
• COVID-19
• SARS-CoV-2

Acknowledgements

The authors acknowledge several Pfizer colleagues who contributed to this work including: David Janetos and Nandini Patel for assistance with formulation studies, Lisa Aschenbrenner, R. Scott Obach, Claire Steppan and Jean Sathish for assistance in research design, Eugene Kadar and Wei Song for bioanalytical support, and Naomi Otis and Jocelyn Rosado for dog pharmacokinetics study support.

Disclosure statement

^§SRG, HE, QY, CG, LB, AD, GW, LD, and ASK are employees of Pfizer and some of the authors are shareholders in Pfizer Inc.

PAXLOVID^TM has not been approved, but has been authorised for emergency use by FDA under an EUA, for the treatment of mild-to moderate COVID-19 in adults and paediatric patients (12 years of age and older weighing at least 40 kg) with positive results of direct SARS CoV-2 viral testing, and who are at high-risk for progression to severe COVID-19, including hospitalisation or death.

The emergency use of PAXLOVID^TM is only authorised for the duration of the declaration that circumstances exist justifying the authorisation of the emergency use of drugs and biological products during the COVID-19 pandemic under Section 564(b)(1) of the Act, 21 U.S.C. § 360bbb-3(b)(1), unless the declaration is terminated or authorisation revoked sooner.

Data availability statement

The authors confirm that the data supporting the findings of this study are available within the article.

Additional information

Funding

This work was supported by the Pfizer Inc.