Science - USA (2021-12-24)

was safe and well tolerated and exhibited a
significant boost in plasma concentrations
when co-administered with RTV. Oral plasma
concentrations of PF-07321332 250 mg with
RTV were considerably above the SARS-COV-2
antiviral EC 90 value (total EC 90 = 292 ng/ml,
unbound EC 90 = 90.5 ng/ml, 181 nM) at 12 hours
after dose, thus increasing confidence in achiev-
ing robust pan-coronavirus antiviral activity
clinically (Fig. 5D). On the basis of these ob-
servations and the high safety margins at the
NOAEL doses in animals, the efficacy of PF-
07321332 in COVID-19 patients will be assessed
with a twice-daily dosing paradigm with the
potential to increase the dose of PF-07321332
as a single agent and/or co-administered
with RTV.

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ACKNOWLEDGMENTS

The authors thank the participants of the first-in-human (FIH)

study. We would also like to acknowledge the many Pfizer

colleagues who have contributed to the COVID-19 oral protease

program across a number of disciplines. In particular, we

acknowledge S. Sakata, J. Arcari, and J. Zhang for external

research resourcing; K. Farley for NMR studies; L. Lanyon for

protease panel data; Y. Lian, E. LaChapelle, S. Wright, S. O’Neil,

E. Yang, J. Humphrey, and B. Boscoe for compound synthesis;

S. Jenkinson for safety pharmacology data; K. Ryan for structural

biology support; E. Collins and C. Allais for FIH-enabling active

pharmaceutical ingredient supply; F. Clark for bioanalysis; H. Shi

for clinical assay support; G. Nucci and A. Bergman for first-in-

human study design and clinical pharmacology; F. Hackman for

1592 24 DECEMBER 2021¥VOL 374 ISSUE 6575 science.orgSCIENCE

0612 18 24

1

10

100

1000

10000

100000

Time (hr)

Mean Plasma

Concentration (ng/mL)

PF-07321332 60 mg/kg PF-07321332 200 mg/kg

PF-07321332 1000 mg/kg EC 90 SARS-COV-2

0 6 12 18 24

1

10

100

1000

10000

100000

1000000

Time (hr)

Mean Plasma

Concentration (ng/mL)

PF-07321332 40 mg/kg (20 mg/kg BID) PF-07321332 100 mg/kg (50 mg/kg BID)

PF-07321332 600 mg/kg (300 mg/kg BID) EC 90 SARS-COV-2

0 6 12 18 24 30 36 42 48

1

10

100

1000

10000

100000

Time (hr)

Median Plasma

Concentration (ng/mL)

PF-07321332 150 mg PF-07321332 250 mg+RTV

NOAEL (Cmax) EC 90 SARS-COV-2

036912

1

10

100

1000

10000

100000

Time (hr)

M

edian P

la

sm

a

Concentration (ng/mL)

PF-07321332 250 mg+RTV EC 90 MERS

NOAEL (Cmax) EC 90 SARS-COV-1

AB

CD

Fig. 5. Preclinical toxicology and healthy adult participant single ascending
dose study exposures for PF-07321332.(A) Rat oral toxicokinetic exposures
(day 14) of once-daily administered PF-07321332 compared with day 3 antiviral
EC 90 values in dNHBE cells. (B) Monkey oral toxicokinetic exposures (day 15)
of twice-daily administered PF-07321332 compared with day 3 antiviral EC 90
in dNHBE. (C) Human plasma concentrations (total) versus time profile after
oral administration (fasted state) of PF-07321332 (150 mg) and PF-07321332
(250 mg) with RTV (100 mg att=–12 hours, 0 hours, and 12 hours) compared
with day 3 antiviral EC 90 in dNHBE. (D) Human antiviral target coverage >EC90,uat
12 hours for SARS-CoV-1, SARS-CoV-2, and MERS-CoV after oral administration of

PF-07321332 (250 mg) and ritonavir (100 mg att=–12 hours, 0 hours, and 12 hours).

The in vitro unbound SARS-COV-2 EC 90 of 181 nM was converted to nanograms

per milliliter using a molecular weight of 499.5 g/mol for PF-07321332. Total

EC 90 was calculated by dividing unbound EC 90 byfu,plasma[rat 0.479, nonhuman

primate (NHP) 0.208, and human 0.310]. This resulted in total EC 90 values of

189, 208, and 292 ng/ml for rat, NHP, and human, respectively. The calculated

total human EC 90 for MERS-CoV and SARS-COV-1 were 566 and 422 ng/ml,

respectively. A human NOAEL of 79,700 ng/ml (forCmax) was estimated from

the ratCmaxvalue of 51,500 ng/ml at the NOAEL dose of 1000 mg/kg,

normalizing for plasma unbound fraction differences.

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