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Generation of a VeroE6 Pgp gene knock out cell line and its use in SARS-CoV-2 antiviral study.
Zhu, Yuao; Binder, Joe; Yurgelonis, Irina; Rai, Devendra K; Lazarro, Sarah; Costales, Chester; Kobylarz, Keith; McMonagle, Patricia; Steppan, Claire M; Aschenbrenner, Lisa; Anderson, Annaliesa S; Cardin, Rhonda D.
  • Zhu Y; Pfizer Inc, 401 N Middletown Rd, Pearl River, NY, 10965, USA. Electronic address: Yuao.Zhu@pfizer.com.
  • Binder J; Pfizer Inc, 401 N Middletown Rd, Pearl River, NY, 10965, USA.
  • Yurgelonis I; Pfizer Inc, 401 N Middletown Rd, Pearl River, NY, 10965, USA.
  • Rai DK; Pfizer Inc, 401 N Middletown Rd, Pearl River, NY, 10965, USA.
  • Lazarro S; Pfizer Inc, 445 Eastern Point Rd, Groton, CT, 06333, USA.
  • Costales C; Pfizer Inc, 445 Eastern Point Rd, Groton, CT, 06333, USA.
  • Kobylarz K; Pfizer Inc, 401 N Middletown Rd, Pearl River, NY, 10965, USA.
  • McMonagle P; Pfizer Inc, 401 N Middletown Rd, Pearl River, NY, 10965, USA.
  • Steppan CM; Pfizer Inc, 445 Eastern Point Rd, Groton, CT, 06333, USA.
  • Aschenbrenner L; Pfizer Inc, 445 Eastern Point Rd, Groton, CT, 06333, USA.
  • Anderson AS; Pfizer Inc, 401 N Middletown Rd, Pearl River, NY, 10965, USA.
  • Cardin RD; Pfizer Inc, 401 N Middletown Rd, Pearl River, NY, 10965, USA.
Antiviral Res ; 208: 105429, 2022 Dec.
Article in English | MEDLINE | ID: covidwho-2104350
ABSTRACT
Vero cells are widely used for antiviral tests and virology research for SARS-CoV-2 as well as viruses from various other families. However, Vero cells generally express high levels of multi-drug resistance 1 (MDR1) or Pgp protein, the efflux transporter of foreign substances including many antiviral compounds, affecting the antiviral activity as well as interpretation of data. To address this, a Pgp gene knockout VeroE6 cell line (VeroE6-Pgp-KO) was generated using CRISPR-CAS9 technology. These cells no longer expressed the Pgp protein as indicated by flow cytometry analysis following staining with a Pgp-specific monoclonal antibody. They also showed significantly reduced efflux transporter activity in the calcein acetoxymethyl ester (calcein AM) assay. The VeroE6-Pgp-KO cells and the parental VeroE6 cells were each infected with SARS-CoV-2 to test antiviral activities of remdesivir and nirmatrelvir, two known Pgp substrates, in the presence or absence of a Pgp inhibitor. The compounds showed antiviral activities in VeroE6-Pgp-KO cells similar to that observed in the presence of the Pgp inhibitor. Thus, the newly established VeroE6-Pgp-KO cell line adds a new in vitro virus infection system for SARS-CoV-2 and possibly other viruses to test antiviral therapies without a need to control the Pgp activity. Removal of the Pgp inhibitor for antiviral assays will lead to less data variation and prevent failed assays.
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Full text: Available Collection: International databases Database: MEDLINE Main subject: SARS-CoV-2 / COVID-19 Drug Treatment Topics: Variants Limits: Animals / Humans Language: English Journal: Antiviral Res Year: 2022 Document Type: Article

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Full text: Available Collection: International databases Database: MEDLINE Main subject: SARS-CoV-2 / COVID-19 Drug Treatment Topics: Variants Limits: Animals / Humans Language: English Journal: Antiviral Res Year: 2022 Document Type: Article