Tropism and Pathogenesis of Influenza Virus and Coronavirus in Human Brain Explant Culture (COVID-19)

Background:

Influenza and coronavirus have been repeatedly causing pandemic recently. Like the Influenza A/H7N9 virus has caused five epidemics in China since its first detection in East China in 2013. In 2017, the previously low pathogenic avian influenza (LPAI) H7N9 virus underwent mutation in its haemagglutinin to give to a highly pathogenic avian influenza (HPAI) virus causing 32 human cases and potentially poses a threat to animal and human health. More recently, the SARS-CoV-2 pandemic has been heavily affecting the world. Therefore an effective risk assessment platform is urgently required for better pandemic preparation.

Hypothesis:

The tissue tropism and pathogenesis of a newly emerged infectious viruses, like the highlypathogenic influenza, like H7N9 and coronavirus, like SARS-CoV-2 would be different from that of their low pathogenic subtype and it would infect and replicate the human respiratory system more efficiently.

Because of its resistance to oseltamivir for influenza and no effective antiviral for coronavirus, investigators therefore propose to set up an novel and effective risk assessment platform for emerging infectious viruses.

Experimental Design:

The tissue tropism and viral replication kinetics of a HPAI and LP influenza and coronavirus will be determined in ex vivo cultures of human brain and compared with their LP subtype. The replication competence and innate immune responses of influenza and coronavirus will be studied and compared with other LP virus in in vitro cultures of human brain cells and human microvascular endothelial cells (HMVEC) both isolated from human brain tissues.

Expected outcomes:

HPAI influenza and coronavirus particularly SARS-CoV-2 will infect and replicate the human brain tissues and cells more efficiently than their LP subtype. Besides, HPAI influenza and SARS-CoV-2 will induce dysregulated host innate immune response than the LP subtype.

Study Overview

• Status: Recruiting
• Conditions: Coronavirus; Influenza Virus

Detailed Description

In this study, 80 subjects who will undergoing elective or emergency craniotomies for intrinsic brain lesions at Prince of Wales Hospital, will be recruited.

This is a prospective and qualitative study. There is no randomization in the study procedure nor therapeutic invention for study subjects. No investigational product is involved.

Brain tissues that are normal discarded during the operation from patients who undergo elective or emergency craniotomies for intrinsic brain lesions will be collected for this study.

A consent for operation and agreement to use of removed tissue for scientific research will be obtained prior to the procedure.

• Study Type: Observational
• Enrollment (Anticipated): 80

Contacts and Locations

• Study Contact: Name: Owen Ho Ko, Ph.D; Phone Number: +852 26352160; Email: ho.ko@cuhk.edu.hk

Study Locations

• Hong Kong
  • NT
    • Shatin, NT, Hong Kong, 00000
      • Recruiting
      • Prince of Wales Hospital
      • Contact: Owen Ko, PhD; Phone Number: 26352160; Email: ho.ko@cuhk.edu.hk
      • Contact: Pauline Kwan, Master; Phone Number: 26352160; Email: paulinekwan@cuhk.edu.hk

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: 1 year to 70 years (Child, Adult, Older Adult)
• Accepts Healthy Volunteers: No
• Genders Eligible for Study: All

• Sampling Method: Non-Probability Sample

Study Population

This study will recruit participants who will undergoing elective or emergency craniotomies for intrinsic brain lesions at Prince of Wales Hospital

Description

Inclusion Criteria:

1. Age: > 1 year old and < 70 years old
2. Undergo elective or emergency craniotomies for intrinsic brain lesions

Exclusion Criteria:

a. Samples containing infected material

Study Plan

How is the study designed?

Design Details

• Observational Models: Case-Only
• Time Perspectives: Prospective

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Replication kinetics of influenza virus using ex vivo cultures of human brain	Tissue fragments will be infected with 106 TCID50/mL for 1h at 37°C and then washed with 3 ml of warm PBS for three times to remove unbound virus. To determine productive viral replication from the infected biopsy specimens, supernatants of the infected cultures will be collected at 1, 24, 48 and 72hpi and virus titers will be determined by TCID50 assay. Explant cultures will be fixed in 10% formalin at 24 and 72hpi and immunohistochemistry staining (IHC) against HB65 antibody (European Veterinary Laboratories), SARS-CoV or SARS-CoV-2 viral antibody will be performed for the detection of virus-infected cells.	baseline
replication efficiency and innate immune responses of influenza virus	Cells will be infected with viruses at either MOI of 0.01 or 2 for 1h. After 1h virus adsorption, viruses will be aspirated and cells will be washed with PBS for three times, then medium will be replenished. For cells infected at MOI of 0.01, cell culture supernatant will be harvested at 1, 24, 48 and 72hpi and viral titers will be determined by TCID50 assay.	baseline
Replication kinetics of coronavirus using ex vivo cultures of human brain	Tissue fragments will be infected with 106 TCID50/mL for 1h at 37°C and then washed with 3 ml of warm PBS for three times to remove unbound virus. To determine productive viral replication from the infected biopsy specimens, supernatants of the infected cultures will be collected at 1, 24, 48 and 72hpi and virus titers will be determined by TCID50 assay. Explant cultures will be fixed in 10% formalin at 24 and 72hpi and immunohistochemistry staining (IHC) against HB65 antibody (European Veterinary Laboratories), SARS-CoV or SARS-CoV-2 viral antibody will be performed for the detection of virus-infected cells.	Baseline
replication efficiency and innate immune responses coronavirus	Cells will be infected with viruses at either MOI of 0.01 or 2 for 1h. After 1h virus adsorption, viruses will be aspirated and cells will be washed with PBS for three times, then medium will be replenished. For cells infected at MOI of 0.01, cell culture supernatant will be harvested at 1, 24, 48 and 72hpi and viral titers will be determined by TCID50 assay.	Baseline

Collaborators and Investigators

• Sponsor: Chinese University of Hong Kong
• Collaborators: The University of Hong Kong
• Investigators: Principal Investigator: Owen Ho Ko, PhD, Chinese University of Hong Kong

Study record dates

Study Major Dates

• Study Start (Actual): May 26, 2022
• Primary Completion (Anticipated): March 31, 2024
• Study Completion (Anticipated): September 30, 2024

Study Registration Dates

• First Submitted: May 12, 2022
• First Submitted That Met QC Criteria: May 12, 2022
• First Posted (Actual): May 16, 2022

Study Record Updates

• Last Update Posted (Actual): August 17, 2022
• Last Update Submitted That Met QC Criteria: August 16, 2022
• Last Verified: August 1, 2022

More Information

Terms related to this study

• Additional Relevant MeSH Terms: Coronaviridae Infections; Nidovirales Infections; RNA Virus Infections; Virus Diseases; Infections; Respiratory Tract Infections; Respiratory Tract Diseases; Orthomyxoviridae Infections; Coronavirus Infections; Influenza, Human
• Other Study ID Numbers: 2022.120

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: No

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No