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Principal Investigator: PHILIPPE ANDRE GALLAY
Organization: SCRIPPS RESEARCH INSTITUTE, THE
Fiscal Year: 2024
Award: $775,538
Funding agency: National Institute of Allergy and Infectious Diseases
While the development of effective vaccines against CoV-2 is cause for optimism, vaccine hesitancy in
developed countries and shortages in low-income countries are jeopardizing efforts to curb the pandemic. Out
of the 6.4 billion people living in low-income countries, only 2% have access to vaccines. Consequently, the
conditions are ripe for continued spike mutation and evolution to increasingly transmissible strains causing more
severe illness. Some of these emerging strains may even challenge the protection of vaccines. In this application,
we propose a novel therapeutic approach for the eradication of CoV-2. We developed a new strategy, which
consists of hijacking the viral replication machinery to trigger the death of CoV-2-infected cells, while preserving
uninfected cells. We propose to administer intranasally human ACE2 transgenic mice and Syrian hamsters a
“tailored” RNA encoding the diphtheria toxin fragment A (DTA) called {CoV-2 Hijack DTA} that is only recognized
and transcribed by the CoV-2 polymerase (Pol/RdRp) present in infected cells, triggering DTA expression and
rapid death of infected cells. Since DTA cannot cross the cellular membrane, it cannot kill uninfected cells.
Because RNA can be easily broken down in the body, it needs to be transported within a protective carrier.
Noninvasive aerosol inhalation is a well-established method of drug delivery to the respiratory tract and
represents an ideal route for nucleic-acid-based therapeutics as demonstrated in various clinical trials. We
propose to design degradable polymer-lipid nanoparticles (LNPs) that can deliver RNAs by nebulization
(inhalation) to the respiratory tract. We propose to synthesize hyperbranched poly-beta amino esters (hPBAEs)
to enable nanoformulation by nebulizer of stable and concentrated polyplexes suitable for inhalation. This
strategy should achieve uniform distribution of RNAs throughout lungs resulting in high levels of proteins of
interest 24h post-inhalation of hPBAE polyplexes without local or systemic toxicity due to rapid degradation of
hPBAE vectors. The safety and antiviral efficacy of nebulized {CoV-2 Hijack DTA} RNA LNPs stably protected
by degradable hPBAEs will be analyzed. Our in vivo imaging IVIS Lumina S5 system permits a daily
bioluminescence (NanoLuc-CoV-2) or fluorescence (mNeonGreen CoV-2) quantification of the {CoV-2 Hijack
DTA} RNA LNPs-mediated killing of infected lungs in live animals. We will investigate the MoA causing the killing
of CoV-2-infected cells by {CoV-2 Hijack DTA}. We will use complementary approaches to determine whether
{CoV-2 Hijack DTA} triggers apoptosis, membrane permeability and/or chromosomal degradation leading to cell
killing. By scRNA-Seq, we will analyze i) the specific killing of infected cells at high resolution on large numbers
of cells exposed to {CoV-2 Hijack DTA}; ii) the global map of apoptotic DNA breakpoints such as DNA
fragmentation; and iii) the phenotype of immunological target cells. We will examine whether {CoV-2 Hijack DTA}
RNA LNPs counteract the deleterious inflammatory response, which occurs during CoV-2 infection including
histopathological lesion development, interstitial pneumonia and cytokine cascade.
Terms: <2019 novel corona virus><2019 novel coronavirus><2019-nCoV><2019-nCoV vaccine><ACE2><Aerosols><Animals><Apoptosis><Apoptosis Pathway><Apoptotic><Bioluminescence><COVID-19><COVID-19 associated death><COVID-19 associated fatality><COVID-19 associated mortality><COVID-19 death><COVID-19 exposure><COVID-19 fatality><COVID-19 induced death><COVID-19 induced fatality><COVID-19 induced mortality><COVID-19 infection><COVID-19 mortality><COVID-19 related death><COVID-19 related fatality><COVID-19 related mortality><COVID-19 therapy><COVID-19 treatment><COVID-19 vaccine><COVID-19 virus><COVID-19 virus infection><COVID19 associated death><COVID19 associated fatality><COVID19 associated mortality><COVID19 death><COVID19 fatality><COVID19 induced death><COVID19 induced fatality><COVID19 induced mortality><COVID19 infection><COVID19 mortality><COVID19 related death><COVID19 related fatality><COVID19 related mortality><COVID19 virus><CV-19><Cell Body><Cell Membrane Permeability><Cells><Cellular Membrane><Cessation of life><Clinical Trials><CoV-2><CoV2><Coronavirus Infectious Disease 2019><Cricetinae><DNA><DNA Fragmentation><Death><Dengue><Deoxyribonucleic Acid><Developed Countries><Development><Drug Delivery><Drug Delivery Systems><Ebola><Esters><Event><Evolution><Exposure to><Fluorescence><Gene Transcription><Genes><Genetic Alteration><Genetic Change><Genetic Transcription><Genetic defect><Goals><Golden Hamsters><Golden Syrian Hamsters><Grippe><HBV><Hamsters><Hamsters Mammals><Health><Hepatitis B Virus><Homologous Serum Hepatitis Virus><Human><Image><Immune Targeting><In Vitro><Industrialized Countries><Industrialized Nations><Infection><Inflammatory Response><Influenza><Influenza A><Influenza A virus><Influenza B><Influenza B Virus><Influenza Virus><Influenza Viruses Type A><Influenza Viruses Type B><Influenzavirus A><Inhalation><Inhalators><Inhaler><Inhaling><Interstitial Lung Inflammation><Interstitial Pneumonia><Interstitial 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