Process for producing poxviruses and poxvirus compositions
Inventors
Malarme, Daniel • Cordier, Yves • Sene, Claude
Assignees
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Publication Number
US-8415133-B2
Publication Date
2013-04-09
Expiration Date
Abstract
The present invention relates to compositions and pharmaceutical compositions comprising poxviruses and more particularly extracellular enveloped viruses. The present invention also relates to a process for producing poxviruses and poxviruses obtained thereof. Moreover, the present invention also relates to the use of said poxvirus and said composition for the preparation of a medicament.
Core Innovation
The invention relates to producing a composition of a wild type, an attenuated, and/or a recombinant poxvirus with no targeted infection specificity. The process includes preparing a cell culture of packaging cells, infecting the cell culture, culturing infected cells to produce poxviral particles, and recovering poxviral particles from the culture supernatant and from packaging cells disrupted by high speed homogenizer to obtain a poxviral particle mixture. The resulting mixture is clarified by depth filtration to remove cellular debris, then concentrated by microfiltration and diafiltrated as part of producing the composition.
The process is free from animal products and does not use nuclease. The produced poxvirus composition comprises intracellular mature virus (IMV) and extracellular enveloped virus (EEV) with more than 1% EEV, and the packaging cells are chicken embryo fibroblast (CEF) prepared by enzymatic digestion.
The description contrasts prior purification that discards EEV with the finding that non-targeted-specificity EEV provides greater therapeutic/prophylactic efficacy than IMV. Preclinical mouse results comparing IMV/EEV versus IMV-only lots for HPV (TG4001) and MUC1 (TG4010) are described as showing improved survival and tumor-free rates with IMV/EEV formulations.
Claims Coverage
The partial content includes one independent claim. Across this claim, the inventive coverage is centered on producing a non-targeted-specificity poxvirus composition containing IMV and EEV above a threshold, using packaging-cell infection and homogenizer-based recovery followed by a defined downstream clarification, concentration, and diafiltration scheme, while being free from animal products and not using nuclease.
Non-targeted-specificity poxvirus production process with IMV and EEV above 1%
A process for producing a composition of a wild type, an attenuated, and/or a recombinant poxvirus with no targeted infection specificity, wherein the poxvirus composition comprises intracellular mature virus (IMV) and extracellular enveloped virus (EEV) with more than 1% EEV.
Packaging-cell infection and homogenizer recovery of poxviral particle mixture
Preparing a cell culture of packaging cells, infecting the cell culture to obtain infected cells, culturing to produce poxviral particles, and recovering poxviral particles from the culture supernatant and from packaging cells disrupted by high speed homogenizer to obtain a poxviral particle mixture.
Depth filtration clarification followed by microfiltration concentration and diafiltration
Clarifying the poxviral particle mixture thereby removing cellular debris, wherein said clarifying comprises depth filtration, concentrating the clarified poxviral particle mixture wherein said concentrating comprises microfiltration to obtain a concentrated poxviral particle mixture, and diafiltrating the concentrated poxviral mixture.
Animal-product-free and nuclease-free process using CEF packaging cells
A process that is free from animal products and does not use nuclease, wherein said packaging cells are chicken embryo fibroblast (CEF) prepared by enzymatic digestion.
EEV proportion above 5%
The process further includes that the poxvirus composition contains more than 5% EEV.
Microfiltration and/or diafiltration with 0.1 µm to 0.15 µm membranes
Performing microfiltration and/or diafiltration using filtration membranes with a pore size between 0.1 µm and 0.15 µm.
The claim coverage centers on an animal-product-free, nuclease-free process that uses packaging-cell infection with homogenizer-based recovery, followed by depth filtration, microfiltration concentration, and diafiltration, to produce a poxvirus composition containing IMV and EEV with stated EEV thresholds. Additional dependent coverage specifies membrane pore size ranges.
Stated Advantages
Greater therapeutic/prophylactic efficacy for non-targeted-specificity EEV than IMV.
Documented Applications
Preclinical mouse use in HPV (TG4001) and MUC1 (TG4010) contexts where IMV/EEV formulations showed improved survival and tumor-free rates compared to IMV-only lots.
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