Method of producing pharmaceutical compositions comprising immunogenic chikungunya virus CHIKV-Delta5NP3
Inventors
FRITZER, ANDREA • Meinke, Andreas • Lundberg, Urban • Nebenführ, Mario • HEINDL-WRUSS, Jürgen • Schiegl, Robert • Leon, Arnaud
Assignees
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Publication Number
US-11357846-B2
Publication Date
2022-06-14
Expiration Date
Abstract
The present invention relates to a process for producing an immunogenic live attenuated Chikungunya virus, as well as pharmaceutical compositions comprising the same.
Core Innovation
The invention relates to immunogenic live attenuated CHIKV-ΔnsP3 vaccine materials produced as CHIKV-ΔnsP3 particles that express an E2 structural protein defined by the amino acid sequence of SEQ ID NO: 2. Serial passaging or adaptation on cell substrates generates reproducible point mutations in E2 that reduce immunogenicity, including especially E168K and also G55R.
The document further describes that the adaptation-associated E2 mutations correspond to decreases in neutralizing immunogenicity and decreases in plaque size. It states that E168K emerges early and frequently, and that immunogenicity is reduced as the frequency of E168K increases.
To address this, the document defines concepts for minimizing such mutations while maintaining high titers for industrial manufacturing, including limiting passage numbers, using controlled low MOI, and characterizing genetic heterogeneity using Sanger sequencing and next generation sequencing (NGS). It also notes mitigation of rescuing mutation A38S in nsP1 that can preserve immunogenicity on the E168K background.
The document additionally describes upstream and downstream manufacturing considerations and characterization of E2 sequence variants using defined SEQ ID NOs to distinguish immunogenic versus non-immunogenic particles or compositions. It characterizes genetic heterogeneity of the particles and supports composition definition for use as a pharmaceutical composition and vaccine.
Claims Coverage
Independent claim clm-00001 defines a pharmaceutical composition or vaccine composition containing CHIKV-ΔnsP3 particles with an E2 structural protein (SEQ ID NO: 2) bearing E168K and/or G55R, with 1–50% of particles expressing the mutated E2. The main inventive features focus on defining the E2 sequence and specifying a particle-level mutated E2 fraction.
Fractional presence of E168K and/or G55R E2 in CHIKV-ΔnsP3 particles
A pharmaceutical composition comprising CHIKV-ΔnsP3 particles expressing an E2 structural protein as defined by the amino acid sequence of SEQ ID NO: 2, where 1–50% of the CHIKV-ΔnsP3 particles present express an E2 structural protein having said E168K and/or G55R mutation.
E2 sequence definition by SEQ ID NO: 2 with E168K and/or G55R
CHIKV-ΔnsP3 particles expressing an E2 structural protein as defined by the amino acid sequence of SEQ ID NO: 2, and the E2 structural protein having an E168K and/or G55R mutation in the amino acid sequence of SEQ ID NO: 2.
The claim coverage centers on a composition-level specification of CHIKV-ΔnsP3 particles whose E2 sequence is defined by SEQ ID NO: 2 and carries E168K and/or G55R, with a defined fraction of particles expressing the mutated E2. Dependent claims further specify the composition as a vaccine and/or a lyophilized form, and introduce additional performance and production constraints based on sequence (SEQ ID NO: 1) and passaging limits.
Stated Advantages
Maintains high titers for industrial manufacturing while minimizing mutations.
Reduces immunogenicity loss associated with adaptation-associated E2 mutations.
Preserves immunogenicity by mitigating rescuing mutation A38S in nsP1 on the E168K background.
Documented Applications
Use as a pharmaceutical composition.
Use as a vaccine.
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