Clonal strains of attenuated vaccinia viruses and methods of use thereof
Inventors
Szalay, Aladar A. • Chen, Nanhai G. • Yu, Yong A. • Zhang, Qian
Assignees
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Publication Number
US-12338464-B2
Publication Date
2025-06-24
Expiration Date
Abstract
Clonal strains of vaccinia viruses are provided. Also provided are methods of identifying and isolating attenuated and oncolytic clonal strains from virus preparations. Modified recombinant forms of the clonal strains also are provided. The clonal strains and virus preparations can be used for diagnostic and therapeutic methods, in particular for therapy and diagnosis or monitoring treatment of proliferative disorders, including neoplastic diseases, such as, but are not limited to, solid tumors and blood cancers.
Core Innovation
The invention provides isolated clonal LIVP strains and recombinant LIVP strains derived from the isolated clonal LIVP strains. The clonal LIVP strains are defined so that the genome is not modified to contain non-viral heterologous nucleic acid containing an open reading frame encoding a non-viral heterologous protein, and the genome includes specified nucleotide segments from SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, or SEQ ID NO:8, or sequences having defined sequence identity to those segments, while further constraining identity to SEQ ID NO:10 and excluding the sequence of nucleotides set forth in SEQ ID NO:10.
A core aspect of the invention is functional improvement relative to the strain designated GLV-1h68. The clonal LIVP strain has greater anti-tumorigenicity or reduced toxicity or both compared to GLV-1h68, and clonal isolates are prepared from an LIVP virus sample, assayed for toxicity and anti-tumorigenicity, and selected for exhibiting greater anti-tumorigenicity and/or reduced toxicity compared to GLV-1h68.
The invention also extends to recombinant LIVP virus strains that are derivatives of a clonal LIVP strain, where the recombinant strain retains the ability to replicate but is modified in its genomic sequence compared to the clonal LIVP strain. In the recombinant strains, nucleic acid is deleted, replaced with heterologous nucleic acid, or heterologous nucleic acid is inserted, including cases where the deletion, insertion, or replacement occurs in a non-essential gene or region.
Claims Coverage
The independent claim coverage is provided for isolated clonal LIVP strains, a cell culture or isolated cell comprising such strains, and recombinant LIVP virus derivatives. Across these independent claims, the inventive features center on genome sequence constraints tied to SEQ ID NOs, comparative anti-tumorigenicity and/or toxicity versus GLV-1h68, absence of non-viral heterologous open reading frames in the clonal strains, and recombinant strains that retain replication while permitting genome modification, with one claim further restricting edits to a non-essential gene or region.
Isolated clonal LIVP strain with constrained genome and improved anti-tumorigenicity or reduced toxicity
The genome is not modified to contain non-viral heterologous nucleic acid containing an open reading frame encoding a non-viral heterologous protein; it comprises specified nucleotide sequences selected from SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, or SEQ ID NO:8, or sequences having at least 95% sequence identity to those segments; the clonal strain has greater anti-tumorigenicity or reduced toxicity or both compared to GLV-1h68; and the genome comprises a sequence with at least 90% sequence identity to SEQ ID NO:10 but does not comprise the sequence of nucleotides set forth in SEQ ID NO:10.
Cell culture or isolated cell comprising clonal LIVP strain with constrained genome and improved anti-tumorigenicity or reduced toxicity
A cell culture or isolated cell comprises a clonal LIVP strain with the same genome constraints, comparative anti-tumorigenicity or toxicity limitation versus GLV-1h68, and selection from clonal isolates prepared from an LIVP virus sample by assaying toxicity and anti-tumorigenicity and selecting a clonal isolate exhibiting greater anti-tumorigenicity and/or reduced toxicity compared to GLV-1h68.
Recombinant LIVP derivative with replication retained and improved anti-tumorigenicity or reduced toxicity
A recombinant LIVP virus strain is a derivative of a clonal LIVP strain, comprises deletion of nucleic acid or replacement of nucleic acid with heterologous nucleic acid or insertion of heterologous nucleic acid in the genome of the clonal LIVP strain, retains the ability to replicate, and has greater anti-tumorigenicity or reduced toxicity or both compared to GLV-1h68.
Recombinant LIVP derivative with deletion, insertion, or replacement in a non-essential gene or region
The recombinant LIVP virus strain is a derivative of a clonal LIVP strain, retains the ability to replicate, is modified in genomic sequence compared to the genome of the clonal LIVP strain, has greater anti-tumorigenicity or reduced toxicity or both compared to GLV-1h68, and the deletion, insertion, or replacement is in a non-essential gene or region in the genome of the clonal LIVP virus strain.
Overall, the independent claims define isolated clonal LIVP strains and recombinant derivatives by specified genome sequence inclusion and exclusion constraints, require greater anti-tumorigenicity and/or reduced toxicity relative to GLV-1h68, and require that clonal strains lack a non-viral heterologous open reading frame encoding a non-viral heterologous protein. Recombinant claims additionally require retention of replication while enabling deletion, replacement, or insertion of nucleic acid, with one claim further requiring modification in a non-essential gene or region.
Stated Advantages
greater anti-tumorigenicity compared to GLV-1h68
reduced toxicity compared to GLV-1h68
greater anti-tumorigenicity or reduced toxicity or both compared to GLV-1h68
Documented Applications
treating a proliferative disorder in a subject by administering a pharmaceutical composition that includes the isolated recombinant strain
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