Furin-knockdown and GM-CSF-augmented (FANG) cancer vaccine
Inventors
Nemunaitis, John J. • Senzer, Neil • Maples, Phillip B. • Rao, Donald
Assignees
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Publication Number
US-9132146-B2
Publication Date
2015-09-15
Expiration Date
Abstract
Compositions and methods for cancer treatment are discloses herein. More specifically the present invention describes an autologous cancer vaccine genetically modified for Furin knockdown and GM-CSF expression. The vaccine described herein attenuates the immunosuppressive activity of TGF-β through the use of bi-functional shRNAs to knock down the expression of furin in cancer cells, and to augment tumor antigen expression, presentation, and processing through expression of the GM-CSF transgene.
Core Innovation
The invention describes an autologous cancer vaccine concept that combines Furin-knockdown with GM-CSF augmentation using a bishRNAfurin/GMCSF expression vector plasmid. The vaccine includes a first nucleic acid insert encoding Granulocyte Macrophage Colony Stimulating Factor (GM-CSF) cDNA and a second nucleic acid insert encoding one or more short hairpin RNAs capable of hybridizing to a region of an mRNA transcript encoding furin.
The shRNA inhibits furin expression via RNA interference while incorporating cleavage dependent siRNA and cleavage independent miRNA motifs as defined by SEQ ID NO:2. The described approach is grounded in a rationale that furin-mediated activation of TGF-β2 isoforms underlies TGF-β2 immunosuppression, such that furin knockdown is intended to attenuate TGF-β2 activity.
In parallel, GM-CSF is used to enhance antigen expression, presentation, and processing in the autologous vaccine setting. The invention further specifies bi-functional shRNAfurin/GMCSF constructs and vector organization, including a first stem-loop structure and a second stem-loop structure with mismatches in a passenger strand capable of hybridizing to furin mRNA.
Claims Coverage
The relevant independent claims are clm-00001, clm-00006, and clm-00007, each directed to an autologous cell or composition for cancer treatment involving GM-CSF and furin inhibition by RNA interference using a defined bishRNAfurin/GMCSF or shRNAfurin construct. Across the independent claims, the core inventive features center on a plasmid or transfection context carrying a GM-CSF cDNA insert and a furin-targeting bi-functional shRNA that incorporates cleavage dependent siRNA and cleavage independent miRNA motifs.
BishRNAfurin/GMCSF expression vector plasmid for an autologous cell vaccine
An autologous cell vaccine comprising a bishRNAfurin/GMCSF expression vector plasmid having a first nucleic acid insert encoding GM-CSF cDNA under a promoter and a second nucleic acid insert encoding one or more shRNAs hybridizing to a region of a furin mRNA transcript to inhibit furin expression via RNA interference, wherein the shRNA incorporates cleavage dependent siRNA and cleavage independent miRNA motifs and comprises SEQ ID NO:2.
Autologous FANG cancer vaccine composition via transfected autologous cancer cells
An autologous furin-knockdown and GM-CSF augmented (FANG) cancer vaccine composition comprising an autologous cancer cell transfected with a vector having a first nucleic acid insert encoding GM-CSF cDNA under a promoter and a second nucleic acid insert encoding one or more shRNAs hybridizing to a furin mRNA region to inhibit furin expression via RNA interference, wherein the shRNA incorporates cleavage dependent siRNA and cleavage independent miRNA motifs of SEQ ID NO:2.
Bi-functional shRNAfurin providing cleavage and sequestration for furin inhibition
An autologous cell vaccine composition for cancer treatment by inhibition of furin expression via RNA interference, comprising a bishRNAfurin/GMCSF expression vector plasmid with a first nucleic acid insert encoding GM-CSF cDNA under a promoter and a second nucleic acid insert encoding bi-functional shRNAfurin providing a single targeted site for both a cleavage and a sequestering mechanism of RNA interference, wherein the bi-functional shRNAfurin includes first and second stem-loop structures with mismatches in a passenger strand capable of hybridizing to a furin mRNA region, and wherein the shRNA comprises SEQ ID NO:2.
The independent claim set consistently covers autologous cancer vaccine compositions using a GM-CSF cDNA insert and a furin-targeting bi-functional shRNA that inhibits furin expression via RNA interference with cleavage dependent siRNA and cleavage independent miRNA motifs. One independent claim further emphasizes a design that provides both cleavage and a sequestering mechanism through the stem-loop configuration and passenger-strand mismatches.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
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