DNA vector and transformed tumor cell vaccines
Inventors
Lawman, Michael J. P. • Lawman, Patricia D. • RAMIYA, VIJAY • GENTILINI, MEGHAN
Assignees
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Publication Number
US-10751400-B2
Publication Date
2020-08-25
Expiration Date
Abstract
Customized whole cell cancer vaccines can be produced from autologous (ex vivo or in situ) or allogeneic human or veterinary patient cell lines. Cells are transformed with S. pyogenes DNA that expresses an Emm protein on the cell surface and cytosol. Treatment of cancer patients with an Emm vector vaccine induces an immunologic response to the cancer by enhancing immunogenicity of a tumor. Emm vaccines can be used in patients where the cancer is not identified due to lower tumor burden or used to treat a specific cancer and subsequently treat for a second type that may have arisen through metastasis.
Core Innovation
The disclosed subject matter relates to membrane anchoring of an M serotype 55 group A Streptococcus protein by expressing a modified DNA construct encoding a membrane anchoring protein. The method inserts a first DNA encoding the amino acid sequence of SEQ ID NO: 7 into the N-terminal signal region of a DNA encoding the M serotype 55 group A Streptococcus protein, and inserts a second DNA encoding the amino acid sequence of SEQ ID NO: 9 into the C-terminal anchor region.
The encoded membrane anchoring protein is expressed from the modified DNA, and the expressed protein is described as a membrane anchoring protein. The modification positions an N-terminal signal region and a C-terminal anchor region so that the encoded protein is expressed in a membrane anchoring form. Dependent refinements include specifying the M serotype 55 group A Streptococcus protein, the Gram-positive bacterial cell wall nature of the protein, and alternative encoded protein sequence content of SEQ ID NO: 5.
The broader document context additionally describes therapeutic DNA vector cancer vaccines using an Emm-based approach with S. pyogenes emm, including Emm55. In this context, Emm expression is positioned to enhance innate and adaptive anti-tumor immunity via APC activation, danger signaling, and cross-priming, with reported immune responses and tumor outcomes associated with in vivo and in situ transformation approaches and whole-cell vaccines derived from transformed tumor cells.
Claims Coverage
The partial content identifies one independent claim with dependent claims. The inventive coverage centers on constructing an M serotype 55 group A Streptococcus protein as a membrane anchoring protein by inserting SEQ ID NO: 7 and SEQ ID NO: 9 into specified regions, with dependent refinements specifying the source/type of the M serotype 55 protein and the anchoring context to a cell membrane.
Membrane anchoring protein by N-terminal and C-terminal sequence insertion
A method for preparing a membrane anchoring protein comprising inserting a first DNA encoding the amino acid sequence of SEQ ID NO: 7 into the N-terminal signal region of a DNA encoding an M serotype 55 group A Streptococcus protein, and inserting a second DNA encoding the amino acid sequence of SEQ ID NO: 9 into the C-terminal anchor region of the DNA encoding said protein, wherein the encoded protein expressed from the modified DNA is a membrane anchoring protein.
Modified M serotype 55 group A Streptococcus protein refinement
The membrane anchoring method wherein the M serotype 55 group A Streptococcus protein is a Gram-positive bacterial cell wall protein.
Alternative encoded protein sequence option
The membrane anchoring method wherein the encoded protein has the amino acid sequence of SEQ ID NO: 5.
Cell membrane anchoring context
The membrane anchoring method wherein the membrane anchoring protein anchors to a cell membrane.
Eukaryotic membrane anchoring context
The membrane anchoring method wherein the cell membrane is a eukaryotic membrane.
Prokaryotic membrane anchoring context
The membrane anchoring method wherein the cell membrane is a prokaryotic membrane.
Overall, the claim coverage centers on preparing a membrane anchoring protein based on an M serotype 55 group A Streptococcus protein by inserting SEQ ID NO: 7 into an N-terminal signal region and inserting SEQ ID NO: 9 into a C-terminal anchor region so that the expressed protein is a membrane anchoring protein. Dependent refinements further specify the Gram-positive bacterial cell wall nature of the protein, an alternative encoded protein sequence, and whether the anchoring membrane is eukaryotic or prokaryotic.
Stated Advantages
Potentially enhances innate and adaptive anti-tumor immunity via APC activation, danger signaling, and cross-priming.
Reported increases in anti-tumor antibody responses.
Reported increases in immune-cell infiltration and lesion regression.
Reported improved survival/QoL.
Documented Applications
In vivo and in situ transformation for Emm-based therapeutic DNA vector cancer vaccines in solid tumors, including direct intratumoral injection of DNA/plasmid and transformation producing autologous or allogeneic whole-cell vaccines.
Whole-cell vaccine preparation using transformed patient or donor cell lines for cancer immunotherapy.
Preclinical/veterinary examples including canine lymphoma, mammary adenocarcinoma, melanoma, and transitional cell carcinoma.
Expression detection and evaluation of Emm55 expression, including detection described with Western blot and flow cytometry.
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