Telomelysin/GFP-expressing recombinant virus
Inventors
Fujiwara, Toshiyoshi • Tanaka, Noriaki • Kyo, Satoru • Mizuguchi, Hiroyuki • Hayakawa, Takao
Assignees
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Publication Number
US-7943373-B2
Publication Date
2011-05-17
Expiration Date
Abstract
The present invention provides a reagent for cancer cell detection or cancer diagnosis. The present invention relates to a reagent for cancer cell detection, comprising a recombinant virus where a replication cassette comprising a promoter from human telomerase, an E1A gene, an IRES sequence and an E1B gene in this order is integrated in E1 region of the viral genome and a labeling cassette comprising a gene encoding a labeling protein and a promoter capable of regulating the expression of the gene encoding the labeling protein is integrated in E3 region of the viral genome.
Core Innovation
The invention relates to a recombinant adenovirus reagent for cancer cell detection, in which a replication cassette is integrated in the E1 region of the adenoviral genome. The replication cassette includes an hTERT promoter, an adenoviral E1A gene, an IRES sequence, and an adenoviral E1B gene in this order, and this arrangement supports telomerase-specific replication driven by the hTERT promoter.
The recombinant adenovirus further includes a labeling cassette integrated into the E3 region, wherein a cytomegalovirus promoter is operably linked to a gene encoding GFP or EGFP. In this labeling cassette design, the region encoding adenovirus death protein (ADP) is removed from the adenoviral genome while the rest of the E3A and E3B regions remain present, and this design delays cytotoxicity to improve fluorescence-based visualization.
The document also describes a related cell-death-inducing version in which an E3-region death-induction gene cassette is included and controlled by a suitable promoter, with death proteins such as PA28, TRAIL, and AU5 cited as examples. Overall, the disclosed constructs are positioned to enable detection, diagnosis, and visualization of cancer cells and tumors using fluorescence, including evaluation in model systems showing GFP fluorescence preferentially in cancer cells and tumors.
Claims Coverage
The document provides coverage through one independent claim directed to a recombinant adenovirus reagent for cancer cell detection, with inventive features centered on the specific arrangement of an hTERT-driven replication cassette in the E1 region and a CMV-driven GFP/EGFP labeling cassette in the E3 region with ADP removed while E3A/E3B remain. Dependent claims further refine use contexts and sequence definition via SEQ ID references.
hTERT-driven replication cassette integrated in the E1 region
A recombinant adenovirus where a replication cassette comprising an hTERT promoter, an adenoviral E1A gene, an IRES sequence and an adenoviral E1B gene in this order is integrated in the E1 region of the adenoviral genome.
CMV-driven GFP/EGFP labeling cassette integrated in the E3 region with ADP removed
A labeling cassette comprising a gene encoding GFP or EGFP and a cytomegalovirus promoter operably linked to the gene encoding GFP or EGFP is integrated into the E3 region of the adenoviral genome such that the region encoding adenovirus death protein (ADP) is removed from the adenoviral genome, wherein the rest of the E3A and E3B region are still present in the adenoviral genome.
In vivo cancer detection, diagnosis, or navigation surgery use
The reagent is used for in vivo cancer detection, in vivo diagnosis, or navigation surgery.
Specific sequence identities for hTERT promoter, E1A, IRES, and E1B via SEQ ID numbers
The hTERT promoter, adenoviral E1A gene, IRES sequence and adenoviral E1B gene are each defined by particular nucleotide sequences using SEQ ID NO:1, SEQ ID NO:2, SEQ ID NO:4 and SEQ ID NO:3, respectively.
Overall claim coverage centers on the structural genome configuration of the recombinant adenovirus: an hTERT promoter–E1A–IRES–E1B replication cassette integrated in the E1 region, together with a CMV promoter–GFP/EGFP labeling cassette integrated in the E3 region where ADP is removed while E3A/E3B remain, enabling fluorescence-based cancer cell detection. Dependent coverage specifies in vivo detection, diagnosis, or navigation surgery and defines key cassette components by SEQ ID numbers.
Stated Advantages
Delays cytotoxicity to improve fluorescence-based visualization.
Provides telomerase-specific replication supporting fluorescence in cancer cells and tumors.
Documented Applications
In vivo cancer detection or diagnosis.
Navigation surgery.
Ex vivo detection in blood by flow cytometry.
In vivo visualization including intratumoral visualization and visualization of lymph nodes, including sentinel lymph node or lymph-node visualization.
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