Individualized cancer therapy
Inventors
Shanahan, David • Nemunaitis, John J. • Senzer, Neil • Maples, Phillip B. • Rao, Donald
Assignees
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Publication Number
US-9382589-B2
Publication Date
2016-07-05
Expiration Date
Abstract
In certain preferred embodiments, the invention provides methods for treating cancer, which comprise (a) obtaining a specimen of cancer tissue from a patient; (b) obtaining a specimen of normal tissue in the proximity of the cancer tissue from such patient; (c) extracting total protein and RNA from the cancer tissue and normal tissue; (d) obtaining a protein expression profile of the cancer tissue and normal tissue using 2D DIGE and mass spectrometry; (e) identifying proteins that are expressed in such cancer tissue at significantly different levels than in the normal tissue; (f) obtaining a gene expression profile of the cancer tissue and normal tissue using microarray technology and comparing the results thereof to the protein expression profile; (g) prioritizing over-expressed proteins by assessing the connectivity thereof to other cancer-related or stimulatory proteins; (h) designing an appropriate RNA interference expression cassette to, directly or indirectly, modulate the expression of genes encoding such prioritized proteins; (i) incorporating said cassette into an appropriate delivery vehicle; and (j) providing the patient with an effective amount of the delivery vehicle to, directly or indirectly, modify the expression (i.e., production) of such proteins.
Core Innovation
A method for treating cancer obtains a specimen of cancer tissue and normal tissue from a patient and extracts total protein and RNA from the cancer tissue and the normal tissue. A protein and an RNA expression profile of the cancer tissue and the normal tissue are obtained and used to compare protein and RNA expression results. Over-expressed proteins are identified in the cancer tissue where a protein is considered over-expressed if the protein is found in the cancer tissue at higher levels than in normal tissue.
At least one prioritized protein for targeting in a cancer associated pathway is identified by connecting the over-expressed protein and RNA to the cancer associated pathway. An RNA interference expression cassette is made to modulate the expression of at least one gene encoding the prioritized protein, and a patient is provided with an effective amount of the RNA interference expression cassette. In related embodiments, the workflow includes obtaining a protein expression profile using 2D DIGE and mass spectrometry and comparing results to a gene expression profile obtained using one or more microarrays.
In further embodiments, the cassette is provided in the form of a plasmid and the plasmid comprises an RNA interference expression cassette encoding one or more enhanced shRNA molecules. In specific embodiments for targeted genes, RNA interference expression cassettes are prepared to encode nucleic acid sequences at least substantially complementary to mRNA transcripts encoded by the Stathmin 1 gene (SEQ ID NO:27) and are provided to a cancer cell. In additional embodiments, the method includes identifying over-expressed proteins and under-expressed proteins in the cancer tissue, and the method provides a standard chemo- or radiation-therapy before making and providing an RNA interference expression cassette that modulates expression of a prioritized protein after the standard chemo- or radiation-therapy.
Claims Coverage
The document includes four independent claims, covering paired cancer and normal tissue multi-omics profiling with pathway-connected prioritized proteins, a 2D DIGE/mass spectrometry and microarray embodiment with an RNAi plasmid encoding enhanced shRNA, Stathmin 1-targeted RNA interference expression cassettes, and a workflow that applies standard chemo- or radiation-therapy followed by RNAi modulation of prioritized pathway proteins identified from paired tissue expression profiling.
Paired cancer/normal tissue profiling to identify over-expressed proteins linked to cancer-associated pathways
The method obtains a specimen of cancer tissue and normal tissue, extracts total protein and RNA, obtains protein and RNA expression profiles, identifies one or more over-expressed proteins in the cancer tissue as higher levels than in normal tissue, compares the protein and RNA expression profile to a gene expression profile, and identifies at least one prioritized protein for targeting in a cancer associated pathway by connecting the over-expressed protein and RNA to the cancer associated pathway.
RNA interference expression cassette administration to modulate prioritized pathway proteins
The method makes an RNA interference expression cassette to modulate the expression of at least one gene encoding the prioritized protein and provides a patient with an effective amount of the RNA interference expression cassette.
2D DIGE and mass spectrometry protein profile with microarray gene profile and RNAi plasmid with enhanced shRNA
The method obtains a specimen of cancer tissue and normal tissue, extracts total protein and RNA, obtains a protein expression profile using 2D DIGE and mass spectrometry, identifies over-expressed proteins where higher levels are found in cancer tissue than in normal tissue, obtains a gene expression profile using one or more microarrays and compares to the protein expression profile, and identifies at least one prioritized protein for targeting by connecting the over-expressed protein to the cancer associated pathway; it makes a plasmid comprising an RNA interference expression cassette that encodes one or more enhanced shRNA molecules and provides the patient with an effective amount of the plasmid.
Stathmin 1 mRNA-targeted RNA interference expression cassettes to reduce growth or induce apoptosis
The method prepares one or more RNA interference expression cassettes encoding nucleic acid sequences at least substantially complementary to mRNA transcripts encoded by the Stathmin 1 gene (SEQ ID NO:27) and provides the one or more RNA interference expression cassettes to a cancer cell.
Standard chemo- or radiation-therapy followed by RNAi modulation of prioritized pathway proteins
The method obtains cancer tissue and normal tissue, extracts total protein and RNA, obtains a protein expression profile, identifies over-expressed proteins and under-expressed proteins where over-expressed means higher levels in cancer tissue than in normal tissue, obtains a gene expression profile and compares it to the protein expression profile, identifies at least one prioritized protein for targeting in a cancer associated pathway by connecting the over-expressed protein to the cancer associated pathway, provides a patient with a standard chemo- or radiation-therapy, makes an RNA interference expression cassette to modulate the expression of at least one prioritized protein of the cancer associated pathway after the standard chemo- or radiation-therapy, and provides the patient with an effective amount of the RNA interference expression cassette after the standard chemo- or radiation-therapy.
Overall, the independent claims converge on using paired cancer/normal tissue expression profiling to identify over-expressed proteins linked to cancer associated pathways and then administering RNA interference expression cassettes, including plasmid-based enhanced shRNA, to modulate prioritized targets; additional coverage includes a Stathmin 1 mRNA-targeting embodiment and an embodiment where RNAi modulation is performed after standard chemo- or radiation-therapy.
Stated Advantages
Not explicitly described in patent.
Documented Applications
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