Individualized cancer therapy

Abstract

In certain embodiments, the invention provides methods for treating cancer, comprising: obtaining a specimen of cancer tissue and normal tissue from a patient; extracting total protein; obtaining a protein expression profile; identifying over-expressed proteins; comparing the protein expression profile to a gene expression profile; identifying at least one prioritized protein target; designing a first RNA interference expression cassette; designing a first RNA interference expression cassette to modulate the expression of at least one gene encoding; incorporating the first cassette into a delivery vehicle; and providing a patient with an effective amount of the first delivery vehicle.

1 . A method for treating cancer, comprising:
 obtaining a specimen of cancer tissue and normal tissue from a patient;   extracting total protein and RNA from the cancer tissue and normal tissue;   obtaining a protein expression profile of the cancer tissue and normal tissue;   identifying over-expressed proteins in the cancer tissue;   comparing the protein expression profile to a gene expression profile;   identifying at least one prioritized protein target;   designing a RNA interference expression cassette to modulate the expression of at least one gene encoding the prioritized target protein;   incorporating the RNA interference expression cassette into a delivery vehicle; and   providing a patient with an effective amount of the delivery vehicle sufficient to treat the cancer.   
     
     
         2 . The method of  claim 1 , wherein the normal tissue is extracted from an area in close proximity to the cancer tissue or from an area of the tissue of origin of the cancer tissue. 
     
     
         3 . The method of  claim 1 , wherein the cancer and normal tissue is extracted using laser capture microdissection. 
     
     
         4 . The method of  claim 1 , wherein the protein expression profile is obtained using at least one of 2D DIGE and mass spectrometry; HPLC and mass spectroscopy; or using one or more microarrays. 
     
     
         5 . The method of  claim 1 , wherein the proteins are considered to be over-expressed if the proteins are found in the cancer tissue at higher levels than in the normal tissue. 
     
     
         6 . The method of  claim 1 , wherein the protein is overexpressed if they are at least two-fold higher in the cancer tissue than in normal tissue. 
     
     
         7 . The method of  claim 1 , wherein the RNA interference expression cassette encodes one or more bifunctional shRNA molecules, shRNA molecules, or siRNA molecules. 
     
     
         8 . The method of  claim 1 , wherein the delivery vehicle is selected from the group consisting of immunoliposomes, immunolipoplexes, small molecule targeted lipoplexes, RGD targeted nanoparticles, RGD targeted liposomes, nanoparticles, aptamers, dendrimers, chitosan, pegylated derivatives thereof, and oncolytic viral vectors. 
     
     
         9 . The method of  claim 1 , further comprising measuring whether the RNA interference expression cassette is capable of suppressing the expression of one or more genes that encode the at least one or more prioritized proteins in vitro prior to providing said delivery vehicle to a patient. 
     
     
         10 . The method of  claim 1 , further comprising the steps of:
 measuring whether the at least one prioritized protein exhibits a reduced expression level after provision of the delivery vehicle to the patient; and   continuing treatment if the expression level of the prioritized protein target decreases or designing a second protein expression inhibitor selected from at least one prioritized protein target identified by connectivity to the prioritized protein.   
     
     
         11 . The method of  claim 1 , wherein the RNA interference expression cassette comprises a tumor-specific promoter. 
     
     
         12 . The method of  claim 1 , wherein the RNA interference expression cassette comprises a bi-functional short hairpin RNA that targets the RNA that expresses the prioritized proteins for degradation and sequestration. 
     
     
         13 . The method of  claim 1 , wherein the one or more RNA interference expression cassettes are provided to cancer cells via a delivery vehicle selected from the group consisting of immunoliposomes, immunolipoplexes, RGD targeted nanoparticles, RGD targeted liposomes, nanoparticles, aptamers, dendrimers, chitosan, pegylated derivatives thereof, and oncolytic viral vectors. 
     
     
         14 . The method of  claim 1 , further comprising the steps of:
 extracting total protein and RNA from the treated cancer tissue;   identifying over-expressed proteins in the treated cancer tissue;   designing a second RNA interference expression cassette to modulate the expression of a second prioritized protein in the treated tissue;   incorporating the second cassette into a second delivery vehicle; and   providing the previously treated patient with an effective amount of the second delivery vehicle in an amount sufficient to treat the cancer.   
     
     
         15 . A method for treating cancer, comprising:
 obtaining a specimen of cancer tissue and normal tissue from a patient;   extracting total protein and RNA from the cancer tissue and normal tissue;   obtaining a protein expression profile of the cancer tissue and normal tissue;   identifying over-expressed proteins in the cancer tissue;   identifying at least one prioritized protein target by assessing connectivity of each over-expressed protein;   designing a first RNA interference expression cassette to modulate the expression of at least one gene encoding the prioritized target protein;   incorporating the first cassette into a first delivery vehicle; and   providing a patient with an effective amount of the first delivery vehicle sufficient to treat the cancer.   
     
     
         16 . The method of  claim 15 , wherein the normal tissue is extracted from an area in close proximity to the cancer tissue or from an area of the tissue of origin of the cancer tissue. 
     
     
         17 . The method of  claim 15 , wherein the cancer and normal tissue is extracted using laser capture microdissection. 
     
     
         18 . The method of  claim 15 , wherein the protein expression profile is obtained using at least one of 2D DIGE and mass spectrometry; HPLC and mass spectroscopy; or using one or more microarrays. 
     
     
         19 . The method of  claim 15 , wherein the proteins are considered to be over-expressed if the proteins are found in the cancer tissue at higher levels than in the normal tissue. 
     
     
         20 . The method of  claim 15 , wherein the protein is overexpressed if they are at least two-fold higher in the cancer tissue than in normal tissue. 
     
     
         21 . The method of  claim 15 , wherein the RNA interference expression cassette encodes one or more bifunctional shRNA molecules, shRNA molecules, or siRNA molecules. 
     
     
         22 . The method of  claim 15 , wherein the delivery vehicle is selected from the group consisting of immunoliposomes, immunolipoplexes, small molecule targeted lipoplexes, RGD targeted nanoparticles, RGD targeted liposomes, nanoparticles, aptamers, dendrimers, chitosan, pegylated derivatives thereof, and oncolytic viral vectors. 
     
     
         23 . The method of  claim 15 , further comprising the steps of:
 measuring whether the at least one prioritized protein exhibits a reduced expression level after provision of the delivery vehicle to the patient; and   continuing treatment if the expression level of the prioritized protein target decreases; or   designing a second protein expression inhibitor selected from at least a second prioritized protein target identified by connectivity to the prioritized protein, wherein the second prioritized protein target is an effector downstream from the first prioritized protein target.   
     
     
         24 . The method of  claim 15 , further comprising the steps of:
 measuring whether the at least one prioritized protein exhibits a reduced expression level after provision of the delivery vehicle to the patient; and   continuing treatment if the expression level of the prioritized protein target decreases; or   designing a second protein expression inhibitor selected from at least a second prioritized protein target identified by connectivity to the prioritized protein, wherein the second prioritized protein target is an upstream effector in the case of feedback loop of the first prioritized protein target.   
     
     
         25 . A method for treating cancer, comprising:
 obtaining a specimen of cancer tissue and normal tissue from a patient;   extracting total protein and RNA from the cancer tissue and normal tissue;   obtaining a protein expression profile of the cancer tissue and normal tissue;   identifying over-expressed proteins in the cancer tissue;   comparing the protein expression profile to a gene expression profile;   identifying at least one prioritized protein target by assessing connectivity of each said over-expressed protein to other cancer-related or stimulatory proteins;   designing a first RNA interference expression cassette to modulate the expression of at least one gene encoding the prioritized target protein;   designing a first RNA interference expression cassette to modulate the expression of at least one gene encoding a protein of higher priority in the signaling pathway in which the first protein is a component;   incorporating the first cassette into a first delivery vehicle;   providing a patient with an effective amount of the first delivery vehicle;   extracting total protein and RNA from the treated cancer tissue;   identifying over-expressed proteins in the treated cancer tissue;   designing a second RNA interference expression cassette to modulate the expression of a second prioritized protein in the treated tissue;   incorporating the second cassette into a second delivery vehicle;   providing the previously treated patient with an effective amount of the second delivery vehicle;   identifying a novel protein signal following prior treatment with protein specific knockdown;   identifying a gene mutation provided by gene sequencing/microarray on assessment of other protein signals; and   identifying of a novel protein signal as a result of determination of the gene mutation and assessment of other protein signals.