Detection of differential expression of protein using gel-free proteomics
OwnerCynthia J. Brameinventor
Life of this patent
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Abstract
Methods and reagents for analyzing differential expression and/or abundance of distinct membrane-associated polypeptide samples, particularly integral membrane polypeptide samples are provided. Also provided are methods for screening pharmaceutical components that can affect expression or abundance of certain membrane-associated polypeptides; methods for identification of drug targets; and methods for diagnosis of certain disease states. Business methods for conducting a pharmaceutical business based on the result of using the above methods are also provided.
Claims17
A method for identifying changes in membrane-associated polypeptides, comprising:
- (i) providing a test sample of membrane-associated polypeptides isolated from a test cell(s);
- (ii) by mass spectrometry using a quantitative mass analyzer, determining the levels of polypeptides in said test sample;
- (iii) comparing the level of one or more of the polypeptides from said test sample with levels of respective polypeptides from a reference sample;
- (iv) identifying the sequences of polypeptides in the test sample which, relative to the reference sample, have altered abundance and/or altered levels of post-translational modification.
A method for identification of membrane-associated polypeptide targets of a compound, comprising:
- (i) providing two test samples of membrane-associated polypeptides isolated from two test cells, wherein one test sample is a reference sample and the other is a sample treated by said compound;
- (ii) by mass spectrometry using a quantitative mass analyzer, determining the levels of polypeptides in said test samples;
- (iii) comparing the level of one or more of the polypeptides from said treated test sample with levels of respective polypeptides from said reference sample;
- (iv) identifying the sequences of polypeptides in said treated sample which, relative to the reference sample, have altered abundance and/or altered levels of post-translational modification(s), thereby identifying the membrane-associated polypeptide targets of said compound.
A method for identifying a compound which alters the abundance of a membrane-associated polypeptide in a sample, comprising:
- (i) providing a reference sample and a plurality of test samples of membrane-associated polypeptides, each isolated from a test cell treated by a specific test compound;
- (ii) by mass spectrometry using a quantitative mass analyzer, determining the levels of said membrane-associated polypeptides in said test samples and said reference samples;
- (iii) comparing the level of one or more of said membrane-associated polypeptides from said test samples with levels of respective polypeptides from said reference sample;
- (iv) identifying the test sample which, relative to the reference sample, have altered abundance, thereby identifying the test compound responsible for the change.
A method for identifying a compound which alters the levels of post-translational modification of a membrane-associated polypeptide in a sample, comprising:
- (i) providing a reference sample and a plurality of test samples of membrane-associated polypeptides, each isolated from a test cell treated by a specific test compound;
- (ii) by mass spectrometry using a quantitative mass analyzer, determining the levels of said membrane-associated polypeptides in said test samples and said reference samples;
- (iii) comparing the level of one or more of said membrane-associated polypeptides from said test samples with levels of respective polypeptides from said reference sample;
- (iv) identifying the test sample which, relative to the reference sample, have altered levels of post-translational modification, thereby identifying the test compound responsible for the change.
A method of conducting a pharmaceutical business, comprising:
- (i) by the above-described method, determining the identity of a target polypeptide isolated on the basis of the polypeptide being (a) having a differential cellular localization of interest;
- (b) having a differential expression pattern of interest;
- (c) having a differential post-translational modification of interest;
- or (d) having a differential abundance of interest;
- (ii) identifying compounds by their ability to alter the abundance or subcellular localization or post-translational modification of the target polypeptide;
- (iii) conducting therapeutic profiling of compounds identified in step (ii), or further analogs thereof, for efficacy and toxicity in animals;
- and,
- (iv) formulating a pharmaceutical preparation including one or more compounds identified in step (iii) as having an acceptable therapeutic profile.
A method of conducting a pharmaceutical business, comprising:
- (i) by the above-described method, determining the identity of a target polypeptide isolated on the basis of the polypeptide:
- (a) having a differential cellular localization of interest, (b) having a differential expression pattern of interest, (c) having a differential post-translational modification of interest, or (d) having a differential abundance of interest;
- (ii) (optionally) conducting therapeutic profiling of the target gene for efficacy and toxicity in animals;
- and
- (iii) licensing, to a third party, the rights for further drug development of inhibitors or activators of the target gene.
Dependencies are inferred from the claim text.