Abstract: Ischemic conditions are a leading cause of death for both men and women. Ischemia, a condition characterized by reduced blood flow and oxygen to an organ. Re-establishment of blood flow, or reperfusion, and re-oxygenation of the affected area following an ischemic episode is critical to limit irreversible damage. However, reperfusion also associates potentially damaging consequences. For instance, increased vascular permeability is an important contributor to edema and tissue damage following ischemic events. Here the inventors shows that genetic inhibition of PI3K-C2? reduces cerebral infarction in two ischemia/reperfusion (I/R) models and improves neurological outcome. The genetic inhibition stabilizes the blood—brain barrier (BBB) after ischemic stroke and reduces inflammation.

Abstract: Ischemic conditions are a leading cause of death for both men and women. Ischemia, a condition characterized by reduced blood flow and oxygen to an organ. Re-establishment of blood flow, or reperfusion, and re-oxygenation of the affected area following an pot ischemic episode is critical to limit irreversible damage. However, reperfusion also associates potentially damaging consequences. For NI instance, increased vascular permeability is an important contributor to edema and tissue damage following ischemic events. Here the inventors shows that genetic inhibition of PI3K-C2? reduces cerebral infarction in two ischemia/reperfusion (UR) models and improves neurological outcome. The genetic inhibition stabilizes the blood-brain barrier (BBB) after ischemic stroke and reduces inflammation.

Abstract: The present invention provides a method for identifying differential activation of a bisubstrate protein modifying enzyme between samples, comprising: (i) incubating a first sample with x different concentrations of the non-protein substrate of said enzyme, wherein x is 2 or greater than 2; (ii) quantifying modification of a polypeptide in said sample at each of the x different concentrations of the non-protein substrate; (iii) determining the affinity of said enzyme for said non-protein substrate; (iv) repeating steps (i) to (iii) for a second or subsequent sample; and (v) comparing the affinity of said enzyme for said non-protein substrate between said samples; wherein a difference in affinity of said enzyme for said non-protein substrate between samples is indicative of differential activation of said enzyme between samples. The present invention also provides a method for identifying an in vivo substrate of a bisubstrate protein modifying enzyme.

Abstract: The present invention provides a method for quantifying modified peptides in a sample, the method comprising: (a) obtaining peptides from the sample; (b) adding reference modified peptides to the peptides obtained in step (a) to produce a mixture of peptides and reference modified peptides; (c) carrying out mass spectrometry (MS) on said mixture of peptides and reference modified peptides to obtain data relating to the peptides in the sample; and (d) comparing the data relating to the peptides in the sample with data in a database of modified peptides using a computer program; wherein the database of modified peptides is compiled by a method comprising: (i) obtaining peptides from a sample; (ii) enriching modified peptides from the peptides obtained in step (i); (iii) carrying out liquid chromatography-tandem mass spectrometry (LC-MS/MS) on the enriched modified peptides obtained in step (ii); (iv) comparing the modified peptides detected in step (iii) to a known reference database in order to identify the mo

Abstract: The present invention provides a method for identifying differential activation of a bisubstrate protein modifying enzyme between samples, comprising: (i) incubating a first sample with x different concentrations of the non-protein substrate of said enzyme, wherein x is 2 or greater than 2; (ii) quantifying modification of a polypeptide in said sample at each of the x different concentrations of the non-protein substrate; (iii) determining the affinity of said enzyme for said non-protein substrate; (iv) repeating steps (i) to (iii) for a second or subsequent sample; and (v) comparing the affinity of said enzyme for said non-protein substrate between said samples; wherein a difference in affinity of said enzyme for said non-protein substrate between samples is indicative of differential activation of said enzyme between samples. The present invention also provides a method for identifying an in vivo substrate of a bisubstrate protein modifying enzyme.

Abstract: The present invention provides a method for quantifying modified peptides in a sample, the method comprising: (a) obtaining peptides from the sample; (b) adding reference modified peptides to the peptides obtained in step (a) to produce a mixture of peptides and reference modified peptides; (c) carrying out mass spectrometry (MS) on said mixture of peptides and reference modified peptides to obtain data relating to the peptides in the sample; and (d) comparing the data relating to the peptides in the sample with data in a database of modified peptides using a computer programme; wherein the database of modified peptides is compiled by a method comprising: (i) obtaining peptides from a sample; (ii) enriching modified peptides from the peptides obtained in step (i); (iii) carrying out liquid chromatography-tandem mass spectrometry (LC-MS/MS) on the enriched modified peptides obtained in step (ii); (iv) comparing the modified peptides detected in step (iii) to a known reference database in order to identify the

Abstract: The disclosure relates to methods of analyzing the enzymatic activity of enzymes in samples containing a plurality of enzymes, using mass spectrometry. Immobilized substrates are employed. Purified enzymes and enzymes from crude cell lysates can be analyzed using the disclosed methods.

Abstract: The disclosure relates to methods of quantitatively analyzing the enzymatic activity of enzymes in samples containing a plurality of enzymes, using mass spectrometry. Isotopically labeled standards are employed. Purified enzymes and enzymes from crude cell lysates may be analyzed using the disclosed methods. As little as 0.02 pg of cell lysate may be detected. Also disclosed are kits for providing compositions so as to practice the disclosed methods.

Abstract: The invention relates to a novel lipid kinase which is part of the PI3 Kinase family. PI3 Kinases catalyze the addition of phosphate to inositol generating inositol mono, di and triphosphate. Inositol phosphates have been implicated in regulating intracellular signaling cascades resulting in alternations in gene expression which, amongst other effects, can result in cytoskeletal remodeling and modulation of cellular motility. More particularly the invention relates to a novel human PI3 Kinase, p110? which interacts with p85, has a broad phosphinositide specificity and is sensitive to the same kinase inhibitors as P13 Kinase p110A. However, in contrast to previously identified PI3 Kinases which show a ubiquitous pattern of expression, p110? is selectively expressed in leucocytes. Importantly, p110? shows enhanced expression in most melanomas tested and therefore may play a crucial role in regulating the metastatic property exhibited by melanomas.

Abstract: The invention relates to a novel lipid kinase which is part of the PI3 Kinase family. PI3 Kinases catalyze the addition of phosphate to inositol generating inositol mono, di and triphosphate. Inositol phosphates have been implicated in regulating intracellular signaling cascades resulting in alternations in gene expression which, amongst other effects, can result in cytoskeletal remodeling and modulation of cellular motility. More particularly the invention relates to a novel human PI3 Kinase, p110? which interacts with p85, has a broad phosphinositide specificity and is sensitive to the same kinase inhibitors as PI3 Kinase p110A. However, in contrast to previously identified PI3 Kinases which show a ubiquitous pattern of expression, p110? is selectively expressed in leucocytes. Importantly, p110? shows enhanced expression in most melanomas tested and therefore may play a crucial role in regulating the metastatic property exhibited by melanomas.

Abstract: The invention relates to a novel lipid kinase which is part of the PI3 Kinase family. PI3 Kinases catalyze the addition of phosphate to inositol generating inositol mono, di and triphosphate. Inositol phosphates have been implicated in regulating intracellular signaling cascades resulting in alternations in gene expression which, amongst other effects, can result in cytoskeletal remodeling and modulation of cellular motility. More particularly the invention relates to a novel human PI3 Kinase, p110&Dgr; which interacts with p85, has a broad phosphinositide specificity and is sensitive to the same kinase inhibitors as PI3 Kinase p110A. However, in contrast to previously identified PI3 Kinases which show a ubiquitous pattern of expression, p110&Dgr; is selectively expressed in leucocytes. Importantly, p110&Dgr; shows enhanced expression in most melanomas tested and therefore may play a crucial role in regulating the metastatic property exhibited by melanomas.

Abstract: The invention relates to a novel lipid kinase which is part of the PI3 Kinase family. PI3 Kinases catalyze the addition of phosphate to inositol generating inositol mono, di and triphosphate. Inositol phosphates have been implicated in regulating intracellular signalling cascades resulting in alternations in gene expression which, amongst other effects, can result in cytoskeletal remodelling and modulation of cellular motility. More particularly the invention relates to a novel human PI3 Kinase, p110&dgr; which interacts with p85, has a broad phosphoinositide specificity and is sensitive to the same kinase inhibitors as PI3 Kinase p110&agr;. However in contrast to previously identified PI3 Kinases which show a ubiquitous pattern of expression, p110&dgr; is selectively expressed in leucocytes. Importantly, p110&dgr; shows enchanced expression in most melanomas tested and therefore may play a crucial role in regulating the metastatic property exhibited by melanomas.