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High Throughput Kinomic Profiling of Human Clear Cell Renal Cell Carcinoma Identifies Kinase Activity Dependent Molecular Subtypes.

Anderson JC, Willey CD, Mehta A, Welaya K, Chen D, Duarte CW, Ghatalia P, Arafat W, Madan A, Sudarshan S, Naik G, Grizzle WE, Choueiri TK, Sonpavde G - PLoS ONE (2015)

Bottom Line: Potential driver kinases implicated include PFTAIRE (PFTK1), PKG1, and SRC, which were identified in groups A, B, and C, respectively.Of the 9 patients who had tumor recurrence, only one was found in Group B.Supervised analysis showed decreased kinase activity of CDK1 and RSK1-4 substrates in those which progressed compared to others.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.

ABSTRACT
Despite the widespread use of kinase-targeted agents in clear cell renal cell carcinoma (CC-RCC), comprehensive kinase activity evaluation (kinomic profiling) of these tumors is lacking. Thus, kinomic profiling of CC-RCC may assist in devising a classification system associated with clinical outcomes, and help identify potential therapeutic targets. Fresh frozen CC-RCC tumor lysates from 41 clinically annotated patients who had localized disease at diagnosis were kinomically profiled using the PamStation®12 high-content phospho-peptide substrate microarray system (PamGene International). Twelve of these patients also had matched normal kidneys available that were also profiled. Unsupervised hierarchical clustering and supervised comparisons based on tumor vs. normal kidney and clinical outcome (tumor recurrence) were performed and coupled with advanced network modeling and upstream kinase prediction methods. Unsupervised clustering analysis of localized CC-RCC tumors identified 3 major kinomic groups associated with inflammation (A), translation initiation (B), and immune response and cell adhesions (C) processes. Potential driver kinases implicated include PFTAIRE (PFTK1), PKG1, and SRC, which were identified in groups A, B, and C, respectively. Of the 9 patients who had tumor recurrence, only one was found in Group B. Supervised analysis showed decreased kinase activity of CDK1 and RSK1-4 substrates in those which progressed compared to others. Twelve tumors with matching normal renal tissue implicated increased PIM's and MAPKAPK's in tumors compared to adjacent normal renal tissue. As such, comprehensive kinase profiling of CC-RCC tumors could provide a functional classification strategy for patients with localized disease and identify potential therapeutic targets.

No MeSH data available.


Related in: MedlinePlus

Kinases altered in CC-RCC and relationship to clinical outcome.CC-RCC tumors that had matched normal fresh frozen material available (n = 12) were directly compared and statistically different phosphopeptides (p<0.001) were identified and are shown in (A). These significant peptides were used to query Kinexus Phosphonet as in Fig 3 (and as described in Materials and Methods). Predicted upstream kinases that distinguish CC-RCC from matched normal kidney (indicated as increased or decreased in CC-RCC relative to normal kidney) are shown in (B). GeneGo MetaCore Network Modeling of the proteins that contain the significantly altered phosphopeptides (Listed as Uniprot ID’s in A) is shown in (C). Red circles indicate increased phosphorylation of the peptide while blue circles indicate decreased substrate phosphorylation. A supervised analysis of the CC-RCC tumors was performed to determine kinomic differences between patients who remained locally controlled after a minimum follow up of 18 months (NonProg) and those who progressed (Prog). Peptides significantly altered between these groups (D) were used to query Kinexus Phosphonet as above and are shown in (E) which were decreased. GeneGo MetaCore Network Modeling of the proteins containing the significantly altered phosphopeptides is shown in (F) where blue circles indicate decreased phosphorylation of the peptide.
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pone.0139267.g004: Kinases altered in CC-RCC and relationship to clinical outcome.CC-RCC tumors that had matched normal fresh frozen material available (n = 12) were directly compared and statistically different phosphopeptides (p<0.001) were identified and are shown in (A). These significant peptides were used to query Kinexus Phosphonet as in Fig 3 (and as described in Materials and Methods). Predicted upstream kinases that distinguish CC-RCC from matched normal kidney (indicated as increased or decreased in CC-RCC relative to normal kidney) are shown in (B). GeneGo MetaCore Network Modeling of the proteins that contain the significantly altered phosphopeptides (Listed as Uniprot ID’s in A) is shown in (C). Red circles indicate increased phosphorylation of the peptide while blue circles indicate decreased substrate phosphorylation. A supervised analysis of the CC-RCC tumors was performed to determine kinomic differences between patients who remained locally controlled after a minimum follow up of 18 months (NonProg) and those who progressed (Prog). Peptides significantly altered between these groups (D) were used to query Kinexus Phosphonet as above and are shown in (E) which were decreased. GeneGo MetaCore Network Modeling of the proteins containing the significantly altered phosphopeptides is shown in (F) where blue circles indicate decreased phosphorylation of the peptide.

Mentions: The kinomic profiles of the 12 normal kidney tissues were directly compared with the matching CC-RCC and significantly altered phosphopeptides (p<0.001) were identified and are displayed in Fig 4A. It should be noted that the phosphopeptide changes were dominated by serine/threonine substrates (STK PamChip). Using a less stringent significance cutoff (p<0.01), 5 tyrosine substrate phosphopeptides (PTK PamChip) were significantly altered in the CC-RCC versus normal kidney (S3 Fig). Top potentially altered kinases included increased PIM’s and MAPKAPK’s and decreased JNK2 and CDK1 in tumors compared to adjacent normal renal tissue (Fig 4B). In addition, we performed network mapping of the altered phosphopeptide substrates which is shown in Fig 4C. Of note, DNA damage repair and cell cycle regulatory components were identified in this mapping.


High Throughput Kinomic Profiling of Human Clear Cell Renal Cell Carcinoma Identifies Kinase Activity Dependent Molecular Subtypes.

Anderson JC, Willey CD, Mehta A, Welaya K, Chen D, Duarte CW, Ghatalia P, Arafat W, Madan A, Sudarshan S, Naik G, Grizzle WE, Choueiri TK, Sonpavde G - PLoS ONE (2015)

Kinases altered in CC-RCC and relationship to clinical outcome.CC-RCC tumors that had matched normal fresh frozen material available (n = 12) were directly compared and statistically different phosphopeptides (p<0.001) were identified and are shown in (A). These significant peptides were used to query Kinexus Phosphonet as in Fig 3 (and as described in Materials and Methods). Predicted upstream kinases that distinguish CC-RCC from matched normal kidney (indicated as increased or decreased in CC-RCC relative to normal kidney) are shown in (B). GeneGo MetaCore Network Modeling of the proteins that contain the significantly altered phosphopeptides (Listed as Uniprot ID’s in A) is shown in (C). Red circles indicate increased phosphorylation of the peptide while blue circles indicate decreased substrate phosphorylation. A supervised analysis of the CC-RCC tumors was performed to determine kinomic differences between patients who remained locally controlled after a minimum follow up of 18 months (NonProg) and those who progressed (Prog). Peptides significantly altered between these groups (D) were used to query Kinexus Phosphonet as above and are shown in (E) which were decreased. GeneGo MetaCore Network Modeling of the proteins containing the significantly altered phosphopeptides is shown in (F) where blue circles indicate decreased phosphorylation of the peptide.
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Related In: Results  -  Collection

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pone.0139267.g004: Kinases altered in CC-RCC and relationship to clinical outcome.CC-RCC tumors that had matched normal fresh frozen material available (n = 12) were directly compared and statistically different phosphopeptides (p<0.001) were identified and are shown in (A). These significant peptides were used to query Kinexus Phosphonet as in Fig 3 (and as described in Materials and Methods). Predicted upstream kinases that distinguish CC-RCC from matched normal kidney (indicated as increased or decreased in CC-RCC relative to normal kidney) are shown in (B). GeneGo MetaCore Network Modeling of the proteins that contain the significantly altered phosphopeptides (Listed as Uniprot ID’s in A) is shown in (C). Red circles indicate increased phosphorylation of the peptide while blue circles indicate decreased substrate phosphorylation. A supervised analysis of the CC-RCC tumors was performed to determine kinomic differences between patients who remained locally controlled after a minimum follow up of 18 months (NonProg) and those who progressed (Prog). Peptides significantly altered between these groups (D) were used to query Kinexus Phosphonet as above and are shown in (E) which were decreased. GeneGo MetaCore Network Modeling of the proteins containing the significantly altered phosphopeptides is shown in (F) where blue circles indicate decreased phosphorylation of the peptide.
Mentions: The kinomic profiles of the 12 normal kidney tissues were directly compared with the matching CC-RCC and significantly altered phosphopeptides (p<0.001) were identified and are displayed in Fig 4A. It should be noted that the phosphopeptide changes were dominated by serine/threonine substrates (STK PamChip). Using a less stringent significance cutoff (p<0.01), 5 tyrosine substrate phosphopeptides (PTK PamChip) were significantly altered in the CC-RCC versus normal kidney (S3 Fig). Top potentially altered kinases included increased PIM’s and MAPKAPK’s and decreased JNK2 and CDK1 in tumors compared to adjacent normal renal tissue (Fig 4B). In addition, we performed network mapping of the altered phosphopeptide substrates which is shown in Fig 4C. Of note, DNA damage repair and cell cycle regulatory components were identified in this mapping.

Bottom Line: Potential driver kinases implicated include PFTAIRE (PFTK1), PKG1, and SRC, which were identified in groups A, B, and C, respectively.Of the 9 patients who had tumor recurrence, only one was found in Group B.Supervised analysis showed decreased kinase activity of CDK1 and RSK1-4 substrates in those which progressed compared to others.

View Article: PubMed Central - PubMed

Affiliation: Department of Radiation Oncology, University of Alabama at Birmingham, Birmingham, Alabama, United States of America.

ABSTRACT
Despite the widespread use of kinase-targeted agents in clear cell renal cell carcinoma (CC-RCC), comprehensive kinase activity evaluation (kinomic profiling) of these tumors is lacking. Thus, kinomic profiling of CC-RCC may assist in devising a classification system associated with clinical outcomes, and help identify potential therapeutic targets. Fresh frozen CC-RCC tumor lysates from 41 clinically annotated patients who had localized disease at diagnosis were kinomically profiled using the PamStation®12 high-content phospho-peptide substrate microarray system (PamGene International). Twelve of these patients also had matched normal kidneys available that were also profiled. Unsupervised hierarchical clustering and supervised comparisons based on tumor vs. normal kidney and clinical outcome (tumor recurrence) were performed and coupled with advanced network modeling and upstream kinase prediction methods. Unsupervised clustering analysis of localized CC-RCC tumors identified 3 major kinomic groups associated with inflammation (A), translation initiation (B), and immune response and cell adhesions (C) processes. Potential driver kinases implicated include PFTAIRE (PFTK1), PKG1, and SRC, which were identified in groups A, B, and C, respectively. Of the 9 patients who had tumor recurrence, only one was found in Group B. Supervised analysis showed decreased kinase activity of CDK1 and RSK1-4 substrates in those which progressed compared to others. Twelve tumors with matching normal renal tissue implicated increased PIM's and MAPKAPK's in tumors compared to adjacent normal renal tissue. As such, comprehensive kinase profiling of CC-RCC tumors could provide a functional classification strategy for patients with localized disease and identify potential therapeutic targets.

No MeSH data available.


Related in: MedlinePlus