1.Quantitative profiling of protein tyrosine kinases in human cancer cell lines by multiplexed parallel reaction monitoring assays.
Kim HJ1, Lin D1, Li M1, Liebler DC2. Mol Cell Proteomics. 2015 Sep 25. pii: mcp.O115.051813. [Epub ahead of print]
Protein tyrosine kinases (PTKs) play key roles in cellular signal transduction, cell cycle regulation, cell division, and cell differentiation. Dysregulation of PTK-activated pathways, often by receptor overexpression, gene amplification, or genetic mutation, is a causal factor underlying numerous cancers. In this study, we have developed a parallel reaction monitoring (PRM)-based assay for quantitative profiling of 83 PTKs. The assay detects 308 proteotypic peptides from 54 receptor tyrosine kinases and 29 nonreceptor tyrosine kinases in a single run. Quantitative comparisons were based on the labeled reference peptide method. We implemented the assay in four cell models: 1) a comparison of proliferating versus epidermal growth factor (EGF)-stimulated A431 cells, 2) a comparison of SW480Null (mutant APC) and SW480APC (APC restored) colon tumor cell lines, and 3) a comparison of 10 colorectal cancer cell lines with different genomic abnormalities, and 4) lung cancer cell lines with either susceptibility (11-18) or acquired resistance (11-18R) to the epidermal growth factor receptor tyrosine kinase inhibitor erlotinib.
2.Combined inhibition of AXL, Lyn and p130Cas kinases block migration of triple negative breast cancer cells.
Pénzes K1, Baumann C, Szabadkai I, Orfi L, Kéri G, Ullrich A, Torka R. Cancer Biol Ther. 2014;15(11):1571-82. doi: 10.4161/15384047.2014.956634.
Blocking the migration of metastatic cancer cells is a major goal in the therapy of cancer. The receptor tyrosine kinase AXL is one of the main triggers for cancer cell migration in neoplasia of breast, colon, skin, thyroid and prostate. In our study we analyzed the effect of AXL inhibition on cell motility and viability in triple negative breast cancer cell lines overexpressing AXL. Thereby we reveal that the compound BMS777607, exhibiting the lowest IC50 values for inhibition of AXL kinase activity in the studied cell lines, attenuates cell motility to a lower extent than the kinase inhibitors MPCD84111 and SKI606. By analyzing the target kinases of MPCD84111 and SKI606 with kinase profiling assays we identified Lyn, a Src family kinase, as a target of both compounds. Knockdown of Lyn and the migration-related CRK-associated substrate (p130Cas), had a significant inhibitory effect on cell migration. Taken together, our findings highlight the importance of combinatorial or multikinase inhibition of non-receptor tyrosine kinases and AXL receptor tyrosine kinase in the therapy of triple negative breast cancer.
3.Characterization of Novel Src Family Kinase Inhibitors to Attenuate Microgliosis.
Manocha GD1, Puig KL1, Austin SA2, Seyb K3, Glicksman MA3, Combs CK1. PLoS One. 2015 Jul 10;10(7):e0132604. doi: 10.1371/journal.pone.0132604. eCollection 2015.
Microgliosis is a major hallmark of Alzheimer's disease (AD) brain pathology. Aβ peptide is hypothesized to act as a stimulus for microglia leading to activation of non-receptor tyrosine kinases and subsequent secretion of pro-inflammatory cytokines. Therefore, the signaling pathways mediating microglial activation may be important therapeutic targets of anti-inflammatory therapy for AD. Four novel compounds were chosen after high throughput screening kinase activity assays determined them as potential Lyn kinase inhibitors. Their kinase inhibitory and anti-inflammatory effect on Aβ-stimulated activation was assessed using the murine microglial cell line, BV2. Cells were treated with the compounds to determine effects on active, phosphorylated levels of Src family kinases, Src and Lyn, as well as MAP kinases ERK, JNK and p38. Only one compound, LDDN-0003499, produced a dose dependent decrease in basal levels of active, phosphorylated Src and Lyn in the BV2 cells.
4.GW5074 and PP2 kinase inhibitors implicate nontraditional c-Raf and Lyn function as drivers of retinoic acid-induced maturation.
Jensen HA1, Bunaciu RP2, Varner JD1, Yen A2. Cell Signal. 2015 Aug;27(8):1666-75. doi: 10.1016/j.cellsig.2015.03.014. Epub 2015 Mar 26.
The multivariate nature of cancer necessitates multi-targeted therapy, and kinase inhibitors account for a vast majority of approved cancer therapeutics. While acute promyelocytic leukemia (APL) patients are highly responsive to retinoic acid (RA) therapy, kinase inhibitors have been gaining momentum as co-treatments with RA for non-APL acute myeloid leukemia (AML) differentiation therapies, especially as a means to treat relapsed or refractory AML patients. In this study GW5074 (a c-Raf inhibitor) and PP2 (a Src-family kinase inhibitor) enhanced RA-induced maturation of t(15;17)-negative myeloblastic leukemia cells and rescued response in RA-resistant cells. PD98059 (a MEK inhibitor) and Akti-1/2 (an Akt inhibitor) were less effective, but did tend to promote maturation-uncoupled G1/G0 arrest, while wortmannin (a PI3K inhibitor) did not enhance differentiation surface marker expression or growth arrest. PD98059 and Akti-1/2 did not enhance differentiation markers and have potential, antagonistic off-targets effects on the aryl hydrocarbon receptor (AhR), but neither could the AhR agonist 6-formylindolo(3,2-b)carbazole (FICZ) rescue differentiation events in the RA-resistant cells.
