Biology subjects

Neel, B. G.

Publications and source records attributed to Neel, B. G..

6 recordsLinked to original sources

bioRxiv2018

SHP2 Inhibition Abrogates MEK inhibitor Resistance in Multiple Cancer Models

Adaptive resistance to MEK inhibitors (MEK-Is) typically occurs via induction of genes for different receptor tyrosine kinases (RTKs) and/or their ligands, even in tumors of the same histotype, making combination strategies challenging. SHP2 (PTPN11) is required for RAS/ERK pathway activation by most RTKs, and might provide a common resistance node. We found that combining the SHP2 inhibitor SHP099 with a MEK-I inhibits proliferation of multiple cancer cells in vitro. PTPN11 knockdown/MEK-I had similar effects, while expressing SHP099-binding mutants conferred resistance, demonstrating that SHP099 was on-target. This combination was efficacious in xenograft and/or genetically engineered models of KRAS-mutant pancreas cancer and ovarian cancer and in wild-type RAS-expressing triple negative breast cancer. Biochemical studies show that SHP099 impedes SOS/RAS/MEK/ERK1/2 reactivation in response to MEK-Is and blocks ERK1/2-dependent transcriptional programs. SHP099 alone also inhibited RAS activation in some, but not all, KRAS-mutant lines. Hence, SHP099/MEK-I combinations could have therapeutic utility in multiple malignancies.\n\nSIGNIFICANCEMEK inhibitors have shown limited efficacy as single agents because of the rapid development of adaptive resistance. We find that combining SHP2 and MEK inhibition abrogates adaptive resistance in multiple cancer models, expressing mutant and wild-type KRAS.

cancer biology↗

bioRxiv2018

Pathological Oxidation of PTPN12 Underlies ABL1 Phosphorylation in HLRCC

Hereditary Leiomyomatosis and Renal Cell Cancer (HLRCC) is an inherited cancer syndrome associated with a highly aggressive form of type 2 papillary renal cell carcinoma (PRCC). Germ line inactivating alterations in Fumarate Hydratase (FH) cause HLRCC, and result in elevated levels of reactive oxygen species (ROS). Recent work indicates that FH -/-PRCC cells have increased ABL1 activation, which promotes tumor growth, but how ABL1 is activated remained unclear. Oxidation can regulate protein-tyrosine phosphatase (PTP) catalytic activity; conceivably, ROS-catalyzed inactivation of an ABL-directed PTP might account for ABL1 activation in this malignancy. Previously, our group developed \"q-oxPTPome,\" a method that can globally monitor the oxidation of classical PTPs. We have now refined the q-oxPTPome approach, increasing its sensitivity by >10X. Applying q-oxPTPome to FH-deficient cell models shows that multiple PTPs are either highly oxidized (including PTPN12) or overexpressed. In general, highly oxidized PTPs were those that have relatively high sensitivity to exogenous H2O2. Most PTP oxidation in FH-deficient cells is reversible, although nearly 40% of PTPN13 is oxidized irreversibly to the sulfonic acid state. Using \"substrate-trapping mutants\", we mapped PTPs to their putative substrates, and found that only PTPN12 could target ABL1. Furthermore, knockdown experiments identify PTPN12 as the major ABL1 phosphatase in HLRCC. Overall, our results show that ROS-induced PTPN12 oxidation accounts for ABL1 phosphorylation in HLRCC-associated PRCC, reveal a novel mechanism for inactivating a tumor suppressor gene product, and establish a direct link between pathological PTP oxidation and neoplastic disease.

cancer biology↗

bioRxiv2018

Off-target inhibition by active site-targeting SHP2 inhibitors

Due to the involvement of SHP2 (SH2 domain-containing protein tyrosine phosphatase) in human disease, including Noonan syndrome and cancer, several inhibitors targeting SHP2 have been developed. Here, we report that the commonly used SHP2 inhibitor NSC-78788 does not exhibit robust inhibitory effects on growth factor-dependent MAPK (mitogen-activated protein kinase) pathway activation, and that the recently developed active site-targeting SHP2 inhibitors IIB-08, 11a-1, and GS-493 show off-target effects on ligand-evoked activation/trans-phosphorylation of the PDGFR{beta} (platelet-derived growth factor receptor {beta}). GS-493 also inhibits purified human PDGFR{beta} and SRC in vitro, whereas PDGFR{beta} inhibition by IIB-08 and 11a-1 occurs only in the cellular context. Our results argue for extreme caution in inferring specific functions for SHP2 based on studies using these inhibitors.

cell biology↗

bioRxiv2017

SHP2 Is Required for BCR-ABL1-Induced Hematologic Neoplasms

BCR-ABL1-targeting tyrosine kinase inhibitors (TKIs) have revolutionized treatment of Philadelphia chromosome-positive (Ph+) hematologic neoplasms. Nevertheless, acquired TKI resistance remains a major problem in chronic myeloid leukemia (CML), and TKIs are less effective against Ph+ B-cell acute lymphoblastic leukemia (B-ALL). GAB2, a scaffolding adaptor that binds and activates SHP2, is essential for leukemogenesis by BCR-ABL1, and a GAB2 mutant lacking SHP2 binding cannot mediate leukemogenesis. Using a genetic loss-of-function approach and bone marrow transplantation (BMT) models for CML and BCR-ABL1+ B-ALL, we show that SHP2 is required for BCR-ABL1-evoked myeloid and lymphoid neoplasia. Ptpn11 deletion impairs initiation and maintenance of CML-like myeloproliferative neoplasm, and compromises induction of BCR-ABL1+ B-ALL. SHP2, and specifically, its SH2 domains, PTP activity and C-terminal tyrosines, is essential for BCR-ABL1+, but not WT, pre-B cell proliferation. The MEK/ERK pathway is regulated by SHP2 in WT and BCR-ABL1+ pre-B cells, but is only required for the proliferation of BCR-ABL1+ cells. SHP2 is required for SRC family kinase (SFK) activation only in BCR-ABL1+ pre-B cells. RNAseq reveals distinct SHP2-dependent transcriptional programs in BCR-ABL1+ and WT pre-B cells. Our results suggest that SHP2, via SFKs and ERK, represses MXD3/4 to facilitate a MYC-dependent proliferation program in BCR-ABL1-transformed pre-B cells.

cancer biology↗

bioRxiv2017

Assay to visualize specific protein oxidation reveals spatio-temporal regulation of SHP2

Reactive oxygen species (ROS) are produced transiently in response to cell stimuli, and function as second messengers that oxidize target proteins. Protein-tyrosine phosphatases (PTPs) are important ROS targets, whose oxidation results in rapid, reversible, catalytic inactivation. Despite increasing evidence for the importance of PTP oxidation in signal transduction, the cell biological details of ROS-catalyzed PTP inactivation have remained largely unclear, due to our inability to visualize PTP oxidation in cells. By combining proximity ligation assay (PLA) with chemical labeling of cysteine residues in the sulfenic acid state, we visualize oxidized Src homology 2 domain-containing protein-tyrosine phosphatase 2 (SHP2). We find that platelet-derived growth factor (PDGF) evokes transient oxidation on or close to RAB5+/EEA1-endosomes. SHP2 oxidation requires NADPH oxidases (NOXs), and oxidized SHP2 co-localizes with PDGF receptor and NOX1/4. Our data demonstrate spatially and temporally limited protein oxidation within cells, and suggest that PDGF-dependent \"redoxosomes,\" contribute to proper signal transduction.

cell biology↗

bioRxiv2017

Cellular Interplay and Cytokine Hierarchy Cause Pathological Cardiac Hypertrophy in RAF1-Mutant Noonan Syndrome

Noonan syndrome (NS) is caused by mutations in RAS/ERK pathway genes, and is characterized by craniofacial, growth, cognitive and cardiac defects. NS patients with kinase-activating RAF1 alleles typically develop pathological left ventricular hypertrophy (LVH), which is reproduced in Raf1L613V/+ knock-in mice. Here, using inducible Raf1L613V expression, we show that LVH results from the interplay of cardiac cell types. Cardiomyocyte Raf1L613V enhances Ca2+ sensitivity and cardiac contractility without causing hypertrophy. Raf1L613V expression in cardiomyocytes or activated fibroblasts exacerbates pressure overload-evoked fibrosis. Endothelial/endocardial (EC) Raf1L613V causes cardiac hypertrophy without affecting contractility. Co-culture and neutralizing antibody experiments reveal a cytokine (TNF/IL6) hierarchy in Raf1L613V-expressing ECs that drives cardiomyocyte hypertrophy in vitro. Furthermore, post-natal TNF inhibition normalizes the increased wall thickness and cardiomyocyte hypertrophy in vivo. We conclude that NS cardiomyopathy involves cardiomyocytes, ECs, and fibroblasts, TNF/IL6 signaling components represent potential therapeutic targets, and abnormal EC signaling might contribute to other forms of LVH.

cell biology↗