A robust optimal control framework for controlling aberrant RTK signaling pathways in esophageal cancer.

This study presents a new framework for obtaining personalized optimal treatment strategies targeting aberrant signaling pathways in esophageal cancer, such as the epidermal growth factor (EGF) and vascular endothelial growth factor (VEGF) signaling pathways. A new pharmacokinetic model is developed taking into account specific heterogeneities of these signaling mechanisms. The optimal therapies are designed to be obtained using a three step process. First, a finite-dimensional constrained optimization problem is solved to obtain the parameters of the pharmacokinetic model, using discrete patient data measurements. Next, a sensitivity analysis is carried out to determine which of the parameters are sensitive to the evolution of the variants of EGF receptors and VEGF receptors. Finally, a second optimal control problem is solved based on the sensitivity analysis results, using a modified pharmacokinetic model that incorporates two representative drugs Trastuzumab and Bevacizumab, targeting EGF and VEGF, respectively. Numerical results with the combination of the two drugs demonstrate the efficiency of the proposed framework.

Muscarinic signaling regulates voltage-gated potassium channel KCNQ2 phosphorylation in the nucleus accumbens via protein kinase C for aversive learning.

The nucleus accumbens (NAc) plays critical roles in emotional behaviors, including aversive learning. Aversive stimuli such as an electric foot shock increase acetylcholine (ACh) in the NAc, and muscarinic signaling appears to increase neuronal excitability and aversive learning. Muscarinic signaling inhibits the voltage-dependent potassium KCNQ current which regulates neuronal excitability, but the regulatory mechanism has not been fully elucidated. Phosphorylation of KCNQ2 at threonine 217 (T217) and its inhibitory effect on channel activity were predicted. However, whether and how muscarinic signaling phosphorylates KCNQ2 in vivo remains unclear. Here, we found that PKC directly phosphorylated KCNQ2 at T217 in vitro. Carbachol and a muscarinic M1 receptor (M1R) agonist facilitated KCNQ2 phosphorylation at T217 in NAc/striatum slices in a PKC-dependent manner. Systemic administration of the cholinesterase inhibitor donepezil, which is commonly used to treat dementia, and electric foot shock to mice induced the phosphorylation of KCNQ2 at T217 in the NAc, whereas phosphorylation was suppressed by an M1R antagonist. Conditional deletion of Kcnq2 in the NAc enhanced electric foot shock induced aversive learning. Our findings indicate that muscarinic signaling induces the phosphorylation of KCNQ2 at T217 via PKC activation for aversive learning.

Rho-Rho-Kinase Regulates Ras-ERK Signaling Through SynGAP1 for Dendritic Spine Morphology.

The structural plasticity of dendritic spines plays a critical role in NMDA-induced long-term potentiation (LTP) in the brain. The small GTPases RhoA and Ras are considered key regulators of spine morphology and enlargement. However, the regulatory interaction between RhoA and Ras underlying NMDA-induced spine enlargement is largely unknown. In this study, we found that Rho-kinase/ROCK, an effector of RhoA, phosphorylated SynGAP1 (a synaptic Ras-GTPase activating protein) at Ser842 and increased its interaction with 14-3-3ζ, thereby activating Ras-ERK signaling in a reconstitution system in HeLa cells. We also found that the stimulation of NMDA receptor by glycine treatment for LTP induction stimulated SynGAP1 phosphorylation, Ras-ERK activation, spine enlargement and SynGAP1 delocalization from the spines in striatal neurons, and these effects were prevented by Rho-kinase inhibition. Rho-kinase-mediated phosphorylation of SynGAP1 appeared to increase its dissociation from PSD95, a postsynaptic scaffolding protein located at postsynaptic density, by forming a complex with 14-3-3ζ. These results suggest that Rho-kinase phosphorylates SynGAP1 at Ser842, thereby activating the Ras-ERK pathway for NMDA-induced morphological changes in dendritic spines.

Rho-Kinase/ROCK Phosphorylates PSD-93 Downstream of NMDARs to Orchestrate Synaptic Plasticity.

The N-methyl-D-aspartate receptor (NMDAR)-mediated structural plasticity of dendritic spines plays an important role in synaptic transmission in the brain during learning and memory formation. The Rho family of small GTPase RhoA and its downstream effector Rho-kinase/ROCK are considered as one of the major regulators of synaptic plasticity and dendritic spine formation, including long-term potentiation (LTP). However, the mechanism by which Rho-kinase regulates synaptic plasticity is not yet fully understood. Here, we found that Rho-kinase directly phosphorylated discs large MAGUK scaffold protein 2 (DLG2/PSD-93), a major postsynaptic scaffold protein that connects postsynaptic proteins with NMDARs; an ionotropic glutamate receptor, which plays a critical role in synaptic plasticity. Stimulation of striatal slices with an NMDAR agonist induced Rho-kinase-mediated phosphorylation of PSD-93 at Thr612. We also identified PSD-93-interacting proteins, including DLG4 (PSD-95), NMDARs, synaptic Ras GTPase-activating protein 1 (SynGAP1), ADAM metallopeptidase domain 22 (ADAM22), and leucine-rich glioma-inactivated 1 (LGI1), by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Among them, Rho-kinase increased the binding of PSD-93 to PSD-95 and NMDARs. Furthermore, we found that chemical-LTP induced by glycine, which activates NMDARs, increased PSD-93 phosphorylation at Thr612, spine size, and PSD-93 colocalization with PSD-95, while these events were blocked by pretreatment with a Rho-kinase inhibitor. These results indicate that Rho-kinase phosphorylates PSD-93 downstream of NMDARs, and suggest that Rho-kinase mediated phosphorylation of PSD-93 increases the association with PSD-95 and NMDARs to regulate structural synaptic plasticity.

Signal flow in the NMDA receptor-dependent phosphoproteome regulates postsynaptic plasticity for aversive learning.

Structural plasticity of dendritic spines in the nucleus accumbens (NAc) is crucial for learning from aversive experiences. Activation of NMDA receptors (NMDARs) stimulates Ca2+-dependent signaling that leads to changes in the actin cytoskeleton, mediated by the Rho family of GTPases, resulting in postsynaptic remodeling essential for learning. We investigated how phosphorylation events downstream of NMDAR activation drive the changes in synaptic morphology that underlie aversive learning. Large-scale phosphoproteomic analyses of protein kinase targets in mouse striatal/accumbal slices revealed that NMDAR activation resulted in the phosphorylation of 194 proteins, including RhoA regulators such as ARHGEF2 and ARHGAP21. Phosphorylation of ARHGEF2 by the Ca2+-dependent protein kinase CaMKII enhanced its RhoGEF activity, thereby activating RhoA and its downstream effector Rho-associated kinase (ROCK/Rho-kinase). Further phosphoproteomic analysis identified 221 ROCK targets, including the postsynaptic scaffolding protein SHANK3, which is crucial for its interaction with NMDARs and other postsynaptic scaffolding proteins. ROCK-mediated phosphorylation of SHANK3 in the NAc was essential for spine growth and aversive learning. These findings demonstrate that NMDAR activation initiates a phosphorylation cascade crucial for learning and memory.

Comparison of the Performance of the TPTest, Tubex, Typhidot and Widal Immunodiagnostic Assays and Blood Cultures in Detecting Patients with Typhoid Fever in Bangladesh, Including Using a Bayesian Latent Class Modeling Approach.

BACKGROUND: There is an urgent need for an improved diagnostic assay for typhoid fever. In this current study, we compared the recently developed TPTest (Typhoid and Paratyphoid Test) with the Widal test, blood culture, and two commonly used commercially available kits, Tubex and Typhidot. METHODOLOGY: For analysis, we categorized 92 Bangladeshi patients with suspected enteric fever into four groups: S. Typhi bacteremic patients (n = 28); patients with a fourfold change in Widal test from day 0 to convalescent period (n = 7); patients with Widal titer ≥1:320 (n = 13) at either acute or convalescent stage of disease; and patients suspected with enteric fever, but with a negative blood culture and Widal titer (n = 44). We also tested healthy endemic zone controls (n = 20) and Bangladeshi patients with other febrile illnesses (n = 15). Sample size was based on convenience to facilitate preliminary analysis. PRINCIPLE FINDINGS: Of 28 S. Typhi bacteremic patients, 28 (100%), 21 (75%) and 18 (64%) patients were positive by TPTest, Tubex and Typhidot, respectively. In healthy endemic zone controls, the TPTest method was negative in all, whereas Tubex and Typhidot were positive in 3 (15%) and 5 (25%), respectively. We then estimated sensitivity and specificity of all diagnostic tests using Bayesian latent class modeling. The sensitivity of TPTest, Tubex and Typhidot were estimated at 96.0% (95% CI: 87.1%-99.8%), 60.2% (95% CI: 49.3%-71.2%), and 59.6% (95% CI: 50.1%-69.3%), respectively. Specificity was estimated at 96.6% (90.7%-99.2%) for TPTest, 89.9% (79.6%-96.8%) for Tubex, and 80.0% (67.7%-89.7%) for Typhidot. CONCLUSION: These results suggest that the TPTest is highly sensitive and specific in diagnosing individuals with typhoid fever in a typhoid endemic setting, outperforming currently available and commonly used alternatives.