Preclinical characterization of tunlametinib, a novel, potent, and selective MEK inhibitor

Background: Aberrant activation of RAS-RAF-MEK-ERK signaling pathway has been implicated in more than one-third of all malignancies. MEK inhibitors are promising therapeutic approaches to target this signaling pathway. Though four MEK inhibitors have been approved by FDA, these compounds possess either limited efficacy or unfavorable PK profiles with toxicity issues, hindering their broadly application in clinic. Our efforts were focused on the design and development of a novel MEK inhibitor, which subsequently led to the discovery of tunlametinib.Methods: This study verified the superiority of tunlametinib over the current MEK inhibitors in preclinical studies. The protein kinase selectivity activity of tunlametinib was evaluated against 77 kinases. Anti-proliferation activity was analyzed using the 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) or (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS) assay. ERK and phospho-ERK levels were evaluated by Western blot analysis. Flow cytometry analysis was employed to investigate cell cycle and arrest. Cell-derived xenograft (CDX) and Patient-derived xenograft (PDX) models were used to evaluate the tumor growth inhibition. The efficacy of tunlametinib as monotherapy treatment was evaluated in KRAS/BRAF mutant or wild type xenograft model. Furthermore, the combination studies of tunlametinib with BRAF/KRASG12C/SHP2 inhibitors or chemotherapeutic agent were conducted by using the cell proliferation assay in vitro and xenograft models in vivo.Results:In vitro, tunlametinib demonstrated high selectivity with approximately 19-fold greater potency against MEK kinase than MEK162, and nearly 10–100-fold greater potency against RAS/RAF mutant cell lines than AZD6244. In vivo, tunlametinib resulted in dramatic tumor suppression and profound inhibition of ERK phosphorylation in tumor tissue. Mechanistic study revealed that tunlametinib induced cell cycle arrest at G0/G1 phase and apoptosis of cells in a dose-proportional manner. In addition, tunlametinib demonstrated a favorable pharmacokinetic profile with dose-proportionality and good oral bioavailability, with minimal drug exposure accumulation. Furthermore, tunlametinib combined with BRAF/KRASG12C/SHP2 inhibitors or docetaxel showed synergistically enhanced response and marked tumor inhibition.Conclusion: Tunlametinib exhibited a promising approach for treating RAS/RAF mutant cancers alone or as combination therapies, supporting the evaluation in clinical trials. Currently, the first-in-human phase 1 study and pivotal clinical trial of tunlametinib as monotherapy have been completed and pivotal trials as combination therapy are ongoing

MAPK1 promotes the metastasis and invasion of gastric cancer as a bidirectional transcription factor

Background: The Mitogen-activated protein kinase 1 (MAPK1) has both independent functions of phosphorylating histones as a kinase and directly binding the promoter regions of genes to regulate gene expression as a transcription factor. Previous studies have identified elevated expression of MAPK1 in human gastric cancer, which is associated with its role as a kinase, facilitating the migration and invasion of gastric cancer cells. However, how MAPK1 binds to its target genes as a transcription factor and whether it modulates related gene expressions in gastric cancer remains unclear. Results: Here, we integrated biochemical assays (protein interactions and chromatin immunoprecipitation (ChIP)), cellular analysis assays (cell proliferation and migration), RNA sequencing, ChIP sequencing, and clinical analysis to investigate the potential genomic recognition patterns of MAPK1 in a human gastric adenocarcinoma cell-line (AGS) and to uncover its regulatory effect on gastric cancer progression. We confirmed that MAPK1 promotes AGS cells invasion and migration by regulating the target genes in different directions, up-regulating seven target genes (KRT13, KRT6A, KRT81, MYH15, STARD4, SYTL4, and TMEM267) and down-regulating one gene (FGG). Among them, five genes (FGG, MYH15, STARD4, SYTL4, and TMEM267) were first associated with cancer procession, while the other three (KRT81, KRT6A, and KRT13) have previously been confirmed to be related to cancer metastasis and migration. Conclusion: Our data showed that MAPK1 can bind to the promoter regions of these target genes to control their transcription as a bidirectional transcription factor, promoting AGS cell motility and invasion. Our research has expanded the understanding of the regulatory roles of MAPK1, enriched our knowledge of transcription factors, and provided novel candidates for cancer therapeutics

Single cell atlas for 11 non-model mammals, reptiles and birds.

The availability of viral entry factors is a prerequisite for the cross-species transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Large-scale single-cell screening of animal cells could reveal the expression patterns of viral entry genes in different hosts. However, such exploration for SARS-CoV-2 remains limited. Here, we perform single-nucleus RNA sequencing for 11 non-model species, including pets (cat, dog, hamster, and lizard), livestock (goat and rabbit), poultry (duck and pigeon), and wildlife (pangolin, tiger, and deer), and investigated the co-expression of ACE2 and TMPRSS2. Furthermore, cross-species analysis of the lung cell atlas of the studied mammals, reptiles, and birds reveals core developmental programs, critical connectomes, and conserved regulatory circuits among these evolutionarily distant species. Overall, our work provides a compendium of gene expression profiles for non-model animals, which could be employed to identify potential SARS-CoV-2 target cells and putative zoonotic reservoirs

The effectiveness of blinatumomab in clearing measurable residual disease in pediatric B‐cell acute lymphoblastic leukemia patients detected by next‐generation sequencing

Abstract Background Blinatumomab improved survival outcomes in B‐cell acute lymphoblastic leukemia (B‐ALL) patients with measurable residual disease (MRD) <10−4. However, data on blinatumomab clearing MRD with high sensitivity of 10−6 remain scarce. This study evaluates the effectiveness of blinatumomab in eradicating extremely low level (up to <10−6) of MRD, as detected by next‐generation sequencing (NGS), in children with B‐ALL. Methods Patients (n = 19) whose MRD was undetectable by multiparameter flow cytometry (MFC) (sensitivity of 10−4) but detectable by NGS after chemotherapy and followed by blinatumomab consolidation were included retrospectively. Results After one course of blinatumomab, 13/19 patients (68%) successfully achieved NGS‐MRD clearance (undetectable). With a median follow‐up of 13.3 months, three of patients who were NGS‐MRD positive relapsed within 1.8 months, while another three remained complete remission. Conclusions Our study was the first to demonstrate that blinatumomab could further eradicate MRD after patients achieve MFC‐MRD undetectable in B‐ALL patients

Distributed Variable Droop Curve Control Strategies in Smart Microgrid

In micro grid (MG), active/reactive power sharing for all dis-patchable units is an important issue. To meet fluctuating loads’ active and reactive power demands, the units generally adopt primary P-f and Q-U droop control methods. However, at different state of charge (SOC) values, the capability of Lead Acid Battery Bank (LABB) based units to take loads varies in a large range; active power should not be shared according to the units P capacities in a constant ratio. Besides, influenced by the output and line impedance between units, reactive power is not able to be shared in proportion to the units Q capacities. Another problem, after MG power balance requirement is satisfied, frequency and voltage are deviating from their rated values thus power quality is reduced. This paper presents a new smart MG which is based on the multi agent system. To solve the problems mentioned above, P-f and Q-U droop curves are adjusted dynamically and autonomously in local agents. To improve the power quality, secondary restoration function is realized in a decentralized way, the computation tasks are assigned to local, the computation capability and communication reliability requirements for central PC are low, and operation reliability is high. Simulation results back the proposed methods

Chemical Characteristics and Controlling Factorsof Groundwater in Chahannur Basin

This paper studies the spatial distribution characteristics and controlling factors of groundwater chemistry in the Chahannur Basin. One hundred and seventy shallow groundwater samples (50 m shallow) are collected, and seven ions, pH, TDS, TH, iron, manganese, COD, barium and other indicators, are detected. Piper triplex graph, Gibbs model, ion ratio, analysis of variance and Kriging interpolation are used to carry out the research. The results show that bicarbonate water is the main water chemical type in the Chahannur Basin, in which bicarbonate water accounts for 65.23%, chloride water accounts for 15.15% and sulfate water accounts for 19.62%. Bicarbonate water is mainly distributed in the mountainous areas in the north and south of the basin, and the main controlling factor is rock weathering. Sulfate-type water is mainly distributed in the lower reaches of the northern mountains of the basin, and the main controlling factors are rock weathering and evaporation concentration. The chloride water is mainly distributed in the Chahannur Lake area and the shallow groundwater buried area in the central region of the basin. The main controlling factors are evaporation concentration and human influence. Na+ is mainly derived from atmospheric precipitation and rock salt leaching, Ca2+ and Mg2+ are mainly derived from carbonate minerals leaching, and silicate minerals leaching is less. The pH of groundwater in the basin ranges from 6.3 to 9.18, with an average value of 7.50. The TDS in the basin ranges from 227 to 22,700 mg/L, with an average of 1661 mg/L. Iron in the catchment ranges from 0.01 to 15.343 mg/L, with a mean of 0.837 mg/L. The manganese content in the basin ranges from 0.005 to 3.802 mg/L, with an average value of 0.254 mg/L. COD in the basin ranges from 0.71–32.72 mg/L, with an average value of 3.49 mg/L. Barium in the basin ranges from 0.005 to 0.312 mg/L, with an average of 0.075 mg/L. The research results provide basic scientific data support for groundwater hydrochemistry research in the Chahannur Basin and show that the types of water chemistry in the study area are complex and diverse, and although the distribution is still controlled by terrain and geological conditions, the area affected by human activities accounts for a high proportion, so attention should be paid to the impact of human activities on groundwater in this area

Comparative study of biomarkers for the early identification of Epstein–Barr virus-associated hemophagocytic lymphohistiocytosis in infectious mononucleosis

Abstract Background and aim Epstein-Barr virus-associated hemophagocytic lymphohistiocytosis (EBV-HLH) and infectious mononucleosis (EBV-IM) share mimic symptoms in the early stages of childhood development. We aimed to examine the clinical features and laboratory indices of these two diseases in children and uncover unique indicators to assist pediatricians in identifying these diseases early. Methods We collected clinical data from 791 pediatric patients diagnosed with EBV-IM or EBV-HLH, compared the clinical traits and laboratory biomarkers presented in the two groups, and constructed predictive models based on them. Results Patients with EBV-IM had greater ratios of cervical lymphadenopathy, eyelid edema, and tonsillitis, whereas individuals with EBV-HLH were more likely to have hepatomegaly and splenomegaly. When using the criteria of interleukin (IL)-10 > 89.6 pg/mL, interferon (IFN)-γ > 45.6 pg/mL, ferritin > 429 μg/L, D-dimer > 3.15 mg/L and triglycerides > 2.1 mmol/L, the sensitivity was 87.9%, 90.7%, 98.1%, 91.1% and 81.5% to predict EBV-HLH, while the specificity was 98.4%, 96.3%, 96.5%, 94.1% and 80.6%, respectively. A logistic regression model based on four parameters (IL-10, ferritin, D-dimer, and triglycerides) was established to distinguish EBV-HLH patients from EBV-IM patients, with a sensitivity of 98.0% and a specificity of 98.2%. Conclusions IL-10, IFN-γ, ferritin and D-dimer levels are significantly different between EBV-HLH and EBV-IM. Predictive models based on clinical signs and laboratory findings provide simple tools to distinguish the two situations

DataSheet1_Preclinical characterization of tunlametinib, a novel, potent, and selective MEK inhibitor.docx

Background: Aberrant activation of RAS-RAF-MEK-ERK signaling pathway has been implicated in more than one-third of all malignancies. MEK inhibitors are promising therapeutic approaches to target this signaling pathway. Though four MEK inhibitors have been approved by FDA, these compounds possess either limited efficacy or unfavorable PK profiles with toxicity issues, hindering their broadly application in clinic. Our efforts were focused on the design and development of a novel MEK inhibitor, which subsequently led to the discovery of tunlametinib.Methods: This study verified the superiority of tunlametinib over the current MEK inhibitors in preclinical studies. The protein kinase selectivity activity of tunlametinib was evaluated against 77 kinases. Anti-proliferation activity was analyzed using the 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) or (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium) (MTS) assay. ERK and phospho-ERK levels were evaluated by Western blot analysis. Flow cytometry analysis was employed to investigate cell cycle and arrest. Cell-derived xenograft (CDX) and Patient-derived xenograft (PDX) models were used to evaluate the tumor growth inhibition. The efficacy of tunlametinib as monotherapy treatment was evaluated in KRAS/BRAF mutant or wild type xenograft model. Furthermore, the combination studies of tunlametinib with BRAF/KRASG12C/SHP2 inhibitors or chemotherapeutic agent were conducted by using the cell proliferation assay in vitro and xenograft models in vivo.Results:In vitro, tunlametinib demonstrated high selectivity with approximately 19-fold greater potency against MEK kinase than MEK162, and nearly 10–100-fold greater potency against RAS/RAF mutant cell lines than AZD6244. In vivo, tunlametinib resulted in dramatic tumor suppression and profound inhibition of ERK phosphorylation in tumor tissue. Mechanistic study revealed that tunlametinib induced cell cycle arrest at G0/G1 phase and apoptosis of cells in a dose-proportional manner. In addition, tunlametinib demonstrated a favorable pharmacokinetic profile with dose-proportionality and good oral bioavailability, with minimal drug exposure accumulation. Furthermore, tunlametinib combined with BRAF/KRASG12C/SHP2 inhibitors or docetaxel showed synergistically enhanced response and marked tumor inhibition.Conclusion: Tunlametinib exhibited a promising approach for treating RAS/RAF mutant cancers alone or as combination therapies, supporting the evaluation in clinical trials. Currently, the first-in-human phase 1 study and pivotal clinical trial of tunlametinib as monotherapy have been completed and pivotal trials as combination therapy are ongoing.</p