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UNLABELLED: We sought to evaluate the efficacy of WEE1 inhibitor adavosertib in patients with solid tumor malignancies (cohort A) and clear cell renal cell carcinoma (ccRCC; cohort B). NCT03284385 was a parallel cohort, Simon two-stage, phase II study of adavosertib (300 mg QDAY by mouth on days 1-5 and 8-12 of each 21-day cycle) in patients with solid tumor malignancies harboring a pathogenic SETD2 mutation. The primary endpoint was the objective response rate. Correlative assays evaluated the loss of H3K36me3 by IHC, a downstream consequence of SETD2 loss, in archival tumor tissue. Eighteen patients were enrolled (9/cohort). The median age was 60 years (range 45-74). The median duration of treatment was 1.28 months (range 0-24+). No objective responses were observed in either cohort; accrual was halted following stage 1. Minor tumor regressions were observed in 4/18 (22%) evaluable patients. Stable disease (SD) was the best overall response in 10/18 (56%) patients, including three patients with SD > 4 months. One patient with ccRCC remains on treatment for >24 months. The most common adverse events of any grade were nausea (59%), anemia (41%), diarrhea (41%), and neutropenia (41%). Nine patients (50%) experienced a Grade ≥3 adverse event. Of eight evaluable archival tissue samples, six (75%) had a loss of H3K36me3 by IHC. Adavosertib failed to exhibit objective responses in SETD2-altered ccRCC and other solid tumor malignancies although prolonged SD was observed in a subset of patients. Combination approaches may yield greater depth of tumor response. SIGNIFICANCE: WEE1 inhibition with adavosertib monotherapy demonstrated limited clinical activity in patients with SETD2-altered solid tumors despite compelling preclinical data indicating a synthetic lethal effect, which did not translate into robust tumor regression. Loss of the H3K36me3 trimethylation mark caused by SETD2-deficiency was confirmed in the majority of evaluable tumors. A subset of patients derived clinical benefit as manifested by minor tumor regressions and prolonged SD.

There is an ongoing need for antifungal agents to treat humans. Identification of new antifungal agents can be based on screening compounds using whole cell assays. Screening compounds that target a particular molecule is possible in budding yeast wherein sophisticated strain engineering allows for controlled expression of endogenous or heterologous genes. We have considered the yeast Mps1 protein kinase as a reasonable target for antifungal agents because mutant or druggable forms of the protein, upon inactivation, cause rapid loss of cell viability. Furthermore, extensive analysis of the Mps1 in budding yeast has offered potential tactics for identifying inhibitors of its enzymatic activity. One such tactic is based on the finding that overexpression of Mps1 leads to cell cycle arrest via activation of the spindle assembly checkpoint. We have endeavored to adapt this assay to be based on the overexpression of Mps1 orthologs from pathogenic yeast in hopes of having a whole-cell assay system to test the activity of these orthologs. Mps1 orthologous genes from seven pathogenic yeast or other pathogenic fungal species were isolated and expressed in budding yeast. Two orthologs clearly produced phenotypes similar to those produced by the overexpression of budding yeast Mps1, indicating that this system for heterologous Mps1 expression has potential as a platform for identifying prospective antifungal agents.

Protein kinases are critical components of a myriad biological processes and strongly associated with various diseases. While kinase research has been a point of focus in biomedical research for several decades, a large portion of the kinome is still considered understudied or "dark," because prior research is targeted towards a subset of kinases with well-established roles in cellular processes. We present an empirical and in-silico hybrid workflow to extend the functional knowledge of understudied kinases. Utilizing multiplex peptide activity arrays and robust in-silico analyses, we extended the functional knowledge of five dark tyrosine kinases (AATK, EPHA6, INSRR, LTK, TNK1) and explored their roles in schizophrenia, Alzheimer's dementia (AD), and major depressive disorder (MDD). Using this hybrid approach, we identified 195 novel kinase-substrate interactions with variable degrees of affinity and linked extended functional networks for these kinases to biological processes that are impaired in psychiatric and neurological disorders. Biochemical assays and mass spectrometry were used to confirm a putative substrate of EPHA6, an understudied dark tyrosine kinase. We examined the EPHA6 network and knowledgebase in schizophrenia using reporter peptides identified and validated from the multi-plex array with high affinity for phosphorylation by EPHA6. Identification and confirmation of putative substrates for understudied kinases provides a wealth of actionable information for the development of new drug treatments as well as exploration of the pathophysiology of disease states using signaling network approaches. [ABSTRACT FROM AUTHOR]

Background Hepatitis B virus–related acute-on-chronic liver failure (HBV-ACLF) has a high short-term mortality. This study aimed to determine the diagnostic and prognostic role of MER tyrosine kinase (MERTK) in patients with HBV-ACLF. Methods Transcriptomics analysis evaluated MERTK expression and function during disease progression. The diagnostic and prognostic significance of MERTK for patients with HBV-ACLF were verified by enzyme-linked immunosorbent assay, area under the receiver operating characteristic curve (AUROC) analysis, and immunohistochemistry (IHC) of liver tissues. Results MERTK mRNA was highly expressed in patients with HBV-ACLF compared to those with liver cirrhosis (LC), chronic hepatitis B (CHB), and normal controls (NC). Elevated MERTK mRNA predicted poor prognosis for HBV-ACLF at 28 and 90 days (AUROC = 0.814 and 0.731, respectively). Functional analysis showed MERTK was significantly associated with toll-like receptor and inflammatory signaling and several key biological processes. External validation with 285 plasma subjects confirmed the high diagnostic accuracy of plasma MERTK for HBV-ACLF (AUROC = 0.859) and potential prognostic value for 28- and 90-day mortality rates (AUROC = 0.673 and 0.644, respectively). Risk stratification analysis indicated higher mortality risk for patients with plasma MERTK level above the cutoff value. Moreover, IHC staining showed increasing MERTK expression from NC, CHB, and LC to HBV-ACLF. Conclusions MERTK shows promise as a candidate biomarker for early diagnosis and prognosis of HBV-ACLF. [ABSTRACT FROM AUTHOR]

The serine/threonine/tyrosine kinase 1 (STYK1) is a receptor protein-tyrosine kinase (RPTK)-like molecule that is detected in several human organs. STYK1 plays an important role in promoting tumorigenesis and metastasis in various cancers. By analyzing the expression of RTKs in immune cells in the database of 2013 Immunological Genome Project, we found that STYK1 was principally expressed in NK cells. In order to investigate the function of STYK1, we used CRISPR/Cas9 technology to generate STYK1-deleted mice, we found STYK1 deletion mice have normal number, development, and function of NK cells in spleen and bone marrow in tumor-free resting state. To examine the tumor surveillance of STYK1 in vivo, we utilized a variety of tumor models, including NK cell-specific target cell (ß2M and RMA-S) clearance experiments in vivo, subcutaneous and intravenous injection of B16F10 melanoma model, and the spontaneous breast cancer model MMTV-PyMT. Surprisingly, we discovered that deletion of the oncogenic STYK1 promoted the four-model tumor progression, and we observed a reduction of NK cell accumulation in the tumor tissues of STYK1 deletion mice compared to WT mice. In order to study the mechanism of STYK1 in NK, RNA sequence of STYK1−/− and WT NK have unveiled a disparity in the signaling pathways linked to migration and adhesion in STYK1−/− NK cells. Further analysis of chemokine receptors associated with NK cell migration revealed that STYK1-deficient NK cells exhibited a significant reduction in CCR2 expression. The STYK1 expression was negatively associated with tumor progression in glioma patients. Overall, our study found the expression of STYK1 in NK cell mediates NK cell anti-tumor response through regulating CCR2 and infiltrating into tumor tissue. [ABSTRACT FROM AUTHOR]

Background: The inability of biologics to pass the plasma membrane prevents their development as therapeutics for intracellular targets. To address the lack of methods for cytosolic protein delivery, we used the type III secretion system (T3SS) of Y. enterocolitica, which naturally injects bacterial proteins into eukaryotic host cells, to deliver monobody proteins into cancer cells. Monobodies are small synthetic binding proteins that can inhibit oncogene signaling in cancer cells with high selectivity upon intracellular expression. Here, we engineered monobodies targeting the BCR::ABL1 tyrosine kinase for efficient delivery by the T3SS, quantified cytosolic delivery and target engagement in cancer cells and monitored inhibition of BCR::ABL1 signaling. Methods: In vitro assays were performed to characterize destabilized monobodies (thermal shift assay and isothermal titration calorimetry) and to assess their secretion by the T3SS. Immunoblot assays were used to study the translocation of monobodies into different cell lines and to determine the intracellular concentration after translocation. Split-Nanoluc assays were performed to understand translocation and degradation kinetics and to evaluate target engagement after translocation. Phospho flow cytometry and apoptosis assays were performed to assess the functional effects of monobody translocation into BCR:ABL1-expressing leukemia cells. Results: To enable efficient translocation of the stable monobody proteins by the T3SS, we engineered destabilized mutant monobodies that retained high affinity target binding and were efficiently injected into different cell lines. After injection, the cytosolic monobody concentrations reached mid-micromolar concentrations considerably exceeding their binding affinity. We found that injected monobodies targeting the BCR::ABL1 tyrosine kinase selectively engaged their target in the cytosol. The translocation resulted in inhibition of oncogenic signaling and specifically induced apoptosis in BCR::ABL1-dependent cells, consistent with the phenotype when the same monobody was intracellularly expressed. Conclusion: Hence, we establish the T3SS of Y. enterocolitica as a highly efficient protein translocation method for monobody delivery, enabling the selective targeting of different oncogenic signaling pathways and providing a foundation for future therapeutic application against intracellular targets. [ABSTRACT FROM AUTHOR]

The fidelity of signal transduction requires the binding of regulatory molecules to their cognate targets. However, the crowded cell interior risks off-target interactions between proteins that are functionally unrelated. How such off-target interactions impact fitness is not generally known. Here, we use Saccharomyces cerevisiae to inducibly express tyrosine kinases. Because yeast lacks bona fide tyrosine kinases, the resulting tyrosine phosphorylation is biologically spurious. We engineered 44 yeast strains each expressing a tyrosine kinase, and quantitatively analysed their phosphoproteomes. This analysis resulted in ~30,000 phosphosites mapping to ~3500 proteins. The number of spurious pY sites generated correlates strongly with decreased growth, and we predict over 1000 pY events to be deleterious. However, we also find that many of the spurious pY sites have a negligible effect on fitness, possibly because of their low stoichiometry. This result is consistent with our evolutionary analyses demonstrating a lack of phosphotyrosine counter-selection in species with tyrosine kinases. Our results suggest that, alongside the risk for toxicity, the cell can tolerate a large degree of non-functional crosstalk as interaction networks evolve. Synopsis: How do off-target interactions between proteins inside crowded cells affect cellular fitness? Expression of human tyrosine kinases in yeast, which lacks general tyrosine phosphorylation, shows that many individual spurious pY sites have no impact, but also a strong negative correlation between phosphorylation and fitness. 20% of pY sites are predicted to destabilise the substrate. Native pS/pT signalling in yeast is likely dysregulated. Many Y sites are likely phosphorylated at low stoichiometry. Spurious phosphorylation is widespread and not restricted to homologs of native substrates. Evolutionary selection to avoid spurious Y phosphorylation appears unlikely. Expression of human tyrosine kinases in yeast shows that while there is a strong negative correlation between phosphorylation and fitness, many individual pY sites have negligible impact. [ABSTRACT FROM AUTHOR]

Statins are one of the most important classes of drugs. In this analytical review, we elucidate the intricate molecular mechanisms and toxicological rationale regarding both the on‐ (targeting 3‐hydroxy‐3‐methylglutaryl‐coenzyme A reductase [HMGCR]) and off‐target effects of statins. Statins interact with a number of membrane kinases, such as epidermal growth factor receptor (EGFR), erb‐b2 receptor tyrosine kinase 2 (HER2) and MET proto‐oncogene, receptor tyrosine kinase (MET), as well as cytosolic kinases, such as SRC proto‐oncogene, non‐receptor tyrosine kinase (Src) and show inhibitory activity at nanomolar concentrations. In addition, they interact with calcium ATPases and peroxisome proliferator‐activated receptor α (PPARα/NR1C1) at higher concentrations. Statins interact with mitochondrial complexes III and IV, and their inhibition of coenzyme Q10 synthesis also impairs the functioning of complexes I and II. Statins act as inhibitors of kinases, calcium ATPases and mitochondrial complexes, while activating PPARα. These off‐target effects likely contribute to the side effects observed in patients undergoing statin therapy, including musculoskeletal symptoms and hepatic effects. Interestingly, some off‐target effects of statins could also be the cause of favourable outcomes, relating to repurposing statins in conditions such as inflammatory disorders and cancer. [ABSTRACT FROM AUTHOR]

Acute myeloid leukemia (AML) patients with the FMS‐related receptor tyrosine kinase 3 internal tandem duplication (FLT3/ITD) mutation have a poorer prognosis, and treatment with FLT3 tyrosine kinase inhibitors (TKIs) has been hindered by resistance mechanisms. One such mechanism is known as adaptive resistance, in which downstream signaling pathways are reactivated after initial inhibition. Past work has shown that FLT3/ITD cells undergo adaptive resistance through the reactivation of extracellular signal‐regulated kinase (ERK) signaling within 24 h of sustained FLT3 inhibition. We investigated the mechanism(s) responsible for this ERK reactivation and hypothesized that targeting tyrosine‐protein kinase receptor UFO (AXL), another receptor tyrosine kinase that has been implicated in cancer resistance, may overcome the adaptive ERK reactivation. Experiments revealed that AXL is upregulated and activated in FLT3/ITD cell lines mere hours after commencing TKI treatment. AXL inhibition combined with FLT3 inhibition to decrease the ERK signal rebound and to exert greater anti‐leukemia effects than with either treatment alone. Finally, we observed that TKI‐induced AXL upregulation occurs in patient samples, and combined inhibition of both AXL and FLT3 increased efficacy in our in vivo models. Taken together, these data suggest that AXL plays a role in adaptive resistance in FLT3/ITD AML and that combined AXL and FLT3 inhibition might improve FLT3/ITD AML patient outcomes. [ABSTRACT FROM AUTHOR]

Translational medicine provides insight into novel drugs and predicts unwanted effects. In well-characterized pathways (e.g., cytokine-Janus kinase [JAK]-signal transducers and activators of transcription [STAT]), a variety of in vitro assessments were used to estimate selectivity of effects on different potential targets (i.e., JAK1, JAK2, JAK3, and tyrosine kinase 2 [TYK2]). Several approved drugs were characterized as selective for the JAK family. These assessments are challenged by a lack of compounds that only inhibit one JAK family member. Deucravacitinib is a first-in-class, oral, selective, allosteric inhibitor of TYK2, a kinase required for IL-12, IL-23, and Type I interferon signaling. Unlike deucravacitinib, which selectively binds to the TYK2 regulatory domain, JAK1,2,3 inhibitors target the catalytic domain, contributing to nonselective targeting of JAK1,2,3. Cytokines associated with JAK1,2,3 signaling are required for both immune and nonimmune functions. A similar laboratory abnormality profile was observed in clinical trials using JAK1,2,3 inhibitors that has not been observed with deucravacitinib. In vitro testing of JAK1,2,3 inhibitors has relied upon assays of signal transduction, such as those measuring STAT phosphorylation, for estimates of potency and selectivity. These assay systems can be effective in estimating in vivo efficacy; however, they may not provide insight into downstream outcomes of receptor signaling, which may be more relevant for evaluating safety aspects. Assay systems assessing functional outcomes from cells may yield a more useful translational evaluation. Here, deucravacitinib was assessed for potency and selectivity versus three representatives of the JAK inhibitor class (tofacitinib, baricitinib, and upadacitinib) based on functional assays. JAK inhibitors had suppressive activity against JAK2-dependent hematopoietic colony-forming assays modeling thrombopoiesis, erythropoiesis, and myelopoiesis; however, deucravacitinib did not. Deucravacitinib had limited potency against NK cells, cytotoxic T cells, Thelper cells, and regulatory T cells activated by JAK1/JAK3-dependent common gamma chain cytokines. These data are consistent with the biologic role of JAK1,2,3 and pharmacodynamic changes in clinical laboratory abnormalities. Against TYK2-dependent cytokines, deucravacitinib selectively inhibited Type I interferon stimulation of monocytes and dendritic cells and was a more potent inhibitor than JAK inhibitors. IL-12 and IL-23 functional outputs were similarly potently inhibited by deucravacitinib. Results are consistent with deucravacitinib selectively inhibiting TYK2. [ABSTRACT FROM AUTHOR]

Neurotrophic receptor tyrosine kinases (TrkA, TrkB, TrkC), despite their homology, contribute to the clinical heterogeneity of the childhood cancer neuroblastoma. TrkA expression is associated with low-stage disease and is often seen with spontaneous tumour regression. Conversely, TrkB is present in unfavourable neuroblastomas that often harbour amplification of the MYCN oncogene. The role of TrkC is less clearly defined, although some studies suggest its association with a favourable outcome. Understanding the differences in activity of Trk receptors that drive divergent clinical phenotypes as well as the influence of MYCN amplification on downstream Trk receptor signalling remains poorly understood. Here, we present a comprehensive label-free mass spectrometry-based total proteomics and phosphoproteomics dataset (432 raw files with FragPipe search outputs; available on PRIDE with accession number PXD054441) where we identified and quantified 4,907 proteins, 16,744 phosphosites and 5,084 phosphoproteins, derived from NGF/BDNF/NT-3 treated TrkA/B/C-overexpressing neuroblastoma cells with differential MYCN status. Analysing our dataset offers valuable insights into TrkA/B/C receptor signalling in neuroblastoma and its modulation by MYCN status; and holds potential for advancing therapeutic strategies in this challenging childhood cancer. [ABSTRACT FROM AUTHOR]

Background: NEK10, a serine/threonine/tyrosine kinase belonging to the NEK (NIMA-related kinases) family, has been associated with diverse cellular processes. However, no specific target pathways have been identified. Our previous work knocking down NEK10 in HeLa cells suggested a functional association with mitochondria, as we observed altered mitochondrial morphology, mitochondrial oxygen consumption, mtDNA integrity, and reactive oxygen species levels. Methods: To better understand this association, we studied human HAP1 cells fully knockout for NEK10 and confirmed that NEK10 has an important role in mitochondrial homeostasis. We performed the study of mitochondrial respiration, mitochondrial morphology, mitochondrial mass, and mtDNA analysis. Additionally, we showed proteome and phosphoproteome data of crude mitochondrial fraction of Parental and NEK10 KO cells using liquid chromatography-mass spectrometry (LC–MS/MS). Results: In the absence of NEK10 several mitochondrial functions were disturbed. Moreover, proteome and phosphoproteome analyses of mitochondrial fractions showed that NEK10 alters the threonine phosphorylation status of several mitochondrial/endoplasmic reticulum components, including HSP60, NDUFB4, and TOM20. These changes impacted the steady-state levels of a larger group of proteins, preferentially involving respiratory complexes and autophagy pathways. Conclusion: We concluded that NEK10 plays a key role in mitochondrial function, possibly by modulating the phosphorylation status of mitochondrial proteins. [ABSTRACT FROM AUTHOR]

Despite our increasing understanding of macrophage heterogeneity, drivers of macrophage phenotypic and functional polarization in the microenvironment are not fully elucidated. Here, our single-cell RNA sequencing data identify a subpopulation of macrophages expressing high levels of the phagocytic receptor MER proto-oncogene tyrosine kinase (MerTK+ macrophages), which is closely associated with melanoma progression and immunotherapy resistance. Adoptive transfer of the MerTK+ macrophages into recipient mice notably accelerated tumor growth regardless of macrophage depletion. Mechanistic studies further revealed that ALK And LTK Ligand 1 (ALKAL1), a target gene of aryl hydrocarbon receptor (AhR), facilitated MerTK phosphorylation, resulting in heightened phagocytic activity of MerTK+ macrophages and their subsequent polarization toward an immunosuppressive phenotype. Specifically targeted delivery of AhR antagonist to tumor-associated macrophages with mannosylated micelles could suppress MerTK expression and improved the therapeutic efficacy of anti-programmed cell death ligand 1 therapy. Our findings shed light on the regulatory mechanism of MerTK+ macrophages and provide strategies for improving the efficacy of melanoma immunotherapy. [ABSTRACT FROM AUTHOR]

Until recently, treatment options for patients diagnosed with Acute Myeloid Leukemia (AML) were limited and predominantly relied on various combinations, dosages, or schedules of traditional chemotherapeutic agents. Patients with advanced age, relapsed/refractory disease or comorbidities were often left without effective treatment options. Novel advances in the understanding of leukemogenesis at the molecular and genetic levels, alongside recent progress in drug development, have resulted in the emergence of novel therapeutic agents and strategies for AML patients. Among these innovations, the receptor tyrosine kinase AXL has been established as a promising therapeutic target for AML. AXL is a key regulator of several cellular functions, including epithelial-to-mesenchymal transition in tumor cells, immune regulation, apoptosis, angiogenesis and the development of chemoresistance. Clinical studies of AXL inhibitors, as single agents and in combination therapy, have demonstrated promising efficacy in treating AML. Additionally, novel AXL-targeted therapies, such as AXL-specific antibodies or antibody fragments, present potential solutions to overcome the limitations associated with traditional small-molecule AXL inhibitors or multikinase inhibitors. This review provides a comprehensive overview of the structure and biological functions of AXL under normal physiological conditions, including its role in immune regulation. We also summarize AXL's involvement in cancer, with a specific emphasis on its role in the pathogenesis of AML, its contribution to immune evasion and drug resistance. Moreover, we discuss the AXL inhibitors currently undergoing (pre)clinical evaluation for the treatment of AML. [ABSTRACT FROM AUTHOR]

Background: Latency remains a major obstacle to finding a cure for HIV despite the availability of antiretroviral therapy. Due to virus dormancy, limited biomarkers are available to identify latent HIV-infected cells. Profiling of individual HIV-infected cells is needed to explore potential latency biomarkers and to study the mechanisms of persistence that maintain the HIV reservoir. Methods: Single cell spatial transcriptomic characterization using the CosMx Spatial Molecular Imager platform was conducted to analyze HIV-infected cells in formalin-fixed paraffin-embedded sections of splenic tissue surgically obtained from an HIV-infected humanized mouse model. Regulation of over a thousand human genes was quantified in both viremic and aviremic specimens. In addition, in situ hybridization and immunohistochemistry were performed in parallel to identify HIV viral RNA- and p24-containing cells, respectively. Finally, initial findings from CosMx gene profiling were confirmed by isolating RNA from CD4 + T cells obtained from a person living with HIV on antiretroviral therapy following either PMA/Ionomycin or DMSO treatment. RNA was quantified using qPCR for a panel of targeted human host genes. Results: Supervised cell typing revealed that most of the HIV-infected cells in the mouse spleen sections were differentiated CD4 + T cells. A significantly higher number of infected cells, 2781 (1.61%) in comparison to 112 (0.06%), and total HIV transcripts per infected cell were observed in viremic samples compared to aviremic samples, respectively, which was consistent with the data obtained from ISH and IHC. Notably, the expression of 55 genes was different in infected cells within tissue from aviremic animals compared to viremic. In particular, both spleen tyrosine kinase (SYK) and CXCL17, were expressed approximately 100-fold higher. This data was further evaluated against bulk RNA isolated from HIV-infected human primary CD4 + T cells. A nearly 6-fold higher expression of SYK mRNA was observed in DMSO-treated CD4 + T cells compared to those stimulated with PMA/Ionomycin. Conclusion: This study found that the CosMx SMI platform is valuable for assessing HIV infection and providing insights into host biomarkers associated with HIV reservoirs. Higher relative expression of the SYK gene in aviremic-infected cells from the humanized mouse HIV model was consistent with levels found in CD4 + T cells of aviremic donors. [ABSTRACT FROM AUTHOR]

7-Azaindoles are compounds of considerable medicinal interest. During development of the structure–activity relationship for inhibitors of the colony stimulated factor 1 receptor tyrosine kinase (CSF1R), a specific 2-aryl-1H-pyrrolo[2,3-b]pyridin-4-amine was needed. Two different synthetic strategies were evaluated, in which the order of the key C-C and C-N cross-coupling steps differed. The best route relied on a chemoselective Suzuki–Miyaura cross-coupling at C-2 on a 2-iodo-4-chloropyrrolopyridine intermediate, and subsequently a Buchwald–Hartwig amination with a secondary amine at C-4. Masking of hydroxyl and pyrroles proved essential to succeed with the latter transformation. The final trimethylsilylethoxymethyl (SEM) deprotection step was challenging, as release of formaldehyde gave rise to different side products, most interestingly a tricyclic eight-membered 7-azaindole. The target 2-aryl-1H-pyrrolo[2,3-b]pyridin-4-amine (compound 3c) proved to be 20-fold less potent than the reference inhibitor, confirming the importance of the N-3 in the pyrrolopyrimidine parent compound for efficient CSF1R inhibition. [ABSTRACT FROM AUTHOR]

Background: Psoriasis, a chronic, immune-mediated, inflammatory disease, affects 2‒3% of the population. Tyrosine kinase 2 (TYK2) mediates cytokine signaling involved in adaptive [interleukin (IL)-12, IL-23] and innate (type-I interferons) immune responses; IL-23–driven T-helper (Th)17 pathways play a key role in chronic inflammation in psoriasis. In a phase 2 trial, deucravacitinib, an oral, selective, allosteric TYK2 inhibitor, reduced IL-23/Th17 and type-I interferon pathway expression in the skin of patients with moderate to severe plaque psoriasis, reductions that were accompanied by clinical improvement of psoriatic lesions. Objectives: The aim of this study was to identify biomarkers of psoriatic disease in serum from patients enrolled in the phase 2 trial and to assess the effects of deucravacitinib on those biomarkers. Methods: Serum biomarkers from Olink proteomics and other quantitative assays were evaluated for a pharmacodynamic response to deucravacitinib treatment and correlation with psoriasis disease activity measures. Results: Serum biomarkers associated with the IL-23/Th17 pathway [IL-17A, IL-17C, IL-19, IL-20, beta-defensin, and peptidase inhibitor 3 (PI3)] were upregulated in patients with psoriasis versus healthy controls. Deucravacitinib treatment reduced IL-17A (adjusted mean change from baseline at Day 85; 12 mg once daily versus placebo; −0.240 versus −0.067), IL-17C (−14.850 versus −1.664), IL-19 (−96.445 versus −8.119), IL-20 (−0.265 versus −0.064), beta-defensin (−65,025.443 versus −7553.961), and PI3 (−14.005 versus −1.360) expression. Reductions in serum biomarker expression occurred in a dose- and time-dependent manner, with significant reductions from baseline seen with deucravacitinib doses ≥ 3 mg twice daily (P ≤ 0.05). Biomarker expression correlated with disease activity measures such as Psoriasis Area and Severity Index (PASI) at baseline. Biomarker expression also correlated with PASI scores at Week 12. Conclusion: IL-23/Th17 pathway expression in the serum of patients with psoriasis is an indicator of disease activity and response to deucravacitinib treatment. Trial registration number: NCT02931838. Plain Language Summary: Plaque psoriasis is a long-term disease that causes inflammation, scaling, and itching of the skin. Compared with healthy volunteers without psoriasis, patients with psoriasis have higher amounts of certain biomarkers (molecules that indicate what is happening in the body) in their blood that are associated with inflammation. Higher amounts of these biomarkers are also associated with more severe psoriasis. In a study of patients with psoriasis, those who received the oral drug deucravacitinib had lower amounts of biomarkers after 12 weeks of treatment compared with patients who received a placebo (a lookalike pill that contains no medicine). Patients who were treated with deucravacitinib also saw an improvement in their psoriasis after 12 weeks compared with patients who received placebo. [ABSTRACT FROM AUTHOR]

Background: Receptor tyrosine kinase (RTK) fusions of RET, NTRK1/3, and ALK are enriched among pediatric thyroid cancer patients with metastatic and persistent disease, and their oncoproteins represent attractive drug targets. Methods: We performed RNA-sequencing in a papillary thyroid cancer (PTC) lacking other frequent driver alterations. Results: We report a novel RTK fusion, TG-insulin-like growth factor 1 receptor gene (IGF1R), in a 17-year-old female patient with angioinvasive follicular variant PTC. The in-frame fusion protein preserves the cholinesterase‐like domain of TG with dimerization properties and the transmembrane and kinase domain of IGF1R. The tumor sample shows increased IGF1R mRNA expression and tyrosine kinase phosphorylation, augmentation of Mitogen activated protein kinase (MAPK) transcriptional output genes, and decreased NIS levels. Conclusions: We reveal a novel targetable kinase fusion oncogene in thyroid cancer which is not incorporated in different thyroid-specific sequencing panels. The integration of IGF1R fusion screening in the next versions of thyroid-specific targeted next-generation sequencing panels may be beneficial to thyroid cancer patients. [ABSTRACT FROM AUTHOR]

Objective: Encouraged by the significant anticancer activities of benzimidazole, piperazine, and 1,2,3-triazole containing derivatives and based on the need for the development of new EGFR targeting antibreast cancer agents, our group now interested to synthesize new benzimidazole-piperazine-1,2,3-triazoles (VIIa–VIIn) as in vitro antibreast cancer and anti-EGFR agents. Methods: In this study, we focused on the synthesis of benzimidazole-piperazine-1,2,3-triazoles. As well, the in vitro antibreast cancer activity of compounds (VIIa–VIIn) against MDA-MB-231 and MCF-7 cell lines, and anti-EGFR activity of few compounds were studied via IC50 values. Results and Discussion: Compounds (1-methyl-1H-benzo[d]imidazol-2-yl)(4-((1-(p-tolyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)methanone (VIIb), (4-((1-(4-fluorophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)(1-methyl-1H-benzo[d]imidazol-2-yl)metha none (VIIe), (4-((1-(3,5-dichlorophenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)(1-methyl-1H-benzo[d]imidazol-2-yl)methanone (VIIh), and (4-((1-(4-chloro-3,5-dimethoxyphenyl)-1H-1,2,3-triazol-4-yl)methyl)piperazin-1-yl)(1-methyl-1H-benzo[d]imidazol-2-yl)methanone (VIIl) displayed higher activity against two cell lines than the Erlotinib. Furthermore, compounds (VIIb), (VIIe), and (VIIh) displayed greater inhibition against tyrosine kinase EGFR than the Erlotinib. The results of molecular docking studies of (VIIb), (VIIe), (VIIh), and (VIIl) as EGFR (PDB ID: 4HJO) targeting agents were found to be supportive with corresponding IC50 data. Conclusions: We propose that mechanism of in vitro antibreast cancer activity of the investigated compounds, possibly will due to the tyrosine kinase EGFR inhibition. [ABSTRACT FROM AUTHOR]

Objective: Midpalatal expansion (MPE) is routinely employed to treat transverse maxillary arch deficiency. Neutrophils are indispensable for recruiting bone marrow stromal cells (BMSCs) at the initial stage of bone regeneration. This study aimed to explore whether neutrophils participate in MPE and how they function during bone formation under mechanical stretching. Materials and Methods: The presence and phenotype of neutrophils in the midpalatal suture during expansion were detected by flow cytometry and immunofluorescence staining. The possible mechanism of neutrophil recruitment and polarization was explored in vitro by exposing vascular endothelial cells (VECs) to cyclic tensile strain. Results: The number of neutrophils in the distracted suture peaked on Day 3, and N2‐type neutrophils significantly increased on Day 5 after force application. The depletion of circulatory neutrophils reduced bone volume by 43.6% after 7‐day expansion. The stretched VECs recruited neutrophils via a CXCR2 mechanism in vitro, which then promoted BMSC osteogenic differentiation through the VEGFA/VEGFR2 axis. Consistently, these neutrophils showed higher expression of canonical N2 phenotype genes, including CD206 and Arg1. Conclusions: These results suggested that neutrophils participated in early bone formation during MPE. Based on these findings, we propose that stretched VECs recruited and polarized neutrophils, which, in turn, induced BMSC osteogenic differentiation. [ABSTRACT FROM AUTHOR]

Spleen tyrosine kinase (SYK), a non-receptor tyrosine kinase that is overexpressed in several solid tumors, presents a promising therapeutic target. SYK activation downstream of various receptors drives signaling pathways that facilitate tumor progression and immune evasion. SYK inhibition enhances the antitumor immune response in solid tumors by reprogramming myeloid and B cells or directly targeting tumor cells. SYK inhibition can overcome resistance to chemotherapy and immunotherapy in solid tumors. Clinical trials targeting SYK in solid tumors are underway. Spleen tyrosine kinase (SYK) is predominantly expressed in hematopoietic cells and has been extensively studied for its pivotal role in B cell malignancies and autoimmune diseases. In epithelial solid tumors, SYK shows a paradoxical role, acting as a tumor suppressor in some cancers while driving tumor growth in others. Recent preclinical studies have identified the role of SYK in the tumor microenvironment (TME), revealing that SYK signaling in immune cells, especially B cells, and myeloid cells, promote immunosuppression, tumor growth, and metastasis across various solid tumors. This review explores the emerging roles of SYK in solid tumors, the mechanisms of SYK activation, and findings from preclinical and clinical studies of SYK inhibitors as either standalone treatments or in combination with immunotherapy or chemotherapy for solid tumors. [ABSTRACT FROM AUTHOR]

Background: Cancer is the predominant cause of mortality and a remarkable obstacle to elevating life anticipation in every nation on globe. Hepatocellular carcinoma (HCC), a hyper-vascular tumour, develops and progresses due to angiogenesis, a key feature of malignancy. HCC exhibits high neoangiogenic activity because of the need to generate new blood vessels for tumour growth. Methods: The present work includes the construction of virtual library of ligands, virtual screening using the Dockthor-VS server, ADMET study using the SwissADME and Osiris property explorer. All the synthesized compounds were characterized by UV, IR, NMR and mass spectroscopic techniques. MTT assay was done to find the IC50 of the synthesized compounds against HepG2 cell line. The more active compound found is subjected to the molecular dynamics simulation study. Results: The ligands exhibited good docking scores, ADMET profile compared to the reference drugs. The target compounds were obtained with the satisfactory yields of 66–82%. The best activity against the HepG2 cancer cell line is observed with the compound SA-4 with IUPAC name (2-(3-methyl-2-oxoquinoxalin-1(2H)-yl)-N-(5-(3-nitrophenyl)-5H-thiazolo[4,3-b] [1,3,4] thiadiazol-2-yl) acetamide). The experimental results obtained show correlation with the in silico results. MD simulation of the compound SA-4 indicates the moderate stability of the protein-ligand complex in real time environment. Conclusion: The results obtained suggest that the compound SA-4 has the potential to be a promising anticancer agent effective against the VEGFR-2 and FGFR-4. [ABSTRACT FROM AUTHOR]

The synthesis of some new quinoxaline-1,2,4-oxadiazole-amide conjugates (6a–n) was described, and their structures were determined using 1HNMR, 13CNMR, and mass spectral analysis. The in vitro anti-cancer activity of the compounds (6a–n) against three human cancer cell lines such as MCF-7 (breast), HepG2 (lung), and DU-145 (prostate) revealed that the compounds 6d, 6e, and 6f exhibited promising activity against three cancer cell lines. Predominantly, compound 6f demonstrated greater activity than the standard drug Etoposide on MCF-7, HepG2, and DU-145 with IC50 values of 0.82 ± 0.01, 1.30 ± 0.02, and 2.12 ± 0.04 µM, respectively. Furthermore, the compounds 6e and 6f displayed promising inhibitory activity over the tyrosine kinase EGFR when compared with the standard Erlotinib. Molecular docking studies carried out on three potent compounds (6d, 6e, and 6f) on the EGFR receptor recommended that the compound 6f strongly binds to protein EGFR (pdbid: 4HJO). In addition, the in silico pharmacokinetic profile was also achieved for the three potent compounds 6d, 6e, and 6f using SWISS/ADME and pk CSM. Results showed that the compounds 6d, 6e, and 6f followed the Lipinski rule, Veber rule, Egan rule, Ghose rule, and Muegge rule without any deviation. [ABSTRACT FROM AUTHOR]

We synthesized a new series of coumarin-piperazine 1,2,3-triazole hybrids (7a–7n) and screened for their in vitro anti-breast cancer activity against MCF-7, MDA-MB-468, and MDA-MB-231 cancer cell lines. The results revealed that compound 7f showed superior activity against all cell lines than the standard drug 5-fluorouracil. As well, compounds 7e, 7h, and 7k showed greater activity against MCF-7 than the positive control. Furthermore, 7f and 7h showed higher potency in inhibiting tyrosine kinase epidermal growth factor receptor (EGFR) than the standard drug erlotinib. Finally, in silico molecular docking studies on EGFR (PDB ID-4HJO) revealed that compounds 7e, 7f, 7h, and 7k possess good binding energies and inhibition constants as compared to erlotinib. [ABSTRACT FROM AUTHOR]

Herein, we synthesized some new 4-azaindole-thiazolidine-2,4-dione-1,2,3-triazole hybrids (6a-6n) via Knoevenagel condensation and copper(I) catalyzed azide-alkyne cycloaddition (CuAAC) as key approaches. These hybrids were then screened for their in vitro anticancer activity against three human cancer cell lines like MCF-7 (breast), A549 (lung) and HepG2 (hepatocellular) using erlotinib as standard drug. Out of all, compounds 6a, 6k and 6m were found to be more active against all cell lines with IC50 values <18 µM. As well, compounds 6a (IC50 = 0.40 µΜ), 6k (IC50 = 0.29 µΜ) and 6m (IC50 = 0.18 µΜ) showed higher potency ininhibiting tyrosine kinase EGFR than the erlotinib (IC50 = 0.41 µΜ). Further, molecular dockingof compounds 6a, 6k and 6 m on EGFR protein revealed that they have good binding energies with the target protein (−9.96 kcal/mol, −9.92 kcal/mol and −10.37 kcal/mol respectively) which were found to be supportive with the corresponding in vitro activities data. [ABSTRACT FROM AUTHOR]

Objective: The 48‐week, phase 2 SLEek study (NCT03978520) evaluated the efficacy and safety of upadacitinib (JAK inhibitor) and elsubrutinib (BTK inhibitor) alone or in combination (ABBV‐599) in adults with moderately to severely active systemic lupus erythematosus (SLE). Methods: Patients were randomized 1:1:1:1:1 to elsubrutinib 60 mg and upadacitinib 30 mg once daily (ABBV‐599 high dose), elsubrutinib 60 mg and upadacitinib 15 mg once daily (ABBV‐599 low dose), elsubrutinib 60 mg once daily (QD), upadacitinib 30 mg QD, or placebo QD. The primary endpoint was the proportion of patients achieving both Systemic Lupus Erythematosus Responder Index 4 (SRI‐4) and glucocorticoid dose ≤10 mg QD at week 24. Additional assessments through week 48 included British Isles Lupus Assessment Group‐Based Composite Lupus Assessment (BICLA) and Lupus Low Disease Activity State (LLDAS) responses, number of flares, time to first flare, and adverse events. Results: The study enrolled 341 patients. The ABBV‐599 low dose and elsubrutinib arms were discontinued after a planned interim analysis showed lack of efficacy (no safety concerns). More patients achieved the primary endpoint with upadacitinib (54.8%; P = 0.028) and ABBV‐599 high dose (48.5%; P = 0.081) versus placebo (37.3%). SRI‐4, BICLA, and LLDAS response rates were higher for both upadacitinib and ABBV‐599 high dose versus placebo at weeks 24 and 48. Flares were reduced, and time to first flare through week 48 was substantially delayed with both upadacitinib and ABBV‐599 high dose versus placebo. No new safety signals were observed beyond those previously reported for upadacitinib or elsubrutinib. Conclusion: Upadacitinib 30 mg alone or in combination with elsubrutinib (ABBV‐599 high dose) demonstrated significant improvements in SLE disease activity and reduced flares and were well tolerated through 48 weeks. [ABSTRACT FROM AUTHOR]

Cytokines function as communication tools of the immune system, serving critical functions in many biological responses and shaping the immune response. When cytokine production or their biological activity goes awry, the homeostatic balance of the immune response is altered, leading to the development of several pathologies such as autoimmune and inflammatory disorders. Cytokines bind to specific receptors on cells, triggering the activation of intracellular enzymes known as Janus kinases (JAKs). The JAK family comprises four members, JAK1, JAK2, JAK3 and tyrosine kinase 2, which are critical for intracellular cytokine signalling. Since the mid-2010s multiple JAK inhibitors have been approved for inflammatory and haematological indications. Currently, approved JAK inhibitors have demonstrated clinical efficacy; however, improved selectivity for specific JAKs is likely to enhance safety profiles, and different strategies have been used to accomplish enhanced JAK selectivity. In this update, we discuss the background of JAK inhibitors, current approved indications and adverse effects, along with new developments in this field. We address the issue of JAK selectivity and its relevance in terms of efficacy, and describe new modalities of JAK targeting, as well as new aspects of JAK inhibitor action. This Review provides an update on developments in Janus kinase inhibitors, including new disease indications and adverse effects. The authors discuss issues surrounding selectivity and efficacy, as well as new routes for administration of Janus kinase inhibitors. Key points: Januse kinase (JAK) inhibitors show equivalent or superior efficacy compared with biologic DMARDs in several autoimmune and inflammatory diseases. The use of JAK inhibitors has been hampered by adverse events, which could be linked to the JAKs that are blocked. Assessment of individual JAK inhibitor selectivity is still a matter of debate, as different assays can yield different results. Selective JAK inhibitors are generally equally effective, but improved, long-term safety has not been fully established. Tissue-targeted JAK inhibitors could circumvent the problem of adverse effects resulting from systemic administration. Alternative strategies for JAK inhibition with small interfering RNAs or metabolite derivatives constitute an exciting area of development. [ABSTRACT FROM AUTHOR]

New generation BCR-ABL1 TKIs raised attention regarding their adverse effects, including hepatotoxicity. Indeed, bosutinib and nilotinib were associated with severe hepatotoxicity compared with imatinib. Moreover, ponatinib has a boxed warning due to its potential to cause inflammatory liver damage, even death. However, the underlying mechanisms remain unclear. This study aimed to investigate the role of NLRP3 inflammasome activation in the underlying mechanism of ponatinib and bosutinib-induced hepatotoxicity. Furthermore, we determined the initiating event of this adverse outcome pathway by measuring the levels of reactive oxygen species as well as mitochondrial membrane potential in AML12 cells. The results demonstrated that ponatinib or bosutinib markedly inhibited cell viability and caused cytosolic membrane damage in cells. Moreover, drugs (IC 50) dramatically induced oxidative stress and mitochondrial membrane potential disruption, which led to upregulation in the expression levels of NLRP3 inflammasome-related genes and proteins, activation of NLRP3 inflammasomes, cleavage of gasdermin-D and caspase-1, secretion of IL-1β, and cytosolic membrane damage. Furthermore, MCC950, a well-known specific inhibitor of NLRP3 inflammasome, and antioxidant N-acetyl-l-cysteine reversed the effects of drugs on the NLRP3 signaling pathway and cytosolic membranes. In summary, NLRP3 inflammasome activation is involved in new-generation BCR-ABL1 TKIs-triggered hepatotoxicity. Mitochondrial damage and reactive oxygen species accumulation were significant upstream signaling events in this signaling pathway. [Display omitted] • NLRP3 signaling pathway is a promising therapeutic target for the prevention of drug toxicity. • NLRP3 inflammasome activation might contribute to the hepatotoxicity effects of ponatinib and bosutinib. • Oxidative stress might be initiating event of ponatinib and bosutinib-induced NLRP3 inflammasome activation. [ABSTRACT FROM AUTHOR]

Colony stimulating factor 1 receptor (CSF1R) is an essential receptor for both colony stimulating factor 1 (CSF1) and interleukin (IL) 34 signaling expressed on monocyte precursors and myeloid cells, including monocytes, dendritic cells (DC), and microglia. In humans, dominant heterozygous pathogenic variants in CSF1R cause a neurological condition known as CSF1R-related disorder (CSF1R-RD), typically with late onset, previously referred to as adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP). CSF1R-RD is characterized by microglia reduction and altered monocyte function; however, the impact of pathogenic CSF1R variants on the human DC lineage remains largely unknown. We previously reported that cord blood CD34+ stem cell-derived DCs generated in vitro originate specifically from CSF1R expressing precursors. In this study, we examined the DC lineage of four unrelated patients with late-onset CSF1R-RD who carried heterozygous missense CSF1R variants (c.2330G>A, c.2375C>A, c.2329C>T, and c.2381T>C) affecting different amino acids in the protein tyrosine kinase domain of CSF1R. CD34+ stem cells and CD14+ monocytes were isolated from peripheral blood and subjected to an in vitro culture protocol to differentiate towards conventional DCs and monocyte-derived DCs, respectively. Flow cytometric analysis revealed that monocytes from patients with late-onset CSF1R-RD were still able to differentiate into monocyte-derived DCs in vitro , whereas the ability of CD34+ stem cells to differentiate into conventional DCs was impaired. Strikingly, the peripheral blood of patients contained all naturally occurring DC subsets. We conclude that the in vitro abrogation of DC-development in patients with heterozygous pathogenic missense CSF1R variants does not translate to an impairment in DC development in vivo and speculate that CSF1R signalling in vivo is compensated, which needs further study. • CSF1R is crucial for in vitro cDC differentiation from CD34+ stem cells but not essential for monocyte-derived DC development. • Late-onset CSF1R-RD patients have all naturally occurring dendritic cell subsets in their peripheral blood despite impaired CSF1R. • In vivo compensation by other cytokines might explain normal DC subsets in late-onset CSF1R-RD patients. [ABSTRACT FROM AUTHOR]

The past year has offered significant advancements in the field of non–small cell lung cancer (NSCLC), both in the early and advanced disease settings. The identification of guideline‐recommended actionable targets has provided the foundation for developing multiple new therapeutic agents. There has been a focus on developing drugs designed to overcome acquired resistance, a limitation of tyrosine kinase inhibitor–based therapy in lung cancer. In addition, there is an emerging trend toward combination therapies for patients in the first‐line setting with the goal of preventing or delaying resistance. Another promising area of development has been the use of antibody–drug conjugates, where there are the initial reports of central nervous system efficacy and activity in patients with genomic alterations. Over the past year, numerous publications and presentations have highlighted multiple therapeutic advances, offering new treatment options for patients with NSCLC. The focus of this review is to summarize the most impactful findings, emphasizing their significance in the evolving treatment landscape for NSCLC. Several landmark trials in lung cancer with practice‐changing clinical implications have been presented and published in 2023. This article reviews a selection of these trials as they relate to early and advanced‐stage oncogene‐driven lung cancer. The ADAURA and ALINA trials, in which targeted therapy given in the adjuvant setting has demonstrated improved clinical outcomes, are reviewed. In the advanced‐stage setting, recent trials in the context of specific oncogene drivers are reviewed, including EGFR, ALK, ROS1, RET, ERBB2 (HER2), BRAF, MET exon 14 skipping (METex14), and KRAS alterations. Also discussed are the results of several trials that have evaluated the use of combination therapies and resistance‐mechanism agnostic treatment strategies. Plain Language Summary: Targeted therapy plays an important role for patients with early and advanced‐stage non–small cell lung cancer carrying specific genetic alterations.New strategies that combine multiple therapies are now being studied in randomized clinical trials, with the goal of enhancing the effectiveness of targeted therapy for patients with advanced lung cancer. Several clinical trials published this past year have contributed to significant advances in thoracic oncology. Here, the focus is on targeted therapy advances reported in 2023 that are impacting clinical decision‐making in everyday practice. [ABSTRACT FROM AUTHOR]

Recent clinical evidence shows that the antibody-drug conjugate (ADC) trastuzumab deruxtecan (T-DXd) can successfully treat patients with advanced HER2-mutant non-small cell lung cancer (NSCLC). We aimed to characterize HER2 mutations in cervical neuroendocrine carcinoma (NEC) among Taiwanese women to provide the rationale for exploring T-DXd as a tumor-agnostic targeted therapy option. We analyzed 12 archived primary cervical NEC samples from Taiwanese patients. Tumor-rich areas were marked for microdissection on 10 μm unstained sections. DNA was extracted, and HER2 hotspots were sequenced using a targeted panel on the Illumina MiSeq. HER2 missense mutations were identified in 5 of 12 cases (41.7%). Of the 5 cases with mutations, 2 patients (40%) had a single mutation, while 3 patients (60%) had double mutations. We detected 4 substitutions outside the tyrosine kinase domain (non-TKD), which were p.P1170A, p.S305C, p.I655V, and a novel T328K alteration. No mutations were found within the tyrosine kinase domain (TKD). The 41.7% HER2 mutation rate warrants expanded screening and future clinical investigation of the T-DXd targeting HER2 mutations in cervical NEC patients. Overall, this study contributes to the molecular understanding of cervical NEC and lays the groundwork for developing more effective treatment strategies. [ABSTRACT FROM AUTHOR]

The plasma proteome can mediate poor oral health problems (POHP)'s link to incident dementia. We screened 37,269 UK Biobank participants 50–74 years old (2006–2010) for prevalent POHP, further tested against 1463 plasma proteins and incident dementia over up to 15 years of follow-up. Total effect (TE) of POHP-dementia through plasma proteomic markers was decomposed into pure indirect effect (PIE), interaction referent (INTREF), controlled direct effect (CDE), or mediated interaction (INTMED). POHP increased the risk of all-cause dementia by 17% (P < 0.05). Growth differentiation factor 15 (GDF15) exhibited the strongest mediating effects (PIE > 0, P < 0.001), explaining 28% the total effect of POHP on dementia, as a pure indirect effect. A first principal component encompassing top 4 mediators (GDF15, IL19, MMP12, and ACVRL1), explained 11% of the POHP-dementia effect as a pure indirect effect. Pathway analysis including all mediators (k = 173 plasma proteins) revealed the involvement of the immune system, signal transduction, metabolism, disease, and gene expression, while STRING analysis indicated that top mediators within the first principal component were also represented in the two largest proteomic clusters. The dominant biological GO pathway for the GDF15 cluster was GO:0007169 labeled as "transmembrane receptor protein tyrosine kinase signaling pathway." Dementia is linked to POHP mediated by GDF15 among several proteomic markers. [ABSTRACT FROM AUTHOR]

Objective: Here, we aimed to investigate the role of glucocorticoid and mineralocorticoid receptors (GRs and MRs, respectively) in the regulation of energy homeostasis. Methods: We used three mouse models with simultaneous deletion of GRs and MRs in either forebrain neurons, the paraventricular nucleus, or corticotropin‐releasing hormone (CRH) neurons and compared them with wild‐type controls or isolated knockout groups. In addition to body weight, food intake, energy expenditure, insulin sensitivity, fat/lean mass distribution, and plasma corticosterone levels, we also performed transcriptomic analysis of CRH neurons and assessed their response to melanocortinergic stimulation. Results: Similar to global double‐knockout models, deletion of GRs and MRs specifically in mature CRH neurons resulted in obesity. Importantly, the latter was accompanied by insulin resistance, but not increased plasma corticosterone levels. Transcriptomic analysis of these neurons revealed upregulation of several genes involved in postsynaptic signal transduction, including the Ptk2b gene, which encodes proline‐rich tyrosine kinase 2. Knockout of both nuclear receptors leads to upregulation of Ptk2b in CRH neurons, which results in their diminished responsiveness to melanocortinergic stimulation. Conclusions: Our data demonstrate the functional redundancy of GRs and MRs in CRH neurons to maintain energy homeostasis and prevent obesity. Simultaneous targeting of both receptors might represent an unprecedented approach to counteract obesity. [ABSTRACT FROM AUTHOR]

Background: Asthma is a common immune disease with high morbidity in children. Type 2 inflammation is the center of asthma development, and mainly mediated by a subset of CD4 + T cells, T helper 2 (Th2) cells. Excess Th2 differentiation was generally associated with asthmatic attack. Casitas B-lineage lymphoma (c-CBL) was reported to involved in T cell development and databank showed its decreased expression in CD4 + T cells from peripheral blood of asthmatic children. This study aims to investigate the role of c-CBL in childhood asthma and Th2 differentiation, and explore the underlying mechanism. Methods: We collected peripheral blood samples from clinical childhood asthma cases and healthy controls, and determined c-CBL expression in CD4 + T cells. Asthma was induced in neonatal mice by ovalbumin (OVA) intraperitoneal injection and aerosol inhalation, and c-CBL expression in CD4 + T cells from peripheral blood and spleen was measured. Gain-of-function experiments was performed to confirm the effects of c-CBL on Th2 differentiation in vitro. Finally, c-CBL was delivered into asthmatic mice via lentivirus infection to verify its effects on experimental asthma. Results: c-CBL was lowly expressed in CD4 + T cells from asthmatic children than those of healthy controls. Similarly, it was downregulated in CD4 + T cells from peripheral blood and spleen of asthma mice. Overexpression of c-CBL restrained lung pathological injury and type 2 inflammation in experimental asthmatic mice. Gain-of-function experiments demonstrated that c-CBL inhibited Th2 differentiation of CD4 + T cells from healthy children, and mediated the ubiquitination of lymphocyte cell-specific protein-tyrosine kinase (LCK). LCK acted as a kinase to phosphorylate and activate c-JUN, which was predicted to bind promoter sequence of CD28 by bioinformatic analysis. Dual-luciferase reporter assay verified that c-JUN and ETS1 synergically enhanced transcription of CD28, and this transcription activation was aggravated by LCK overexpression. Conclusion: c-CBL alleviated asthma and suppressed Th2 differentiation by facilitating LCK ubiquitination, interrupting c-JUN activation and CD28 expression in vivo and in vitro. c-CBL/LCK/c-JUN/ETS1/CD28 axis was partially involved in childhood asthma, and may provide novel insights for clinical treatment for asthma. [ABSTRACT FROM AUTHOR]

Glioblastoma (GBM), a primary brain tumor, exhibits remarkable invasiveness and is characterized by its intricate location, infiltrative behavior, the presence of both the blood–brain barrier (BBB) and the blood–brain tumor barrier (BBTB), phenotypic diversity, an immunosuppressive microenvironment with limited development yet rich vascularity, as well as the resistant nature of glioblastoma stem cells (GSCs) towards traditional chemotherapy and radiotherapy. These formidable factors present substantial obstacles in the quest for effective GBM treatments. Following extensive research spanning three decades, the hepatocellular receptor A2 (EphA2) receptor tyrosine kinase has emerged as a promising molecular target with translational potential in the realm of cancer therapy. Numerous compounds aimed at targeting EphA2 have undergone rigorous evaluation and clinical investigation. This article provides a comprehensive account of the distinctive roles played by canonical and non-canonical EphA2 signaling in various contexts, while also exploring the involvement of the EphA2-ephrin A1 signaling axis in GBM pathogenesis. Additionally, the review offers an overview of completed clinical trials targeting EphA2 for GBM treatment, shedding light on both the prospects and challenges associated with EphA2-directed interventions in the domain of cancer therapeutics. [ABSTRACT FROM AUTHOR]

Developmental biology-inspired strategies for tissue-building have extraordinary promise for regenerative medicine, spurring interest in the relationship between cell biophysical properties and morphological transitions. However, mapping gene or protein expression data to cell biophysical properties to physical morphogenesis remains challenging with current techniques. Here, we present multiplexed adhesion and traction of cells at high yield (MATCHY). MATCHY advances the multiplexing and throughput capabilities of existing traction force and cell-cell adhesion assays using microfabrication and a semiautomated computation scheme with machine learning-driven cell segmentation. Both biophysical assays are coupled with serial downstream immunofluorescence to extract cell type/signaling state information. MATCHY is especially suited to complex primary tissue-, organoid-, or biopsy-derived cell mixtures since it does not rely on a priori knowledge of cell surface markers, cell sorting, or use of lineage-specific reporter animals. We first validate MATCHY on canine kidney epithelial cells engineered for rearranged during transfection (RET) tyrosine kinase expression and quantify a relationship between downstream signaling and cell traction. We then use MATCHY to create a biophysical atlas of mouse embryonic kidney primary cells and identify distinct biophysical states along the nephron differentiation trajectory. Our data complement expression-level knowledge of adhesion molecule changes that accompany nephron differentiation with quantitative biophysical information. These data reveal an "energetic ratchet" that accounts for spatial trends in nephron progenitor cell condensation as they differentiate into early nephron structures, which we validate through agent-based computational simulation. MATCHY offers semiautomated cell biophysical characterization at >10,000-cell throughput, an advance benefiting fundamental studies and new synthetic tissue strategies for regenerative medicine. [ABSTRACT FROM AUTHOR]

Tumor extracellular matrices (ECM) exhibit aberrant changes in composition and mechanics compared to normal tissues. Proteoglycans (PG) are vital regulators of cellular signaling in the ECM with the ability to modulate receptor tyrosine kinase (RTK) activation via their sulfated glycosaminoglycan (sGAG) side chains. However, their role on tumor cell behavior is controversial. Here, it is demonstrated that PGs are heavily expressed in lung adenocarcinoma (LUAD) patients in correlation with invasive phenotype and poor prognosis. A bioengineered human lung tumor model that recapitulates the increase of sGAGs in tumors in an organotypic matrix with independent control of stiffness, viscoelasticity, ligand density, and porosity, is developed. This model reveals that increased sulfation stimulates extensive proliferation, epithelial‐mesenchymal transition (EMT), and stemness in cancer cells. The focal adhesion kinase (FAK)‐phosphatidylinositol 3‐kinase (PI3K) signaling axis is identified as a mediator of sulfation‐induced molecular changes in cells upon activation of a distinct set of RTKs within tumor‐mimetic hydrogels. The study shows that the transcriptomic landscape of tumor cells in response to increased sulfation resembles native PG‐rich patient tumors by employing integrative omics and network modeling approaches. [ABSTRACT FROM AUTHOR]

Osimertinib, a third-generation inhibitor of epidermal growth factor receptor (EGFR) tyrosine kinase, exhibits remarkable efficacy in prolonging the survival of patients with non-small cell lung cancer (NSCLC) carrying EGFR mutations, surpassing the efficacy of first- and second-generation EGFR tyrosine kinases. Nevertheless, the emergence of osimertinib resistance is inevitable, necessitating an investigation into the underlying mechanisms. Increasing evidence has revealed that non-coding RNAs (ncRNAs), including microRNAs, long ncRNAs, and circular RNAs, play a significant role in the development and progression of lung cancer. These ncRNAs regulate essential signaling pathways, offering a novel avenue for understanding the fundamental mechanisms of osimertinib resistance. Recent studies have reported the significant impact of ncRNAs on osimertinib resistance, achieved through various mechanisms that modulate treatment sensitivity. We provide a concise overview of the functions and underlying mechanisms of extensively researched ncRNAs in the development of osimertinib resistance and emphasize their potential clinical application in EGFR-mutated NSCLC resistant to osimertinib. Finally, we discuss the obstacles that must be addressed to effectively translate ncRNA-based approaches into clinical practice. [ABSTRACT FROM AUTHOR]

AbstractRosmarinic acid (RosA), a hydrophilic phenolic compound found in various plants, has several biological effects such as anti-inflammatory and anti-apoptosis activities. However, its potential impact on chronic obstructive pulmonary disease (COPD) and its underlying mechanism has not been investigated. In this study, we explored the potential therapeutic effects and mechanism of RosA on COPD airway inflammation and alveolar epithelial apoptosis in vivo and in vitro. Our data suggested that RosA may be a therapeutic candidate for COPD with low toxicity. The corresponding mechanism lies in its anti-inflammatory effect on macrophage and bronchial epithelial cells, as well as protective effect on lung epithelial apoptosis via the jointly cross-target spleen tyrosine kinase (Syk). [ABSTRACT FROM AUTHOR]

As one of the factors regulating tumour angiogenesis, angiopoietin-4 (ANGPT4), which plays an important role in promoting tumour proliferation, survival, expansion and angiogenesis, is highly expressed in some tumours, such as lung adenocarcinoma, glioblastoma and ovarian cancer. This may be related to the fact that ANGPT4 affects the blood vessels and lymphatic system of the tumour. Specifically, ANGPT4 could play an effective role in promoting cancer by affecting the tyrosine kinase receptor TIE2, ERK1/2 and PI3K/AKT signalling pathways. Therefore, ANGPT4 may be an important biomarker for the occurrence and development of cancer and poor prognosis. In addition, the inhibition of ANGPT4 may be a useful cancer treatment. This paper reviews the latest preclinical research on ANGPT4, emphasizes its role in tumourigenesis and broadens our understanding of the carcinogenic function of ANGPT4 and the development of ANGPT4 inhibitors. This is the latest version of the revised version of the previous article. [ABSTRACT FROM AUTHOR]

ABSTRACT A general strategy was developed for the synthesis of new fully decorated 1,2,3‐triazoles (4a–4m and 5a–5g) containing quinazolines from 1‐(4‐nitrophenyl)‐2‐(quinazolin‐8‐ylsulfonyl) ethan‐1‐one and several azides using Ramachary organocatalytic azide‐ketone cycloaddition method. This reaction is reported for the synthesis of fully substituted sulfonyl‐1,2,3‐triazolyl quinazolins at a temperature of 100°C and the yields of the products produced are satisfactory to excellent. In vitro anticancer activity of all these derivatives demonstrated that six compounds, 4d, 4f, 4i, 4j, 5d, and 5e, were effective against two human breast cancer cell lines, MCF‐7 and MDA‐MB‐231. Compounds 4f, 4j, and 5d had more action against both cell lines than Erlotinib. Later, the findings of inhibitory assays of potent compounds 4d, 4f, 4i, 4j, 5d, and 5e against the tyrosine kinase EGFR revealed that compound 5d proved more potent than the reference erlotinib, while 4f and 4j had comparable efficacy. In silico molecular docking studies were also performed on six strong medicines to identify interactions with the EGFR receptor, and the energy estimations were shown to be comparable with the observed IC50 values. Ultimately, using SWISS/ADME and pkCSM, the in silico pharmacokinetic profile of potent compounds 4d, 4f, 4i, 4j, 5d, and 5e was predicted. All of the compounds precisely followed the principles established by Lipinski, Veber, Egan, and Muegge. [ABSTRACT FROM AUTHOR]

Lung cancer remains a formidable health challenge due to its high mortality and morbidity rates. Non-small cell lung cancer (NSCLC) constitutes approximately 85% of all lung cancer cases, with small cell lung cancer (SCLC) accounting for the remainder. Both NSCLC and SCLC cells express receptor tyrosine kinases, which may be overexpressed or mutated in lung cancer, leading to increased activation. The c-Met receptor tyrosine kinase, crucial for cell transformation and tumor growth, invasion, and metastasis, became the focus of our study. We used an E1B55KD-deleted, replication-selective oncolytic adenovirus (Ad.What), driven by the c-Met promoter, targeting lung cancer cells with c-Met overexpression, thus sparing normal cells. Previous studies have shown the enhanced antitumor efficacy of oncolytic adenoviruses when combined with chemotherapeutic agents. We explored combining rapamycin, a selective mTOR inhibitor with promising clinical trial outcomes for various cancers, with Ad.What. This combination increased infectivity by augmenting the expression of coxsackievirus and adenovirus receptors and αV integrin on cancer cells and induced autophagy. Our findings suggest that combining a c-Met promoter-driven oncolytic adenovirus with rapamycin could be an effective lung cancer treatment strategy, offering a targeted approach to exploit lung cancer cells' vulnerabilities, potentially marking a significant advancement in managing this deadly disease. [ABSTRACT FROM AUTHOR]

Simple Summary: Protein tyrosine kinase 7 (PTK7) is a conserved catalytically defective transmembrane receptor that is a potential therapeutic target in triple-negative breast cancer, ovarian cancer, and non-small cell lung cancer. The aim of this retrospective study was to determine if PTK7 expression levels in breast tumours were associated with the clinical outcome of breast cancer patients. We determined PTK7 expression in patient tumours using immunohistochemistry and assessed PTK7 mRNA expression in the METABRIC and TCGA cohorts. We determined that PTK7 expression was not clearly linked with survival in unselected patient cohorts; however, it was associated with breast cancer-specific survival of patients with poor and moderate Nottingham Prognostic Index values. Protein tyrosine kinase 7 (PTK7), originally known as colon carcinoma kinase (CCK4), is an evolutionary conserved, catalytically defective transmembrane receptor involved in Wnt signalling. PTK7 has been identified as a potential therapeutic target, and a PTK7 antibody drug conjugate (PF-06647020; cofetuzumab pelidotin) has been investigated in phase I clinical trials for triple-negative breast cancer, ovarian cancer, and non-small cell lung cancer. PTK7 protein expression was evaluated in 1136 early-stage invasive breast tumours by immunohistochemistry. In addition, PTK7 mRNA expression in the METABRIC (n = 1980) and the TCGA breast cancer cohorts (n = 1082) was evaluated. Associations between PTK7 expression and clinicopathological criteria and patient outcome were determined. No association between PTK7 protein expression and breast cancer-specific survival was observed; however, PTK7 mRNA expression in the METABRIC cohort was associated with breast cancer-specific survival (p < 0.001). PTK7 protein and mRNA expression were associated with breast cancer-specific survival of patients with a poor prognostic Nottingham Prognostic Index (NPI) and a moderate prognostic NPI, respectively. Taken together, these data indicate that PTK7 expression is associated with patient outcome in subgroups of breast cancer patients. [ABSTRACT FROM AUTHOR]

Background: Doxorubicin is an important anticancer drug, however, elicits dose-dependently cardiomyopathy. Given its mode of action, i.e. topoisomerase inhibition and DNA damage, we investigated genetic events associated with cardiomyopathy and searched for mechanism-based possibilities to alleviate cardiotoxicity. We treated rats at clinically relevant doses of doxorubicin. Histopathology and transmission electron microscopy (TEM) defined cardiac lesions, and transcriptomics unveiled cardiomyopathy-associated gene regulations. Genomic-footprints revealed critical components of Abl1-p53-signaling, and EMSA-assays evidenced Abl1 DNA-binding activity. Gene reporter assays confirmed Abl1 activity on p53-targets while immunohistochemistry/immunofluorescence microscopy demonstrated Abl1, p53&p73 signaling. Results: Doxorubicin treatment caused dose-dependently toxic cardiomyopathy, and TEM evidenced damaged mitochondria and myofibrillar disarray. Surviving cardiomyocytes repressed Parkin-1 and Bnip3-mediated mitophagy, stimulated dynamin-1-like dependent mitochondrial fission and induced anti-apoptotic Bag1 signaling. Thus, we observed induced mitochondrial biogenesis. Transcriptomics discovered heterogeneity in cellular responses with minimal overlap between treatments, and the data are highly suggestive for distinct cardiomyocyte (sub)populations which differed in their resilience and reparative capacity. Genome-wide footprints revealed Abl1 and p53 enriched binding sites in doxorubicin-regulated genes, and we confirmed Abl1 DNA-binding activity in EMSA-assays. Extraordinarily, Abl1 signaling differed in the heart with highly significant regulations of Abl1, p53 and p73 in atrial cardiomyocytes. Conversely, in ventricular cardiomyocytes, Abl1 solely-modulated p53-signaling that was BAX transcription-independent. Gene reporter assays established Abl1 cofactor activity for the p53-reporter PG13-luc, and ectopic Abl1 expression stimulated p53-mediated apoptosis. Conclusions: The tyrosine kinase Abl1 is of critical importance in doxorubicin induced cardiomyopathy, and we propose its inhibition as means to diminish risk of cardiotoxicity. [ABSTRACT FROM AUTHOR]

Background: Currently, the prognosis of advanced intrahepatic cholangiocarcinoma (ICC) is poor, and the current treatmentmethods are not effective. Objective: The aim of this study was to evaluate the anticancer efficacy of chemotherapy combined with PD-1 inhibitors and tyrosine kinase inhibitors (TKIs) in patients with ICC. Methods: We retrospectively screened patients with advanced intrahepatic cholangiocarcinoma (ICC) who received chemotherapy combined with lenvatinib and PD-1. We evaluated overall survival (OS), progression-free survival (PFS), the objective response rate (ORR), the disease control rate (DCR), the tumor shrinkage rate, and safety. Results: We enrolled 95 patients with ICC and divided them into three groups with a median follow-up duration of 15.1 months. The chemotherapy group (chemo-regimen group), chemotherapy combined with immune checkpoint inhibitors (dual-regimen group), and chemotherapy combined with lenvatinib (triple-regimen group) had median OS times of 13.1 months, 20.8 months, and 39.6 months, respectively. Notably, the triple-regimen group had a significantly longer OS than did the chemo-regimen and dual-regimen groups. The chemoregimen group, dual-regimen group, and triple-regimen group reported median PFS durations of 4.8 months, 11.9 months, and 23.4 months, respectively. Both combination groups exhibited significantly longer PFS than the chemotherapyonly group (P<0.05). The ORRs of the chemo-regimen, dual-regimen, and tripleregimen groups were 18.2%, 55.5%, and 54.7%, respectively. The DCRs were 72.7%, 90%, and 96.2%, respectively, indicating significantly better outcomes in the combination therapy groups. Conclusion: The combination of chemotherapy with PD-1 inhibitors and lenvatinib demonstrates considerable efficacy and tolerability as a treatment strategy for patients with advanced ICC. [ABSTRACT FROM AUTHOR]

Dysregulation of master regulator c-MYC (MYC) plays a central role in hepatocellular carcinoma (HCC) and other cancers but remains an elusive target for therapeutic intervention. MYC expression is epigenetically modulated within naturally occurring DNA loop structures, Insulated Genomic Domains (IGDs). We present a therapeutic approach using an epigenomic controller (EC), a programmable epigenomic mRNA medicine, to precisely modify MYC IGD sub-elements, leading to methylation of MYC regulatory elements and durable downregulation of MYC mRNA transcription. Significant antitumor activity is observed in preclinical models of HCC treated with the MYC-targeted EC, as monotherapy or in combination with tyrosine kinase or immune checkpoint inhibitors. These findings pave the way for clinical development of MYC-targeting epigenomic controllers in HCC patients and provide a framework for programmable epigenomic mRNA therapeutics for cancer and other diseases. Development of targeted MYC inhibitors for cancer therapy remains challenging. Here, the authors design an mRNA medicine which downregulates MYC gene transcription via epigenetic modification of MYC regulatory elements, showing significant antitumor activity in preclinical models of hepatocellular carcinoma. [ABSTRACT FROM AUTHOR]

We identified a novel mouse plasmacytoid dendritic cell (pDC) lineage derived from the common lymphoid progenitors (CLPs) that is dependent on expression of Bcl11a. These CLPderived pDCs, which we refer to as ‘B-pDCs’, have a unique gene expression profile that includes hallmark B cell genes, normally not expressed in conventional pDCs. Despite expressing most classical pDC markers such as SIGLEC-H and PDCA1, B-pDCs lack IFN-α secretion, exhibiting a distinct inflammatory profile. Functionally, B-pDCs induce T cell proliferation more robustly than canonical pDCs following Toll-like receptor 9 (TLR9) engagement. B-pDCs, along with another homogeneous subpopulation of myeloid-derived pDCs, display elevated levels of the cell surface receptor tyrosine kinase AXL, mirroring human AXL+ transitional DCs in function and transcriptional profile. Murine B-pDCs therefore represent a phenotypically and functionally distinct CLP-derived DC lineage specialized in T cell activation and previously not described in mice. [ABSTRACT FROM AUTHOR]

MET is a receptor tyrosine kinase (RTK) responsible for initiating signaling pathways involved in development and wound repair. MET activation relies on ligand binding to the extracellular receptor, which prompts dimerization, intracellular phosphorylation, and recruitment of associated signaling proteins. Mutations, which are predominantly observed clinically in the intracellular juxtamembrane and kinase domains, can disrupt typical MET regulatory mechanisms. Understanding how juxtamembrane variants, such as exon 14 skipping (METΔEx14), and rare kinase domain mutations can increase signaling, often leading to cancer, remains a challenge. Here, we perform a parallel deep mutational scan (DMS) of the MET intracellular kinase domain in two fusion protein backgrounds: wild-type and METΔEx14. Our comparative approach has revealed a critical hydrophobic interaction between a juxtamembrane segment and the kinase ⍺C-helix, pointing to potential differences in regulatory mechanisms between MET and other RTKs. Additionally, we have uncovered a β5 motif that acts as a structural pivot for the kinase domain in MET and other TAM family of kinases. We also describe a number of previously unknown activating mutations, aiding the effort to annotate driver, passenger, and drug resistance mutations in the MET kinase domain. [ABSTRACT FROM AUTHOR]

PURPOSE: MDM2, a negative regulator of the TP53 tumor suppressor, is oncogenic when amplified. MDM2 amplification (MDM2amp) is mutually exclusive with TP53 mutation and is seen in 6% of patients with lung adenocarcinoma (LUAD), with significant enrichment in subsets with receptor tyrosine kinase (RTK) driver alterations. Recent studies have shown synergistic activity of MDM2 and MEK inhibition in patient-derived LUAD models with MDM2amp and RTK driver alterations. However, the combination of MDM2 and RTK inhibitors in LUAD has not been studied. METHODS: We evaluated the combination of MDM2 and RTK inhibition in patient-derived models of LUAD. RESULTS: In a RET-fusion LUAD patient-derived model with MDM2amp, MDM2 inhibition with either milademetan or AMG232 combined with selpercatinib resulted in long-term in vivo tumor control markedly superior to either agent alone. Similarly, in an EGFR-mutated model with MDM2amp, combining either milademetan or AMG232 with osimertinib resulted in long-term in vivo tumor control, which was strikingly superior to either agent alone. CONCLUSION: These preclinical in vivo data provide a rationale for further clinical development of this combinatorial targeted therapy approach. Preclinical in vivo data support personalized genomically informed combination targeted therapy in lung cancer. [ABSTRACT FROM AUTHOR]

Cipangopaludina chinensis is a common freshwater mollusk that is widely distributed worldwide, especially in China. In our research, 1,382 proteins and 1,039 phosphorylated proteins were identified from C. chinensis carcasses, and 690 differentially expressed proteins (DEPs) were quantified. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses revealed that the DEPs are involved in cellular processes, single-organism processes, metabolic processes, developmental processes, localization, and biological regulation. The phosphorylated proteins were found to be related to the Rap1 signaling pathway, Ras signaling pathway, calcium signaling pathway, and longevity-regulating pathways. Moreover, we also identified important regulatory enzymes, such as guanylate cyclase, tyrosine protein kinase, receptor protein tyrosine kinase, and glyoxylate reductase/hydroxypyruvate reductase. Notably, we found guanylate cyclase to be present in multiple signaling pathways, including the Rap1 signaling pathway, calcium signaling pathway, Ras signaling pathway, insulin secretion, longevity regulating pathway, glutamatergic synapse, circadian entrainment, and gap junction. This enzyme may play a crucial role in regulating molecular mechanisms in C. chinensis. In summary, proteomic and phosphoproteomic analyses of C. chinensis carcasses displayed significant differences among different geographical isolates, which helps enhance our understanding of food nutrition, signaling pathways, and metabolic mechanisms in C. chinensis. [ABSTRACT FROM AUTHOR]