Protein Kinase Research Articles

Unconventional Histopathological Features With KIAA1549::BRAF Fusion in Advanced Prostatic Adenocarcinoma.

This report delineates an intriguing example of advanced prostatic adenocarcinoma displaying distinctive histopathological characteristics associated with a KIAA1549::BRAF fusion, a genomic anomaly predominantly identified in central nervous system tumors. A 66-year-old man, presenting with acute renal failure, exhibited metastatic disease involving pelvic soft tissue, bladder, liver, and bone. Histological examination revealed a markedly unconventional morphology within the prostate, characterized by infiltrative tumor sheets exhibiting abundant vacuolated cytoplasm, hyperchromatic nuclei, and irregular nuclear membranes, distinct from typical prostatic adenocarcinoma. Immunophenotyping confirmed NKX3.1 positivity and GATA3 negativity. Molecular analysis revealed the rare KIAA1549::BRAF fusion alongside pertinent mutations in phosphatidylinositol 3-kinase, phosphatase and tensin homolog, and tumor protein 53 genes. Despite diverse therapeutic interventions targeting mitogen-activated protein kinase signaling and subsequent clinical trial enrollment, disease progression remained relentless, culminating in the patient's demise within 4 years of diagnosis. This report highlights the exceptional histopathological presentation associated with KIAA1549::BRAF fusion in prostatic adenocarcinoma, emphasizing the need for a deeper comprehension of its implications on disease behavior and therapeutic responsiveness in similar instances.

Abstract 4134426: Targeting the TRIM29-PERK signaling axis for treating viral myocarditis

Introduction: Viral myocarditis is an inflammatory disease of the myocardium that significantly contributes to sudden death in children and young adults. The COVID-19 pandemic underscores the critical need to understand the pathogenesis and develop effective treatment strategies for viral myocarditis. Methods: To investigate the role of the novel E3 ligase TRIM29 in viral myocarditis, we used wild-type and TRIM29 knockout mice infected with coxsackievirus B3 (CVB3) and encephalomyocarditis virus (EMCV), to induce the myocarditis. Additionally, wild-type mice infected with CVB3 were treated with the protein kinase RNA-like endoplasmic reticulum kinase (PERK) inhibitor GSK2656157 or a DMSO control to evaluate potential therapeutic interventions. Mice survival and heart function were monitored using transthoracic echocardiography, and hearts were harvested for histological and immunohistochemical analysis. Techniques including real-time PCR, western blotting, co-immunoprecipitation, enzyme-linked immunoassay, flow cytometry, over-expression, and knockdown were employed to elucidate the pathogenic mechanisms by which TRIM29 regulates the endoplasmic reticulum stress response after viral infection. Results: We found that TRIM29 was highly induced by cardiotropic viruses, including CVB3 and EMCV, and promoted PERK-mediated endoplasmic reticulum (ER) stress, apoptosis, and reactive oxygen species (ROS) responses, which restricted viral replication in cardiomyocytes in vitro . TRIM29 deficiency protected mice from viral myocarditis by enhancing cardiac antiviral functions and reducing PERK-mediated inflammation and immunosuppressive cells in vivo . Mechanistically, TRIM29 interacted with PERK to catalyze the SUMOylation of PERK, maintaining its stability and thereby promoting PERK-mediated signaling pathways. Furthermore, we demonstrated that the PERK inhibitor GSK2656157 mitigated viral myocarditis by disrupting the TRIM29-PERK interaction, which improved cardiac function, enhanced cardiac antiviral responses, and reduced inflammation and immunosuppressive cells in vivo . Conclusions: Our findings provide the first evidence that the TRIM29-PERK signaling axis can be targeted for the treatment of viral myocarditis. This suggests that targeting the TRIM29-PERK axis could effectively mitigate viral myocarditis and associated cardiovascular diseases.

Role and Interplay of Different Signaling Pathways Involved in Sciatic Nerve Regeneration.

Regeneration of the sciatic nerve is a sophisticated process that involves the interplay of several signaling pathways that orchestrate the cellular responses critical to regeneration. Among the key pathways are the mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3K)/AKT, cyclic adenosine monophosphate (cAMP), and Janus kinase/signal transducers and transcription activators (JAK/STAT) pathways. In particular, the cAMP pathway modulates neuronal survival and axonal regrowth. It influences various cellular behaviors and gene expression that are essential for nerve regeneration. MAPK is indispensable for Schwann cell differentiation and myelination, whereas PI3K/AKT is integral to the transcription, translation, and cell survival processes that are vital for nerve regeneration. Furthermore, GTP-binding proteins, including those of the Ras homolog gene family (Rho), regulate neural cell adhesion, migration, and survival. Notch signaling also appears to be effective in the early stages of nerve regeneration and in preventing skeletal muscle fibrosis after injury. Understanding the intricate mechanisms and interactions of these pathways is vital for the development of effective therapeutic strategies for sciatic nerve injuries. This review underscores the need for further research to fill existing knowledge gaps and improve therapeutic outcomes.

M6A-modified circCacna1c regulates necroptosis and ischemic myocardial injury by inhibiting Hnrnpf entry into the nucleus.

Circular RNAs (circRNAs) are differentially expressed in various cardiovascular diseases, including myocardial infarction (MI) injury. However, their functional role in necroptosis-induced loss of cardiomyocytes remains unclear. We identified a cardiac necroptosis-associated circRNA transcribed from the Cacna1c gene (circCacna1c) to investigate the involvement of circRNAs in cardiomyocyte necroptosis. To investigate the role of circCacna1c during oxidative stress, H9c2 cells and neonatal rat cardiomyocytes were treated with hydrogen peroxide (H2O2) to induce reactive oxygen species (ROS)-induced cardiomyocyte death. The N6-methyladenosine (m6A) modification level of circCacna1c was determined by methylated RNA immunoprecipitation quantitative polymerase chain reaction (MeRIP-qPCR) analysis. Additionally, an RNA pull-down assay was performed to identify interacting proteins of circCacna1c in cardiomyocytes, and the regulatory role of circCacna1c in target protein expression was tested using a western blotting assay. Furthermore, theMI mouse model was constructed to analyze the effect of circCacna1c on heart function and cardiomyocyte necroptosis. The expression of circCacna1c was found to be reduced in cardiomyocytes exposed to oxidative stress and in mouse hearts injured by MI. Overexpression of circCacna1c inhibited necroptosis of cardiomyocytes induced by hydrogen peroxide and MI injury, resulting in a significant reduction in myocardial infarction size and improved cardiac function. Mechanistically, circCacna1c directly interacts with heterogeneous nuclear ribonucleoprotein F (Hnrnpf) in the cytoplasm, preventing its nuclear translocation and leading to reduced Hnrnpf levels within the nucleus. This subsequently suppresses Hnrnpf-dependent receptor-interacting protein kinase 1 (RIPK1) expression. Furthermore, fat mass and obesity-associated protein (FTO) mediates demethylation of m6A modification on circCacna1c during necrosis and facilitates degradation of circCacna1c. Our study demonstrates that circCacna1c can improve cardiac function following MI-induced heart injury by inhibiting the Hnrnpf/RIPK1-mediated cardiomyocyte necroptosis. Therefore, the FTO/circCacna1c/Hnrnpf/RIPK1 axis holds great potential as an effective target for attenuating cardiac injury caused by necroptosis in ischemic heart disease.

Antimicrobial Peptide Pro10-1D Exhibits Anti-Allergic Activity: A Promising Therapeutic Candidate

Although antimicrobial peptides (AMPs) exhibit a range of biological functions, reports on AMPs with therapeutic effects in allergic disorders are limited. In this study, we investigated the anti-allergic effects of Pro10-1D, a 10-meric AMP derived from insect defensin protaetiamycine. Our findings demonstrate that Pro10-1D effectively inhibits antigen-induced degranulation of mast cells (MCs) with IC50 values of approximately 11.6 μM for RBL-2H3 cells and 2.7 μM for bone marrow-derived MCs. Furthermore, Pro10-1D suppressed the secretion of cytokines with IC50 values of approximately 2.8 μM for IL-4 and approximately 8.6 μM for TNF-α. Mechanistically, Pro10-1D inhibited the Syk-LAT-PLCγ1 signaling pathway in MCs and decreased the activation of mitogen-activated protein kinases (MAPKs). Pro10-1D demonstrated a dose-dependent reduction in IgE-mediated passive cutaneous anaphylaxis in mice with an ED50 value of approximately 7.6 mg/kg. Further investigation revealed that Pro10-1D significantly reduced the activity of key kinases Fyn and Lyn, which are critical in the initial phase of the FcεRI-mediated signaling pathway, with IC50 values of approximately 22.6 μM for Fyn and approximately 1.5 μM for Lyn. Collectively, these findings suggest that Pro10-1D represents a novel therapeutic candidate for the treatment of IgE-mediated allergic disorders by targeting the Lyn/Fyn Src family kinases in MCs.

Glucose Upregulates ChREBP via Phosphorylation of AKT and AMPK to Modulate MALT1 and WISP1 Expression.

Glucose can activate the carbohydrate response element binding protein (ChREBP) transcription factor to control gene expressions in the metabolic pathways. The way of ChREBP involvement in human prostate cancer development remains undetermined. This study examined the interactions between prostate fibroblasts and cancer cells under the influences of ChREBP. Results showed that high glucose (30 mM) increased the phosphorylation of AKT at S473 and AMP-activated protein kinase (AMPK) at S485 in human prostate fibroblast (HPrF) cells and prostate cancer PC-3 cells. High glucose enhanced the expression of ChREBP, which increased the expressions of fibronectin, alpha-smooth muscle actin (α-SMA), and WNT1 inducible signaling pathway protein 1 (WISP1), magnifying the cell growth and contraction in HPrF cells in vitro. The cell proliferation, invasion, and tumor growth in prostate cancer PC-3 cells were enhanced by inducing the expressions of ChREBP, mucosa-associated lymphoid tissue 1 (MALT1), and epithelial-mesenchymal transition markers with high glucose treatment. Moreover, ectopic ChREBP overexpression induced NF-κB signaling activities via upregulating MALT1 expression in PC-3 cells. Our findings illustrated that ChREBP is an oncogene in the human prostate. High glucose condition induces a glucose/ChREBP/MALT1/NF-κB axis which links the glucose metabolism to the NF-κB activation in prostate cancer cells, and a glucose/ChREBP/WISP1 axis mediating autocrine and paracrine signaling between fibroblasts and cancer cells to promote cell migration, contraction, growth, and invasion of the human prostate.

Role of Mast Cell in Hyperalgesic Priming and the Preventive Effect of Electroacupuncture on the Transition from Acute to Chronic Pain

Objective: Injury can lead to long-term changes that increase the sensitivity of afferent nerve endings to subsequent stimulation and pain can transition from acute to chronic. This phenomenon is known as hyperalgesic priming (HP). This study aimed to understand the mechanisms underlying the effect of Electroacupuncture (EA) on HP and optimize acupoint selection for EA to prevent pain transition. Methods: A rat HP model was established using sequential intraplantar injections of carrageenan (Cg) and prostaglandin E2 (PGE2). The pain thresholds were measured using von Frey filaments. EA on bilateral Zusanli(ST36) and Kunlun(BL60) was used to prevent pain transition. The number of mast cells in the ipsilateral hindpaw skin was determined using toluidine blue or fluorescence-labeled avidin staining. The protein expression levels of protein kinase C (PKCε) in the lumbar dorsal root ganglions (DRGs) were detected by western blotting 24 h after PGE2 injection. Serial pharmacological experiments were conducted to evaluate the relationship between mast cells and pain transition. Finally, EA on the bilateral ST36 and Chongyang(ST42) or a novel combination (ST36 and ST42 on the ipsilateral side, and ST36 and BL60 on the contralateral side) was used to prevent pain transition. Results: Although EA applied to ST36 and BL60 alleviated acute pain induced by Cg injection, it failed to prevent the pain transition caused by PGE2 injection. Mast cell accumulation in the ipsilateral hind paw was observed 7 days after Cg injection. Furthermore, mast cell degranulation may be responsible for PKCε activation in the DRG, a marker of pain transition. EA significantly decreased the number of mast cells in the skin of the ipsilateral hind paw when applied at ST36 and ST42, but not when applied at ST36 and BL60. Furthermore, EA employed to ST36 and ST42 significantly reversed long-term hyperalgesia induced by PGE2 injection, even when administered before injection. However, EA did not alleviate acute pain caused by Cg injection. By using a novel acupoint combination, EA simultaneously alleviated acute pain and prevented pain transition. Conclusions: Our study suggests that mast cells play a critical role in both HP and the transition from acute to chronic pain, whereas EA can prevent pain transition by decreasing the number of mast cells in the local tissue. Graphical abstract: http://links.lww.com/AHM/A144.

Astaxanthin has a beneficial influence on pain-related symptoms and opioid-induced hyperalgesia in mice with diabetic neuropathy-evidence from behavioral studies.

The treatment of painful diabetic neuropathy is still a clinical problem. The aim of this study was to determine whether astaxanthin, a substance that inhibits mitogen-activated protein kinases, activates nuclear factor erythroid 2-related factor 2 and influences N-methyl-D-aspartate receptor, affects nociceptive transmission in mice with diabetic neuropathy. The studies were performed on streptozotocin-induced mouse diabetic neuropathic pain model. Single intrathecal and intraperitoneal administrations of astaxanthin at various doses were conducted in both males and females. Additionally, repeated twice-daily treatment with astaxanthin (25mg/kg) and morphine (30mg/kg) were performed. Hypersensitivity was evaluated with von Frey and cold plate tests. This behavioral study provides the first evidence that in a mouse model of diabetic neuropathy, single injections of astaxanthin similarly reduce tactile and thermal hypersensitivity in both male and female mice, regardless of the route of administration. Moreover, repeated administration of astaxanthin slightly delays the development of morphine tolerance and significantly suppresses the occurrence of opioid-induced hyperalgesia, although it does not affect blood glucose levels, body weight, or motor coordination. Surprisingly, astaxanthin administered repeatedly produces a better analgesic effect when administered alone than in combination with morphine, and its potency becomes even more pronounced over time. These behavioral results provide a basis for further evaluation of the potential use of astaxanthin in the clinical treatment of diabetic neuropathy and suggest that the multidirectional action of this substance may have positive effects on relieving neuropathic pain in diabetes.

Divergent roles of RIPK3 and MLKL in high-fat diet-induced obesity and MAFLD in mice.

Cell death frequently occurs in the pathogenesis of obesity and metabolic dysfunction-associated fatty liver disease (MAFLD). However, the exact contribution of core cell death machinery to disease manifestations remains ill-defined. Here, we show via the direct comparison of mice genetically deficient in the essential necroptotic regulators, receptor-interacting protein kinase-3 (RIPK3) and mixed lineage kinase domain-like (MLKL), as well as mice lacking apoptotic caspase-8 in myeloid cells combined with RIPK3 loss, that RIPK3/caspase-8 signaling regulates macrophage inflammatory responses and drives adipose tissue inflammation and MAFLD upon high-fat diet feeding. In contrast, MLKL, divergent to RIPK3, contributes to both obesity and MAFLD in a manner largely independent of inflammation. We also uncover that MLKL regulates the expression of molecules involved in lipid uptake, transport, and metabolism, and congruent with this, we discover a shift in the hepatic lipidome upon MLKL deletion. Collectively, these findings highlight MLKL as an attractive therapeutic target to combat the growing obesity pandemic and metabolic disease.

Mining of Candidate Genes Associated with Leaf Shape Traits in Grapes

As the most important organ for photosynthesis, leaves provide the main energy source for plant growth. Leaf traits affect light energy utilization and, thus, plant development and biomass. Given the high morphological variability of leaves between and within grape genotypes, phenotypic analysis is challenging. This study first evaluated leaf shape trait parameters using a specific leaf profile and area analyzer, along with genome-wide association study (GWAS) analyses, to identify additional candidate genes related to grape leaf shape traits. In the two-year analysis, 89 single-nucleotide polymorphisms (SNPs) were found to be significantly associated with leaf shape traits. These SNP loci were distributed on 18 chromosomes, of which chromosome 15 had the most relevant SNPs. We found that leaf shape-associated genes included mainly plant hormone-, ubiquitin ligase-, serine/threonine protein kinase-, transcription factor-, and cell wall metabolism-related genes. By analyzing the expression of these candidate genes on the chip, we found that they exhibited diverse expression levels in leaves at different developmental stages (young, mature, and senescent). This suggests that these genes could be considered candidates for grape leaf improvement.

The long noncoding RNA ELFN1-AS1 promotes gastric cancer growth and metastasis by interacting with TAOK1 to inhibit the Hippo signaling pathway.

Gastric cancer (GC) is a common digestive malignancy that causes numerous cancer-related deaths. Long noncoding RNAs (lncRNAs) play a crucial role in the development of various tumors, including GC. In this study, we revealed that ELFN1-AS1, a lncRNA with aberrantly high expression, contributes to the proliferation and metastasis of GC. Mechanically, ELFN1-AS1 plays an oncogenic role by binding to the protein kinase domain of thousand and one amino acid protein kinase (TAOK1), a tumor suppressor in GC, and disrupting the TAOK1-STK3 interaction, leading to decreased STK3 phosphorylation. This decrease is accompanied by attenuation of the Hippo kinase cascade, resulting in reduced YAP1 phosphorylation, a crucial effector of the Hippo signaling pathway. Subsequently, the reduced YAP1 phosphorylation promotes its nuclear translocation, thereby enhancing the expression of MYC, a downstream target of the pathway and well-known oncogene. Taken together, the ELFN1-AS1/TAOK1/STK3/YAP1 axis may promote GC progression and is a promising target for GC treatment.

Expression and clinical significance of PIKFYVE gene in hepatocellular carcinoma analyzed based on TCGA database and experimental validation

Objective: To investigate the expression and clinical significance of human FYVE finger-containing phosphoinositide kinase (PIKFYVE) in hepatocellular carcinoma (HCC) on the basis of cancer genome atlas (The cancer genome atlas, TCGA) database analysis and clinical samples experimental validation. Methods: Based on the data information of 424 clinical samples (including 374 cases of HCC tissues and 50 cases of nontumorous liver tissues) in the TCGA database, Cox regression analysis and Kaplan-Meier method were used to analyse the relationship between the PIKFYVE mRNA expression and the clinical characteristics, prognosis for survival of HCC patients. The relationship between the PIKFYVE gene and immune cell infiltration was examined by correlation analysis between the PIKFYVE gene and 24 immune cells. In addition, we analysed the correlation between the mRNA expression of PIKFYVE gene and RAC-alpha serine/threonine-protein kinase (AKT1), phosphatase and tensin homolog (PTEN), protein kinase C, alpha (PRKCA), inositol polyphosphate-5-phosphatase (INPP5D), phosphoinositide-3-kinase regulatory subunit 1(PIK3R1), Inositol Polyphosphate 4-phosphatase Type II (INPP4B) and phospholipase-C4 gene (PLCB4) in HCC tissues. Meanwhile, paraffin sections of highly differentiated, moderately differentiated, poorly differentiated, and nontumorous liver tissue in the Department of Pathology of the First Affiliated Hospital of Xinjiang Medical University were collected, each of which was 30 cases, and the histopathological observation was carried out by HE staining, and the expression levels of PIKFYVE and Ki67 proteins were verified by immunohistochemistry in each clinical sample. Results: The expression level of PIKFYVE gene in HCC tumours was significantly higher than that in normal liver tissues (P=0.000 2, P<0.01), and the overall survival of patients in the low PIKFYVE expression group was significantly longer than that in the high expression group (HR=1.57, 95%CI: 1.10～2.25, P=0.014). The results of Univariate Cox regression analysis showed that there was an effect of TNM stage, pathological stage, tumour status and residual tumour on Overall survival (OS) (P<0.05), and the expression level of PIKFYVE had an effect on OS survival (P<0.05); the PIKFYVE prognostic risk model score ratio was HR=1.533 (1.077-2.181, P=0.018). Multivariate Cox regression analysis showed a PIKFYVE prognostic risk model score ratio HR=1.481 (0.886-2.476, P=0.134) and an area under the Receiver Operating Characteristic curve of 0.640, which was greater than 0.5, suggesting that the PIKFYVE prognostic risk model has a predictive value in survival prediction. Correlation analysis showed that the expression level of PIKFYVE was highly correlated with immune cell infiltration and TP53 (P<0.01). The immunohistochemistry staining results showed that the expression of PIKFYVE in HCC tissues was significantly higher than that of nontumorous tissues (P<0.05), and there was a negative correlation with the degree of differentiation. Conclusion: PIKFYVE, as an independent risk factor for HCC, is expected to be developed as a clinical diagnostic biomarker for HCC, which will provide a reference for new drugs for the treatment of HCC.

Avenanthramides Ameliorate Insulin Resistance by Modulating Gluconeogenesis and Glycogen Synthesis in HepG2 Cells.

Diabetes mellitus (DM) is a multifaceted metabolic condition, mainly defined by elevated blood glucose levels. A feature of type 2 DM includes insulin resistance (IR), which involves impairments within the insulin signaling pathways. Avenanthramides (AVNs) are phenolic alkaloids found in Avena sativa L. The major AVNs are AVN A, AVN B, and AVN C. They have been reported to offer benefits in preventing inflammation, cancer, and cardiovascular diseases. However, the effects of AVNs on the liver glucose metabolism pathways remain unknown. This study examined the effects and underlying mechanisms through which AVNs alleviate IR induced by free fatty acid (FFA) in HepG2 cells. The results indicated that FFA treatment significantly decreased glucose consumption by 34.54% compared to the control. However, treatments with AVN A, B, and C at 100 μM increased glucose uptake by 57.93%, 58.28%, and 53.10%, respectively, compared to FFA treatment alone. This effect occurs through the increased expression of glucose transporter 4. Furthermore, AVNs significantly enhanced the glycogen content. AVNs induced increased phosphorylation of insulin receptor substrate-1 (IRS-1), phosphatidylinositol-3-kinase (PI3K), and protein kinase B (Akt). AVNs treatment decreased the levels of phosphoenolpyruvate carboxykinase and glucose-6-phosphatase in HepG2 cells. This effect was attributed to AMP-activated protein kinase activation and inhibition of forkhead box protein O1. Collectively, these results suggest that AVNs regulate glucose metabolism by activating the IRS-1/PI3K/Akt pathway, which is related to glycogen synthesis, and by inhibiting key molecules that promote gluconeogenesis.

CSIG-30. EXPLORING THE ROLE OF CAPICUA (CIC) IN RESISTANCE MECHANISMS OF MEK INHIBITORS IN GLIOBLASTOMA

Abstract Glioblastoma multiforme (GBM) is the most common and fatal primary brain tumor, resistant to conventional therapy. Despite its heterogeneous nature, a key feature of GBM is aberrant kinase signaling. Specifically, hyperactivation of receptor tyrosine kinases (RTK) and their downstream tumorigenic effectors Ras/MEK/Extracellular Signal Regulated Kinase (ERK) pathway are attributed to the aggressive nature of GBM. Capicua (CIC) is a High Mobility Group (HMG) box transcriptional repressor that directly binds to DNA and counteracts the transcription of genes typically expressed only in response to RTK activation such as oncogenic transcription factors ETV1/4/5. Hyperactive MEK/ERK signaling in GBM causes CIC degradation. The mechanism of CIC’s repressor function is not well understood, however, transcriptional repressors (like CIC) typically form co-repressor complexes to repress target genes. We discovered a novel interaction between CIC and the transcriptional regulator yin yang 1 (YY1), a context-dependent transcription factor that can repress or activate gene expression. We uncovered that 90-kDa ribosome S6 protein kinase (p90RSK), a downstream effector of ERK and a known CIC regulator, mediates the CIC/YY1 interaction. Interestingly, we found re-activation of p90RSK following MEK/ERK inhibition implicating p90RSK in the resistance mechanism of RTK/MEK/ERK inhibition in GBM. We hypothesize that p90RSK inhibition will stabilize the CIC/YY1 co-repressor complex and re-sensitize towards MEK/ERK inhibition. Patient-derived glioma stem-like cells (GSCs) were treated with increasing doses of BI-D1870 or LJI308 (p90RSK inhibitors) with and without selumetinib (MEK inhibitor) and various functional assays were performed to examine the effect of p90RSK inhibition on the CIC/YY1 complex and sensitization to MEK/ERK inhibition. Treatment with BI-D1870 and LJI308 markedly reduced viability and mRNA expression of ETV1/4/5 indicating that p90RSK inhibition may mediate CIC /YY1 DNA binding ability. Importantly, inhibiting p90RSK sensitized GSCs to MEK/ERK inhibition. These results highlight the importance of downstream p90RSK inhibition in stabilizing CIC/YY1 repressor complex function and sensitizing cells to RTK/MEK/ERK inhibition.

Neutrophil and mononuclear leukocyte pathways and upstream regulators revealed by serum proteomics of adult and juvenile dermatomyositis.

Serum protein abundance was assessed in adult and juvenile dermatomyositis (DM and JDM) patients to determine differentially regulated proteins, altered pathways, and candidate disease activity biomarkers. Serum protein expression from 17 active adult DM and JDM patients each was compared to matched, healthy control subjects by a multiplex immunoassay. Pathway analysis and protein clustering of the differentially regulated proteins were examined to assess underlying mechanisms. Candidate disease activity biomarkers were identified by correlating protein expression with disease activity measures. Seventy-eight of 172 proteins were differentially expressed in the sera of DM and JDM patients compared to healthy controls. Forty-eight proteins were differentially expressed in DM, 32 proteins in JDM, and 14 proteins in both DM and JDM. Twelve additional differentially expressed proteins were identified after combining the DM and JDM cohorts. C-X-C motif chemokine ligand 10 (CXCL10) was the most strongly upregulated protein in both DM and JDM sera. Other highly upregulated proteins in DM included S100 calcium binding protein A12 (S100A12), CXCL9, and nicotinamide phosphoribosyltransferase (NAMPT), while highly upregulated proteins in JDM included matrix metallopeptidase 3 (MMP3), growth differentiation factor 15 (GDF15), and von Willebrand factor (vWF). Pathway analysis indicated that phosphoinositide 3-kinase (PI3K), p38 mitogen-activated protein kinase (MAPK), and toll-like receptor 7 (TLR7) signaling were activated in DM and JDM. Additional pathways specific to DM or JDM were identified. A protein cluster associated with neutrophils and mononuclear leukocytes and a cluster of interferon-associated proteins were observed in both DM and JDM. Twenty-two proteins in DM and 24 proteins in JDM sera correlated with global, muscle, and/or skin disease activity. Seven proteins correlated with disease activity measures in both DM and JDM sera. IL-1 receptor like 1 (IL1RL1) emerged as a candidate global disease activity biomarker in DM and JDM. Coordinate analysis of protein expression in DM and JDM patient sera by a multiplex immunoassay validated previous gene expression studies and identified novel dysregulated proteins, altered signaling pathways, and candidate disease activity biomarkers. These findings may further inform the assessment of DM and JDM patients and aid in the identification of potential therapeutic targets.

Utility of patient-derived xenografts to evaluate drug sensitivity and select optimal treatments for individual non-small-cell lung cancer patients.

Patient-derived xenograft (PDX) is currently considered a preferred preclinical model to evaluate drug sensitivity, explore drug resistance mechanisms, and select individualized treatment regimens. Histopathological examination, immunohistochemistry and whole-exome sequencing confirmed similarity between our PDX tumors and primary tumors in terms of morphology and genetic characteristics. The drug reactivity of the PDX tumor was validated in vivo. The mechanisms of acquired resistance to Osimertinib PDX tumors were investigated by WES and WB. We successfully established 13 NSCLC-PDXs derived from 62 patients, including eight adenocarcinomas, four squamous-cell carcinoma, and one large-cell neuroendocrine carcinoma. Histological subtype and clinical stage were significant factors affecting the successful PDXs establishment. The treatment responses to conventional chemotherapy in PDXs were entirely consistent with that of their corresponding patients. According to the genetic status of tumors, more appropriate targeted agents were selected in PDXs for their corresponding patients as alternative treatment options. In addition, a PDX model with acquired resistance to osimertinib was induced, and the overactivation of RAS mitogen-activated protein kinase (MAPK)-extracellular signal-regulated kinase (ERK) signaling pathway caused by the dual-specificity phosphatase 6 (DUSP6) M62I mutation was found to play a key role in the development of osimertinib resistance. Trametinib, a specific inhibitor of the MAPK-ERK pathway significantly slowed down the tumor growth in osimertinib-resistant PDX models, providing an alternative treatment in patients after osimertinib failure.

DNAR-16. TARGETING APOBEC CYTIDINE DEAMINASES TO ENHANCE RADIOSENSITIVITY IN GLIOMA

Abstract Glioma is the most common and aggressive primary adult brain tumor. Radiation therapy (RT) is a critical part of glioma treatment, but radioresistance severely limits its therapeutic efficacy. To better understand how radioresistance develops, we analyzed 212 paired primary and recurrent gliomas from the Glioma Longitudinal Analysis Consortium. We found a significant increase in mutational signatures associated with the activity of the Apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) family of cytidine deaminases in RT-treated recurrent gliomas (p&amp;lt;0.001, paired Wilcoxon signed-rank test). Further analysis of post-treatment mutations showed that RT increased APOBEC mutational signatures independent of other factors (p&amp;lt;0.001, multivariable log-linear regression), suggesting that APOBEC proteins become activated by radiation. However, their role in promoting tumor evolution and treatment resistance is currently unknown. Further analyses of extended signature contexts and expression data nominated APOBEC3B (A3B) and APOBEC3G (A3G) as candidates for creating these mutational signatures. To investigate their roles in RT response, we knocked out (KO) A3B and A3G individually in patient-derived glioma cell lines before RT. Compared to Control cells, irradiated A3B-KO and A3G-KO cells acquired significantly fewer APOBEC mutational signatures by whole genome sequencing (p&amp;lt;0.05, Kruskal–Wallis test) and exhibited enhanced radiosensitivity in vitro. Interestingly, the KO cells displayed a significant decrease in mutational signatures associated with Non-Homologous End-Joining (NHEJ), one of the main pathways for repairing the RT-induced DNA damage (p&amp;lt;0.05, Kruskal–Wallis test), suggesting that APOBEC deletion impaired DNA repair. This was further supported by persistent gH2AX DNA damage foci 48h post-RT and reduced autophosphorylation of DNA-dependent protein kinase (DNA-PK), a vital component of the NHEJ pathway, particularly in A3G KO cells. Overall, our results identify APOBEC proteins as potential targets for radiosensitizing gliomas and highlight the clinical utility of using mutational signatures to predict and monitor response to treatment.

CSIG-16. A CX43-WNK1-C-MYC SIGNALING AXIS DRIVES GLIOBLASTOMA CANCER STEM CELL SURVIVAL

Abstract Tumors represent a highly dynamic system that relies on coordinated signaling to drive growth and adapt to selective pressures, including those induced by anti-cancer therapies. Given the need for tumors to engage in rapid and coordinated cell-cell communication, mechanisms such as gap junction intracellular communication (GJIC) should be essential. However, the connexin proteins that make up gap junctions have traditionally been considered tumor suppressors based on the frequent loss of GJIC and lower connexin expression in tumor cells compared to non-neoplastic tissue. For this reason, connexin 43 (Cx43) has been proposed to suppress the growth of glioblastoma (GBM), the most common primary malignant brain tumor. However, recent data suggest that this tumor-suppressive effect is context dependent and that connexins can also function in a tumor-promoting role. Using next-generation sequencing techniques, we found Cx43 expressed at high levels in a panel of GBM patient-derived xenograft (PDX) CSCs and that these models rely on Cx43 for their survival and self-renewal. Mechanistically, depletion of Cx43 led to a dramatic loss of c-MYC expression through reduced phosphorylation of its upstream mediator WNK lysine-deficient protein kinase 1 (WNK1). Depletion of WNK1 phenocopied knockdown of Cx43 and reduced MYC protein and mRNA as well as tumor growth in vivo. Together, this work defines a novel signaling axis downstream of gap junction protein expression that promotes tumor growth and cancer stem cell phenotypes in GBM. Due to the difficulty in targeting both Cx43 and MYC, the identification of intermediate targetable signaling nodes may lead to improved therapies for patients with GBM.

INNV-12. CUTANEOUS TOXICITIES OF MITOGEN-ACTIVATED PROTEIN KINASE INHIBITORS IN CHILDREN AND YOUNG ADULTS WITH NEUROFIBROMATOSIS-1

Abstract BACKGROUND Neurofibromatosis type 1 (NF1) is an autosomal dominant disorder which commonly causes neoplasms leading to disfigurement or dysfunction. Mitogen-activated protein kinase inhibitors (MEKi) are generally well-tolerated treatments which target neural tumor progression in patients with NF1. Cutaneous adverse events (CAEs) are common and may hinder patients’ abilities to remain on treatment, particularly in children. We aim to characterize CAEs secondary to MEKi treatment in pediatric and young adult patients with NF1. METHODS We reviewed institutional medical records of patients under 30 years with a diagnosis of “NF1,” “NF2,” or “other neurofibromatoses” on MEKi therapy between January 1, 2019 and June 1, 2022. We recorded the time-to-onset, type, and distribution of CAEs, non-cutaneous adverse events (AEs), AE management, and tumor response. RESULTS The median age of 40 patients with NF1 was 14 years old. Tumor types included low-grade gliomas (51%) and plexiform neurofibromas (38%). MEKi used included selumetinib (69%), trametinib (25%), and mirdametinib (6%) (Table 1). A total of 74 CAEs occurred, with 28 cases of acneiform rash (38%) (Figure 1). Twenty-three out of the 28 cases of acneiform eruption were in post-pubertal patients (82.1%). Other common CAEs were paronychia, seborrheic dermatitis, eczema, xerosis, and oral mucositis. The most common treatments included oral antibiotics and topical corticosteroids. Most patients had clinical (stable or improved) tumor response (71%) while 29% had tumor progression while on a MEKi (Table 2). There was no significant association between CAE presence and tumor response (p=0.39). CONCLUSIONS MEK inhibitors have been demonstrated as a valuable treatment, but CAEs complicate the ability to stay on the treatment. Most patients in our cohort developed a CAE, demonstrating its prevalence in this population. Improvement in characterization of MEKi toxicities and their management is important to develop treatment guidelines for pediatric and young adult patients with NF1 on MEKi therapy.

PATH-45. TREATMENT OF FGFR1 – ALTERED WHO GRADE 1 GLIONEURONAL TUMORS WITH FGFR INHIBITOR ERDAFITINIB

Abstract INTRODUCTION Low-grade mitogen-activated protein kinase (MAPK) pathway-altered glioneuronal tumors carry distinct molecular alterations, which may represent therapeutic opportunities to avoid or delay toxic treatments like radiation. Erdafitinib is a pan-FGFR tyrosine kinase inhibitor, regulatorily approved in recurrent urothelial carcinoma, and is under clinical investigation for recurrent FGFR-altered gliomas (NCT05859334). METHODS We report two patients with FGFR1-altered rosette-forming glioneuronal tumors (RGNT) treated with erdafitinib. RESULTS Patient 1 is a 32-year-old woman who presented with obstructive hydrocephalus and a non-enhancing pineal tumor. She underwent a subtotal resection, with histopathology initially consistent with a low grade dysembryoplastic neuroepithelial tumor. She was under observant management for 7 years, and developed worsening symptoms with slow non-enhancing growth, although stable per RANO low grade criteria. The initial tissue was sent for next-generation sequencing (NGS) and DNA methylation profiling, revealing an FGFR1 pathogenic variant with DNA methylation profiling matching with RGNT. Patient began Erdafitinib 8 mg daily, and had stable disease per RANO at 6 months. She developed CTCAE grade 1 toxicities of subretinal fluid, fatigue, dry skin, and paronychia. Patient 2 is a 34-year-old man diagnosed 1 year prior with a tectal tumor, who underwent biopsy due to non-measurable/ non-enhancing progression. Histopathology was consistent with RGNT. NGS revealed FGFR1 mutation, PIK3CA mutation, and NF1 pathogenic variant, described in the literature as characteristic of RGNT. There was insufficient tissue for DNA methylation profiling. Patient started erdafitinib 8 mg daily and remains clinically stable for 1 month. He developed asymptomatic hyperphosphatemia (CTCAE grade 1). Both patients have tolerated treatment without interruptions or dose reductions. CONCLUSION Erdafitinib is a well-tolerated and potentially efficacious treatment of FGFR1- altered RGNT, and may avoid the need for radiation. Diagnosis of MAPK- altered low-grade glioneuronal tumors, specifically RGNT, is increasingly reliant on molecular data, and can significantly impact treatment options.