Focal Adhesion Kinase Inhibitor Research Articles

COL12A1 Promotes Osteosarcoma Progression via the FAK/PI3K/AKT/mTOR Pathway.

Osteosarcoma (OS) is a common malignancy among adolescents and children, characterized by a high propensity for metastasis and resistance to chemotherapy. This study aimed to investigate the role of COL12A1, a gene often overexpressed in various cancers and associated with poor prognosis, in the progression of OS and explore the underlying mechanisms. The expression pattern and potential function of COL12A1 in OS were evaluated using bioinformatics analyses, clinical sample examination, and OS cell lines. Various assays, including transwell, CCK-8, flow cytometry, and wound healing, were performed to assess the impact of COL12A1 on OS cell growth, cell cycle progression, apoptosis, invasion, and migration. Western blot analysis was conducted to investigate markers associated with the FAK/PI3K/AKT/mTOR pathway. COL12A1 expression was significantly elevated in OS tissues and cells. Upregulation of COL12A1 promoted cell growth, accelerated cell cycle progression, and enhanced migration and invasion while inhibiting apoptosis. Conversely, the knockdown of COL12A1 had the opposite effect. Additionally, COL12A1 overexpression increased the phosphorylation of components in the FAK/PI3K/AKT/mTOR pathway. The FAK inhibitor Y15 mitigated the effects of COL12A1 overexpression on cell apoptosis, invasion, proliferation, and the FAK/PI3K/AKT/mTOR pathway in OS. Our findings indicated that COL12A1 enhanced OS development by activating the FAK/PI3K/AKT/mTOR pathway, suggesting that COL12A1 could serve as a valuable biomarker for the prediction and identification of OS patients.

Synergistic effects of combined BET and FAK inhibition against Vestibular Schwannomas in NF2-related Schwannomatosis.

Neurofibromatosis type 2 (NF2) is a rare disorder that causes vestibular schwannomas (VS), meningiomas and ependymomas. To date, there is no FDA approved drug-based treatment for NF2. We have previously identified that BET inhibition can selectively reduce growth of the NF2-null schwannoma and Schwann cells in vitro and tumorigenesis in vivo and, separately, reported that inhibition of Focal Adhesion Kinase 1 (FAK1) via crizotinib has antiproliferative effects in NF2-null Schwann cells. The current study was aimed at determining whether combined BET and FAK inhibition can synergize and to identify the mechanisms of action. A panel of normal and NF2-null Schwann and schwannoma cell lines were used to characterize the effects of combined BET and FAK inhibition in vitro and in vivo using pharmacological and genetic approaches. The mechanism of action was explored by chromatin immunoprecipitation, ChIP-PCR, western blotting, and functional approaches. We find that combined BET and FAK inhibition are synergistic and inhibit the proliferation of NF2-null schwannoma and Schwann cell lines in vitro and in vivo, by arresting cells in the G1/S and G2/M phases of the cell cycle. Further, we identify the mechanism of action through the downregulation of FAK1 transcription by BET inhibition, which potentiates inhibition of FAK by 100-fold. Our findings suggest that combined targeting of BET and FAK1 may offer a potential therapeutic option for the treatment of NF2-related schwannomas.

The Impact of the VEGF/VEGFR2/PI3K/AKT Signaling Axis on the Proliferation and Migration Abilities of Human Dental Pulp Stem Cells.

The potential therapeutic benefits of human dental pulp stem cells (HDPSCs) in dental regenerative medicine have been demonstrated. However, little is known about the molecular mechanisms regulating the biological characteristics of HDPSCs. The experiment aims to explore whether VEGF activates signaling pathways such as FAK, PI3K, Akt, and p38 in HDPSCs, and to investigate the molecular mechanisms by which VEGF influences proliferation and migration of HDPSCs. Normal and inflamed human dental pulp (HDP) samples were collected, and the levels of VEGF in HDP were assessed. HDPSCs were cultured and purified. HDPSCs were stimulated with lipopolysaccharide (LPS) at gradient concentrations, and real-time quantitative polymerase chain reaction (qPCR) was used to assess changes in VEGF mRNA. Gradient concentrations of VEGF were used to stimulate HDPSCs, and cell migration ability was evaluated through scratch assays and Transwell chamber experiments. Phosphorylation levels of FAK, AKT, and P38 were assessed using Western blotting. Inhibitors of VEGFR2, FAK, AKT, P38, and VEGF were separately applied to HDPSCs, and cell migration ability and phosphorylation levels of FAK, AKT, and P38 were determined. The results indicated significant differences in VEGF levels between normal and inflamed HDP tissues, with levels in the inflamed state reaching 435% of normal levels (normal: 87.91 ng/mL, inflamed: 382.76 ng/mL, P < 0.05). LPS stimulation of HDPSCs showed a significant increase in VEGF mRNA expression with increasing LPS concentrations (LPS concentrations of 0.01, 0.1, 1, and 10 μg/mL resulted in VEGF mRNA expressions of 181.2%, 274.2%, 345.8%, and 460.9%, respectively, P < 0.05). VEGF treatment significantly enhanced the migration ability of HDPSCs in Transwell chamber experiments, with migration rates increasing with VEGF concentrations (VEGF concentrations of 0, 1, 10, 20, 50, and 100 ng/mL resulted in migration rates of 8.41%, 9.34%, 21.33%, 28.41%, 42.87%, and 63.15%, respectively, P < 0.05). Inhibitors of VEGFR2, FAK, AKT, P38, and combined VEGF stimulation demonstrated significant migration inhibition, with migration rates decreasing to 8.31%, 12.64%, 13.43%, 18.32%, and 74.17%, respectively. The migration rate with combined VEGF stimulation showed a significant difference (P < 0.05). The analysis of phosphorylation levels revealed that VEGF stimulation significantly activated phosphorylation of FAK, AKT, and P38, with phosphorylation levels increasing with VEGF concentrations (P < 0.05). The VEGF/VEGFR2 signaling axis regulated the migration ability of HDPSCs through the FAK/PI3K/AKT and P38MAPK pathways. This finding highlighted not only the crucial role of VEGF in injury repair of HDPSCs but also provided important clues for a comprehensive understanding of the potential applications of this signaling axis in dental regenerative medicine.

New 2,4-diphenylaminopyrimidinothiophene derivatives: Synthesis, Spectroscopic analysis, X-ray, DFT calculation, ADMET prediction, and biological activity

Pyrimidine derivatives have garnered significant attention due to their wide range of biological activities, including anticancer, antimicrobial, and antihypertensive effects. Leveraging the diverse pharmacological potential of pyrimidines, we designed and synthesized a series of novel 2,4-diphenylaminopyrimidinothiophene derivatives. The structures of these target compounds 2a-2h were confirmed using NMR and HRMS techniques, with compound 2 g further characterized by single-crystal X-ray diffraction and FT-IR analysis. Additionally, Density Functional Theory (DFT) calculations for compound 2g were conducted at the B3LYP/6–311++G(d,p) level. Focal adhesion kinase (FAK) inhibitory activity and in vitro antitumor efficacy of compounds 2a-2h against U87-MG cell lines were assessed, revealing that certain compounds exhibited significant antitumor activity. Molecular docking studies indicated that compound 2g effectively occupies the ATP binding site of FAK. Furthermore, the ADMET properties of compounds 2a-2h were predicted, providing valuable insights into their pharmacokinetic profiles. These findings lay a foundation for further exploration of compound 2g as a potential FAK inhibitor.

Fitness Screens Map State-Specific Glioblastoma Stem Cell Vulnerabilities.

Glioblastoma (GBM) is the most common and lethal primary brain tumor in adults and is driven by self-renewing glioblastoma stem cells (GSCs) that persist after therapy and seed treatment refractory recurrent tumors. GBM tumors display a high degree of intra- and inter-tumoral heterogeneity that is a prominent barrier to targeted treatment strategies. This heterogeneity extends to GSCs that exist on a gradient between two transcriptional states or subtypes termed developmental and injury-response. Drug targets for each subtype are needed to effectively target GBM. To identify conserved and subtype-specific genetic dependencies across a large and heterogeneous panel of GSCs, we designed the GBM5K targeted gRNA library and performed fitness screens in a total of 30 patient-derived GSC cultures. The focused CRISPR screens identified the most conserved subtype-specific vulnerabilities in GSCs and elucidated the functional dependency gradient existing between the developmental and injury-response states. Developmental-specific fitness genes were enriched for transcriptional regulators of neurodevelopment, whereas injury-response-specific fitness genes were highlighted by several genes implicated in integrin and focal adhesion signaling. These context-specific vulnerabilities conferred differential sensitivity to inhibitors of β1 integrin, FAK, MEK and OLIG2. Interestingly, the screens revealed that the subtype-specific signaling pathways drive differential cyclin D (CCND1 vs. CCND2) dependencies between subtypes. These data provide biological insight and mechanistic understanding of GBM heterogeneity and point to opportunities for precision targeting of defined GBM and GSC subtypes to tackle heterogeneity.

Exploiting mechanoregulation via FAK/YAP to overcome platinum resistance in ovarian cancer

Cancer cells mechanically interact with the tumor microenvironment during cancer development. Mechano-reciprocity has emerged as a crucial factor affecting anti-cancer drug resistance during adjuvant therapy. Here, we investigated the focal adhesion kinase (FAK)/Yes-associated protein (YAP) signaling axis as a prospective strategy for circumventing cisplatin resistance in ovarian cancer (OC). The Cancer Genome Atlas (TCGA) data analysis revealed that FAK overexpression significantly correlated with unfavorable clinical outcomes in patients with ovarian cancer. AFM indentation experiments showed that cell elasticity depends on FAK activity. Notably, the combination of FAK inhibition and cisplatin treatment led to a 69 % reduction in the IC50 of cisplatin. This combined treatment also increased apoptosis compared to the individual treatments, along with the upregulation of the pro-apoptotic factor BAX and cleaved PARP. Suppressing FAK expression sequestered YAP in the cytosol, potentially reducing cellular proliferation and promoting apoptosis. Moreover, reduced FAK expression sensitized drug-resistant OC cells to cisplatin treatment owing to a decrease in nuclear tension, allowing the relocation of YAP to the cytosol. In a mouse model, the co-administration of an FAK inhibitor and cisplatin significantly suppressed tumor growth and increased apoptotic events and DNA fragmentation. Our findings suggest that drug resistance can be attributed to the perturbation of mechanosensing signaling pathways, which drive the mechanical reinforcement of cancer cells. OC cells can restore their sensitivity to cisplatin treatment by strategically reducing YAP localization in the nucleus through FAK downregulation.

High expression of CX3CL1/CX3CR1 at the mother-fetus interface of preeclampsia inhibits trophoblast invasion and migration

IntroductionPreeclampsia is associated with maternal inflammatory overreaction and imbalanced immunity at the mother-fetus interface. The pro-inflammatory chemokine fractalkine (CX3CL1) is recently recognized apart from imbalanced immunity. In this study, CX3CL1- CX3C chemokine receptor 1(CX3CR1) regulation of decidual macrophage function and trophoblast invasion ability in preeclampsia was initially explored. MethodsThe study comprised 60 women allocated to NP group (normotensive pregnant woman, n = 30) and sPE group (woman with severe preeclampsia, n = 30). After the delivery, the expression of CX3CL1 in placental tissues of the two groups was detected by immunohistochemical analysis. The protein level of CX3CL1 in placental tissue and CX3CR1 in decidua tissue was detected by Western Blot and the localization of CX3CR1 expression in decidua was detected by immunofluorescence. Macrophages were polarized into classically activated (M1) macrophages. M1 were treat with PBS (control group), recombinant human CX3CL1 (CX3CL1 group), recombinant human CX3CL1+ selective CX3CR1 antagonist-JMS-17-2 (CX3CL1+anti-CX3CR1 group) and recombinant human CX3CL1 + selective CX3CR1 antagonist-JMS-17-2 + VS-6063 (CX3CL1+anti-CX3CR1+ FAK inhibitor group). M1 and HTR8/SVneo cells were co-cultured as described previously to assess invasion and migration capacity by transwell assays and Wound-healing assay. ResultsIn this study, CX3CL1 expression is high in the placental tissues of severe preeclampsia (sPE) patients than in normotensive pregnancies (NP). CX3CR1 expression is high in the decidual tissues of severe preeclampsia patients and mainly expressed in macrophages of decidual tissues. CX3CL1/CX3CR1 decreased VEGF expression in M1 macrophages and reduced the invasion and migration function of HTR-8/SVneo through the FAK signaling pathway. DiscussionThese findings revealed that CX3CL1-CX3CR1 regulate the trophoblast function by FAK and provided new insights into the pathogenesis of preeclampsia.

A novel selective FAK inhibitor E2 inhibits ovarian cancer metastasis and growth by inducing cytotoxic autophagy

Ovarian cancer (OC) is the deadliest form of the gynecologic malignancies and effective therapeutic drugs are urgently needed. Focal adhesion kinase (FAK) is overexpressed in various solid tumors, and could serve as a potential biomarker of ovarian cancer. However, there are no launched drugs targeting FAK. Hence, the development of the novel FAK inhibitors is an emerging approach for the treatment of ovarian cancer. In this work, we characterized a selective FAK inhibitor E2, with a high inhibitory potency toward FAK. Moreover, E2 had cytotoxic, anti-invasion and anti-migration activity on ovarian cancer cells. Mechanistically, after treatment with E2, FAK downstream signaling cascades (e.g., Src and AKT) were suppressed, thus resulting in the ovarian cancer cell arrest at G0/G1 phase and the induction of cytotoxic autophagy. In addition, E2 attenuated the tumor growth of PA-1 and ES-2 ovarian cancer subcutaneous xenografts, as well as suppressed peritoneal metastasis of OVCAR3-luc. Furthermore, E2 exhibited favorable pharmacokinetic properties. Altogether, these findings demonstrate that E2 is a selective FAK inhibitor with potent anti-ovarian cancer activities both in vivo and in vitro, offering new possibilities for OC treatment strategies.

Exploring the therapeutic potential of a novel series of imidazothiadiazoles targeting focal adhesion kinase (FAK) for pancreatic cancer treatment: synthesis, mechanistic insights and promising antitumor and safety profile

Focal Adhesion Kinase (FAK) is a non-receptor protein tyrosine kinase that plays a crucial role in various oncogenic processes related to cell adhesion, migration, proliferation, and survival. The strategic targeting of FAK represents a burgeoning approach to address resistant tumours, such as pancreatic ductal adenocarcinoma (PDAC). Herein, we report a new series of twenty imidazo[2,1-b][1, 3, 4]thiadiazole derivatives assayed for their antiproliferative activity against the National Cancer Institute (NCI-60) panel and a wide panel of PDAC models. Lead compound 10l exhibited effective antiproliferative activity against immortalised (SUIT-2, CAPAN-1, PANC-1, PATU-T, BxPC-3), primary (PDAC-3) and gemcitabine-resistant clone (PANC-1-GR) PDAC cells, eliciting IC50 values in the low micromolar range (1.04–3.44 µM), associated with a significant reduction in cell-migration and spheroid shrinkage in vitro. High-throughput kinase arrays revealed a significant inhibition of the FAK signalling network, associated to induction of cell cycle arrest in G2/M phase, suppression of tumour cell invasion and apoptosis induction. The high selectivity index/toxicity prompted studies using PDAC mouse xenografts, demonstrating significant inhibition of tumour growth and safety. In conclusion, compound 10l displayed antitumor activity and safety in both in vitro and in vivo models, emerging as a highly promising lead for the development of FAK inhibitors in PDAC.

Defactinib inhibits FAK phosphorylation and regulates psoriasis via attenuating hyperproliferation of keratinocytes.

Excessive proliferation of keratinocytes is a crucial pathological risk feature of psoriasis. Focal adhesion kinase (FAK) is a non-receptor protein that primarily regulates cell proliferation and migration. However, the expression and regulatory mechanism of FAK in psoriasis remains unclear. This study aimed to investigate the regulation of FAK in psoriasis and examined the potential impact of FAK inhibitor on psoriasis. A small molecular selective FAK inhibitor, defactinib, was used to evaluate the effect of FAK on psoriasis in invitro and invivo functional assays. In our experiments, imiquimod (IMQ)-induced psoriasis mice and human keratinocytes cells were used to study the potential roles and mechanisms of FAK in psoriasis. FAK phosphorylation has been weakly detected in normal intact skin and is markedly elevated upon IMQ treatment. By reducing FAK phosphorylation (p-FAK),defactinib treatment could attenuate psoriasiform inflammation and epidermal hyperplasia in IMQ-treated mice compared with IMQ-induced mice treated with the vehicle. In invitro studies, resiquimod (R848) increased (p-FAK) and promoted cell proliferation in human keratinocytes cells, while defactinib reversed this effect. Mechanistically, defactinib can alleviate the proliferation via JNK/YB1 pathway invitro and invivo. Defactinib significantly attenuates psoriasiform inflammation and epidermal hyperproliferation through the inhibition of the FAK-mediated axis. The downregulation of phosphorylated FAK then suppressed the activation of JNK/YB1 protein signaling pathway in psoriasis. Our work highlights targeting FAK as a potentially effective strategy for the treatment of psoriasis.

Protective Effect of Caffeine and Chlorogenic Acids of Coffee in Liver Disease.

Coffee is one of the most widely consumed beverages in the world due to its unique aroma and psychostimulant effects, mainly due to the presence of caffeine. In recent years, experimental evidence has shown that the moderate consumption of coffee (3/4 cups per day) is safe and beneficial to human health, revealing protective effects against numerous chronic metabolic diseases such as diabetes, cardiovascular, neurodegenerative, and hepatic diseases. This review focuses on two of coffee's main bioactive compounds, i.e., caffeine and chlorogenic acids, and their effects on the progression of chronic liver diseases, demonstrating that regular coffee consumption correlates with a lower risk of the development and progression of non-alcoholic steatohepatitis, viral hepatitis, liver cirrhosis, and hepatocellular carcinoma. In particular, this review analyzes caffeine and chlorogenic acid from a pharmacological point of view and explores the molecular mechanism through which these compounds are responsible for the protective role of coffee. Both bioactive compounds, therefore, have antifibrotic effects on hepatic stellate cells and hepatocytes, induce a decrease in connective tissue growth factor, stimulate increased apoptosis with anti-cancer effects, and promote a major inhibition of focal adhesion kinase, actin, and protocollagen synthesis. In conclusion, coffee shows many beneficial effects, and experimental data in favor of coffee consumption in patients with liver diseases are encouraging, but further prospective studies are needed to demonstrate its preventive and therapeutic role in chronic liver diseases.

Focal adhesion kinase signaling - tumor vulnerabilities and clinical opportunities.

Focal adhesion kinase (FAK; encoded by PTK2) was discovered over 30 years ago as a cytoplasmic protein tyrosine kinase that is localized to cell adhesion sites, where it is activated by integrin receptor binding to extracellular matrix proteins. FAK is ubiquitously expressed and functions as a signaling scaffold for a variety of proteins at adhesions and in the cell cytoplasm, and with transcription factors in the nucleus. FAK expression and intrinsic activity are essential for mouse development, with molecular connections to cell motility, cell survival and gene expression. Notably, elevated FAK tyrosine phosphorylation is common in tumors, including pancreatic and ovarian cancers, where it is associated with decreased survival. Small molecule and orally available FAK inhibitors show on-target inhibition in tumor and stromal cells with effects on chemotherapy resistance, stromal fibrosis and tumor microenvironment immune function. Herein, we discuss recent insights regarding mechanisms of FAK activation and signaling, its roles as a cytoplasmic and nuclear scaffold, and the tumor-intrinsic and -extrinsic effects of FAK inhibitors. We also discuss results from ongoing and advanced clinical trials targeting FAK in low- and high-grade serous ovarian cancers, where FAK acts as a master regulator of drug resistance. Although FAK is not known to be mutationally activated, preventing FAK activity has revealed multiple tumor vulnerabilities that support expanding clinical combinatorial targeting possibilities.

Recurrent RhoGAP gene fusion CLDN18-ARHGAP26 promotes RHOA activation and focal adhesion kinase and YAP-TEAD signalling in diffuse gastric cancer

ObjectiveGenomic studies of gastric cancer have identified highly recurrent genomic alterations impacting RHO signalling, especially in the diffuse gastric cancer (DGC) histological subtype. Among these alterations are interchromosomal translations leading...

Schisandra chinensis Bee Pollen Extract Inhibits Proliferation and Migration of Hepatocellular Carcinoma HepG2 Cells via Ferroptosis-, Wnt-, and Focal Adhesion-Signaling Pathways.

Bee pollen possesses favorable anticancer activities. As a medicinal plant source, Schisandra chinensis bee pollen (SCBP) possesses potential pharmacological properties, such as reducing cisplatin-induced liver injury, but its anti-liver cancer effect is still rarely reported. This paper aims to investigate the effect and mechanism of SCBP extract (SCBPE) on hepatocellular carcinoma HepG2 cells. The effect of SCBPE on cell proliferation and migration of HepG2 cells was evaluated based on MTT assay, morphology observation, or scratching assay. Furthermore, tandem mass tag-based quantitative proteomics was used to study the effect mechanisms. The mRNA expression levels of identified proteins were verified by RT-qPCR. Tandem mass tag-based quantitative proteomics showed that 61 differentially expressed proteins were obtained in the SCBPE group compared with the negative-control group: 18 significantly downregulated and 43 significantly upregulated proteins. Bioinformatic analysis showed the significantly enriched KEGG pathways were predominantly ferroptosis-, Wnt-, and hepatocellular carcinoma-signaling ones. Protein-protein interaction network analysis and RT-qPCR validation revealed SCBPE also downregulated the focal adhesion-signaling pathway, which is abrogated by PF-562271, a well-known inhibitor of FAK. This study confirmed SCBPE suppressed the cell proliferation and migration of hepatocellular carcinoma HepG2 cells, mainly through modulation of ferroptosis-, Wnt-, hepatocellular carcinoma-, and focal adhesion-signaling pathways, providing scientific data supporting adjuvant treatment of hepatocellular carcinoma using SCBP.

SPP1 promotes tumor progression in esophageal carcinoma by activating focal adhesion pathway.

Recurrence and metastasis are the major obstacles affecting the therapeutic efficacy and clinical outcomes for patients with esophageal carcinoma (ESCA). Secreted phosphoprotein 1 (SPP1) is considered as a hub gene in ESCA and is negatively associated with disease-free survival (DFS) in ESCA. However, the exact roles and underlying mechanisms remain elusive. This study aims to examine the roles of SPP1 on ESCA, and elucidate the potential mechanisms. Bioinformatics were used to analyze the expression of SPP1 in ESCA tissues, and its relations with clinicopathological characteristics and clinical prognosis in patients with ESCA based on The Cancer Genome Atlas (TCGA) dataset. Loss-of-function was conducted to examine the roles of SPP1 on malignant behaviors of ESCA cells by cell counting kit-8 (CCK8), plate clone, wound healing, and transwell assays. Gene set enrichment analysis (GSEA) was conducted to screen the pathways associated with SPP1 in ESCA. Then, the enriched pathway and the underlying mechanism were elucidated by western blotting, cell adhesion, and cell spreading assays. Lastly, Y15 [a specific inhibitor of focal adhesion kinase (FAK)] was used to examine its potential to inhibit tumor growth in ESCA cells. SPP1 was upregulated in ESCA tissues compared to the adjacent nontumorous tissues, which was closely associated with clinical stage, lymph node metastasis, histological subtype, and p53 mutation. A high expression of SPP1 indicated a poor clinical prognosis in patients with ESCA. The knockdown of SPP1 inhibited cell proliferative, migratory, and invasive capacities in ESCA cells. GSEA indicated that the focal adhesion pathway was closely related with SPP1 in ESCA. Further studies confirmed that the knockdown of SPP1 suppressed cell adhesion ability and reduced the expression of p-FAK and p-Erk in ESCA cells. In addition, Y15 inhibited FAK autophosphorylation and dramatically inhibited cell proliferation, migration, and invasion in ESCA cells. SPP1 promotes tumor progression in ESCA by activating FAK/Erk pathway, and FAK is a potential therapeutic target to overcome tumor recurrence and metastasis of ESCA.

Regulation of intracellular cAMP levels in osteocytes by mechano-sensitive focal adhesion kinase via PDE8A.

Osteocytes are the primary mechano-sensitive cell type in bone. Mechanical loading is sensed across the dendritic projections of osteocytes leading to transient reductions in focal adhesion kinase (FAK) activity. Knowledge regarding the signaling pathways downstream of FAK in osteocytes is incomplete. We performed tyrosine-focused phospho-proteomic profiling in osteocyte-like Ocy454 cells to identify FAK substrates. Gsα, parathyroid hormone receptor (PTH1R), and phosphodiesterase 8A (PDE8A), all proteins associated with cAMP signaling, were found as potential FAK targets based on their reduced tyrosine phosphorylation in both FAK- deficient or FAK inhibitor treated cells. Real time monitoring of intracellular cAMP levels revealed that FAK pharmacologic inhibition or gene deletion increased basal and GPCR ligand-stimulated cAMP levels and downstream phosphorylation of protein kinase A substrates. Mutating FAK phospho-acceptor sites in Gsα and PTH1R had no effect on PTH- or FAK inhibitor-stimulated cAMP levels. Since FAK inhibitor treatment augmented cAMP levels even in the presence of forskolin, we focused on potential FAK substrates downstream of cAMP generation. Indeed, PDE8A inhibition mimicked FAK inhibition at the level of increased cAMP, PKA activity, and expression of cAMP-regulated target genes. In vitro kinase assay showed that PDE8A is directly phosphorylated by FAK while immunoprecipitation assays revealed intracellular association between FAK and PDE8A. Thus, FAK inhibition in osteocytes acts synergistically with signals that activate adenylate cyclase to increase intracellular cAMP. Mechanically-regulated FAK can modulate intracellular cAMP levels via effects on PDE8A. These data suggest a novel signal transduction mechanism that mediates crosstalk between mechanical and cAMP-linked hormonal signaling in osteocytes.

Adipocyte-specific FAK deletion promotes pancreatic β-cell apoptosis via adipose inflammatory response to exacerbate diabetes mellitus.

White adipose tissue (WAT) has a key role in maintaining energy balance throughout the body, and their dysfunction take part in the regulation of diabetes mellitus. However, the internal regulatory mechanisms underlying are still unknown. We generated adipocyte-specific FAK KO (FAK-AKO) mice and investigated their phenotype. The cascade of adipocyte, macrophage in adipocyte tissues, and pancreatic β-cells were proposed in FAK-AKO mice and validated by cell line studies using 3T3-L1, Raw264.7 and Min6. The FAK-AKO mice exhibited glucose intolerance, reduced adipose tissue mass and increased apoptosis, lipolysis and inflammatory response in adipose tissue. We further demonstrate that adipocyte FAK deletion increases β cell apoptosis and inflammatory infiltrates into islets, which is potentiated if mice were treated with STZ. In the STZ-induced diabetes model, FAK AKO mice exhibit less serum insulin content and pancreatic β cell area. Moreover, serum pro-inflammatory factors increased and insulin levels decreased after glucose stimulation in FAK AKO mice. In a parallel vitro experiment, knockdown or inhibition of FAK during differentiation also increased apoptosis, lipolysis and inflammatory in 3T3-L1 adipocytes, whereas the opposite was observed upon overexpression of FAK. Moreover, coculturing LPS-treated RAW264.7 macrophages with knockdown FAK of 3T3-L1 adipocytes increased macrophage pro-inflammatory response. Furthermore, conditioned medium from above stimulated Min6 cells apoptosis (with or without STZ), whereas the opposite was observed upon overexpression of FAK. Mechanistically, FAK protein interact with TRAF6 in adipocytes and knockdown or inhibition of FAK activated TRAF6/TAK1/NF-κB signaling, which exacerbates inflammation of adipocytes themselves. Adipocyte FAK deletion promotes both adipocyte apoptosis and adipose tissue inflammation. Pro-inflammatory factors released by the FAK-null adipose tissue further trigger apoptosis in pancreatic islets induced by the administration of STZ, thereby exacerbating the diabetes mellitus. This study reveals a link between FAK-mediated adipose inflammation and diabetes mellitus, a mechanism that has not been previously recognized.

Injectable hybrid hydrogels enable enhanced combination chemotherapy and roused anti-tumor immunity in the synergistic treatment of pancreatic ductal adenocarcinoma

Chemotherapy and immunotherapy have shown no significant outcome for unresectable pancreatic ductal adenocarcinoma (PDAC). Multi-drug combination therapy has become a consensus in clinical trials to explore how to arouse anti-tumor immunity and meanwhile overcome the poorly tumoricidal effect and the stroma barrier that greatly hinders drug penetration. To address this challenge, a comprehensive strategy is proposed to fully utilize both the ferroptotic vulnerability of PDAC to potently irritate anti-tumor immunity and the desmoplasia-associated focal adhesion kinase (FAK) to wholly improve the immunosuppressive microenvironment via sustained release of drugs in an injectable hydrogel for increasing drug penetration in tumor location and averting systematic toxicity. The injectable hydrogel ED-M@CS/MC is hybridized with micelles loaded with erastin that exclusively induces ferroptosis and a FAK inhibitor defactinib for inhibiting stroma formation, and achieves sustained release of the drugs for up to 12 days. With only a single intratumoral injection, the combination treatment with erastin and defactinib produces further anti-tumor performance both in xenograft and KrasG12D-engineered primary PDAC mice and synergistically promotes the infiltration of CD8+ cytotoxic T cells and the reduction of type II macrophages. The findings may provide a novel promising strategy for the clinical treatment of PDAC.

Plexin D1 negatively regulates macrophage-derived foam cell migration via the focal adhesion kinase/Paxillin pathway

BackgroundMacrophage-derived foam cell formation is a hallmark of atherosclerosis and is retained during plaque formation. Strategies to inhibit the accumulation of these cells hold promise as viable options for treating atherosclerosis. Plexin D1 (PLXND1), a member of the Plexin family, has elevated expression in atherosclerotic plaques and correlates with cell migration; however, its role in macrophages remains unclear. We hypothesize that the guidance receptor PLXND1 negatively regulating macrophage mobility to promote the progression of atherosclerosis. MethodsWe utilized a mouse model of atherosclerosis based on a high-fat diet and an ox-LDL- induced foam cell model to assess PLXND1 levels and their impact on cell migration. Through western blotting, Transwell assays, and immunofluorescence staining, we explored the potential mechanism by which PLXND1 mediates foam cell motility in atherosclerosis. ResultsOur study identifies a critical role for PLXND1 in atherosclerosis plaques and in a low-migration capacity foam cell model induced by ox-LDL. In the aortic sinus plaques of ApoE−/− mice, immunofluorescence staining revealed significant upregulation of PLXND1 and Sema3E, with colocalization in macrophages. In macrophages treated with ox-LDL, increased expression of PLXND1 led to reduced pseudopodia formation and decreased migratory capacity. PLXND1 is involved in regulating macrophage migration by modulating the phosphorylation levels of FAK/Paxillin and downstream CDC42/PAK. Additionally, FAK inhibitors counteract the ox-LDL-induced migration suppression by modulating the phosphorylation states of FAK, Paxillin and their downstream effectors CDC42 and PAK. ConclusionOur findings indicate that PLXND1 plays a role in regulating macrophage migration by modulating the phosphorylation levels of FAK/Paxillin and downstream CDC42/PAK to promoting atherosclerosis.

A reanalysis and integration of transcriptomics and proteomics datasets unveil novel drug targets for Mekong schistosomiasis

Schistosomiasis, caused by Schistosoma trematodes, is a significant global health concern, particularly affecting millions in Africa and Southeast Asia. Despite efforts to combat it, the rise of praziquantel (PZQ) resistance underscores the need for new treatment options. Protein kinases (PKs) are vital in cellular signaling and offer potential as drug targets. This study focused on focal adhesion kinase (FAK) as a candidate for anti-schistosomal therapy. Transcriptomic and proteomic analyses of adult S. mekongi worms identified FAK as a promising target due to its upregulation and essential role in cellular processes. Molecular docking simulations assessed the binding energy of FAK inhibitors to Schistosoma FAK versus human FAK. FAK inhibitor 14 and PF-03814735 exhibited strong binding to Schistosoma FAK with minimal binding for human FAK. In vitro assays confirmed significant anti-parasitic activity against S. mekongi, S. mansoni, and S. japonicum, comparable to PZQ, with low toxicity in human cells, indicating potential safety. These findings highlight FAK as a promising target for novel anti-schistosomal therapies. However, further research, including in vivo studies, is necessary to validate efficacy and safety before clinical use. This study offers a hopeful strategy to combat schistosomiasis and reduce its global impact.