Activin Receptor-like Kinase Research Articles

Abstract TMP3: Overexpression of Alk1 in Brain Endothelial Cells Alleviates the Severity of Brain Arteriovenous Malformations in Alk1 Mutant Mice

Background and purpose: Brain arteriovenous malformation (bAVM) can cause life threatening intra-cranial hemorrhage. Currently, there is no effective medical therapy available to treat bAVM patients. Mutation of activin receptor-like kinase 1 ( Alk1 also called ACVRL1 ) causes a familiar form of bAVM. Genetically overexpression of Alk1 in endothelial cells (ECs) recurs the phenotype of Alk1 deficient mice. We hypothesize that transduction of Alk1 gene into brain ECs using an adeno associated viral vector (AAV) may lead to cure of bAVM in Alk1 mutant mice. Methods: To identify a AAV capsid that can transduce brain ECs after intravenous (i.v.) delivery, several engineered AAV capsids were screened. A single-stranded CBA promoter driven tdTomato transgene was packaged in these capsids and delivered i.v. to wild-type mice. Tissues were collected 4 weeks later. After identifying a capsid, cc84, that can transduce brain ECs effectively after i.v. deliver, we package CMV-Alk1 transgene into AAV.cc84. Brain AVMs were induced on PdgfbiCre; Alk1 f/f mice through intra-brain injection of AAV1-VEGF followed by intraperitoneal injection of 1.25 mg/25g tamoxifen two weeks later. AAV.cc84-Alk1 was delivered to PdgfbiCre; Alk1 f/f mice through i.v. injection at the time of tamoxifen treatment Histological analyses were used to analyze viral transduction in the brain and liver, vessel density, and abnormal vessels in the brain and liver inflammation. Liver and kidney function were analyzed by colorimetric assays for Alkaline Phosphatase Activity (ALP), Alanine Transaminase Activity ALT), and Creatinine levels (Cr). Results: After i.v. injection, AAV-cc84 transduced a high percentage of brain ECs and a few hepatocytes. AAV.cc84-Alk1 treatment reduced vessel density (p=0.008), abnormal vessels (p=0.034), mouse mortality rates (20% vs 60% respectively) compared to the AAV.cc84-RFP (control vector) treated PdgfbiCre; Alk1 f/f mice. The levels of ALP, ALT and Cr were similar between AAV injected and uninjected mice. Conclusion: AAV.cc84 mediated EC overexpression of Alk1 reduced bAVM severity in Alk1 deficient mice indicating that AAV.cc84 is a promising gene therapy vector to treat bAVMs.

Impact of heterozygous ALK1 mutations on the transcriptomic response to BMP9 and BMP10 in endothelial cells from hereditary hemorrhagic telangiectasia and pulmonary arterial hypertension donors

Heterozygous activin receptor-like kinase 1 (ALK1) mutations are associated with two vascular diseases: hereditary hemorrhagic telangiectasia (HHT) and more rarely pulmonary arterial hypertension (PAH). Here, we aimed to understand the impact of ALK1 mutations on BMP9 and BMP10 transcriptomic responses in endothelial cells. Endothelial colony-forming cells (ECFCs) and microvascular endothelial cells (HMVECs) carrying loss of function ALK1 mutations were isolated from newborn HHT and adult PAH donors, respectively. RNA-sequencing was performed on each type of cells compared to controls following an 18 h stimulation with BMP9 or BMP10. In control ECFCs, BMP9 and BMP10 stimulations induced similar transcriptomic responses with around 800 differentially expressed genes (DEGs). ALK1-mutated ECFCs unexpectedly revealed highly similar transcriptomic profiles to controls, both at the baseline and upon stimulation, and normal activation of Smad1/5 that could not be explained by a compensation in cell-surface ALK1 level. Conversely, PAH HMVECs revealed strong transcriptional dysregulations compared to controls with > 1200 DEGs at the baseline. Consequently, because our study involved two variables, ALK1 genotype and BMP stimulation, we performed two-factor differential expression analysis and identified 44 BMP9-dysregulated genes in mutated HMVECs, but none in ECFCs. Yet, the impaired regulation of at least one hit, namely lunatic fringe (LFNG), was validated by RT-qPCR in three different ALK1-mutated endothelial models. In conclusion, ALK1 heterozygosity only modified the BMP9/BMP10 regulation of few genes, including LFNG involved in NOTCH signaling. Future studies will uncover whether dysregulations in such hits are enough to promote HHT/PAH pathogenesis, making them potential therapeutic targets, or if second hits are necessary.

Gastric and cardiac inflammatory myofibroblastic tumor: an extremely rare case

BackgroundInflammatory myofibroblastic tumor (IMT) is a unique, rarely metastatic tumor composed of myofibroblasts and fibrous spindle cells with inflammatory cell infiltration that can affect any organ in the human body. By reviewing the relevant literature on PubMed, we found that this is the first case report of IMT with both gastric and cardiac involvement.Case presentationA 57-year-old male patient was admitted to the hospital with complaints of malaise, poor appetite, and epigastric pain with black stools. We found a mass in the patient’s stomach and left atrium by contrast-enhanced computed tomography, 18 F-fluorodeoxyglucose positron emission tomography/computed tomography, and other tests. The patient underwent laparoscopic Billroth II subtotal gastrectomy and Braun’s gastrointestinal reconstruction under general anesthesia. On the 46th day following stomach surgery, the cardiac tumor was removed under general anesthesia. The patient has treated with doxorubicin 70 mg of D1 chemotherapy two months after cardiac surgery. Postoperative pathological immunohistochemistry of the mass confirmed the diagnosis of an IMT. His review three months after the cardiac surgery suggested the progression of the left atrial mass, but he declined further treatment and finally died one month after the review.ConclusionsAs a unique class of tumors that rarely metastasize, IMTs have an unknown etiology and pathogenesis, and distant metastasis is primarily observed in patients with negative activin receptor-like kinase (ALK) expression. The preferred treatment for IMT is complete surgical resection, and the effectiveness of adjuvant therapy for patients with distant metastases is still being determined. The clinical presentation of IMT lacks specificity and is often related to the location of tumor growth, which poses a diagnostic challenge. Pathological immunohistochemistry is the only way to confirm the diagnosis at present. Our case report reminds clinicians that a category of ALK-negative IMT with a tendency toward distant metastasis should not be ignored.

Fibronectin mediates activin A-promoted human trophoblast migration and acquisition of endothelial-like phenotype

BackgroundDuring human early placentation, a proportion of extravillous trophoblasts (EVTs) migrate to the maternal decidua, differentiating into endovascular EVTs to remodel spiral arteries and ensure the establishment of blood circulation at the maternal-fetal interface. Inadequate EVT migration and endovascular differentiation are closely associated with adverse pregnancy outcomes such as miscarriage. Activin A and fibronectin are both secretory molecules abundantly expressed at the maternal-fetal interface. Activin A has been reported to regulate EVT biological functions. However, whether fibronectin mediates activin A-promoted EVT migration and acquisition of endothelial-like phenotype as well as the underlying molecular mechanisms remain unknown. Additionally, the role of fibronectin in pregnancy establishment and maintenance warrants further investigation.MethodsPrimary and immortalized (HTR8/SVneo) human EVTs were used as in vitro study models. Cultured human first-trimester chorionic villous explants were utilized for ex vivo validation. A local fibronectin knockdown model in ICR mouse uteri, achieved by nonviral in vivo transfection with small interfering RNA (siRNA) targeting fibronectin 1 (si-Fn1), was employed to explore the roles of fibronectin in the establishment and maintenance of early pregnancy.ResultsOur results showed that activin A treatment significantly induced fibronectin 1 (FN1) mRNA expression and fibronectin protein production, which is essential for human trophoblast migration and endothelial-like tube formation. Both basal and activin A-upregulated fibronectin expression were abolished by the TGF-β type I receptor inhibitor SB431542 or siRNA-mediated knockdown of activin receptor-like kinase (ALK4) or SMAD4. Moreover, activin A-increased trophoblast migration and endothelial-like tube formation were attenuated following the depletion of fibronectin. Fibronectin knockdown via intrauterine siRNA administration reduced CD31 and cytokeratin 8 (CK8) expression at the maternal-fetal interface, resulting in a decrease in the number of implantation sites and embryos.ConclusionsOur study demonstrates that activin A promotes trophoblast cell migration and acquisition of endothelial-like phenotype via ALK4-SMAD2/3-SMAD4-mediated fibronectin upregulation. Furthermore, through a local fibronectin knockdown model in mouse uteri, we found that the absence of fibronectin at the maternal-fetal interface impedes endovascular migration of trophoblasts and decidual vascularization, thereby interfering with early embryo implantation and the maintenance of pregnancy. These findings provide novel insights into placental development during early pregnancy establishment and contribute to the advancement of therapeutic approaches for managing pregnancy complications related to trophoblast dysfunction.

Lrg1 silencing attenuates ischemia-reperfusion renal injury by regulating autophagy and apoptosis through the TGFβ1- Smad1/5 signaling pathway

BackgroundDysfunction in the processes of autophagy and apoptosis within renal tubular epithelial cells (RTEc) contributes to renal ischemia-reperfusion injury (IRI). However, the factors influencing this dysfunction remain unclear. Leucine-rich alpha-2-glycoprotein 1 (Lrg1) plays a role in the progression of diabetic nephropathy and kidney fibrosis by modulating the activin receptor-like kinase 1 (ALK1)-Smad1/5/8 and TGF-β1/Smad3 pathways, respectively. Therefore, we aimed to investigate whether Lrg1 is involved in the pathological mechanisms of renal IRI and whether its effects are related to the dysregulation of autophagy and apoptosis in RTEc. MethodsWe conducted in vitro and in vivo experiments using CoCl2-induced hypoxic human kidney-2 (HK-2) cells and mice with renal IRI, respectively. Lrg1 was silenced using siRNA and lentiviral vectors in HK-2 cells and mouse kidneys. Rapamycin (Rapa) and methyladenine were applied to regulate autophagy in renal IRI models. ResultsIncreased Lrg1 expression was observed in hypoxic HK-2 cells and in the kidneys of mice with renal IRI. Silencing of Lrg1 through siRNA and lentiviral approaches restored autophagy and suppressed apoptosis in CoCl2-induced hypoxic HK-2 cells and renal IRI models. Additionally, reduced Lrg1 expression alleviated kidney damage caused by renal IRI. The downregulation of Lrg1 expression restrained the TGFβ-Smad1/5 signaling pathway in hypoxic-induced HK-2 cells and renal IRI by reducing ALK1 expression. Lastly, the enhancement of autophagy, achieved through Rapa treatment, provided protection against renal IRI in mice. ConclusionsOur findings suggest that Lrg1 silencing can be applied as a potential therapeutic target to inhibit the TGFβ1-Smad1/5 pathway, thereby enhancing autophagy and decreasing apoptosis in patients with acute kidney injury.

Increased Collagen I/Collagen III Ratio Is Associated with Hemorrhage in Brain Arteriovenous Malformations in Human and Mouse.

Background: The increase in the collagen I (COL I)/COL III ratio enhances vessel wall stiffness and renders vessels less resistant to blood flow and pressure changes. Activated microglia enhance inflammation-induced fibrosis. Hypotheses: The COL I/COL III ratio in human and mouse brain arteriovenous malformations (bAVMs) is associated with bAVM hemorrhage, and the depletion of microglia decreases the COL I/COL III ratio and hemorrhage. Method: COL I, COL III, and hemorrhages were analyzed in 12 human bAVMs and 6 control brains, and mouse bAVMs induced in three mouse lines with activin receptor-like kinase 1 (n = 7) or endoglin (n = 7) deleted in the endothelial cells or brain focally (n = 5). The controls for the mouse study were no-gene-deleted litter mates. Mouse bAVMs were used to test the relationships between the Col I/Col III ratio and hemorrhage and whether the transient depletion of microglia reduces the Col I/Col III ratio and hemorrhage. Results: The COL I/COL III ratio was higher in the human and mouse bAVMs than in controls. The microhemorrhage in mouse bAVMs was positively correlated with the Col I/Col III ratio. Transient depletion of microglia reduced the Col I/Col III ratio and microhemorrhage. Conclusions: The COL I/COL III ratio in the bAVMs was associated with bAVM hemorrhage. The depletion of microglia reduced the bAVM Col I/Col III ratio and hemorrhage.

Pathogenic Variant Frequencies in Hereditary Haemorrhagic Telangiectasia Support Clinical Evidence of Protection from Myocardial Infarction.

Hereditary haemorrhagic telangiectasia (HHT) is a vascular dysplasia inherited as an autosomal dominant trait, due to a single heterozygous loss-of-function variant, usually in ACVRL1 (encoding activin receptor-like kinase 1 [ALK1]), ENG (encoding endoglin [CD105]), or SMAD4. In a consecutive single-centre series of 37 positive clinical genetic tests performed in 2021-2023, a skewed distribution pattern was noted, with 30 of 32 variants reported only once, but ACVRL1 c.1231C>T (p.Arg411Trp) identified as the disease-causal gene in five different HHT families. In the same centre's non-overlapping 1992-2020 series where 110/134 (82.1%) HHT-causal variants were reported only once, ACVRL1 c.1231C>T (p.Arg411Trp) was identified in nine further families. In a 14-country, four-continent HHT Mutation Database where 181/250 (72.4%) HHT-causal variants were reported only once, ACVRL1 c.1231C>T (p.Arg411Trp) was reported by 12 different laboratories, the adjacent ACVRL1 c.1232G>A (p.Arg411Gln) by 14, and ACVRL1 c.1120C>T (p.Arg374Trp) by 18. Unlike the majority of HHT-causal ACVRL1 variants, these encode ALK1 protein that reaches the endothelial cell surface but fails to signal. Six variants of this type were present in the three series and were reported 6.8-25.5 (mean 8.9) times more frequently than the other ACVRL1 missense variants (all p-values < 0.0039). Noting lower rates of myocardial infarction reported in HHT, we explore potential mechanisms, including a selective paradigm relevant to ALK1's role in the initiating event of atherosclerosis, where a plausible dominant negative effect of these specific variants can be proposed. In conclusion, there is an ~9-fold excess of kinase-inactive, cell surface-expressed ACVRL1/ALK1 pathogenic missense variants in HHT. The findings support further examination of differential clinical and cellular phenotypes by HHT causal gene molecular subtypes.

An integrated docking and molecular dynamics simulation approach to discover potential inhibitors of activin receptor-like kinase 1.

Activin receptor-like kinase 1 (ALK1) is a transmembrane receptor involved in crucial signaling pathways associated with angiogenesis and vascular development. Inhibition of ALK1 signaling has emerged as a promising therapeutic strategy for various angiogenesis-related diseases, including cancer and hereditary hemorrhagic telangiectasia. This study aimed to investigate the potential of phytoconstituents as inhibitors of ALK1 using a combined approach of virtual screening and molecular dynamics (MDs) simulations. Phytoconstituents from the IMPPAT 2.0 database underwent virtual screening to identify potential inhibitors of ALK1. The compounds were initially filtered based on physicochemical parameters, following Lipinski's rules and the PAINS filter. Subsequently, compounds demonstrating high binding affinities in docking analysis were further analyzed. Additional assessments, including ADMET, PAINS, and PASS evaluations, were conducted to identify more potent hits. Through interaction analysis, a phytoconstituent, Candidine, exhibited appreciable affinity and specific interactions with the ALK1 active site. To validate the results, MD simulations and principal components analysis were performed. The MD simulations demonstrated that Candidine stabilized the ALK1 structure and reduced conformational fluctuations. In conclusion, Candidine shows promising potential as binding partners of ALK1. These findings provide a foundation for further exploration and development of Candidine as a lead molecule for therapeutic interventions targeting ALK1-associated diseases.

Phase 1/2 Study of the Activin Receptor-like Kinase-2 Inhibitor Zilurgisertib (INCB000928, LIMBER-104) As Monotherapy or with Ruxolitinib in Patients with Anemia Due to Myelofibrosis

Introduction:Elevated levels of the iron regulator hepcidin can cause functional iron deficiency anemia. Hepcidin dysregulation is central to anemia of chronic inflammation observed in several malignancies including myelofibrosis (MF). Activin receptor-like kinase-2 (ALK2 [ie, ACVR1]) contributes to MF-associated anemia via hepcidin upregulation. We evaluated the safety, efficacy, pharmacokinetics (PK), and pharmacodynamics (PD) of zilurgisertib-a potent and selective oral ALK2 inhibitor-in patients (pts) with anemia due to MF. Methods:This ongoing open-label, multicenter, phase 1/2 dose-escalation/expansion INCB 00928-104 study (NCT04455841) is evaluating zilurgisertib alone (treatment group A [TGA]) or with ruxolitinib (RUX; treatment group B [TGB]). Eligible pts are ≥18 years old with primary/secondary MF of intermediate (Int)-1 (TGB only) or Int-2/high-risk (TGA and TGB) per the Dynamic International Prognostic Scoring System (DIPSS) and with anemia, including pts who are transfusion dependent (≥4 units of red blood cell transfusions during the 28 days or 8 weeks before Cycle [C] 1 Day [D] 1 for hemoglobin [Hb] &amp;lt;8.5 g/dL in the absence of bleeding or treatment-induced anemia) or are nontransfusion dependent with symptomatic anemia (Hb &amp;lt;10 g/dL during screening on 3 separate occasions ≥7 days apart and not meeting criteria for transfusion dependence). The zilurgisertib starting dose in TGA was 50 mg once daily (qd), with dose increases of ≤2-fold performed until a grade ≥2 toxicity with reasonable probability of being related to the treatment group was observed; subsequent dose increases were limited to ≤50% until the maximum tolerated dose (MTD) was reached or the recommended doses(s) for expansion were identified. In TGB, pts were on a stable regimen of RUX for ≥12 weeks before initiating zilurgisertib at 100 mg qd. The primary endpoints are safety and tolerability, including determination of dose-limiting toxicities (DLTs) and MTD. Secondary endpoints include efficacy (per anemia response parameters), PK, and PD (hepcidin measurement). Results:A total of36 pts were enrolled at the time of analysis (data cutoff, February 15, 2023), with 20 in TGA and 16 in TGB. Median (range) age was 73 (53-84) years for TGA and 75.5 (54-85) years for TGB; 65% of TGA pts had received prior RUX therapy. Overall, 89% had Int-2 risk DIPSS, and the remainder had high-risk DIPSS (1 in TGA, 3 in TGB). In total, 55% of pts in TGA and 25% in TGB were transfusion dependent at time of enrollment. Median (range) baseline Hb was 7.7 (7-10) g/dL in TGA and 8.0 (5-9) g/dL in TGB. Baseline median (range) hepcidin was high in both cohorts (TGA, 171 [18-535] ng/mL; TGB, 126 [7-421] ng/mL; normal range, 0-50 ng/mL). At the time of analysis, dose escalation was ongoing in both treatment groups. No DLTs occurred in either treatment group, and the MTD had not been reached. Treatment-emergent adverse events (TEAEs) were mainly low grade without apparent dose dependence. Grade ≥3 TEAEs occurred in 11 pts, with thrombocytopenia the most common (n=4). The PK profile for zilurgisertib at steady state was consistent with that observed with previous healthy volunteer studies, with observed time of maximum concentration at 2-4 hours after administration and predicted half-life of approximately 24 hours. Maximum reduction in hepcidin following zilurgisertib dosing in C1D1 was observed at 6-8 hours postdose; the highest doses tested in both TGA and TGB demonstrated more effective maintenance of hepcidin suppression over time compared with lower doses. Among non-transfusion-dependent pts, anemia improvement (Hb increase of ≥1.5 g/dL vs baseline) was observed in both treatment groups (Figure). Among transfusion-dependent pts, anemia response of transfusion independence had not been observed at data cutoff. All pts with anemia improvement have maintained Hb increase in the absence of transfusion and remain on study. Conclusions:Treatment with zilurgisertib monotherapy or in combination with RUX in this pt population was generally well tolerated, with predominantly grade 1/2 TEAEs. Reduced hepcidin levels were observed at all dose levels with both monotherapy and in combination with RUX, with greater control of hepcidin over time observed at higher zilurgisertib doses. Preliminary improvements in anemia were observed in non-transfusion-dependent pts during dose escalation, suggesting potential for therapeutic activity.

P-212 - In vitro and in vivo evaluation of [11C]M4K2127 for PET imaging activin receptor-like kinase 2

P-212 - In vitro and in vivo evaluation of [11C]M4K2127 for PET imaging activin receptor-like kinase 2

Oocyte-derived growth differentiation factor 9 suppresses the expression of CYP17A1 and androgen production in human theca cells

ObjectiveTo investigate the direct effect of growth differentiation factor 9 (GDF9) on androgen production in human theca cells. DesignExperimental study SettingTertiary hospital-based research laboratory Patient(s)Women underwent IVF/intracytoplasmic sperm injection (ICSI) at our clinic were included in this study. Intervention(s)Primary cultured human theca cells from women undergoing IVF/ICSI treatment were treated with GDF9, Activin receptor-like kinase 5 (ALK5) inhibitor, and SMAD4 agonist. Main Outcome Measure(s)The expression of androgen synthesis-related gene StAR, CYP17A1, and LHCGR, levels of androstenedione and testosterone, phosphorylation of SMAD2/3 and the interaction between bone morphogenetic protein receptor II (BMPRII) and ALK5 were evaluated by RT-qPCR, Western blot, enzyme-linked immunosorbent assays, and co-immunoprecipitation assays respectively. ResultsGDF9 decreased StAR, CYP17A1, and LHCGR expression levels in human theca cells, which was prevented by the treatment with the ALK5 inhibitor, and suppressed production of androgen in human theca cells. GDF9 increased SMAD2/3 phosphorylation, and the ALK5 inhibitor also suppressed this effect. BMPRII and ALK5 bound to each other following GDF9 stimulation. The SMAD4 agonist kartogenin also decreased mRNA levels of StAR and CYP17A1, and protein levels of StAR in human theca cells. ConclusionGDF9 can activate the BMPRII-ALK5-SMAD2/3 signaling pathway, suppress CYP17A1 expression and androgen production in human theca cells.

The effects of ALK5 inhibition and simultaneous inhibition or activation of HIF-1α in melanoma tumor growth and angiogenesis.

Hypoxia is the most common signature of the tumor microenvironment that drives tumorigenesis through the complex crosstalk of a family of transcription factors called hypoxia-inducible factors (HIFs), with other intercellular signaling networks. Hypoxia increases transforming growth factor-beta (TGF-β) expression. TGF-β and HIF-1α play critical roles in several malignancies and their interactions in melanoma progression remain unknown. Therefore, the aim of this study was to assess the impact of inhibiting activin receptor-like kinase-5 (ALK5), a TGF-β receptor, on the response to HIF-1α activation or inhibition in melanoma tumor progression. Tumors were induced in C57BL/6J mice by subcutaneous inoculation with B16F10 melanoma cells. Mice were divided into HIF-1α inhibitor, ALK5 inhibitor (1 mg/kg) and HIF-1α inhibitor (100 mg/kg), ALK5 inhibitor, HIF-1α activator (1000 mg/kg), HIF-1α activator and ALK5 inhibitor, and control groups to receive inhibitors and activators through intraperitoneal injection. The expression of E-cadherin was evaluated by RT-qPCR. Vessel density and platelet-derived growth factor receptor beta (PDGFR)-β+ cells around the vessels were investigated using immunohistochemistry. The groups receiving HIF-1α inhibitor and activator showed lower and higher tumor growth compared to the control group, respectively. E-cadherin expression decreased in all groups compared to the control group, illustrating the dual function of E-cadherin in the tumor microenvironment. Vascular density was reduced in the groups given HIF-1α inhibitor, ALK5 inhibitor, and ALK5 and HIF-1α inhibitor simultaneously. The percentage of PDGFR-β+ cells was reduced in the presence of HIF-1α inhibitor, ALK5 inhibitor, HIF-1α and ALK5 inhibitors, and upon simultaneous treatment with HIF-1α activator and ALK5 inhibitor. Despite increased expression and interaction between TGF-β and HIF-1α pathways in some cancers, in melanoma, inhibition of either pathway alone may have a stronger effect on tumor inhibition than simultaneous inhibition of both pathways. The synergistic effects may be context-dependent and should be further evaluated in different cancer types.

Activin receptor-like-kinase-2 (ALK2)-Snail pathway drives endothelial to mesenchymal transition (EndoMT) in feto-placental endothelial cells (fpEC) of early onset preeclampsia (EO-PE)

Activin receptor-like-kinase-2 (ALK2)-Snail pathway drives endothelial to mesenchymal transition (EndoMT) in feto-placental endothelial cells (fpEC) of early onset preeclampsia (EO-PE)

POSTER: MPN-548 Phase I/II Study of the Activin Receptor-like Kinase 2 (ALK2) Inhibitor Zilurgisertib (INCB000928, LIMBER-104) as Monotherapy or With Ruxolitinib in Patients With Anemia Due to Myelofibrosis

POSTER: MPN-548 Phase I/II Study of the Activin Receptor-like Kinase 2 (ALK2) Inhibitor Zilurgisertib (INCB000928, LIMBER-104) as Monotherapy or With Ruxolitinib in Patients With Anemia Due to Myelofibrosis

MPN-548 Phase I/II Study of the Activin Receptor-Like Kinase 2 (ALK2) Inhibitor Zilurgisertib (INCB000928, LIMBER-104) as Monotherapy or With Ruxolitinib in Patients With Anemia Due to Myelofibrosis

MPN-548 Phase I/II Study of the Activin Receptor-Like Kinase 2 (ALK2) Inhibitor Zilurgisertib (INCB000928, LIMBER-104) as Monotherapy or With Ruxolitinib in Patients With Anemia Due to Myelofibrosis

The LRG-TGF-β-Alk-1/TGFßRII-Smads as Predictive Biomarkers of Chronic Hydrocephalus after Aneurysmal Subarachnoid Hemorrhage.

Chronic hydrocephalus is a common complication of aneurysmal subarachnoid hemorrhage (aSAH); however, the risk factors and the mechanisms underlying its occurrence have yet to be fully elucidated. The purpose of this study was to identify biomarkers that could be used to predict chronic hydrocephalus after aSAH and to investigate the relationships. We analyzed cerebrospinal fluid (CSF) samples from 19 patients with chronic hydrocephalus after aSAH and 44 controls without hydrocephalus after aSAH. Enzyme-linked immunosorbent assay was used to determine the levels of leucine-rich alpha-2-glycoprotein (LRG), transforming growth factor-β (TGF-β), Smad1, Smad4, Smad5, Smad8, activin receptor-like kinase 1 (Alk-1), activin receptor-like kinase 5 (Alk-5), P38, and TGF-β type II receptor (TGFßRII) in CSF samples. In the CSF of patients with chronic hydrocephalus after aSAH, the levels of LRG, TGF-β, Alk-1, Smad5, and TGFßRII were significantly increased (p < 0.05) and the levels of Smad1, Smad4, and Smad8 were significantly decreased (p < 0.05). There were no significant differences between the two groups concerning the levels of P38 and Alk-5 (p > 0.05). The analysis also identified significant correlations between specific biomarkers: LRG and Smad1, LRG and Smad5, TGF-β and Alk-1, and Alk-1 and Smad4 (p < 0.05); the Pearson's correlation coefficients for these relationships were -0.341, 0.257, 0.256, and -0.424, respectively. The levels of LRG, TGF-β, Alk-1, TGFßRII, Smad1/5/8, and Smad4 in the CSF are potentially helpful as predictive biomarkers of chronic hydrocephalus after aSAH. Moreover, the LRG-TGF-β-Alk-1/TGFßRII-Smad1/5/8-Smad4 signaling pathway is highly likely to be involved in the pathogenic process of chronic hydrocephalus after aSAH.

Recommendations for the healthcare of patients with FOP

Fibrodysplasia ossificans progressiva (FOP) is avery rare, severe genetic disorder triggered by again-of-function mutation in the ACVR1 gene that codes for the typeI bone morphogenetic protein (BMP) receptor ACVR1 (activinAreceptor-type1), also known as ALK2 (activin receptor-like kinase-2). It leads to the onset and progression of heterotopic ossification (HO) in soft and connective tissue. HO is often preceded by episodes of soft tissue swelling or flare-ups. Flare-ups, characteristic of FOP, may be induced by trauma, infection, vaccination, or other medications, as well as surgical procedures or may occur spontaneously. As patients age, they develop severe mobility limitations due to progressive HO formation, including immobility, causing ashortened life expectancy. FOP's first characteristic clinical sign is the congenital malformation of one or both big toes with valgus axis deviation, which is present in almost all patients. To confirm the diagnosis, molecular genetic analysis of the ACVR1 gene is possible. This white paper aims to provide an overview of the necessary prerequisites and conditions for the care of patients with FOP and positively contribute to patients with FOP by improving the overall availability of knowledge. To achieve this, relevant aspects of the care of the very rare disease FOP are presented, from the initial diagnosis to the care in regular care based on the authors' knowledge (German FOP network) and the international FOP Treatment Guidelines. The recommendations presented here are addressed to all actors and decision-makers in the health care system and are also intended to inform patients and the public.

Pacritinib is a potent ACVR1 inhibitor with significant anemia benefit in patients with myelofibrosis

AbstractIn patients with cytopenic myelofibrosis, treatment with the JAK2/IRAK1 inhibitor pacritinib was associated with anemia benefit in the phase 3 PERSIST-2 study. The impact of pacritinib on transfusion independence (TI) has not been previously described, nor has the mechanism by which pacritinib improves anemia been elucidated. Because it has been previously postulated that inhibition of activin A receptor, type 1 (ACVR1)/activin receptor-like kinase-2 improves anemia in patients with myelofibrosis via suppression of hepcidin production, we assessed the relative inhibitory potency of pacritinib compared with other JAK2 inhibitors against ACVR1. Pacritinib inhibited ACVR1 with greater potency (half-maximal inhibitory concentration [IC50] = 16.7 nM; Cmax:IC50 = 12.7) than momelotinib (IC50 = 52.5 nM; Cmax:IC50 = 3.2), fedratinib (IC50 = 273 nM; Cmax:IC50 = 1.0), or ruxolitinib (IC50 > 1000; Cmax:IC50 < 0.01). Pacritinib’s inhibitory activity against ACVR1 was corroborated via inhibition of downstream SMAD signaling in conjunction with marked suppression of hepcidin production. Among patients on PERSIST-2 who were not transfusion independent at baseline based on Gale criteria, a significantly greater proportion achieved TI on pacritinib compared with those treated on best available therapy (37% vs 7%, P = .001), and significantly more had a ≥50% reduction in transfusion burden (49% vs 9%, P < .0001). These data indicate that the anemia benefit of the JAK2/IRAK1 inhibitor pacritinib may be a function of potent ACVR1 inhibition.

Synthesis of and anti-fibrotic effect of pyrazole derivative J-1048: Inhibition of ALK5 as a novel approach to liver fibrosis targeting inflammation

Liver fibrosis is a worldwide challenge of health issue. Developing effective new drugs for treating liver fibrosis is of great importance. In recent years, chemically synthesized drugs have significant advantages in treating liver fibrosis. Small molecule pyrazole derivatives as activin receptor-like kinase 5 (ALK5) inhibitors have also shown anti-fibrotic and tumor growth inhibitory effects. To develop the candidate with anti-fibrotic effect, we synthesized a novel pyrazole derivative, J-1048. The inhibitory effect of J-1048 on ALK5 and p38α mitogen-activated protein (MAP) kinase activity was assessed by enzymatic assays. We established an in vivo liver fibrosis model by injecting thioacetamide (TAA) into mice and in vitro model of TGF-β stimulated hepatic stellated cells to explore the inhibition mechanisms and therapeutic potential of J-1048 as an ALK5 inhibitor in liver fibrosis. Our data showed that J-1048 inhibited TAA-induced liver fibrosis in mice by explicitly blocking the TGF-β/Smad signaling pathway. Additionally, J-1048 inhibited the production of inflammatory cytokine Interleukin-1β (IL-1β) by inhibiting the purinergic ligand-gated ion channel 7 receptor (P2X7r) -Nucleotide-binding domain-(NOD-)like receptor protein 3 (NLRP3) axis, thereby alleviating liver fibrosis. Our findings demonstrated that a novel small molecule ALK5 inhibitor, J-1048, exhibited strong potential as a clinical therapeutic candidate for liver fibrosis.

Transmembrane protein 150b attenuates BMP signaling in the Xenopus organizer.

The vertebrate organizer is a specified embryonic tissue that regulates dorsoventral patterning and axis formation. Although numerous cellular signaling pathways have been identified as regulators of the organizer's dynamic functions, the process remains incompletely understood, and as-yet unknown pathways remain to be explored for sophisticated mechanistic understanding of the vertebrate organizer. To identify new potential key factors of the organizer, we performed complementary DNA(cDNA) microarray screening using organizer-mimicking Xenopus laevis tissue. This analysis yielded a list of prospective organizer genes, and we determined the role of six-transmembrane domain containing transmembrane protein 150b (Tmem150b) in organizer function. Tmem150b was expressed in the organizer region and induced by Activin/Nodal signaling. In X. laevis, Tmem150b knockdown resulted in head defects and a shortened body axis. Moreover, Tmem150b negatively regulated bone morphogenetic protein (BMP) signaling, likely via physical interaction with activin receptor-like kinase 2 (ALK2). These findings demonstrated that Tmem150b functions as a novel membrane regulatory factor of BMP signaling with antagonistic effects, contributing to the understanding of regulatory molecular mechanisms of organizer axis function. Investigation of additional candidate genes identified in the cDNA microarray analysis could further delineate the genetic networks of the organizer during vertebrate embryogenesis.