Dihydroergotamine ameliorates liver fibrosis by targeting transforming growth factor β type Ⅱ receptor

BACKGROUND The transforming growth factor β(TGFβ) signaling pathway plays a crucial role in the development of liver fibrosis by activating TGFβ type Ⅱ receptor(TGFβR2), followed by the recruitment of TGFβR1 finally triggering downstream signaling pathway.AIM To find drugs targeting TGFβR2 that inhibit TGFβR1/TGFβR2 complex formation, theoretically inhibit TGFβ signaling pathway, and thereby ameliorate liver fibrosis.METHODS Food and Drug Administration-approved drugs were screened for binding affinity with TGFβR2 by virtual molecular docking. We identified 6 candidates and further explored their potential by Cell Counting Kit-8(CCK-8) cell cytotoxic experiment to validate toxicity and titrated the best cellular working concentrations. Next, we further demonstrated the detailed molecular working mechanisms using mutagenesis analysis. Finally, we used a mouse model to investigate its potential anti-liver fibrosis effect.RESULTS We identified 6 drug candidates. Among these 6 drugs, dihydroergotamine(DHE) shows great ability in reducing fibrotic gene expressions such as collagen, p-SMAD3, and α-SMA in TGFβ induced cellular model of liver fibrosis in LX-2 cells. Furthermore, we demonstrated that DHE binds to TGFβR2. Moreover, mutation of Leu27, Phe30, Thr51, Ser52, Ile53, and Glu55 of TGFβR2 disrupted the binding of TGFβR2 with DHE. In addition, DHE significantly improved liver fibrosis, as evidenced by Masson’s trichrome staining of liver sections. This is further supported by the width and the velocity of the portal vein, and serum markers of liver function. In line with those observations, DHE also decreased macrophages infiltration and extracellular matrix deposition in the liver.CONCLUSION DHE alleviates liver fibrosis by binding to TGFβR2 thereby suppressing TGFβ signaling pathway. We show here that as far as drug repurposing, DHE has great potential to treat liver fibrosis.

Therapeutic strategies targeting the epidermal growth factor receptor signaling pathway in metastatic colorectal cancer

More than 1.9 million new colorectal cancer(CRC)cases and 935000 deaths were estimated to occur worldwide in 2020,representing about one in ten cancer cases and deaths.Overall,colorectal ranks third in incidence,but second in mortality.More than half of the patients are in advanced stages at diagnosis.Treatment options are complex because of the heterogeneity of the patient population,including different molecular subtypes.Treatments have included conventional fluorouracil-based chemotherapy,targeted therapy,immunotherapy,etc.In recent years,with the development of genetic testing technology,more and more targeted drugs have been applied to the treatment of CRC,which has further prolonged the survival of metastatic CRC patients.

PRaG 3.0 therapy for human epidermal growth factor receptor 2-positive metastatic pancreatic ductal adenocarcinoma:A case report

BACKGROUND Pancreatic ductal adenocarcinoma(PDAC)is a highly fatal disease with limited effective treatment especially after first-line chemotherapy.The human epidermal growth factor receptor 2(HER-2)immunohistochemistry(IHC)positive is associated with more aggressive clinical behavior and shorter overall survival in PDAC.CASE SUMMARY We present a case of multiple metastatic PDAC with IHC mismatch repair proficient but HER-2 IHC weakly positive at diagnosis that didn’t have tumor regression after first-line nab-paclitaxel plus gemcitabine and PD-1 inhibitor treatment.A novel combination therapy PRaG 3.0 of RC48(HER2-antibody-drug conjugate),radio-therapy,PD-1 inhibitor,granulocyte-macrophage colony-stimulating factor and interleukin-2 was then applied as second-line therapy and the patient had confirmed good partial response with progress-free-survival of 6.5 months and overall survival of 14.2 month.She had not developed any grade 2 or above treatment-related adverse events at any point.Percentage of peripheral CD8^(+) Temra and CD4^(+) Temra were increased during first two activation cycles of PRaG 3.0 treatment containing radiotherapy but deceased to the baseline during the maintenance cycles containing no radiotherapy.CONCLUSION PRaG 3.0 might be a novel strategy for HER2-positive metastatic PDAC patients who failed from previous first-line approach and even PD-1 immunotherapy but needs more data in prospective trials.

Advances in targeted therapy for human epidermal growth factor receptor 2 positive in advanced gastric cancer

Emerging therapeutic methods represented by targeted therapy are effective supplements to traditional first-line chemoradiotherapy resistance.Human epidermal growth factor receptor 2(HER2)is one of the most important targets in targeted therapy for gastric cancer.Trastuzumab combined with chemotherapy has been used as the first-line treatment for advanced gastric cancer.The safety and efficacy of pertuzumab and margetuximab in the treatment of gastric cancer have been verified.However,monoclonal antibodies,due to their large molecular weight,inability to penetrate the blood-brain barrier,and drug resistance,lead to decreased therapeutic efficacy,so it is necessary to explore the efficacy of other HER2-targeting therapies in gastric cancer.Small-molecule tyrosine kinase inhibitors,such as lapatinib and pyrrotinib,have the advantages of small molecular weight,penetrating the blood-brain barrier and high oral bioavailability,and are expected to become the drugs of choice for perioperative treatment and neoadjuvant therapy of gastric cancer after validation by large-scale clinical trials in the future.Antibo-drug conjugate,such as T-DM1 and T-DXd,can overcome the resistance of monoclonal antibodies despite their different mechanisms of tumor killing,and are a supplement for the treatment of patients who have failed the treatment of monoclonal antibodies such as trastuzumab.Therefore,after more detailed stratification of gastric cancer patients,various gastric cancer drugs targeting HER2 are expected to play a more significant role.

Concomitant epidermal growth factor receptor mutation/c-ros oncogene 1 rearrangement in non-small cell lung cancer: A case report

BACKGROUND Epidermal growth factor receptor(EGFR)mutation and c-ros oncogene 1(ROS1)rearrangement are key genetic alterations and predictive tumor markers for non-small cell lung cancer(NSCLC)and are typically considered to be mutually exc-lusive.EGFR/ROS1 co-mutation is a rare event,and the standard treatment appr-oach for such cases is still equivocal.CASE SUMMARY Herein,we report the case of a 64-year-old woman diagnosed with lung adenocar-cinoma,with concomitant EGFR L858R mutation and ROS1 rearrangement.The patient received two cycles of chemotherapy after surgery,but the disease prog-ressed.Following 1-month treatment with gefitinib,the disease progressed again.However,after switching to crizotinib,the lesion became stable.Currently,crizotinib has been administered for over 53 months with a remarkable treatment effect.CONCLUSION The efficacy of EGFR tyrosine kinase inhibitors and crizotinib was vastly different in this NSCLC patient with EGFR/ROS1 co-mutation.This report will aid future treatment of such patients.

Human epidermal growth factor receptor 2 expression level and combined positive score can evaluate efficacy of advanced gastric cancer

BACKGROUND Although treatment options for gastric cancer(GC)continue to advance,the overall prognosis for patients with GC remains poor.At present,the predictors of treatment efficacy remain controversial except for high microsatellite instability.AIM To develop methods to identify groups of patients with GC who would benefit the most from receiving the combination of a programmed cell death protein 1(PD-1)inhibitor and chemotherapy.METHODS We acquired data from 63 patients with human epidermal growth factor receptor 2(HER2)-negative GC with a histological diagnosis of GC at the Cancer Hospital,Chinese Academy of Medical Sciences between November 2020 and October 2022.All of the patients screened received a PD-1 inhibitor combined with chemotherapy as the first-line treatment.RESULTS As of July 1,2023,the objective response rate was 61.9%,and the disease control rate was 96.8%.The median progression-free survival(mPFS)for all patients was 6.3 months.The median overall survival was not achieved.Survival analysis showed that patients with a combined positive score(CPS)≥1 exhibited an extended trend in progression-free survival(PFS)when compared to patients with a CPS of 0 after receiving a PD-1 inhibitor combined with oxaliplatin and tegafur as the first-line treatment.PFS exhibited a trend for prolongation as the expression level of HER2 increased.Based on PFS,we divided patients into two groups:A treatment group with excellent efficacy and a treatment group with poor efficacy.The mPFS of the excellent efficacy group was 8 months,with a mPFS of 9.1 months after excluding a cohort of patients who received interrupted therapy due to surgery.The mPFS was 4.5 months in patients in the group with poor efficacy who did not receive surgery.Using good/poor efficacy as the endpoint of our study,univariate analysis revealed that both CPS score(P=0.004)and HER2 expression level(P=0.015)were both factors that exerted significant influence on the efficacy of treatment the combination of a PD-1 inhibitor and chemotherapy in patients with advanced GC(AGC).Finally,multivariate analysis confirmed that CPS score was a significant influencing factor.CONCLUSION CPS score and HER2 expression both impacted the efficacy of immunotherapy combined with chemotherapy in AGC patients who were non-positive for HER2.

Correlation between expression of two transforming growth factor-beta 1 receptors and microvascular density in a rat model of cerebral ischemia and reperfusion injury

The effects of transforming growth factor-β1 （TGF-β1） are currently controversial. Whether TGF-β1 promotes or inhibits revascularization under different conditions remains poorly understood. Based on previous studies, the current experiment established rat models of cerebral ischemia and reperfusion injury （IRI）, and demonstrated that pathological and functional damage was also increased after IRI. The most serious damage was observed at 3 days after reperfusion, at which time microvascular density fell to its lowest level. Soon afterwards, microvascular density increased, new collateral circulation was gradually established at 4 to 7 days after reperfusion, and pathological damage and neurological deficits were improved. TGF-β1, activin receptor-like kinase 5 （ALK5） mRNA and protein expression levels increased gradually over time. In contrast, ALK1 mRNA and protein expression decreased over the same period. A significant negative correlation was detected between microvascular density and expression of the ALK5 gene transcript. There was no correlation between microvascular density and ALK1 gene transcriptional expression following cerebral IRI in a rat model. These findings suggest that ALK5, rather than ALK1, is the critical receptor in the TGF-β1 signal pathways after cerebral IRI.

Stage-specific localization of transforming growth factor β1 and β3 and their receptors during spermatogenesis in men

Aim: To investigate the stage-specific localization of transforming growth factor (TGF) β1 and β3 during spermatogenesis in adult human testis. Methods: The localization of TGFβ1 and β3 was investigated by immunohis tochemical staining method employing specific polyclonal antibodies. Results: Both TGFβ1 and β3 and their recep tors were preponderant in the Leydig celis. TGFβ1 could not be detected in the seminiferous tubules. TGFβ3 and TGFβ-Receptor (R) I were mainly seen in the elongated spermatids, while TGFβ-RⅡ in the pachytene spermatocytes and weak in the spermatogonia, spermatids and Sertoli celis. Only TGFβ-RⅡ was detected in the Sertoli celis. TGFβ3, TGFβ-RⅠ and TGFβ-RⅡ showed a staining pattern dependent upon the stages of the seminiferous epithelium cycle. Conclusion: TGFβ isoforms and their receptors are present in the somatic and germ celis of the adult human testis, suggesting their involvement in the regulation of spermatogenesis.

Expression patterns of transforming growth factor-beta and its receptors in gastric mucosa of patients with refractory gastric ulcer

AIM: Transforming growth factor-β (TGF-β) plays a regulatory role in tissue repair. In a previous study, we found that TGF-β and its receptors were expressed in gastric mucosa of patients with well-healed gastric ulcers, as demonstrated by immunohistochemistry. To further characterize the role of TGF-β and its receptors in repairing gastric ulcers, we investigated the expression patterns of TGF-β and its receptors in gastric mucosa by in situ hybridization and reverse transcriptase-polymerase chain reaction (RT-PCR).METHODS: Seventy-four patients with endoscopically proven gastric ulcers were eligible for participation in this study. All patients had routine biopsies on initial endoscopy and were then treated for 12 wk with an H2 blocker. Repeat endoscopy was then performed. There were 8 patients with poorly healed ulcers, and biopsies were taken from the margin of the residual ulcers. These tissue samples, along with biopsy of gastric mucosa near the original ulcers from 8 randomly selected patients with well-healed ulcers were examined for TGF-β and TGF-β receptor Ⅱ mRNA by RT-PCR and in situ hybridization, as well as immunohistochemistry.RESULTS: TGF-β and TGF-β receptor Ⅱ were strongly expressed in tissues from patients with well-healed ulcers.Four of the 8 patients with poor healing had low or absent expression of TGF-β or TGF-β receptor Ⅱ mRNA. All cases positive by RT-PCR assay were confirmed by in situ hybridization as well as immunohistochemistry.CONCLUSION: It is suggested that TGF-β and its receptors are important for gastric ulcer healing. These results may have implications for further investigation of the healing process and in predicting response to therapy.

Transforming growth factor-β and toll-like receptor-4 polymorphisms are not associated with fibrosis in haemochromatosis

AIM:To investigate the role of genetic polymorphisms in the progression of hepatic fibrosis in hereditary haemochromatosis.METHODS:A cohort of 245 well-characterised C282Y homozygous patients with haemochromatosis was studied,with all subjects having liver biopsy data and DNA available for testing.This study assessed the association of eight single nucleotide polymorphisms(SNPs)in a total of six genes including toll-like receptor 4(TLR4),transforming growth factor-beta(TGF-β),oxoguanine DNA glycosylase,monocyte chemoattractant protein 1,chemokine C-C motif receptor 2 and interleukin-10 with liver disease severity.Genotyping was performed using high resolution melt analysis and sequencing.The results were analysed in relation to the stage of hepatic fibrosis in multivariate analysis incorporating other cofactors including alcohol consumption and hepatic iron concentration.RESULTS:There were significant associations between the cofactors of male gender(P=0.0001),increasing age(P=0.006),alcohol consumption(P=0.0001),steatosis(P=0.03),hepatic iron concentration(P<0.0001)and the presence of hepatic fibrosis.Of the candidate gene polymorphisms studied,none showed a significant association with hepatic fibrosis in univariate or multivariate analysis incorporating cofactors.We also specifically studied patients with hepatic iron loading above threshold levels for cirrhosis and compared the genetic polymorphisms between those with no fibrosis vs cirrhosis however there was no significant effect from any of the candidate genes studied.Importantly,in this large,well characterised cohort of patients there was no association between SNPs for TGF-βor TLR4and the presence of fibrosis,cirrhosis or increasing fibrosis stage in multivariate analysis.CONCLUSION:In our large,well characterised group of haemochromatosis subjects we did not demonstrate any relationship between candidate gene polymorphisms and hepatic fibrosis or cirrhosis.

Detection of Frameshift Mutations of the Transforming Growth Factor p ReceptorⅡin Gastric Cancers with Microsatellite Instability

OBJECTIVE To study the relationship among microsatellite instability （MSI）, frameshift mutations （FM） of the transforming growth factor β receptor Ⅱ （TGF β R Ⅱ）, methylation state of the hMLH1 promoter and hMLH1 protein expression level in gastric cancers, and to explore their relationship to gastric carcinogenesis. METHODS DNA was isolated from 101 gastric specimens and 5 microsatellite loci were detected. PCR, electrophoresis on denatured polyacrylamide gels and silver staining were performed to detect the MSI. The FMs of TGFβR Ⅱ were also screened with the same method. HMLH1 methylation was analyzed by methylation specific PCR （MSP） and sequencing. HMLH1 protein expression was detected using immunohistochemistry. RESULTS The incidence of MSIs was 53.7% and 0% in the cancers and normal tissues, respectively, with the frequency of MSIs being significantly higher in the gastric cancers compared to the normal gastric tissues （P〈0.05）. The frequency of hMLH1 methylation was 41.5%（17/41） in the gastric cancers and 0%（0/60） in the normal group. Decreased hMLH1 expression was observed in 94.1%（16/17） of cases exhibiting methylation. FMs of TGFβR Ⅱ were identified in 5 （62.5%） of the 8 samples with MSIH. In contrast, FMs were not found in MSI-L or microsatellite stable （MSS） cases. CONCLUSION MSIs and FMs of TGFβR Ⅱ may play an important role in gastric carcinogenesis. HMLH1 methylation is an important modification of the DNA which results in inactivation of hMLH1 and mismatch repair defects which lead to MSls and FMs of TGFβR Ⅱ.

Transforming growth factor β receptor Ⅱ expression inexperimental cryptorchidism and apoptosisin spermatogenic cells in rats

Objective: To study the transforming growth factor β receptor Ⅱ (TGFβ-R Ⅱ) expression in experimental cryptorchidism and apoptosis in spermatogenic cells in rats. Methods: The apoptosis of spermatogenic cells was detected by means of the terminal deoxynucleotldyl transferase mediated dUTP nick end lableling method (TUNEL) and the TGFβ-R Ⅱ expression was observed with the immunohistochemistry SABC methods. Results: There was a significant increase in the TGFβ-R Ⅱ expression in unilateral undescended testes (UUTs) compared with that in contralateral descended testes (CDTs, P<0.01). However, there was a significant and time-dependent increase in the mean apoptotic index in UUTs than in CDTs. Conclusion: TGFβ-R Ⅱ may play an important role in spermatogenic cell apoptosis.

EXPRESSION OF EPIDERMAL GROWTH FACTOR, TRANSFORMINGGROWTH FACTOR-α AND THEIR RECEPTOR IN HUMAN PITUITARY TUMORS

Objective: To explore the role of growth factor autocrine stimulation in the pathogenesis of human pituitary tumors. Methods: The expression of EGF, TGF-( and EGFR were studied by immunohisto-chemical method on paraffin-embedded sections of 30 cases pituitary tumor. Results: EGFR and its ligands EGF, TGF-( expressed in majority of pituitary tumors. The expression of EGFR and its ligands varied with cells' intensity, density and type. Conclusion: The EGF autocrine stimulating exerted in the pituitary tumor development process, that tyrosine kinases inhibitors may be useful for pituitary tumors treatment.

Expression of transforming growth factor β receptors in CCI_4, -induced cirrhotic liver in rats

Objective: To investigate the expression and the role of transforming growth factor 6 (TGF-β) receptors in CCI4 --Induced cirrhotic liver. Methods: In situ hybridization was used. Results: The TGF--β type I receptors mRNA was mainly expressed in Ito cells, endothelial cells and myofibroblasts. Only a few hepatocytes expressed it. However. the TGF-β type Ⅱ receptors was mostly localized in endothelial cells and Ito cells but few hepatocytes. The expression of mRNA of both the 2 types of receptors were significantly increased in the cirrhotic liver than in the control. Conclusion: The autocrine and paracrine effects of TGF-β on matrix production and activation of Ito cells might be an important factor of fibrogenesis in CCI4 cirrhotic ever.

Expression of transforming growth factor-β_1 and its typeⅠ receptor in different phases of post-burn hypertrophic scars

Objective: To analyze and compare the expression pattern of the transforming growth factor-β1(TGF-β1) and its type I receptor (TGF-β RI ) in nounal human skin and various phases of post-burn hypertrophic scars (HTS). Method: The immunohistochemical ABC method was employed. Results: In nounal human skin, no evident immunoreactivity of TGF-β1 and TGF-β R I was observed. In activation phase of post-burn HTS, TGF-β R I and TGF-β1 were highly expressed in most dermal fibroblasts which seemed to be the same subset. However, in remission phase, no staining was seen in der mal fibroblasts. Conclusion: The formation of all may involve the increase of TGF-β responsiveness in fibroblasts The ac cumulation at the wound site and failure of apoptosis of over-resposive fibroblasts may contribute to the formation of HTS.

Ontogeny of expression of transforming growth factor-βand its receptors and their possible relationship with scarless or scar-forming healing in human fetal and postnatal skins

Fetal eutaneous wounds that oeeur in earlygestation heal without sear formation.Althoughmueh work has been done to eharaeterize the roleof transforming growth

Inhibition of Ubiquitin-specific Protease 4 Attenuates Epithelial-Mesenchymal Transition of Renal Tubular Epithelial Cells via Transforming Growth Factor Beta Receptor Type Ⅰ

Objective Ubiquitin-specific protease 4(USP4)facilitates the development of transforming growth factor-beta 1(TGF-β1)-induced epithelial-mesenchymal transition(EMT)in various cancer cells.Moreover,EMT of renal tubular epithelial cells(RTECs)is required for the progression of renal interstitial fibrosis.However,the role of USP4 in EMT of RTECs remains unknown.The present study aimed to explore the effect of USP4 on the EMT of RTECs as well as the involved mechanism.Methods In established unilateral ureteral obstruction(UUO)rats and NRK-52E cells,immunohistochemistry and Western blot assays were performed.Results USP4 expression was increased significantly with obstruction time.In NRK-52E cells stimulated by TGF-β1,USP4 expression was increased in a time-dependent manner.In addition,USP4 silencing with specific siRNA indicated that USP4 protein was suppressed effectively.Meanwhile,USP4 siRNA treatment restored E-cadherin and weakened alpha smooth muscle actin(α-SMA)expression,indicating that USP4 may promote EMT.After treatment with USP4 siRNA and TGF-β1 for 24 h,the expression of TGF-β1 receptor type I(TβRI)was decreased.Conclusion USP4 promotes the EMT of RTECs through upregulating TβRI,thereby facilitating renal interstitial fibrosis.These findings may provide a potential target of USP4 in the treatment of renal fibrosis.

Exosome-transported IncRNA H19 regulates insulin-like growth factor-1 via the H19/let-7a/insulin-like growth factor-1 receptor axis in ischemic stroke

LncRNA(long non-coding RNA) H19 is a transcript of the H19 gene that is expressed during embryogenesis.We previously discove red a role for circular lncRNA H19 in the onset and prognosis of cerebral ischemic stroke.In this study,we used serum from patients with ischemic stroke,and mouse and cell culture models to elucidate the roles of plasma and neuronal exosomes in the regulatory effect of lncRNA H19 on insulin-like growth factor-1 and its mechanism in ischemic stroke,using western blotting,quantitative real-time polymerase chain reaction,and enzyme-linked immunosorbent assays.Plasma exosomal IncRNA H19 was negatively associated with blood levels of insulin-like growth factor-1 in samples from patients with cerebral ischemic stroke.In a mouse model,levels of exosomal IncRNA H19 were positively correlated with plasma and cerebral lncRNA H19.In a cell co-culture model,we confirmed that IncRNA H19 was transported from neuro ns to astrocytes by exosomes to induce downregulation of insulin-like growth factor-1 through the H19/let-7 a/insulin-like growth factor-1 receptor axis.This study provides the first evidence for the transpo rtation of IncRNA H19 by exosomes and the relationship between IncRNA H19 and insulinlike growth factor-1.

Organ fibrosis inhibited by blocking transforming growth factor-β signaling via peroxisome proliferator-activated receptor γ agonists

BACKGROUND:Organ fibrosis has been viewed as one of the major medical problems, which can lead to progressive dysfunction of the liver, lung, kidney, skin, heart, and eventually death of patients. Fibrosis is initiated by a variety of pathological, physiological, biochemical, and physical factors. Regardless of their different etiologies, they all share a common pathogenetic process: excessive activation of the key profibrotic cytokine, transforming growth factor-β (TGF-β). Peroxisome proliferator-activated receptor γ (PPARγ), a ligand-activated transcription factor of the nuclear receptor superfamily, has received particular attention in recent years, because the activation of PPARγ by both natural and synthetic agonists could effectively inhibit TGF-β-induced profibrotic effects in many organs. DATA SOURCES: The English-language medical databases, PubMed, Elsevier and SpringerLink were searched for articles on PPARγ, TGF-β, and fibrosis, and related topics. RESULTS: TGF-β is recognized as a key profibrotic cytokine. Excessive activation of TGF-β increases synthesis of extracellular matrix proteins and decreases their degradation, associated with a gradual destruction of normal tissue architecture and function, whereas PPARγ agonists inhibit TGF-β signal transduction and are effective antifibrogenic agents in many organs including the liver, lung, kidney, skin and heart. CONCLUSIONS: The main antifibrotic activity of PPARγ agonists is to suppress the TGF-β signaling pathway by so-called PPARγ-dependent effect. In addition, PPARγ agonists, especially 15d-PGJ2, also exert potentially antifibrotic activity independent of PPARγ activation. TGF-β1/Smads signaling not only plays many essential roles in multiple developmental processes, butalso forms cross-talk networks with other signal pathways, and their inhibition by PPARγ agonists certainly affects the cytokine networks and causes non-suspected side-effects. Anti-TGF-β therapies with PPARγ agonists may have to be carefully tailored to be tissue-and target gene-specific to minimize side-effects, indicating a great challenge to the medical research at present.