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This article reports on two cases of testicular sclerosing stromal tumors (SSTs) that show GLI1 gene alterations. SSTs are neoplasms that were first described in the ovary in 1973 and are characterized by lobulated microscopic appearance. The patients in these cases were aged 35 and 65 years and presented with testicular masses. The tumors demonstrated large areas of hypocellular myxoid/oedematous stroma with scattered hypercellular areas. Both tumors expressed nuclear GLI1. The article suggests that these findings support the existence of rare SSTs in the testis and their potential pathogenesis related to ovarian SSTs. [Extracted from the article]

Pancreatic cancer (PC) is an aggressive and metastatic gastrointestinal tumor with a poor prognosis. Persistent activation of the TGF-β/Smad signaling induces PC cell (PCC) invasion and infiltration via epithelial-to-mesenchymal transition (EMT). Hedgehog signaling is a crucial pathway for the development of PC via the transcription factors Gli1/2/3. This study aimed to investigate the underlying molecular mechanisms of action of hedgehog activation in TGF-β1-triggered EMT in PCCs (PANC-1 and BxPc-3). In addition, overexpression and shRNA techniques were used to evaluate the role of Smad4 in TGF-β1-treated PCCs. Our data showed that TGF-β1 promoted PCC invasion and infiltration via Smad2/3-dependent EMT. Hedgehog-Gli signaling axis in PCCs was activated upon TGF-β1 stimulation. Inhibition of hedgehog with cyclopamine effectively antagonized TGF-β1-induced EMT, thereby suggesting that the hedgehog signaling may act as a downstream cascade signaling of TGF-β1. As a key protein that assists the nuclear translocation of Smad2/3, Smad4 was highly expressed in PANC-1 cells, but not in BxPc-3 cells. Conversely, Gli1 expression was low in PANC-1 cells, but high in BxPc-3 cells. Furthermore, knockdown of Smad4 in PANC-1 cells by shRNA inhibited TGF-β1-mediated EMT and collagen deposition. Overexpression of Smad4 did not affect TGF-β1-mediated EMT due to the lack of significant increase in nuclear expression of Smad4. Importantly, Gli1 activity was upregulated by Smad4 knockdown in PANC-1 cells and downregulated by Smad4 overexpression in BxPc-3 cells, indicating that Gli1 may be a negative target protein downstream of Smad4. Thus, Smad4 regulates TGF-β1-mediated hedgehog activation to promote EMT in PCCs by suppressing Gli1 activity.

Bone tissue provides structural support for our bodies, with the inner bone marrow (BM) acting as a hematopoietic organ. Within the BM tissue, two types of stem cells play crucial roles: mesenchymal stem cells (MSCs) (or skeletal stem cells) and hematopoietic stem cells (HSCs). These stem cells are intricately connected, where BM-MSCs give rise to bone-forming osteoblasts and serve as essential components in the BM microenvironment for sustaining HSCs. Despite the mid-20th century proposal of BM-MSCs, their in vivo identification remained elusive owing to a lack of tools for analyzing stemness, specifically self-renewal and multipotency. To address this challenge, Cre/loxP-based cell lineage tracing analyses are being employed. This technology facilitated the in vivo labeling of specific cells, enabling the tracking of their lineage, determining their stemness, and providing a deeper understanding of the in vivo dynamics governing stem cell populations responsible for maintaining hard tissues. This review delves into cell lineage tracing studies conducted using commonly employed genetically modified mice expressing Cre under the influence of LepR, Gli1, and Axin2 genes. These studies focus on research fields spanning long bones and oral/maxillofacial hard tissues, offering insights into the in vivo dynamics of stem cell populations crucial for hard tissue homeostasis.

Abstract Background Melanoma progression is based on a close interaction between cancer cells and immune cells in the tumor microenvironment (TME). Thus, a better understanding of the mechanisms controlling TME dynamics and composition will help improve the management of this dismal disease. Work from our and other groups has reported the requirement of an active Hedgehog-GLI (HH-GLI) signaling for melanoma growth and stemness. However, the role of the downstream GLI1 transcription factor in melanoma TME remains largely unexplored. Methods The immune-modulatory activity of GLI1 was evaluated in a syngeneic B16F10 melanoma mouse model assessing immune populations by flow cytometry. Murine polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) were differentiated from bone marrow cells and their immunosuppressive ability was assessed by inhibition of T cells. Conditioned media (CM) from GLI1-overexpressing mouse melanoma cells was used to culture PMN-MDSCs, and the effects of CM were evaluated by Transwell invasion assay and T cell inhibition. Cytokine array analysis, qPCR and chromatin immunoprecipitation were performed to explore the regulation of CX3CL1 expression by GLI1. Human monocyte-derived dendritic cells (moDCs) were cultured in CM from GLI1-silenced patient-derived melanoma cells to assess their activation and recruitment. Blocking antibodies anti-CX3CL1, anti-CCL7 and anti-CXCL8 were used for in vitro functional assays. Results Melanoma cell-intrinsic activation of GLI1 promotes changes in the infiltration of immune cells, leading to accumulation of immunosuppressive PMN-MDSCs and regulatory T cells, and to decreased infiltration of dendric cells (DCs), CD8 + and CD4 + T cells in the TME. In addition, we show that ectopic expression of GLI1 in melanoma cells enables PMN-MDSC expansion and recruitment, and increases their ability to inhibit T cells. The chemokine CX3CL1, a direct transcriptional target of GLI1, contributes to PMN-MDSC expansion and recruitment. Finally, silencing of GLI1 in patient-derived melanoma cells promotes the activation of human monocyte-derived dendritic cells (moDCs), increasing cytoskeleton remodeling and invasion ability. This phenotype is partially prevented by blocking the chemokine CCL7, but not CXCL8. Conclusion Our findings highlight the relevance of tumor-derived GLI1 in promoting an immune-suppressive TME, which allows melanoma cells to evade the immune system, and pave the way for the design of new combination treatments targeting GLI1.

Hypertrophy of ligamentum flavum (LF) is a significant contributing factor to lumbar spinal canal stenosis (LSCS). lncRNA plays a vital role in organ fibrosis, but its role in LF fibrosis remains unclear. Our previous findings have demonstrated that Hedgehog- Gli1 signaling is a critical driver leading to LF hypertrophy. Through the RIP experiment, our group found lnc-RMRP was physically associated with Gli1 and exhibited enrichment in Gli1-activated LF cells. Histological studies revealed elevated expression of RMRP in hypertrophic LF. In vitro experiments further confirmed that RMRP promoted Gli1 SUMO modification and nucleus transfer. Mechanistically, RMRP induced GSDMD-mediated pyroptosis, proinflammatory activation, and collagen expression through the Hedgehog pathway. Notably, the mechanical stress-induced hypertrophy of LF in rabbit exhibited analogous pathological changes of LF fibrosis occurred in human and showed enhanced levels of collagen and α-SMA. Knockdown of RMRP resulted in the decreased expression of fibrosis and pyroptosis-related proteins, ultimately ameliorating fibrosis. The above data concluded that RMRP exerts a crucial role in regulating GSDMD-mediated pyroptosis of LF cells via Gli1 SUMOylation, thus indicating that targeting RMRP could serve as a potential and effective therapeutic strategy for LF hypertrophy and fibrosis. [ABSTRACT FROM AUTHOR]

Background: Melanoma progression is based on a close interaction between cancer cells and immune cells in the tumor microenvironment (TME). Thus, a better understanding of the mechanisms controlling TME dynamics and composition will help improve the management of this dismal disease. Work from our and other groups has reported the requirement of an active Hedgehog-GLI (HH-GLI) signaling for melanoma growth and stemness. However, the role of the downstream GLI1 transcription factor in melanoma TME remains largely unexplored. Methods: The immune-modulatory activity of GLI1 was evaluated in a syngeneic B16F10 melanoma mouse model assessing immune populations by flow cytometry. Murine polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) were differentiated from bone marrow cells and their immunosuppressive ability was assessed by inhibition of T cells. Conditioned media (CM) from GLI1-overexpressing mouse melanoma cells was used to culture PMN-MDSCs, and the effects of CM were evaluated by Transwell invasion assay and T cell inhibition. Cytokine array analysis, qPCR and chromatin immunoprecipitation were performed to explore the regulation of CX3CL1 expression by GLI1. Human monocyte-derived dendritic cells (moDCs) were cultured in CM from GLI1-silenced patient-derived melanoma cells to assess their activation and recruitment. Blocking antibodies anti-CX3CL1, anti-CCL7 and anti-CXCL8 were used for in vitro functional assays. Results: Melanoma cell-intrinsic activation of GLI1 promotes changes in the infiltration of immune cells, leading to accumulation of immunosuppressive PMN-MDSCs and regulatory T cells, and to decreased infiltration of dendric cells (DCs), CD8 + and CD4 + T cells in the TME. In addition, we show that ectopic expression of GLI1 in melanoma cells enables PMN-MDSC expansion and recruitment, and increases their ability to inhibit T cells. The chemokine CX3CL1, a direct transcriptional target of GLI1, contributes to PMN-MDSC expansion and recruitment. Finally, silencing of GLI1 in patient-derived melanoma cells promotes the activation of human monocyte-derived dendritic cells (moDCs), increasing cytoskeleton remodeling and invasion ability. This phenotype is partially prevented by blocking the chemokine CCL7, but not CXCL8. Conclusion: Our findings highlight the relevance of tumor-derived GLI1 in promoting an immune-suppressive TME, which allows melanoma cells to evade the immune system, and pave the way for the design of new combination treatments targeting GLI1. [ABSTRACT FROM AUTHOR]

Synovial sarcoma (SS) is a rare soft-tissue tumor characterized by a monomorphic blue spindle cell histology and variable epithelial differentiation. Morphologically, SSs may be confused with other sarcomas. Systemic treatment is more effective for patients with high-risk SSs, patients with advanced disease, and younger patients. However, further studies are required to find new prognostic biomarkers. Herein, we describe the morphological, molecular, and clinical findings, using a wide immunohistochemical panel, of a series of SS cases. We studied 52 cases confirmed as SSs by morphological diagnosis and/or molecular studies. Clinical data (gender, age, tumor size, tumor location, resection margins, adjuvant treatment, recurrences, metastasis, and survival) were also retrieved for each patient. All the available H&E slides were examined by four pathologists. Three tissue microarrays (TMAs) were constructed for each of the tumors, and a wide immunohistochemical panel was performed. For time-to-event variables, survival analysis was performed using Kaplan–Meier curves and log-rank testing, or Cox regression. Statistical significance was considered at p < 0.05. The mean age of our patients was 40.33, and the median was 40.5 years. We found a predominance of males versus females (1.7:1). The most frequent morphological subtype was monophasic. TRPS1, SS18-SSX, and SSX-C-terminus were positive in 96% of cases. GLI1 expression was strong in six and focal (cytoplasmic) in twenty patients. Moreover, BCOR was expressed in more than half of SSs. Positive expression of both proteins, BCOR and GLI1, was correlated with a worse prognosis. Multivariate analysis was also performed, but only BCOR expression appeared to be significant. The combination of GLI1 and BCOR antibodies can be used to group SSs into three risk groups (low, intermediate, and high risk). We hypothesize that these findings could identify which patients would benefit from receiving adjuvant treatment and which would not. Moreover, these markers could represent therapeutic targets in advanced stages. However, further, larger series of SSs and molecular studies are necessary to corroborate our present findings. [ABSTRACT FROM AUTHOR]

Aberrant activation of hedgehog (Hh) signaling has been implicated in various cancers. Current FDA-approved inhibitors target the seven-transmembrane receptor Smoothened, but resistance to these drugs has been observed. It has been proposed that a more promising strategy to target this pathway is at the GLI1 transcription factor level. GANT61 was the first small molecule identified to directly suppress GLI-mediated activity; however, its development as a potential anticancer agent has been hindered by its modest activity and aqueous chemical instability. Our study aimed to identify novel GLI1 inhibitors. JChem searches identified fifty-two compounds similar to GANT61 and its active metabolite, GANT61-D. We combined high-throughput cell-based assays and molecular docking to evaluate these analogs. Five of the fifty-two GANT61 analogs inhibited activity in Hh-responsive C3H10T1/2 and Gli-reporter NIH3T3 cellular assays without cytotoxicity. Two of the GANT61 analogs, BAS 07019774 and Z27610715, reduced Gli1 mRNA expression in C3H10T1/2 cells. Treatment with BAS 07019774 significantly reduced cell viability in Hh-dependent glioblastoma and lung cancer cell lines. Molecular docking indicated that BAS 07019774 is predicted to bind to the ZF4 region of GLI1, potentially interfering with its ability to bind DNA. Our findings show promise in developing more effective and potent GLI inhibitors. [ABSTRACT FROM AUTHOR]

GLI1, a key transcription factor of the Hedgehog (Hh) signaling pathway, plays an important role in the development of cancer. However, the function and mechanisms by which GLI1 regulates gene transcription are not fully understood in gastric cancer (GC). Here, we found that GLI1 induced the proliferation and metastasis of GC cells, accompanied by transcriptional upregulation of INHBA. This increased INHBA expression exerted a promoting activity on Smads signaling and then transcriptionally activated GLI1 expression. Notably, our results demonstrate that disrupting the interaction between GLI1 and INHBA could inhibit GC tumorigenesis in vivo. More intriguingly, we confirmed the N6‐methyladenosine (m6A) activation mechanism of the Helicobacter pylori/FTO/YTHDF2/GLI1 pathway in GC cells. In conclusion, our study confirmed that the GLI1/INHBA positive feedback loop influences GC progression and revealed the mechanism by which H. pylori upregulates GLI1 expression through m6A modification. This positive GLI1/INHBA feedback loop suggests a novel noncanonical mechanism of GLI1 activity in GC and provides potential therapeutic targets for GC treatment. [ABSTRACT FROM AUTHOR]

The Hedgehog (Hh) signaling pathway orchestrates its influence through a dynamic interplay of Hh proteins, the cell surface receptor Ptch1, Smo, and Gli transcription factors, contributing to a myriad of developmental events. Indian Hedgehog (Ihh) and Gli zinc finger transcription factor 1 (Gli1) play crucial roles in developmental regulation within the Hh signaling pathway. Ihh regulates chondrocyte proliferation, differentiation, and bone formation, impacting the development of cranial bones, cartilage, and the temporomandibular joint (TMJ). Losing Ihh results in cranial bone malformation and decreased ossification and affects the formation of cranial base cartilage unions, TMJ condyles, and joint discs. Gli1 is predominantly expressed during early craniofacial development, and Gli1+ cells are identified as the primary mesenchymal stem cells (MSCs) for craniofacial bones, crucial for cell differentiation and morphogenesis. In addition, a complex mutual regulatory mechanism exists between Gli1 and Ihh, ensuring the normal function of the Hh signaling pathway by directly or indirectly regulating each other's expression levels. And the interaction between Ihh and Gli1 significantly impacts the normal development of craniofacial tissues. This review summarizes the pivotal roles of Gli1 and Ihh in the intricate landscape of mammalian craniofacial development and outlines the molecular regulatory mechanisms and intricate interactions governing the growth of bone and cartilage exhibited by Gli1 and Ihh, which provides new insights into potential therapeutic strategies for related diseases or researches of tissue regeneration. [ABSTRACT FROM AUTHOR]

Background: One of the most prevalent cancers worldwide is HCC, which has put patient health at risk. Increasing evidence indicated that messenger RNAs (mRNAs) played significant roles in modulating tumorigenesis. It has been established that Gli1 acts as an oncogene in a number of malignancies. However, more research was necessary to understand the Gli1 regulation mechanism in HCC. Methods: Microarray technology was used to evaluate the expression of mRNAs. RT-qPCR was utilized to evaluate Gli1 and miR-875-5p expression. To investigate the role of Gli1, tests using CCK-8, EdU, transwell, immunofluorescence, and Western blot analysis was performed. RIP, RNA pull down, and luciferase reporter assays were employed to verify the interaction between Gli1 and miR-875-5p. Results: In tissues and cells of HCC, Gli1 expression appeared to be upregulated, especially in metastatic samples and advanced stages of the disease. A worse outcome was predicted by elevated Gli1 expression. Additionally, in HCC, Gli1 inhibition impeded the growth, migration, and development of the EMT. Since miR-875-5p was shown to have a molecular target in Gli1, miR-875-5p mediated the negative regulation of Gli1. In HCC tissues, its expression pattern was less prominent. In HCC tissues, there was an inverse relationship between Gli1 expression and miR-875-5p expression. Overexpressing Gli1 helped to partially counteract the suppression of HCC migration, proliferation, and EMT formation by miR-875-5p overexpression. Conclusions: MiR-875-5p in HCC suppresses tumors by downregulating Gli1, which supplies a novel treatment for HCC patients.

Objective To explore the effect of Sufu on lung adenocarcinoma by observing its mutation and the changes in Gli1 transcription after its mutation. Methods cBioPortal for Cancer Genomics was used to obtain the mutation status of Sufu in lung cancer. Four Sufu single base mutation vectors were constructed and transfected into lung adenocarcinoma cell lines A549 and H1975 to establish cells with Sufu overexpression and knockout. Wounding healing assay was employed to determine the effect of Sufu on cell migration, and RT-qPCR, immunofluorescence assay and dual luciferase reporter gene assay were utilized to explore the changes in Gli1 transcription after Sufu mutation. Results cBioPortal data showed that the mutation rate of Sufu in lung cancer was 11.76% in squamous cell carcinoma (2/17), 0.96% in adenocarcinoma (6/625), 1.01% in small cell lung cancer (4/396), and 0.69% in non-small cell lung cancer (77/11 227). The median survival of lung cancer patients was significantly decreased in those with Sufu mutation than those without (38.00 vs 54.34 months, HR=1.943, P < 0.05). A549 cells were sensitive to Sufu knockout, and it resulted in significant increase of the transcription level of Gli1 (P < 0.05). Overexpression of Sufu inhibited cell migration ability in both A549 and H1975 cells (P < 0.05). Sufu mutation had no significant influence on the location of Gli1 in the cells. The Sufu-T411M mutation up-regulated the the transcription level of Gli1 (P < 0.05). Conclusion Sufu has an inhibitory effect on lung adenocarcinoma metastasis, and can be regarded as a potential target for lung adenocarcinoma metastasis.

Abstract Background Tumor angiogenesis inhibitors have been applied for non-small cell lung cancer (NSCLC) therapy. However, the drug resistance hinders their further development. Intercellular crosstalk between lung cancer cells and vascular cells was crucial for anti-angiogenenic resistance (AAD). However, the understanding of this crosstalk is still rudimentary. Our previous study showed that Glioma-associated oncogene 1 (Gli1) is a driver of NSCLC metastasis, but its role in lung cancer cell-vascular cell crosstalk remains unclear. Methods Conditioned medium (CM) from Gli1-overexpressing or Gli1-knockdown NSCLC cells was used to educate endothelia cells and pericytes, and the effects of these media on angiogenesis and the maturation of new blood vessels were evaluated via wound healing assays, Transwell migration and invasion assays, tube formation assays and 3D coculture assays. The xenograft model was conducted to establish the effect of Gli1 on tumor angiogenesis and growth. Angiogenic antibody microarray analysis, ELISA, luciferase reporte, chromatin immunoprecipitation (ChIP), bFGF protein stability and ubiquitination assay were performed to explore how Gli1 regulate bFGF expression. Results Gli1 overexpression in NSCLC cells enhanced the endothelial cell and pericyte motility required for angiogenesis required for angiogenesis. However, Gli1 knockout in NSCLC cells had opposite effect on this process. bFGF was critical for the enhancement effect on tumor angiogenesis. bFGF treatment reversed the Gli1 knockdown-mediated inhibition of angiogenesis. Mechanistically, Gli1 increased the bFGF protein level by promoting bFGF transcriptional activity and protein stability. Importantly, suppressing Gli1 with GANT-61 obviously inhibited angiogenesis. Conclusion The Gli1-bFGF axis is crucial for the crosstalk between lung cancer cells and vascular cells. Targeting Gli1 is a potential therapeutic approach for NSCLC angiogenesis.

Objective: Gastroblastoma is an extremely rare gastric tumor. Its pathogenesis remains unclear and there is a lack of specific clinical symptoms. The aim of this paper is to report a case of gastroblastoma and provide references for the diagnosis, treatment, and prognosis of this disease. Methods: The diagnosis and treatment of a 51-year-old female patient with gastroblastoma were retrospectively reported. Analyzing this case by combining the clinical data such as imaging and pathological results of patients with the relevant literature. Results: The patient's chief complaint was the presence of melena persisted for over two weeks. Abdominal contrast-enhanced CT showed gastric antral nodules, and micro-probe endoscopic ultrasonography was considered as "gastric antral protruding lesions". The initial diagnosis of "gastric stromal tumor" was made after admission, and surgical treatment was performed on September 23, 2021. Postoperative pathology showed that gastric mixed epithelial and stromal tumor, combined with immunohistochemical staining, was suggestive of gastroblastoma. No signs of tumor recurrence or metastasis were observed during the 2-year follow-up. Conclusion: Combined with the existing literature reports, the incidence of gastroblastoma is mainly higher in young men, and the predilection site is gastric antrum. The biological behavior of the tumor tends to be indolent, and the prognosis of most cases is favorable. However, due to the extremely small number of cases, this conclusion still needs a large number of cases and followup data to support. Postoperative pathological and immunohistochemical examination results are the only methods for definite diagnosis at present, and surgery is the first choice for treatment. [ABSTRACT FROM AUTHOR]

The development of musculoskeletal tissues such as tendon, enthesis, and bone relies on proliferation and differentiation of mesenchymal progenitor cells. Gli1+ cells have been described as putative stem cells in several tissues and are presumed to play critical roles in tissue formation and maintenance. For example, the enthesis, a fibrocartilage tissue that connects tendon to bone, is mineralized postnatally by a pool of Gli1+ progenitor cells. These cells are regulated by hedgehog signaling, but it is unclear if TGFβ signaling, necessary for tenogenesis, also plays a role in their behavior. To examine the role of TGFβ signaling in Gli1+ cell function, the receptor for TGFβ, TbR2, was deleted in Gli1‐lineage cells in mice at P5. Decreased TGFβ signaling in these cells led to defects in tendon enthesis formation by P56, including defective bone morphometry underlying the enthesis and decreased mechanical properties. Immunohistochemical staining of these Gli1+ cells showed that loss of TGFβ signaling reduced proliferation and increased apoptosis. In vitro experiments using Gli1+ cells isolated from mouse tail tendons demonstrated that TGFβ controls cell proliferation and differentiation through canonical and non‐canonical pathways and that TGFβ directly controls the tendon transcription factor scleraxis by binding to its distant enhancer. These results have implications in the development of treatments for tendon and enthesis pathologies. [ABSTRACT FROM AUTHOR]

Background: Tumor angiogenesis inhibitors have been applied for non-small cell lung cancer (NSCLC) therapy. However, the drug resistance hinders their further development. Intercellular crosstalk between lung cancer cells and vascular cells was crucial for anti-angiogenenic resistance (AAD). However, the understanding of this crosstalk is still rudimentary. Our previous study showed that Glioma-associated oncogene 1 (Gli1) is a driver of NSCLC metastasis, but its role in lung cancer cell-vascular cell crosstalk remains unclear. Methods: Conditioned medium (CM) from Gli1-overexpressing or Gli1-knockdown NSCLC cells was used to educate endothelia cells and pericytes, and the effects of these media on angiogenesis and the maturation of new blood vessels were evaluated via wound healing assays, Transwell migration and invasion assays, tube formation assays and 3D coculture assays. The xenograft model was conducted to establish the effect of Gli1 on tumor angiogenesis and growth. Angiogenic antibody microarray analysis, ELISA, luciferase reporte, chromatin immunoprecipitation (ChIP), bFGF protein stability and ubiquitination assay were performed to explore how Gli1 regulate bFGF expression. Results: Gli1 overexpression in NSCLC cells enhanced the endothelial cell and pericyte motility required for angiogenesis required for angiogenesis. However, Gli1 knockout in NSCLC cells had opposite effect on this process. bFGF was critical for the enhancement effect on tumor angiogenesis. bFGF treatment reversed the Gli1 knockdown-mediated inhibition of angiogenesis. Mechanistically, Gli1 increased the bFGF protein level by promoting bFGF transcriptional activity and protein stability. Importantly, suppressing Gli1 with GANT-61 obviously inhibited angiogenesis. Conclusion: The Gli1-bFGF axis is crucial for the crosstalk between lung cancer cells and vascular cells. Targeting Gli1 is a potential therapeutic approach for NSCLC angiogenesis. [ABSTRACT FROM AUTHOR]

This article discusses a rare neoplasm called GLI1-rearranged gastric tumor or gastroblastoma. The article highlights the association of somatic molecular events involving the GLI1 gene with indolent soft-tissue tumors and tumors of the stomach wall. The article presents a case report of a 28-year-old man with a GLI1-rearranged gastric tumor and discusses the long-term outcomes of previously reported GLI1-rearranged enteric tumors. The article emphasizes the need for accurate classification and complete surgical resection of these rare tumors, as well as further research to clarify their metastatic potential. [Extracted from the article]

Prostate cancer (PCa) is the most prevalent cause of death in the male population worldwide. The G Protein-Coupled Estrogen Receptor (GPER) has been gaining relevance in the development of PCa. Hedgehog (Hh) pathway activation is associated with aggressiveness, metastasis, and relapse in PCa patients. To date, no studies have evaluated the crosstalk between the GPER and the Hh pathway along different group grades in PCa. We conducted an analysis of paraffinembedded tissues derived from patients with different prognostic grade of PCa using immunohistochemistry. Expression and correlation between GPER and glioma associated oncogene homologue (GLI) transcriptional factors in the parenchyma and stroma of PCa tumors were evaluated. Our results indicate that GPER is highly expressed in the nucleus and increases with higher grade groups. Additionally, GPER's expression correlates with pGLI3 nuclear expression across different grade groups in PCa tissues; however, whether the receptor induces the activation of GLI transcriptional factors, or the latter modulate the expression of GPER is yet to be discovered, as well as the functional consequence of this correlation. [ABSTRACT FROM AUTHOR]

In this study, we investigated the effect of cycloastragenol (CAG), a triterpenoid isolated from Astragalus membranaceus roots, on regulating the adipogenesis and fat accumulation in vitro and in vivo. During the adipogenesis of 3T3-L1 cells, CAG inhibited lipid accumulation and the expression of key adipogenic factors, proliferator-activated receptor γ (PPARγ) and CCAAT enhancer binding protein α (C/EBPα) and increased the expression of Gli1, a key mediator in Hedgehog (Hh) signaling. In HFD-induced animal experiment, CAG significantly reduced body weight gain without affecting brown fat weight. In addition, CAG regulated the expression of PPARγ, C/EBPα, and Gli1 in visceral white adipose tissue (vWAT). We also confirmed the inhibitory effect of CAG on specifically targeting white adipose tissue (WAT) formation in stromal vascular fraction (SVF) cell differentiation. Taken together, these results suggest that CAG may be a potent phytochemical preventing adipogenesis and obesity via Hh signaling. [ABSTRACT FROM AUTHOR]

Constitutive activation of Hedgehog (Hh) signaling has been implicated in many cancers including hepatocellular carcinoma (HCC). Among them, the terminal glioma-associated oncogene homolog 1 (Gli1) regulates the expression of critical genes in the Hh pathway. The current study aims to evaluate the anti-HCC effect of the Gli1 inhibitor, GANT61. In vitro analysis including cell counting kit-8 (CCK-8) assay, flow cytometry, and migration and invasion assay were adopted to evaluate the effect of GANT61 on HCC cell lines. In vivo, xenograft studies were also performed to verify the effect of GANT61 on HCC. By CCK-8 assay, we found that GANT61 could significantly reduce the growth of HCC cell lines Huh7 and hemophagocytic lymphohistiocytosis (HLE), and their IC50 concentrations were 4.481 and 6.734 μM, respectively. Flow cytometry shows that GANT61 induced cell cycle arrest in the G2/M phase and accelerated apoptosis of both HLE and Huh7 cells. While migration and invasion assay shows that GANT61 weakens cells' migration and invasion ability. Besides that, GANT61 inhibits the expression of Gli1, FoxM1, CyclinD1, and Bcl-2, upregulates the level of Bax protein, and also reverses the epithelial–mesenchymal transition program by downregulating the expression of Vimentin and N-Cadherin and upregulating the expression of epithelial E-Cadherin expression. Furthermore, GANT61 inhibits the growth of subcutaneous xenografts of Huh7 cells in nude mice. Overall, this study suggests that Gli1 is a potential target for therapy and GANT61 shows promising therapeutic potential for future treatment in HCC. [ABSTRACT FROM AUTHOR]

Simple Summary: The chromosomal region 12q13-15 is rich in oncogenes (MDM2, CDK4, STAT6, DDIT3, and GLI1). Amplification of MDM2 and CDK4 genes can be detected in various mesenchymal and nonmesenchymal neoplasms. Therefore, gene amplification alone is not entirely specific for making a definitive diagnosis and requires the integration of clinical, radiological, morphological, and immunohistochemical findings. Despite the diagnostic implications that the overlap of genetic alterations in neoplasms with changes in genes within the 12q13-15 region could create, the discovery of coamplifications of MDM2 with CDK4 and GLI1 offers new therapeutic targets in neoplasms with MDM2/CDK4 amplification. In this review, we delve into the diagnosis and therapeutic implications of neoplasms with genetic alterations involving the chromosomal region 12q13-15, mainly MDM2, CDK4, and GLI1. The chromosomal region 12q13-15 is rich in oncogenes and contains several genes involved in the pathogenesis of various mesenchymal neoplasms. Notable genes in this region include MDM2, CDK4, STAT6, DDIT3, and GLI1. Amplification of MDM2 and CDK4 genes can be detected in various mesenchymal and nonmesenchymal neoplasms. Therefore, gene amplification alone is not entirely specific for making a definitive diagnosis and requires the integration of clinical, radiological, morphological, and immunohistochemical findings. Neoplasms with GLI1 alterations may exhibit either GLI1 rearrangements or amplifications of this gene. Despite the diagnostic implications that the overlap of genetic alterations in neoplasms with changes in genes within the 12q13-15 region could create, the discovery of coamplifications of MDM2 with CDK4 and GLI1 offers new therapeutic targets in neoplasms with MDM2/CDK4 amplification. Lastly, it is worth noting that MDM2 or CDK4 amplification is not exclusive to mesenchymal neoplasms; this genetic alteration has also been observed in other epithelial neoplasms or melanomas. This suggests the potential use of MDM2 or CDK4 inhibitors in neoplasms where alterations in these genes do not aid the pathological diagnosis but may help identify potential therapeutic targets. In this review, we delve into the diagnosis and therapeutic implications of tumors with genetic alterations involving the chromosomal region 12q13-15, mainly MDM2, CDK4, and GLI1. [ABSTRACT FROM AUTHOR]

Meningioma is a common brain tumour which has neither a specific detection nor treatment method. The Sonic hedgehog (Shh) cell signaling pathway is a crucial regulatory pathway of mammalian organogenesis and tumorigenesis including meningioma. Shh cell signalling pathway cascade function by main transcription factor Gli1 and which further regulates in its downstream to Pax6 and Nkx2.2. This current study is aimed to explore the regulation of the Sonic hedgehog-Gli1 cell signaling pathway and its potential downstream targets in meningioma samples. A total of 24 surgically resected meningioma samples were used in this current study.Cytological changes were assessed using electron microscopic techniques as well as hematoxylin & eosin and DAPI staining. The expression pattern of Gli1, Nkx2.2 and Pax6 transcription factors were determined by using immunohistochemistry. The mRNA expression was assessed using RT-qPCR assays. Later, the whole transcriptome analysis of samples was performed with the amploseq technique. Results were compared with those obtained in normal human brain tissue (or normal meninges). Compared to the normal human brain tissue, meningioma samples showed crowded nuclei with morphological changes. Transcription factor Nkx2.2 expressed highly in all samples (24/24, 100%). Twenty-one of the 24 meningiomas (88%) showed high Gli1 and Pax6 expression. Whole transcriptome analysis of two meningioma samples also exhibited a very high increase in Gli1 expression signal in meningioma samples as compare to normal control. Hence, we may conclude that the Shh–Gli1 pathway is aberrantly activated in meningioma cells and is canonically upregulating the expression of transcription factors Pax6 and Nkx2.2. [ABSTRACT FROM AUTHOR]

Gastroblastoma is an extremely rare biphasic tumor that typically occurs in the stomach in patients between the ages of 10 and 30. Only 16 cases have been reported previously. These tumors are important to diagnose and distinguish from more aggressive neoplasms; although numbers are small, prognosis appears excellent overall with complete excision, with only occasional metastasis and/or local recurrence. We report a case of gastroblastoma in a 26-year-old male arising from the pylorus and extending through the first and second portions of the duodenum. This is the first case to be reported from this specific location. [ABSTRACT FROM AUTHOR]

Esophageal squamous cell carcinoma (ESCC) is a frequently seen esophageal tumor type in China. Activation of signaling proteins and relevant molecular mechanisms in ESCC are partially explored, impairing the antitumor efficiency of targeted therapy in ESCC treatment. Tumor‐associated macrophages (TAMs)‐released C‐C motif chemokine 22 (CCL22) can activate intratumoral focal adhesion kinase (FAK), thus promoting the progression of ESCC. Here, we demonstrated that highly secreted CCL22 by TAMs (CCL22‐positive TAMs) induced ESCC cell stemness and invasion through facilitating transcriptional activity of intratumoral glioma‐associated oncogene 1 (Gli1), a downstream effector for Hedgehog (HH) pathway. Mechanistically, FAK‐activated protein kinase B (AKT) mediated Gli1 phosphorylation at its Ser112/Thr115/Ser116 sites and released Gli1 from suppressor of fused homolog, the endogenous inhibitor of Gli1 to activate downstream stemness‐associated factors, such as SRY‐box transcription factor 2 (SOX2), Nanog homeobox (Nanog), or POU class 5 homeobox (OCT4). Furthermore, inhibition of FAK activity by VS‐4718, the FAK inhibitor, enhanced antitumor effect of GDC‐0449, the HH inhibitor, both in xenografted models and in vitro assays. Clinically, CCL22/Gli1 axis is used to evaluate ESCC prognosis. Overall, our study establishes the communication of FAK with HH pathway and offers the novel mechanism related to Gli1 activation independent of Smoothened as well as the rationale for the anti‐ESCC combination treatment. [ABSTRACT FROM AUTHOR]

Abstract Background Regulator of chromosome condensation 2 (RCC2) was a telophase disk-binding protein on mitosis, and functions as an oncogene in many human cancers. However, its role on prostate cancer (PCa) was unknown. The goal of this study is to explore the function of RCC 2 on PCa development. Methods The expression of RCC2 and its methylation level, its correlation with lymph node metastasis or disease-free survival (DFS) was analyzed using TCGA database. The effect of RCC2 on PCa cell proliferation, migration and invasion were detected using CCK-8, cell colony formation, Transwell and wood healing assays. RNA-seq and GSEA analysis were used to search the downstream genes and pathways of RCC2 in mediated PCa progression. Western blot was used to detect the proteins in PCa cells transfected with indicated siRNAs or plasmids. Results RCC2 had high expression and low promoter methylation level in PCa, and its expression was correlated with regional node metastasis and disease-free survival. Cell proliferation, migration, invasion and EMT of PCa cells in vitro were greatly enhanced after RCC2 overexpression, while the RCC2 knockdown suppressed these processes. RNA-seq and GSEA results showed the Hedgehog signaling regulator Gli1 and Gli3 were involved in RCC2 knockdown DU145 cells. Gli1 was also a marker of cancer stem-like cells (CSCs). Mechanistically, RCC2 induced cell growth, EMT, CSCs markers through Gli1; inhibiting Gli1 expression using siGli1 or GLI inhibitor suppressed cell progression in vitro and tumor growth in vivo. Conclusion In summary, RCC2 promoted PCa development through Hh/Gli1 signaling pathway via regulating EMT and CSCs.

Abstract Kidney renal clear cell carcinoma (KIRC) is the most common type of kidney cancer and its pathogenesis is strongly associated with VHL–HIF–VEGF signaling. SHH ligand is the upstream SHH pathway regulator, while GLI1 is its major effector that stimulates as a transcription factor, i.a. expression of VEGFA gene. The aim of present study was to assess the prognostic significance of SHH, GLI1 and VEGFA immunoreactivity in KIRC tissues. The analysis included paired tumor and normal samples from 34 patients with KIRC. The immunoreactivity of SHH, GLI1 and VEGFA proteins was determined by immunohistochemical (IHC) renal tissues staining. The IHC staining results were assessed using the immunoreactive score (IRS) method which takes into account the number of cells showing a positive reaction and the intensity of the reaction. Increased GLI1 protein immunoreactivity was observed in KIRC tissues, especially in early-stage tumors, according to the TNM classification. Elevated expression of the VEGFA protein was noted primarily in high-grade KIRC samples according to the Fuhrman/WHO/ISUP scale. Moreover, a directly proportional correlation was observed between SHH and VEGFA immunoreactivity in TNM 3 + 4 and Fuhrman/ISUP/WHO 3 + 4 tumor tissues as well as in samples of patients with shorter survival. We also observed an association between shorter patient survival as well as increased and decreased immunoreactivity, of the VEGFA and GLI1, respectively. The aforementioned findings suggest that the expression pattern of SHH, GLI1 and VEGFA demonstrates prognostic potential in KIRC.

Triple-negative breast cancer (TNBC) lacks effective therapeutic targets and has a poor prognosis, easy recurrence and metastasis. It is urgent and important to explore TNBC treatment targets. Through mass spectrometry combined with qRT-PCR validation in luminal A cells and TNBC cells, high-content screening and clinical sample analysis, FUNDC2 was discovered as a novel target. The function of the outer mitochondrial membrane protein FUNDC2 in breast cancer is still unclear. In this study, we find that FUNDC2 expression in TNBC tissues is significantly higher than that in luminal subtype breast cancer tissues. FUNDC2 silencing in TNBC cells significantly reduces cell proliferation, migration and invasion. As demonstrated in vivo using subcutaneous tumor xenografts in mice, FUNDC2 suppression significantly inhibits tumor growth. The underlying mechanism might be mediated by inactivating its downstream signal AKT/GSK3β and GLI1, a key factor of the Hedgehog signaling pathway. Therefore, FUNDC2 may promote TNBC progression and provide a therapeutic target for treating TNBC.

Abstract Objective To report a rare gastroblastoma; discuss its clinical features, histopathological morphology, diagnosis, differential diagnosis, treatment, and prognosis; and so as to improve the understanding on this disease and provide reference for its diagnosis, treatment, and prognosis. Methods The diagnosis and treatment, imaging examination, pathological, and genetic data of a 19-year-old young female patient with gastroblastoma were analyzed retrospectively, and the relevant literature was reviewed and summarized. Results The patient was found to have a “gastrointestinal stromal tumor” for 3 days by physical examination in another hospital. Abdominal CT and MRI considered “solid pseudopapilloma of pancreas” and clinically planned to perform “radical pancreatoduodenectomy.” During the operation, the tumor was observed to bulge from the posterior wall of the gastric antrum, and the root was located in the gastric antrum, so it was changed to “partial gastrectomy + Ronx-y gastrojejunal anastomosis.” The postoperative pathology showed that the tumor was bi-differentiated between gastric epithelium and mesenchymal. Combined with the results of IHC and the opinions of several consultation units, the diagnosis of gastric blastoma (low-grade malignancy) was supported. However, the fracture rearrangement of GLI1 and EWSR1 genes was not detected by FISH. After 19 months of follow-up, no signs of tumor recurrence and metastasis were found. Conclusion Combined with existing literature reports, gastroblastoma occurs in young people, equally in men and women, and tends to occur in the gastric antrum. The biological behavior of the tumor tends to be inert, and the prognosis of most cases is good. Postoperative pathology and IHC are reliable methods for the diagnosis of gastric blastoma, and surgical resection of the lesion is the preferred treatment.

Synovial sarcoma (SS) is a rare soft-tissue tumor characterized by a monomorphic blue spindle cell histology and variable epithelial differentiation. Morphologically, SSs may be confused with other sarcomas. Systemic treatment is more effective for patients with high-risk SSs, patients with advanced disease, and younger patients. However, further studies are required to find new prognostic biomarkers. Herein, we describe the morphological, molecular, and clinical findings, using a wide immunohistochemical panel, of a series of SS cases. We studied 52 cases confirmed as SSs by morphological diagnosis and/or molecular studies. Clinical data (gender, age, tumor size, tumor location, resection margins, adjuvant treatment, recurrences, metastasis, and survival) were also retrieved for each patient. All the available H&E slides were examined by four pathologists. Three tissue microarrays (TMAs) were constructed for each of the tumors, and a wide immunohistochemical panel was performed. For time-to-event variables, survival analysis was performed using Kaplan–Meier curves and log-rank testing, or Cox regression. Statistical significance was considered at p < 0.05. The mean age of our patients was 40.33, and the median was 40.5 years. We found a predominance of males versus females (1.7:1). The most frequent morphological subtype was monophasic. TRPS1, SS18-SSX, and SSX-C-terminus were positive in 96% of cases. GLI1 expression was strong in six and focal (cytoplasmic) in twenty patients. Moreover, BCOR was expressed in more than half of SSs. Positive expression of both proteins, BCOR and GLI1, was correlated with a worse prognosis. Multivariate analysis was also performed, but only BCOR expression appeared to be significant. The combination of GLI1 and BCOR antibodies can be used to group SSs into three risk groups (low, intermediate, and high risk). We hypothesize that these findings could identify which patients would benefit from receiving adjuvant treatment and which would not. Moreover, these markers could represent therapeutic targets in advanced stages. However, further, larger series of SSs and molecular studies are necessary to corroborate our present findings.

Aberrant activation of hedgehog (Hh) signaling has been implicated in various cancers. Current FDA-approved inhibitors target the seven-transmembrane receptor Smoothened, but resistance to these drugs has been observed. It has been proposed that a more promising strategy to target this pathway is at the GLI1 transcription factor level. GANT61 was the first small molecule identified to directly suppress GLI-mediated activity; however, its development as a potential anti-cancer agent has been hindered by its modest activity and aqueous chemical instability. Our study aimed to identify novel GLI1 inhibitors. JChem searches identified fifty-two compounds similar to GANT61 and its active metabolite, GANT61-D. We combined high-throughput cell-based assays and molecular docking to evaluate these analogs. Five of the fifty-two GANT61 analogs inhibited activity in Hh-responsive C3H10T1/2 and Gli-reporter NIH3T3 cellular assays without cytotoxicity. Two of the GANT61 analogs, BAS 07019774 and Z27610715, reduced Gli1 mRNA expression in C3H10T1/2 cells. Treatment with BAS 07019774 significantly reduced cell viability in Hh-dependent glioblastoma and lung cancer cell lines. Molecular docking indicated that BAS 07019774 is predicted to bind to the ZF4 region of GLI1, potentially interfering with its ability to bind DNA. Our findings show promise in developing more effective and potent GLI inhibitors.

Introduction Basal cell carcinoma of the prostate is rare, with no established treatment for its recurrence or metastasis. We report a case involving basal cell carcinoma of the prostate controlled using radiotherapy. Case presentation A 57‐year‐old man complained of perineal pain. Although his prostate‐specific antigen was 0.657 ng/mL, a digital rectal examination revealed his prostate was stone hard. Prostate needle biopsy showed basal cell carcinoma of the prostate. The patient then underwent radical prostatectomy. Local recurrence and sacral bone metastasis appeared 2 months after surgery. OncoGuide™ NCC Oncopanel System showed deletion of SMARCB1; however no recommended treatment was identified. Thus, we decided to perform radiotherapy, which reduced all lesions. Conclusion Basal cell carcinoma of the prostate may have a poor prognosis with recurrence or metastasis, hence evaluation of prognostic factors is important. In this case, the genomic profiling test suggested that SMARCB1 deletion may be a prognostic factor associated with disease progression.

Prostate cancer (PCa) is the most prevalent cause of death in the male population worldwide. The G Protein-Coupled Estrogen Receptor (GPER) has been gaining relevance in the development of PCa. Hedgehog (Hh) pathway activation is associated with aggressiveness, metastasis, and relapse in PCa patients. To date, no studies have evaluated the crosstalk between the GPER and the Hh pathway along different group grades in PCa. We conducted an analysis of paraffin-embedded tissues derived from patients with different prognostic grade of PCa using immunohistochemistry. Expression and correlation between GPER and glioma associated oncogene homologue (GLI) transcriptional factors in the parenchyma and stroma of PCa tumors were evaluated. Our results indicate that GPER is highly expressed in the nucleus and increases with higher grade groups. Additionally, GPER’s expression correlates with pGLI3 nuclear expression across different grade groups in PCa tissues; however, whether the receptor induces the activation of GLI transcriptional factors, or the latter modulate the expression of GPER is yet to be discovered, as well as the functional consequence of this correlation.

Abstract Esophageal squamous cell carcinoma (ESCC) is a frequently seen esophageal tumor type in China. Activation of signaling proteins and relevant molecular mechanisms in ESCC are partially explored, impairing the antitumor efficiency of targeted therapy in ESCC treatment. Tumor‐associated macrophages (TAMs)‐released C‐C motif chemokine 22 (CCL22) can activate intratumoral focal adhesion kinase (FAK), thus promoting the progression of ESCC. Here, we demonstrated that highly secreted CCL22 by TAMs (CCL22‐positive TAMs) induced ESCC cell stemness and invasion through facilitating transcriptional activity of intratumoral glioma‐associated oncogene 1 (Gli1), a downstream effector for Hedgehog (HH) pathway. Mechanistically, FAK‐activated protein kinase B (AKT) mediated Gli1 phosphorylation at its Ser112/Thr115/Ser116 sites and released Gli1 from suppressor of fused homolog, the endogenous inhibitor of Gli1 to activate downstream stemness‐associated factors, such as SRY‐box transcription factor 2 (SOX2), Nanog homeobox (Nanog), or POU class 5 homeobox (OCT4). Furthermore, inhibition of FAK activity by VS‐4718, the FAK inhibitor, enhanced antitumor effect of GDC‐0449, the HH inhibitor, both in xenografted models and in vitro assays. Clinically, CCL22/Gli1 axis is used to evaluate ESCC prognosis. Overall, our study establishes the communication of FAK with HH pathway and offers the novel mechanism related to Gli1 activation independent of Smoothened as well as the rationale for the anti‐ESCC combination treatment.

Background: Regulator of chromosome condensation 2 (RCC2) was a telophase disk-binding protein on mitosis, and functions as an oncogene in many human cancers. However, its role on prostate cancer (PCa) was unknown. The goal of this study is to explore the function of RCC 2 on PCa development. Methods: The expression of RCC2 and its methylation level, its correlation with lymph node metastasis or disease-free survival (DFS) was analyzed using TCGA database. The effect of RCC2 on PCa cell proliferation, migration and invasion were detected using CCK-8, cell colony formation, Transwell and wood healing assays. RNA-seq and GSEA analysis were used to search the downstream genes and pathways of RCC2 in mediated PCa progression. Western blot was used to detect the proteins in PCa cells transfected with indicated siRNAs or plasmids. Results: RCC2 had high expression and low promoter methylation level in PCa, and its expression was correlated with regional node metastasis and disease-free survival. Cell proliferation, migration, invasion and EMT of PCa cells in vitro were greatly enhanced after RCC2 overexpression, while the RCC2 knockdown suppressed these processes. RNA-seq and GSEA results showed the Hedgehog signaling regulator Gli1 and Gli3 were involved in RCC2 knockdown DU145 cells. Gli1 was also a marker of cancer stem-like cells (CSCs). Mechanistically, RCC2 induced cell growth, EMT, CSCs markers through Gli1; inhibiting Gli1 expression using siGli1 or GLI inhibitor suppressed cell progression in vitro and tumor growth in vivo. Conclusion: In summary, RCC2 promoted PCa development through Hh/Gli1 signaling pathway via regulating EMT and CSCs. [ABSTRACT FROM AUTHOR]

Kidney renal clear cell carcinoma (KIRC) is the most common type of kidney cancer and its pathogenesis is strongly associated with VHL–HIF–VEGF signaling. SHH ligand is the upstream SHH pathway regulator, while GLI1 is its major effector that stimulates as a transcription factor, i.a. expression of VEGFA gene. The aim of present study was to assess the prognostic significance of SHH, GLI1 and VEGFA immunoreactivity in KIRC tissues. The analysis included paired tumor and normal samples from 34 patients with KIRC. The immunoreactivity of SHH, GLI1 and VEGFA proteins was determined by immunohistochemical (IHC) renal tissues staining. The IHC staining results were assessed using the immunoreactive score (IRS) method which takes into account the number of cells showing a positive reaction and the intensity of the reaction. Increased GLI1 protein immunoreactivity was observed in KIRC tissues, especially in early-stage tumors, according to the TNM classification. Elevated expression of the VEGFA protein was noted primarily in high-grade KIRC samples according to the Fuhrman/WHO/ISUP scale. Moreover, a directly proportional correlation was observed between SHH and VEGFA immunoreactivity in TNM 3 + 4 and Fuhrman/ISUP/WHO 3 + 4 tumor tissues as well as in samples of patients with shorter survival. We also observed an association between shorter patient survival as well as increased and decreased immunoreactivity, of the VEGFA and GLI1, respectively. The aforementioned findings suggest that the expression pattern of SHH, GLI1 and VEGFA demonstrates prognostic potential in KIRC. [ABSTRACT FROM AUTHOR]

B-Raf kinase inhibitors such as vemurafenib (PLX4032) and dabrafenib have limited therapeutic efficacy on BRAF-mutated thyroid cancer. Cancer stem cells (CSCs) play important roles in tumor recurrence, drug resistance, and metastasis. Whether CSCs play a role in dampening the antitumor activity of B-Raf kinase inhibitors remains unknown. Here, we report that vemurafenib (PLX4032) induced the expression of several stemnessrelated genes including Gli1, Snail, BMI1, and SOX2 in two anaplastic thyroid cancer cell lines, SW1736 and 8505C, but decreased the expression of these genes in A375 cells, a human melanoma cell line. PLX4032 promoted thyroid cancer stem cell self-renewal, as evidenced by increased numbers of aldehyde dehydrogenase-positive cells and thyrospheres. Mechanistically, PLX4032 activates the PI-3 and mitogen-activated protein kinase pathways through HER3 to cross-activate Gli1, a transcription factor of the sonic hedgehog (Shh) pathway. GANT61, a specific inhibitor of Gli1, blocked the expression of the stemness-related genes in PLX4032-treated thyroid cancer cells in vitro and in vivo in two thyroid cancer xenograft models. GANT61 treatment alone weakly inhibited SW1736 tumor growth but enhanced the antitumor activity of PLX4032 when used in combination. Our study provides mechanistic insights into how thyroid cancer poorly responds to B-Raf kinase inhibitors and suggests that targeting B-Raf and the Shh pathway in combination may overcome thyroid cancer drug resistance. [ABSTRACT FROM AUTHOR]

Glioma-associated oncogene homolog 1 (Gli1) is a transcriptional activator of hedgehog (Hh) signaling that regulates target gene expression and several cellular biological processes. Cell lineage tracing techniques have highlighted Gli1 as an ideal marker for mesenchymal stem cells (MSCs) in vivo. Gli1+ MSCs are critical for the osteogenesis of the craniofacial bone; however, the regulatory mechanism by which Gli1+ MSCs mediate the bone development and tissue regeneration of craniofacial bone has not been systematically outlined. This review comprehensively elucidates the specific roles of Gli1+ MSCs in craniofacial bone osteogenesis. In addition to governing craniofacial bone development, Gli1+ MSCs are associated with the tissue repair of craniofacial bone under pathological conditions. Gli1+ MSCs promote intramembranous and endochondral ossification of the craniofacial bones, and assist the osteogenesis of the craniofacial bone by improving angiopoiesis. This review summarizes the novel role of Gli1+ MSCs in bone development and tissue repair in craniofacial bones, which offers new insights into bone regeneration therapy. [ABSTRACT FROM AUTHOR]

Objective: To report a rare gastroblastoma; discuss its clinical features, histopathological morphology, diagnosis, differential diagnosis, treatment, and prognosis; and so as to improve the understanding on this disease and provide reference for its diagnosis, treatment, and prognosis. Methods: The diagnosis and treatment, imaging examination, pathological, and genetic data of a 19-year-old young female patient with gastroblastoma were analyzed retrospectively, and the relevant literature was reviewed and summarized. Results: The patient was found to have a "gastrointestinal stromal tumor" for 3 days by physical examination in another hospital. Abdominal CT and MRI considered "solid pseudopapilloma of pancreas" and clinically planned to perform "radical pancreatoduodenectomy." During the operation, the tumor was observed to bulge from the posterior wall of the gastric antrum, and the root was located in the gastric antrum, so it was changed to "partial gastrectomy + Ronx-y gastrojejunal anastomosis." The postoperative pathology showed that the tumor was bi-differentiated between gastric epithelium and mesenchymal. Combined with the results of IHC and the opinions of several consultation units, the diagnosis of gastric blastoma (low-grade malignancy) was supported. However, the fracture rearrangement of GLI1 and EWSR1 genes was not detected by FISH. After 19 months of follow-up, no signs of tumor recurrence and metastasis were found. Conclusion: Combined with existing literature reports, gastroblastoma occurs in young people, equally in men and women, and tends to occur in the gastric antrum. The biological behavior of the tumor tends to be inert, and the prognosis of most cases is good. Postoperative pathology and IHC are reliable methods for the diagnosis of gastric blastoma, and surgical resection of the lesion is the preferred treatment. [ABSTRACT FROM AUTHOR]

Cutaneous T cell lymphomas (CTCLs) are a heterogeneous group of debilitating, incurable malignancies. Mycosis fungoides (MF) and Sézary syndrome (SS) are the most common subtypes, accounting for ~65% of CTCL cases. Patients with advanced disease have a poor prognosis and low median survival rates of four years. CTCLs develop from malignant skin-homing CD4+ T cells that spread to lymph nodes, blood, bone marrow and viscera in advanced stages. Current treatments options for refractory or advanced CTCL, including chemotherapeutic and biological approaches, rarely lead to durable responses. The exact molecular mechanisms of CTCL pathology remain unclear despite numerous genomic and gene expression profile studies. However, apoptosis resistance is thought to play a major role in the accumulation of malignant T cells. Here we show that NT1721, a synthetic epidithiodiketopiperazine based on a natural product, reduced cell viability at nanomolar concentrations in CTCL cell lines, while largely sparing normal CD4+ cells. Treatment of CTCL cells with NT1721 reduced proliferation and potently induced apoptosis. NT1721 mediated the downregulation of GLI1 transcription factor, which was associated with decreased STAT3 activation and the reduced expression of downstream antiapoptotic proteins (BCL2 and BCL-xL). Importantly, NT1721, which is orally available, reduced tumor growth in two CTCL mouse models significantly better than two clinically used drugs (romidepsin, gemcitabine). Moreover, a combination of NT1721 with gemcitabine reduced the tumor growth significantly better than the single drugs. Taken together, these results suggest that NT1721 may be a promising new agent for the treatment of CTCLs.

Background: Intervertebral disc degeneration (IDD) is the most common chronic disease. Oxidative stress and apoptosis of nucleus pulposus (NP) cells disrupt intervertebral disc (IVD) homeostasis, which is the main cause of IDD. Glioma-associated oncogene 1 (Gli1) is an important transcription factor in the Hedgehog (Hh) pathway. Depletion of Gli1 accelerates the occurrence and development of degenerative diseases. This study aimed to explore the role of aging related Gli1 depletion in the progression of IDD. Methods: The relationship between aging related Gli1 depletion and IDD was studied in the NP tissues of human and rats of different ages, and the levels of oxidative stress and NP cell apoptosis during IDD were explored. Gli1 depletion of NP cells were established by targeting inhibitor GANT61 or lentivirus-coated Gli1 sh-RNA (sh-Gli1) to explore the role of Gli1 in NP cells and underlying mechanism. Exogenous Gli1 depletion induced IDD of rats was established by intraperitoneal injection of GANT61. Also, the roles of Fos in the Gli1 depletion induced NP cell oxidative stress, apoptosis and IDD were investigated. Results: Gli1 was down-regulated in the tissues of degenerative NP, and the level of Gli1 was negatively correlated with the severity of aging related IDD in human and rats. Furthermore, we found enhanced oxidative stress and apoptosis in degenerative NP tissues. Gli1 depletion promoted oxidative stress and apoptosis of NP cells and resulted in the degradation of extracellular matrix (ECM) and decreased ECM synthesis. Transcriptome sequencing showed that Gli1 depletion caused Fos activation in NP cells. the effect of Gli1 depletion on the oxidative stress and apoptosis of NP cells were retarded by Fos inhibitor. In vivo, Fos inhibition alleviated the IDD induced by exogenous Gli1 depletion. Conclusions: This study revealed for the first time that Gli1 is gradually depleted in NP with IDD progression. Exogenous Gli1 depletion causes oxidative stress and apoptosis of NP cells both in vivo and in vitro. Fos suppression effectively retards the destructive effects of Gli1 depletion on IVD homoeostasis.The translational potential of this article: This study may provide new potential targets for preventing and reversing IDD. Maintaining Gli1 expression in NP and suppressing Fos activation may be an effective treatment strategy for IDD.

Abstract Background The long non-coding RNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is a cancer biomarker. Furthermore, fusion of the MALAT1 gene with glioma-associated oncogene 1 (GLI1) is a diagnostic marker of plexiform fibromyxoma and gastroblastoma; however, the function of this fusion gene remains unexplored. Method In this study, we elucidate the structure and function of the MALAT1::GLI1 fusion gene. To this end, we determined a transcriptional start site (TSS) and promoter region for truncated GLI1 expression using rapid amplification of the 5' cDNA end and a luciferase reporter assay in cultured cells transfected with a plasmid harboring the MALAT1::GLI1 fusion gene. Results We found that the TATA box, ETS1 motif, and TSS were located in MALAT1 and that MALAT1 exhibited transcriptional activity and induced expression of GLI1 from the MALAT1::GLI1 fusion gene. Truncated GLI1, lacking SUMOylation and SUFU binding sites and located in the nucleus, upregulated mRNA expression of GLI1 target genes in the hedgehog signaling pathway. Conclusions We demonstrate a distinct and alternative function of MALAT1 as a transcriptional promoter for expression of the MALAT1::GLI1 fusion gene. Our findings will aid future research on MALAT1 and its fusion gene partners.

Rheumatoid arthritis (RA) is characterized by joint synovitis and bone destruction, the etiology of which remains to be explored. Many types of cells are involved in the progression of RA joint inflammation, among which the overactivation of M1 macrophages and osteoclasts has been thought to be an essential cause of joint inflammation and bone destruction. Glioma-associated oncogene homolog 1 (GLI1) has been revealed to be closely linked to bone metabolism. In this study, GLI1 expression in the synovial tissue of RA patients was positively correlated with RA-related scores and was highly expressed in collagen-induced arthritis (CIA) mouse articular macrophage-like cells. The decreased expression and inhibition of nuclear transfer of GLI1 downregulated macrophage M1 polarization and osteoclast activation, the effect of which was achieved by modulation of DNA methyltransferases (DNMTs) via transcriptional regulation and protein interactions. By pharmacological inhibition of GLI1, the proportion of proinflammatory macrophages and the number of osteoclasts were significantly reduced, and the joint inflammatory response and bone destruction in CIA mice were alleviated. This study clarified the mechanism of GLI1 in macrophage phenotypic changes and activation of osteoclasts, suggesting potential applications of GLI1 inhibitors in the clinical treatment of RA.

Gastric cancer (GC) is a highly prevalent and aggressive malignancy with a poor prognosis. Recent evidence suggested that metallothionein 1 M (MT1M) may play a critical role in cancer development, progression, and drug resistance; however, its role in GC remains largely unknown. In this study, we investigated the expression and function of MT1M in GC both in vitro and in vivo. We found that MT1M expression was significantly downregulated in GC tissues and cell lines. Decreased expression of MT1M was associated with worse clinical prognosis, particularly in patients treated with 5-fluorouracil. Low expression of MT1M was indicative of poor overall survival (OS, HR 0.56 [95% CI 0.37–0.84], P < 0.005), first progression survival (FP, HR 0.54 [95% CI 0.36–0.79], P < 0.005), and post-progression survival (PPS, HR 0.65 [95% CI 0.45–0.94], P < 0.05). We also demonstrated that overexpression of MT1M inhibited cell proliferation and induced apoptosis in GC cells and in tumor xenografts, and it improved chemosensitivity to 5-fluorouracil. Furthermore, we found that MT1M overexpression could inhibit stem cell characteristics by targeting GLI1 and affecting GLI1 ubiquitination. Collectively, these findings indicated that MT1M may act as a tumor suppressor in GC and could serve as a potential therapeutic target to attenuate stemness and chemotherapy resistance of GC. [Display omitted] • MT1M is downregulated in GC tissues and is associated with poor prognosis. • Overexpression of MT1M inhibits malignant phenotypes in GC cells. • MT1M may suppress GC progression and stemness by inhibiting GLI1. • MT1M overexpression could be a potential therapeutic target for GC. [ABSTRACT FROM AUTHOR]

To investigate the role of GLI1 on skin proliferation and neovascularization during skin expansion in mice. We constructed GLI1‐cre/R26‐Tdtomato and GLI1‐cre/R26‐mtmg gene‐tagged skin expansion mouse models. Using a two‐photon in vivo imaging instrument to observe the changes in the number and distribution of GLI1(+) cells during the expansion process and to clarify the spatial relationship between GLI1(+) cells and blood vessels during the expansion process. In vitro proliferation assays were performed to further validate the effects of SHH (sonic hedgehog) and its downstream component GLI1 on cell proliferation viability. Finally, qRT‐PCR was used to verify the changes in proliferation, angiogenesis‐related factors, SHH signalling pathway‐related factors, and the role of GLI1 cells in the process of skin expansion in mice. The number of GLI1(+) cells increased during dilation and were attached to the outer membrane of the vessel. The epidermis was thickened and the dermis thinned after the dilated skin was taken, while the epidermal thickening was suppressed and the dermis became thinner after the GLI1 cells were inhibited. The non‐inhibited group showed a significant increase in PCNA positivity with prolonged dilation compared to the GANT61(GLI specificity inhibitor) inhibited group; CD31 immunofluorescence showed a significant increase in the number of dilated skin vessels and a significant decrease in the number of vessels after treatment with GANT61 inhibitor. In vitro proliferation results showed that SHH signalling activator significantly increased the proliferation viability of GLI1(+) hair follicle mesenchymal stem cells, while GNAT61 significantly inhibited the proliferation viability of GLI1(+) hair follicle mesenchymal stem cells. GLI1 is necessary for proliferation and neovascularization in expansion skin of mice through activation of the SHH signalling pathway. [ABSTRACT FROM AUTHOR]

The Hedgehog pathway gene Gli1 has been proposed to mark a subpopulation of skeletal stem cells (SSCs) in craniofacial bone. Skeletal stem cells (SSCs) are multi‐potent cells crucial for the development and homeostasis of bone. Recent studies on long bones have suggested that skeletal stem cells in endochondral or intramembranous ossification sites have different differentiation capacities. However, this has not been well‐defined in neural crest derived bones. Generally, the long bones are derived from mesoderm and follow an endochondral ossification model, while most of the cranial bones are neural crest (NC) in origin and follow an intramembranous ossification model. The mandible is unique: It is derived from the neural crest lineage but makes use of both modes of ossification. Early in fetal development, the mandibular body is generated by intramembranous ossification with subsequent endochondral ossification forming the condyle. The identities and properties for SSCs in these two sites remain unknown. Here, we use genetic lineage tracing in mouse to identify cells expressing the Hedgehog responsive gene Gli1, which is thought to mark the tissue resident SSCs. We track the Gli1+ cells, comparing cells within the perichondrium to those in the periosteum covering the mandibular body. In juvenile mice, these have distinct differentiation and proliferative potential. We also assess the presence of Sox10+ cells, thought to mark neural crest stem cells, but find no substantial population associated with the mandibular skeleton, suggesting that Sox10+ cells have limited contribution to maintaining postnatal mandibular bone. All together, our study indicates that the Gli1+ cells display distinct and limited differentiation capacity dependent on their regional associations. [ABSTRACT FROM AUTHOR]

Background and aims: Active intestinal stem cells are prone to injury by ionizing radiation. We previously showed that upon radiation-induced injury, normally quiescent reserve intestinal stem cells (rISCs) (marked by BMI1) are activated by Musashi-1 (MSI1) and exit from the quiescent state to regenerate the intestinal epithelium. This study aims to further establish the mechanism that regulates activation of Bmi1-CreER;Rosa26eYFP (Bmi1-CreER) rISCs following γ radiation–induced injury. Methods: Bmi1-CreER mice were treated with tamoxifen to initiate lineage tracing of BMI1 (eYFP+) cells and exposed to 12 Gy of total body γ irradiation or sham. Intestinal tissues were collected and analyzed by immunofluorescence, Western blot, reverse-transcription quantitative polymerase chain reaction, enzyme-linked immunosorbent assay, and chromatin immunoprecipitation real-time polymerase chain reaction. Results: After irradiation, increased expression of Msi1 in eYFP+ cells was accompanied by increased expression of Axin2, a WNT marker. Promoter studies of the Msi1 gene indicated that Msi1 is a WNT target gene. Coculture of stromal cells isolated from irradiated mice stimulated Bmi1-CreER–derived organoid regeneration more effectively than those from sham mice. Expression of WNT ligands, including Wnt2b, Wnt4, Wnt5a, and Rspo3, was increased in irradiated stromal cells compared with sham-treated stromal cells. Moreover, expression of the Sonic hedgehog (SHH) effector Gli1 was increased in stromal cells from irradiated mice. This was correlated with an increased expression of SHH in epithelial cells postirradiation, indicating epithelial-stromal interaction. Finally, preinjury treatment with SHH inhibitor cyclopamine significantly reduced intestinal epithelial regeneration and Msi1 expression postirradiation. Conclusions: Upon ionizing radiation-induced injury, intestinal epithelial cells increase SHH secretion, stimulating stromal cells to secrete WNT ligands. WNT activators induce Msi1 expression in the Bmi1-CreER cells. This stromal-epithelial interaction leads to Bmi1-CreER rISCs induction and epithelial regeneration.

Abstract Background Medulloblastoma (MB) is one of the most common malignant pediatric brain tumors. Metastasis and relapse are the leading causes of death in MB patients. The initiation of the SHH subgroup of MB (SHH-MB) is due to the aberrant activation of Sonic Hedgehog (Shh) signaling. However, the mechanisms for its metastasis are still unknown. Results AMP-dependent protein kinase (AMPK) restrains the activation of Shh signaling pathway, thereby impeding the proliferation of SHH-MB cells. More importantly, AMPK also hinders the growth and metastasis of SHH-MB cells by regulating NF-κB signaling pathway. Furthermore, Vismodegib and TPCA-1, which block the Shh and NF-κB pathways, respectively, synergistically restrained the growth, migration, and invasion of SHH-MB cells. Conclusions This work demonstrates that AMPK functions through two signaling pathways, SHH-GLI1 and NF-κB. AMPK-NF-κB axis is a potential target for molecular therapy of SHH-MB, and the combinational blockade of NF-κB and Shh pathways confers synergy for SHH-MB therapy.

BackgroundPlexiform fibromyxoma (PF) is a rare gastric tumor often confused with gastrointestinal stromal tumor. These so-called "benign" tumors often present with upper GI bleeding and gastric outlet obstruction. It was recently demonstrated that approximately one-third of PF have activation of the GLI1 oncogene, a transcription factor in the hedgehog (Hh) pathway, via a MALAT1-GLI1 fusion protein or GLI1 up-regulation. Despite this discovery, the biology of most PFs remains unknown.MethodsNext generation sequencing (NGS) was performed on formalin-fixed paraffin-embedded (FFPE) samples of PF specimens collected from three institutions (UCSD, NCI and OHSU). Fresh frozen tissue from one tumor was utilized for in vitro assays, including quantitative RT-PCR and cell viability assays following drug treatment.ResultsEight patients with PF were identified and 5 patients' tumors were analyzed by NGS. An index case had a mono-allelic PTCH1 deletion of exons 15-24 and a second case, identified in a validation cohort, also had a PTCH1 gene loss associated with a suspected long-range chromosome 9 deletion. Building on the role of Hh signaling in PF, PTCH1, a tumor suppressor protein, functions upstream of GLI1. Loss of PTCH1 induces GLI1 activation and downstream gene transcription. Utilizing fresh tissue from the index PF case, RT-qPCR analysis demonstrated expression of Hh pathway components, SMO and GLI1, as well as GLI1 transcriptional targets, CCND1 and HHIP. In turn, short-term in vitro treatment with a Hh pathway inhibitor, sonidegib, resulted in dose-dependent cell killing.ConclusionsFor the first time, we report a novel association between PTCH1 inactivation and the development of plexiform fibromyxoma. Hh pathway inhibition with SMO antagonists may represent a target to study for treating a subset of plexiform fibromyxomas.