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BACKGROUND: Molecular tumor boards (MTBs) are essential for selecting therapies for patients with rare and advanced cancers. We hypothesized that integrating biomarkers beyond targeted DNA/RNA next-generation sequencing (NGS) could increase actionable findings. Human epidermal growth factor receptor 2 (HER2)-low status has emerged as a critical biomarker in breast cancer, with potential relevance across other tumor types. Homologous recombination deficiency (HRD) is pivotal for the application of Poly(ADP-Ribose)-Polymerase (PARP) inhibitors in ovarian and breast cancer, although its role in other malignancies remains unclear. Antibody-drug conjugates (ADCs) are expanding precision oncology, with promising biomarkers like Trop-2, Nectin-4, and folate receptor alpha (FRα) showing potential across multiple tumor entities. METHODS: Tumors were analyzed using the TSO500® panel, enabling tumor mutational burden (TMB) readout. HER2 status was assessed via immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH), alongside antibody-drug conjugate (ADC) IHC, microsatellite instability (MSI) polymerase chain reaction (PCR), mismatch repair (MMR) IHC, programmed death-ligand 1 (PD-L1) IHC, and HRD analysis. Cases were discussed weekly, and outcomes were systematically tracked. Data analysis evaluated the benefit of additional biomarker assessments. RESULTS: Among 658 patients, 329 received therapy recommendations, 182 based on supplementary biomarker analyses. One hundred recommendations were implemented, with 37% attributed to supplementary diagnostics. Among 64 response-evaluable patients, the clinical benefit rate (complete response + partial response + stable disease) was 45.3%. HER2-low status notably expanded targeted therapy options across tumor types, with similar implementation rates for HER2-low and HER2-amplified tumors. HRD analysis refined stratification in tumors with mutations in homologous recombination repair (HRR) genes beyond BRCA1/2, including PALB2, ATM, and CHEK2. ADC IHC supported 20 recommendations and two therapy implementations. CONCLUSIONS: The integration of additional biomarker assessments into MTB workflows enhances precision oncology by expanding the pool of patients eligible for targeted therapies.
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BACKGROUND AND AIMS: Hepatoblastoma (HB) is the predominant primary malignant liver tumor in childhood. Concomitant Yes-associated protein (YAP) and ß-Catenin activation occurs in most HB. However, the signaling pathways distinctively regulated by YAP and ß-Catenin protooncogenes in HB remain unexplored. APPROACH AND RESULTS: We engineered an inducible HB murine model using hydrodynamic injection to deliver transposon plasmids encoding constitutive YAP and doxycycline (Dox)-inducible ΔN90-ß-Catenin(YAP/TRE-ß-Catenin). Gene expression patterns in mouse HB lesions were analyzed upon short-term Dox withdrawal, that is, tumors still existed but ΔN90-ß-Catenin was turned off. The mice rapidly developed aggressive HB lesions when fed Dox. However, upon Dox withdrawal, HB regressed, although tumors did not completely disappear over a long time. At the molecular level, YAP and ß-Catenin were found to regulate distinct gene expression programs in HB. Specifically, YAP controls the Hippo and metabolism-related pathways, whereas ß-Catenin modulates immune-related pathways, contributing to immune exclusion in the tumor microenvironment. Furthermore, we identified the transcription factor ONECUT1 as a tumor suppressor gene downregulated by activated ß-Catenin in HB. Low ONECUT1 expression also characterizes human HB, and the co-expression of ONECUT1 strongly suppressed YAP/ß-Catenin-driven HB formation in the mice. Mechanistically, ONECUT1 functions downstream of activated ß-Catenin and negatively regulates tumor cell glycolysis. CONCLUSIONS: We show that suppressing activated ß-Catenin could hamper HB progression in vivo by affecting pathways distinct from those regulated by YAP in HB. The inhibition of ONECUT1 expression by ß-Catenin might represent a critical molecular event leading to HB formation.
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Proteínas Adaptadoras de Transdução de Sinal , Hepatoblastoma , Neoplasias Hepáticas , Fatores de Transcrição , beta Catenina , beta Catenina/metabolismo , beta Catenina/genética , Animais , Hepatoblastoma/genética , Hepatoblastoma/patologia , Hepatoblastoma/metabolismo , Camundongos , Humanos , Proteínas de Sinalização YAP , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Fatores de Transcrição/metabolismo , Fatores de Transcrição/genética , Transdução de Sinais , Regulação Neoplásica da Expressão Gênica , Modelos Animais de Doenças , Proteínas de Ciclo CelularRESUMO
Epithelial-to-mesenchymal transition (EMT) is an early event during tumor metastasis. Here, the relevance of EMT in liver carcinogenesis and metastasis is sought to be determined in a murine mixed typical intrahepatic cholangiocarcinoma (iCCA)/sarcomatoid iCCA liver tumor model using CRISPR/Cas9-based gene deletion of p19 (sgP19) in combination with transposon-based expression of the activated form of pCaggs-kRASG12D (kRAS) in the mouse liver (sgP19/kRAS mixed model). It discovered that metastasis in the lymph node, lung, or kidney occurred in the sgP19/kRAS mixed model. Both typical iCCA tumor cells with epithelial features and sarcomatoid tumor cells with mesenchymal features could be detected in this model. Lineage tracing technology is applied to confirm the metastasis induced in the sgP19/kRAS model. Subsequently, the gain of expression of mesenchymal marker vimentin in tumor cells revealed the induction of EMT in the sgP19/kRAS model, and it is induced by activating the TGFß/ZEB1 signaling pathway. Altogether, the study suggests that TGFß/ZEB1 mediates the induction of EMT in iCCA, while targeting EMT failed to inhibit iCCA development or tumor metastasis, disputing the claims that EMT is a major molecular event leading to tumor metastasis.
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Recent studies have shown that pulmonary neuroendocrine tumor (NET) subgroups, defined by the transcription factors OTP and ASCL1, correlate with age, sex, and tumor location. Their relationships with histology and hormone production, however, remain unclear. We analyzed 170 pulmonary NETs classified by OTP (O) and ASCL1 (A) expression into four groups: O + /A + , O + /A-, O-/A + , and O-/A-. Subgroups were assessed for histology, hormone expression, therapy-related markers, outcomes, and matched metastases. Among 152 resected primaries, O + /A + tumors (38%) were most frequent, occurring mainly in females (median age 72 years), and typically showed central or peripheral location, solid/spindle morphology with diffuse gastrin-releasing peptide (GRP), and focal ACTH/calcitonin. They also showed strong DLL3 expression and pronounced neuroendocrine cell hyperplasia. O + /A- tumors (23%) occurred predominantly in females (median age 56 years) with solid/trabecular patterns, occasional/ACTH, and strong SSTR2A/5 expression. O-/A- tumors (25%) were more common in males (median age 70 years), often central with solid/trabecular or oncocytic histology, serotonin expression (24%), and frequently SSTR2A-positivity. O-/A + tumors (14%) occurred across both sexes (median age 58 years), were centrally located, and solid, sometimes oncocytic features with moderate DLL3/SSTR2A expression. Metastases mirrored their primaries in transcription factor and hormone profiles. In the univariate analysis, OTP-negative tumors were associated with poorer disease-free survival (DFS). However, the multivariate analysis identified Ki67-based WHO grades (G1-G3) as the only independent prognostic factors. In conclusion, integrating OTP and ASCL1 refines pulmonary NET classification into four histologically and biologically distinct subgroups, providing additional insight into tumor heterogeneity. O + /A + tumors showed solid-spindle features and diffuse GRP and frequent ACTH expression, trabecular patterns characterized ASCL1-negative tumors, while oncocytic histology predominated in OTP-negative tumors, highlighting their role in defining tumor heterogeneity.
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Proteínas Hélice-Alça-Hélice Básicas , Biomarcadores Tumorais , Neoplasias Pulmonares , Tumores Neuroendócrinos , Humanos , Feminino , Tumores Neuroendócrinos/patologia , Tumores Neuroendócrinos/metabolismo , Tumores Neuroendócrinos/classificação , Masculino , Idoso , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/classificação , Neoplasias Pulmonares/mortalidade , Pessoa de Meia-Idade , Proteínas Hélice-Alça-Hélice Básicas/biossíntese , Proteínas Hélice-Alça-Hélice Básicas/análise , Biomarcadores Tumorais/análise , Adulto , Idoso de 80 Anos ou mais , Fenótipo , Proteínas do Tecido Nervoso , Proteínas de HomeodomínioRESUMO
Aberrant activation of AKT is a key oncogenic driver in hepatocellular carcinoma (HCC). As AKT activates multiple downstream signaling pathways, the key mechanisms mediating AKT-driven tumorigenesis must be elucidated to develop optimal treatment strategies. Using an Akt/NRas-induced HCC mouse model, we found that AKT promotes tumorigenesis by targeting tuberous sclerosis complex 2 (TSC2) and glycogen synthase kinase 3 α/ß (GSK3α/ß) rather than forkhead box O protein family members. Loss of either TSC2, leading to mTORC1 activation, or both GSK3 isoforms cooperated with activated NRAS to promote HCC formation in vivo, albeit with different latencies. Simultaneous TSC2 and GSK3α/ß deletion cooperated with NRAS to rapidly induce HCC formation, mirroring observations from the Akt/NRas HCC model. RNA sequencing studies indicated distinct pathways regulated by TSC2/mTORC1 and GSK3α/ß during hepatocarcinogenesis, with FOXM1 functioning as a major effector of GSK3. In summary, these findings uncover AKT's role in suppressing the TSC complex and GSK3 to drive HCC, offering mechanistic insights into oncogenic signaling and potential therapeutic targets. SIGNIFICANCE: GSK3α/ß and TSC2/mTORC1 are key downstream effectors of AKT in liver tumorigenesis that can potentially be targeted to benefit patients with hepatocellular carcinoma harboring aberrantly activated AKT.
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Carcinoma Hepatocelular , Quinase 3 da Glicogênio Sintase , Neoplasias Hepáticas , Proteínas Proto-Oncogênicas c-akt , Proteína 2 do Complexo Esclerose Tuberosa , Animais , Proteína 2 do Complexo Esclerose Tuberosa/metabolismo , Proteína 2 do Complexo Esclerose Tuberosa/genética , Proteína 2 do Complexo Esclerose Tuberosa/antagonistas & inibidores , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Camundongos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Humanos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Transdução de Sinais , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Quinase 3 da Glicogênio Sintase/metabolismo , Quinase 3 da Glicogênio Sintase/genética , Carcinogênese , Proteína Forkhead Box M1/metabolismoRESUMO
Calprotectin, a calcium- and zinc-binding protein composed of the subunits S100A8 and S100A9, has been extensively studied as a biomarker of gastrointestinal (GI) inflammation through fecal and serum analyses. However, its role in intestinal tissue remains poorly understood due to limited availability of biopsies. In this study, we analyzed S100A8 and S100A9 mRNA expression in 579 intestinal biopsies from allogeneic stem cell transplant (ASCT) patients and observed a strong association with acute GI graft-versus-host disease (aGI-GvHD) (p<0.001). Neutrophil infiltration correlated with the severity of aGI-GvHD (p<0.001), and calprotectin expression was strongly linked to Toll-like receptor 4 (TLR4) (p<0.001) and TLR2 (p<0.001) expression. TLR4 and aGI-GvHD were associated with elevated calprotectin mRNA levels (p<0.001). When patients received broad-spectrum antibiotics at disease onset, expression of calprotectin was suppressed (S100A8, p=0.001; S100A9, p=0.01). Gastrointestinal site-specific differences in calprotectin expression were identified: during severe aGI-GvHD, levels increased up to 30-fold in the small intestine and up to 5-fold in the large intestine with respect to mild/no aGI-GVHD, while under homeostasis, the large intestine exhibited higher baseline calprotectin (p=0.001). The high clinical relevance is evident from the observation that calprotectin expression was prognostic for transplant-related mortality (TRM). Our study suggests that (a) calprotectin is a potential biopsy biomarker in aGI-GvHD, (b) calprotectin expression and neutrophil infiltration possibly indicate translocation of microbiota, which (c) may be modulated by antibiotics.
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Intrahepatic cholangiocarcinoma (iCCA) is the second most common primary liver tumor. Due to its aggressive nature and resistance to conventional treatments, there is a pressing need to develop novel and more effective therapies for this deadly malignancy. Here, we explored the therapeutic potential of the DNA minor groove binders trabectedin (TRB) and lurbinectedin (LUR) for the treatment of iCCA using cell lines, spheroids, cancer-associated fibroblasts (CAFs), patient-derived tumor organoids (PDOs), and the chicken chorioallantoic membrane (CAM) in vivo model. TRB and, more substantially, LUR, significantly inhibited cell growth in iCCA cell lines, spheroids, CAFs, and PDOs at very low nanomolar concentrations. Specifically, the two drugs significantly reduced proliferation, triggered apoptosis, and caused DNA damage in iCCA cells. At the metabolic level, TRB and LUR decreased mitochondrial respiration and glycolysis. At the molecular level, the two compounds effectively downregulated the mammalian target of rapamycin complex 1 (mTORC1) and Hippo/YAP pathways and suppressed the expression of yes-associated protein 1 (YAP1), cellular myelocytomatosis oncogene (c-Myc), E2F transcription factor 1 (E2F1), Bromodomain-containing protein 4 (BRD4), TEA domain transcription factor 4 (TEAD4), and cluster of differentiation 7 (CD7) proto-oncogenes. Furthermore, LUR significantly restrained the in vivo growth of iCCA cells in the CAM model. Our data indicate that TRB and LUR possess strong anti-proliferative and pro-apoptotic activities and could represent promising therapeutic agents for the treatment of iCCA.
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Antineoplásicos , Neoplasias dos Ductos Biliares , Carbolinas , Colangiocarcinoma , Compostos Heterocíclicos de 4 ou mais Anéis , Trabectedina , Trabectedina/farmacologia , Humanos , Colangiocarcinoma/tratamento farmacológico , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patologia , Animais , Linhagem Celular Tumoral , Carbolinas/farmacologia , Proliferação de Células/efeitos dos fármacos , Apoptose/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Neoplasias dos Ductos Biliares/tratamento farmacológico , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/patologia , Antineoplásicos/farmacologia , Embrião de GalinhaRESUMO
Background & aims: Intrahepatic cholangiocarcinoma (iCCA) is the second most frequent primary liver tumor, characterized by clinical aggressiveness, dismal outcome, and limited therapeutic options. Thus, innovative treatments are urgently required to improve the prognosis of patients with iCCA. Methods: In this study, we determined the pathogenetic and therapeutic role of eukaryotic initiation factor 4A1 (EIF4A1), a subunit of the eIF4F complex involved in translation initiation, in human iCCA. Results: Preinvasive (n = 12), invasive (n = 162), and metastatic (n = 14) iCCA lesions exhibited ubiquitous eIF4A1 upregulation. In addition, eIF4A1 mRNA levels from 42 specimens showed a significantly higher expression in iCCA samples compared with non-tumorous tissues (p <0.0001) or large duct-type lesions (p = 0.020). Furthermore, eIF4A1 expression was inversely associated with patient prognosis (p <0.001). Moreover, zotatifin, an eIF4A1-specific inhibitor in clinical trials, significantly reduced the growth of iCCA cell lines, iCCA cancer-associated fibroblasts (CAFs), and patient-derived tumor organoids. At the metabolic level, zotatifin decreased glycolysis of iCCA cells without affecting mitochondrial respiration. Moreover, the Bcl-xl inhibitors A-1155463 and DT2216 profoundly augmented apoptotic cell death when administered in association with zotatifin. Conclusions: The data highlight eIF4A1 as a potential target for treating iCCA. Combined inhibition of eIF4A1 and Bcl-xl could offer an effective therapeutic strategy against this deadly disease. Impact and implications: Dysregulation of the translational machinery is a hallmark of cancer, often linked to tumor progression and poor prognosis. This study underscores the potential of zotatifin, a specific inhibitor of EIF4A1 (an essential component of translation initiation) to inhibit the growth of iCCA cells. In addition, zotatifin demonstrated a synergistic effect when used in combination with the Bcl-xl inhibitors A-1155463 and DT2216, significantly enhancing cell apoptosis. Although this investigation did not include an in vivo model, its results, derived from iCCA cell lines, patient-derived organoids, and CAFs, are consistent with the encouraging preliminary results of zotatifin in clinical trials. From a clinical standpoint, these results suggest that zotatifin improves patient outcomes by inhibiting iCCA growth and reducing tumor aggressiveness. Furthermore, combining zotatifin with other drugs could represent a promising therapeutic strategy for targeting iCCA.
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The literature emphasizes that pulmonary ectopic Cushing syndrome (ECS) is associated not only with neuroendocrine tumors (NETs), but also with small cell lung carcinomas (SCLCs). This statement is debatable, because extrapulmonary ECS is associated with NETs in the vast majority of cases and very rarely with neuroendocrine carcinomas (NECs). Therefore, we critically reviewed the literature on SCLCs associated with ECS (ECS-SCLC) and performed immunohistochemical analysis of ACTH expression in 155 resected SCLCs and 158 pulmonary NETs. The literature search revealed that 90% of the 205 ECS-SCLC patients identified between 1952 and 2023 had no or poor-quality histologic images, so the diagnosis of SCLC could not be confirmed. Review of the 20 reports (10%) with histologic images revealed that 18/20 (90%) had to be reclassified as "probable NET", of which 5/18 (28%) showed spindle cell morphology, while only 2 cases were qualified as "SCLC compatible" due to their pleomorphic cell features. Immunohistochemically, 5/155 (3%) resected SCLCs, all without ECS, showed weak single cell ACTH expression, whereas in the NET cohort, 61/158 (39%) tumors expressed ACTH, of which 4 (3%) were associated with ECS. Both observations, the literature review, which casts doubt on previously reported data regarding the frequency of SCLC in ECS, and the immunohistochemical study, suggest that there is limited evidence that SCLC is the cause of ECS.
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Síndrome de ACTH Ectópico , Hormônio Adrenocorticotrópico , Síndrome de Cushing , Neoplasias Pulmonares , Carcinoma de Pequenas Células do Pulmão , Humanos , Neoplasias Pulmonares/complicações , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Carcinoma de Pequenas Células do Pulmão/complicações , Carcinoma de Pequenas Células do Pulmão/metabolismo , Carcinoma de Pequenas Células do Pulmão/patologia , Hormônio Adrenocorticotrópico/biossíntese , Síndrome de ACTH Ectópico/etiologia , Tumores Neuroendócrinos/complicações , Tumores Neuroendócrinos/patologia , Tumores Neuroendócrinos/metabolismo , Carcinoma Neuroendócrino/complicações , Carcinoma Neuroendócrino/patologia , Carcinoma Neuroendócrino/metabolismo , Masculino , FemininoRESUMO
Delta-like ligand 3 (DLL3) is frequently expressed in pulmonary small cell neuroendocrine carcinoma (SCNEC) and has emerged as a promising therapeutic target. However, limited data on DLL3 expression in other neuroendocrine neoplasms (NEN), such as extrapulmonary SCNEC, large cell neuroendocrine carcinomas (LCNEC), mixed neuroendocrine-non-neuroendocrine neoplasms (MiNEN), gastroenteropancreatic neuroendocrine tumours (GEP-NET), and pulmonary carcinoids, impedes an estimation if other types of NEN might be suitable candidates for anti-DLL3 therapies. We evaluated DLL3 expression in 1294 NEN and 479 non-neuroendocrine carcinomas, correlating the findings with histological subtypes, tumour localisation, and overall survival (OS). Furthermore, we explored the concordance of DLL3 expression during metastatic progression in 67 paired primary NEN and metastases. DLL3 expression was significantly higher in NEC (64.0%) compared to GEP-NET and pulmonary carcinoids (10.1%, p < 0.001), particularly in SCNEC (80.4%), followed by LCNEC (62.6%) and MiNEN (28.6%). DLL3 was common in pulmonary carcinoids (41.5%), but rare in GEP-NET (5.1%) and non-neuroendocrine carcinomas (1.3%). Overall DLL3 expression was highly concordant between metastases and corresponding primary NEN (92.5%, p < 0.001). In univariable analyses, DLL3-expressing pulmonary carcinoids (p = 0.005) and GEP-NET (p = 0.018) were associated with decreased OS, but this was not retained in multivariable analyses adjusting for stage and grade (p = n. s.). No prognostic impact was observed in pulmonary (p = 0.708) or GEP-NEC (p = 0.87). Our study highlights significant differences in DLL3 expression across NEN subtypes and localisations, with largely concordant expression in metastases. DLL3-based therapies may be effective in many NEC and pulmonary carcinoids, while DLL3 appears to be a minor therapeutic target for GEP-NET and non-neuroendocrine carcinomas.
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Biomarcadores Tumorais , Carcinoma Neuroendócrino , Peptídeos e Proteínas de Sinalização Intracelular , Neoplasias Pulmonares , Proteínas de Membrana , Tumores Neuroendócrinos , Humanos , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/mortalidade , Masculino , Feminino , Tumores Neuroendócrinos/patologia , Tumores Neuroendócrinos/metabolismo , Tumores Neuroendócrinos/mortalidade , Proteínas de Membrana/biossíntese , Proteínas de Membrana/análise , Pessoa de Meia-Idade , Idoso , Carcinoma Neuroendócrino/patologia , Carcinoma Neuroendócrino/metabolismo , Carcinoma Neuroendócrino/mortalidade , Biomarcadores Tumorais/análise , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Peptídeos e Proteínas de Sinalização Intracelular/análise , Adulto , Estudos de Coortes , Idoso de 80 Anos ou maisRESUMO
Prostate cancer (PC) is the most diagnosed cancer in males across the globe. Following the formation of metastasis, PC is linked to a notable decline in both prognosis and survival rates. Three-dimensional multicellular spheroids (MCSs) of a prostate adenocarcinoma cell line were generated in a three-day simulated microgravity environment (s-µg) to serve as a model for metastasis and to derive transcriptional and epigenetic PC candidates from molecular biological changes. With an FDR of 10-3, we detected the most differentially expressed genes in the two comparisons' adherent cells (AD) to MCSs (N = 751 genes) and 1g control cells to MCSs (N = 662 genes). In these two comparisons, genes related to cell cycle, angiogenesis, cell adhesion, and extracellular space were consistently found to be significantly enriched in GO annotations. Furthermore, at a 5% FDR significance level, we were able to identify 11,090 genome-wide differentially methylated positions (DMPs) and one differentially methylated region in the SRMS gene in the 1g vs. AD comparison, as well as an additional 10,797 DMPs in the 1g vs. MCSs comparison. Finally, we identified five s-µg-related positive enrichments of transcription factor binding sites for AR, IRF1, IRF2, STAT1, STAT2, and FOXJ3 close to the DMPs.
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Neoplasias da Próstata , Simulação de Ausência de Peso , Humanos , Masculino , Neoplasias da Próstata/genética , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Metilação de DNA , Epigênese Genética , Esferoides Celulares/metabolismoRESUMO
Changes in the intestinal microbiome and microbiota-derived metabolites predict clinical outcomes after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Here, we report that desaminotyrosine (DAT), a product of bacterial flavonoid metabolism, correlates with improved overall survival and reduced relapse rates in patients receiving allo-HSCT. In preclinical mouse models, treatment with synthetic DAT prevents graft-versus-host disease by protecting the intestinal barrier and promoting intestinal regeneration and contributes to graft-vs.-leukemia responses. DAT´s beneficial effects on intestinal regeneration remain effective despite broad-spectrum antibiotics-induced dysbiosis, also when administered by fecal microbiota transfer with flavonoid-degrading F. plautii. Mechanistically, DAT promotes mTORC1-dependent activation and proliferation of intestinal stem cells, with concomitant engagement of the innate immune receptor STING required to mitigate metabolic stress and maintain an undifferentiated stem cell state independently of type-I interferon responses. Additionally, DAT can skew T cells towards an effector phenotype to modulate graft-versus-leukemia responses. Our data uncover DAT's dual, tissue- and immune-modulating properties and underscore its potential in precision microbiome-based therapies to improve tissue regeneration and minimize immune-mediated side effects.
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Microbioma Gastrointestinal , Doença Enxerto-Hospedeiro , Intestinos , Alvo Mecanístico do Complexo 1 de Rapamicina , Proteínas de Membrana , Regeneração , Tirosina , Animais , Doença Enxerto-Hospedeiro/prevenção & controle , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Microbioma Gastrointestinal/efeitos dos fármacos , Camundongos , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Intestinos/fisiologia , Intestinos/efeitos dos fármacos , Proteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Regeneração/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Tirosina/análogos & derivados , Tirosina/farmacologia , Tirosina/metabolismo , Masculino , Feminino , Transplante de Microbiota Fecal , Transplante HomólogoRESUMO
Mitotic arrest-deficient 2 like 1 (MAD2L1) is a component of the mitotic spindle assembly checkpoint implicated in cancer cell proliferation and tumorigenesis. The functional role of MAD2L1 in hepatocellular carcinoma (HCC) has not been adequately investigated, especially in vivo. In the current manuscript, we sought to address the function of MAD2L1 in hepatocarcinogenesis. We found that MAD2L1 expression is upregulated in human HCCs, where its expression is associated with higher aggressive tumor grade, elevated proliferative activity, and poor prognosis. In human HCC cell lines, MAD2L1 knockdown led to decreased cell growth. Moreover, RNA-seq results demonstrated that MAD2L1 silencing induces the expression of genes associated with cell cycle, DNA replication, and various cancer-related pathways, supporting the critical role of MAD2L1 during HCC growth and differentiation. In a c-MYC-induced mouse HCC model, we revealed an increased expression of Mad2l1. Furthermore, Mad2l1 CRIPSR-mediated silencing prevented c-MYC-driven mouse liver development. Altogether, our study suggests that MAD2L1 plays a crucial role in hepatocarcinogenesis, and that its suppression could be a promising therapeutic strategy for treating human HCC. MAD2L1 plays a critical role in liver cancer development, silencing MAD2L1 reduced cell growth in vitro and inhibited c-MYC-driven liver cancer development in vivo. MAD2L1 suppression might be a promising therapeutic approach for treating human liver cancer.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Proteínas Mad2 , Proteínas Proto-Oncogênicas c-myc , Animais , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Proteínas Mad2/genética , Proteínas Mad2/metabolismo , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Camundongos , Prognóstico , Proliferação de Células , Linhagem Celular Tumoral , Masculino , Regulação Neoplásica da Expressão Gênica , Carcinogênese/genética , FemininoRESUMO
BACKGROUND: Intrahepatic cholangiocarcinoma (iCCA) is a lethal primary liver tumor characterized by clinical aggressiveness, poor prognosis, and scarce therapeutic possibilities. Therefore, new treatments are urgently needed to render this disease curable. Since cumulating evidence supports the oncogenic properties of the Heat Shock Factor 1 (HSF1) transcription factor in various cancer types, we investigated its pathogenetic and therapeutic relevance in iCCA. METHODS: Levels of HSF1 were evaluated in a vast collection of iCCA specimens. The effects of HSF1 inactivation on iCCA development in vivo were investigated using three established oncogene-driven iCCA mouse models. In addition, the impact of HSF1 suppression on tumor cells and tumor stroma was assessed in iCCA cell lines, human iCCA cancer-associated fibroblasts (hCAFs), and patient-derived organoids. RESULTS: Human preinvasive, invasive, and metastatic iCCAs displayed widespread HSF1 upregulation, which was associated with a dismal prognosis of the patients. In addition, hydrodynamic injection of a dominant-negative form of HSF1 (HSF1dn), which suppresses HSF1 activity, significantly delayed cholangiocarcinogenesis in AKT/NICD, AKT/YAP, and AKT/TAZ mice. In iCCA cell lines, iCCA hCAFs, and patient-derived organoids, administration of the HSF1 inhibitor KRIBB-11 significantly reduced proliferation and induced apoptosis. Cell death was profoundly augmented by concomitant administration of the Bcl-xL/Bcl2/Bcl-w inhibitor ABT-263. Furthermore, KRIBB-11 reduced mitochondrial bioenergetics and glycolysis of iCCA cells. CONCLUSIONS: The present data underscore the critical pathogenetic, prognostic, and therapeutic role of HSF1 in cholangiocarcinogenesis.
Assuntos
Neoplasias dos Ductos Biliares , Colangiocarcinoma , Fatores de Transcrição de Choque Térmico , Colangiocarcinoma/metabolismo , Colangiocarcinoma/patologia , Colangiocarcinoma/genética , Colangiocarcinoma/tratamento farmacológico , Humanos , Animais , Camundongos , Prognóstico , Fatores de Transcrição de Choque Térmico/metabolismo , Fatores de Transcrição de Choque Térmico/genética , Neoplasias dos Ductos Biliares/metabolismo , Neoplasias dos Ductos Biliares/patologia , Neoplasias dos Ductos Biliares/tratamento farmacológico , Neoplasias dos Ductos Biliares/genética , Linhagem Celular Tumoral , Modelos Animais de Doenças , Proliferação de CélulasRESUMO
Activated mTORC2/AKT signaling plays a role in hepatocellular carcinoma (HCC). Research has shown that TSC/mTORC1 and FOXO1 are distinct downstream effectors of AKT signaling in liver regeneration and metabolism. However, the mechanisms by which these pathways mediate mTORC2/AKT activation in HCC are not yet fully understood. Amplification and activation of c-MYC are key molecular events in HCC. In this study, we explored the roles of tuberous sclerosis complex/mTORC1 (TSC/mTORC1) and FOXO1 as downstream effectors of mTORC2/AKT1 in c-MYC-induced hepatocarcinogenesis. Using various genetic approaches in mice, we found that manipulating the FOXO pathway had a minimal effect on c-MYC-induced HCC. In contrast, loss of mTORC2 inhibited c-MYC-induced HCC, an effect that was completely reversed by ablation of TSC2, which activated mTORC1. Additionally, we discovered that p70/RPS6 and 4EBP1/eIF4E acted downstream of mTORC1, regulating distinct molecular pathways. Notably, the 4EBP1/eIF4E cascade is crucial for cell proliferation and glycolysis in c-MYC-induced HCC. We also identified centromere protein M (CENPM) as a downstream target of the TSC2/mTORC1 pathway in c-MYC-driven hepatocarcinogenesis, and its ablation entirely inhibited c-MYC-dependent HCC formation. Our findings demonstrate that the TSC/mTORC1/CENPM pathway, rather than the FOXO cascade, is the primary signaling pathway regulating c-MYC-driven hepatocarcinogenesis. Targeting CENPM holds therapeutic potential for treating c-MYC-driven HCC.
Assuntos
Carcinoma Hepatocelular , Alvo Mecanístico do Complexo 1 de Rapamicina , Alvo Mecanístico do Complexo 2 de Rapamicina , Proteínas Proto-Oncogênicas c-akt , Proteínas Proto-Oncogênicas c-myc , Transdução de Sinais , Proteína 2 do Complexo Esclerose Tuberosa , Animais , Camundongos , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-akt/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteína 2 do Complexo Esclerose Tuberosa/genética , Proteína 2 do Complexo Esclerose Tuberosa/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/genética , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/genética , Complexos Multiproteicos/genética , Complexos Multiproteicos/metabolismo , Proteínas Cromossômicas não Histona/metabolismo , Proteínas Cromossômicas não Histona/genética , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Neoplasias Hepáticas/genética , Proteínas Supressoras de Tumor/metabolismo , Proteínas Supressoras de Tumor/genética , Proteína Forkhead Box O1/metabolismo , Proteína Forkhead Box O1/genética , Serina-Treonina Quinases TOR/metabolismo , Serina-Treonina Quinases TOR/genética , Neoplasias Hepáticas Experimentais/metabolismo , Neoplasias Hepáticas Experimentais/patologia , Neoplasias Hepáticas Experimentais/genética , Camundongos Knockout , Carcinogênese/metabolismo , Carcinogênese/genética , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ciclo Celular/genéticaRESUMO
Many pancreatic neuroendocrine tumors (PanNETs) fall into 2 major prognostic subtypes based on DAXX/ATRX-induced alternative lengthening of telomerase phenotype and alpha- and beta-cell-like epigenomic profiles. However, these PanNETs are still flanked by other PanNETs that do not fit into either subtype. Furthermore, despite advanced genotyping, PanNETs are generally not well-characterized in terms of their histologic and hormonal phenotypes. We aimed to identify new subgroups of PanNETs by extending the currently used transcription factor signatures and investigating their correlation with histologic, hormonal, molecular, and prognostic findings. One hundred eighty-five PanNETs (nonfunctioning 165 and functioning 20), resected between 1996 and 2023, were classified into 5 subgroups (A1, A2, B, C, and D) by cluster analysis based on ARX, PDX1, islet-1 (ISL1), and CDX2 expressions and correlated with trabecular vs solid histology, expression of insulin, glucagon, polypeptide (PP), somatostatin, serotonin, gastrin, calcitonin, adrenocorticotropic hormone (ACTH), DAXX/ATRX, MEN1, and alternative lengthening of telomerase status by fluorescence in situ hybridization, and disease-free survival. A1 (46%, ARX+/ISL1+/PDX1-/CDX2-) and A2 (15%, ARX+/ISL1+/PDX1+/CDX2-) showed trabecular histology and glucagon/PP expression, with A2 also showing gastrin expression. B (18%, PDX1+/ISL1+/ARX-/CDX2-) showed solid histology, insulin, and somatostatin expression (P < .001). It included all insulinomas and had the best outcome (P < .01). C (15%, ARX-/PDX1-/ISL1-/CDX2-) showed solid histology and frequent expression of serotonin, calcitonin, and ACTH. D (5%, PDX1+/CDX2+/ISL1-/ARX-) showed solid histology, expressed ACTH/serotonin, and was an independent poor prognosticator (P < .01). Differential expressions of ARX, PDX1, ISL1, and CDX2 stratified PanNETs into 5 subgroups with different histologies, hormone expressions, and outcomes. Subgroups A1 and A2 resembled the alpha-cell-like type, and subgroup B, the beta-cell-like type. Subgroup C with almost no transcription factor signature was unclear in cell lineage, whereas the PDX+/CDX2+ signature of subgroup D suggested a pancreatic/intestinal cell lineage. Assigning PanNETs to the subgroups may help establish the diagnosis, predict the outcome, and guide the treatment.
Assuntos
Biomarcadores Tumorais , Fator de Transcrição CDX2 , Proteínas de Homeodomínio , Proteínas com Homeodomínio LIM , Tumores Neuroendócrinos , Neoplasias Pancreáticas , Transativadores , Fatores de Transcrição , Humanos , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/metabolismo , Tumores Neuroendócrinos/patologia , Tumores Neuroendócrinos/metabolismo , Proteínas de Homeodomínio/metabolismo , Masculino , Feminino , Pessoa de Meia-Idade , Biomarcadores Tumorais/análise , Biomarcadores Tumorais/metabolismo , Idoso , Adulto , Fator de Transcrição CDX2/metabolismo , Proteínas com Homeodomínio LIM/metabolismo , Proteínas com Homeodomínio LIM/genética , Proteínas com Homeodomínio LIM/análise , Proteínas com Homeodomínio LIM/biossíntese , Prognóstico , Hormônios Pancreáticos/metabolismo , Idoso de 80 Anos ou mais , Adulto JovemRESUMO
Background and Objectives: Aberrant upregulation of fatty acid synthase (FASN), catalyzing de novo synthesis of fatty acids, occurs in various tumor types, including human hepatocellular carcinoma (HCC). Although FASN oncogenic activity seems to reside in its pro-lipogenic function, cumulating evidence suggests that FASN's tumor-supporting role might also be metabolic-independent. Materials and Methods: In the present study, we show that FASN inactivation by specific small interfering RNA (siRNA) promoted the downregulation of the S-phase kinase associated-protein kinase 2 (SKP2) and the consequent induction of p27KIP1 in HCC cell lines. Results: Expression levels of FASN and SKP2 directly correlated in human HCC specimens and predicted a dismal outcome. In addition, forced overexpression of SKP2 rendered HCC cells resistant to the treatment with the FASN inhibitor C75. Furthermore, FASN deletion was paralleled by SKP2 downregulation and p27KIP1 induction in the AKT-driven HCC preclinical mouse model. Moreover, forced overexpression of an SKP2 dominant negative form or a p27KIP1 non-phosphorylatable (p27KIP1-T187A) construct completely abolished AKT-dependent hepatocarcinogenesis in vitro and in vivo. Conclusions: In conclusion, the present data indicate that SKP2 is a critical downstream effector of FASN and AKT-dependent hepatocarcinogenesis in liver cancer, envisaging the possibility of effectively targeting FASN-positive liver tumors with SKP2 inhibitors or p27KIP1 activators.
Assuntos
Carcinoma Hepatocelular , Inibidor de Quinase Dependente de Ciclina p27 , Neoplasias Hepáticas , Proteínas Quinases Associadas a Fase S , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Proteínas Quinases Associadas a Fase S/metabolismo , Proteínas Quinases Associadas a Fase S/genética , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Humanos , Animais , Camundongos , Linhagem Celular Tumoral , Ácido Graxo Sintases/metabolismo , Ácido Graxo Sintase Tipo I/metabolismo , Ácido Graxo Sintase Tipo I/genética , Regulação para Baixo , MasculinoRESUMO
TBX3 behaves as a tumor suppressor or oncoprotein across cancer. However, TBX3 function remains undetermined in intrahepatic cholangiocarcinoma (iCCA), a deadly primary liver malignancy with few systemic treatment options. This study sought to investigate the impact of TBX3 on iCCA. We found that overexpression of TBX3 strongly inhibited human iCCA cell growth. In the Akt/FBXW7ΔF mouse iCCA model, overexpression of Tbx3 reduced cholangiocarcinogenesis in vivo, while inducible genetic knockout of Tbx3 accelerated iCCA growth. RNA-seq identified MAD2L1 as a downregulated gene in TBX3-overexpressing cells, and ChIP confirmed that TBX3 binds to the MAD2L1 promoter. CRISPR-mediated knockdown of Mad2l1 significantly reduced the growth of two iCCA models in vivo. Finally, we found that TBX3 expression is upregulated in ~20% of human iCCA samples, and its high expression is associated with less proliferation and better survival. MAD2L1 expression is upregulated in most human iCCA samples and negatively correlated with TBX3 expression. Altogether, our findings suggest that overexpression of TBX3 suppresses CCA progression via repressing MAD2L1 expression.