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Introduction: Ex vivo organ cultures (EVOC) were recently optimized to sustain cancer tissue for 5 days with its complete microenvironment. We examined the ability of an EVOC platform to predict patient response to cancer therapy. Methods: A multicenter, prospective, single-arm observational trial. Samples were obtained from patients with newly diagnosed bladder cancer who underwent transurethral resection of bladder tumor and from core needle biopsies of patients with metastatic cancer. The tumors were cut into 250 µM slices and cultured within 24 h, then incubated for 96 h with vehicle or intended to treat drug. The cultures were then fixed and stained to analyze their morphology and cell viability. Each EVOC was given a score based on cell viability, level of damage, and Ki67 proliferation, and the scores were correlated with the patients' clinical response assessed by pathology or Response Evaluation Criteria in Solid Tumors (RECIST). Results: The cancer tissue and microenvironment, including endothelial and immune cells, were preserved at high viability with continued cell division for 5 days, demonstrating active cell signaling dynamics. A total of 34 cancer samples were tested by the platform and were correlated with clinical results. A higher EVOC score was correlated with better clinical response. The EVOC system showed a predictive specificity of 77.7% (7/9, 95% CI 0.4-0.97) and a sensitivity of 96% (24/25, 95% CI 0.80-0.99). Conclusion: EVOC cultured for 5 days showed high sensitivity and specificity for predicting clinical response to therapy among patients with muscle-invasive bladder cancer and other solid tumors.
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The core Hippo pathway module consists of a tumour-suppressive kinase cascade that inhibits the transcriptional coactivators Yes-associated protein (YAP) and WW domain-containing transcription regulator protein 1 (WWTR1; also known as TAZ). When the Hippo pathway is downregulated, as often occurs in breast cancer, YAP/TAZ activity is induced. To elaborate the roles of TAZ in triple-negative breast cancer (TNBC), we depleted Taz in murine TNBC 4T1 cells, using either CRISPR/Cas9 or small hairpin RNA (shRNA). TAZ-depleted cells and their controls, harbouring wild-type levels of TAZ, were orthotopically injected into the mammary fat pads of syngeneic BALB/c female mice, and mice were monitored for tumour growth. TAZ depletion resulted in smaller tumours compared to the tumours generated by control cells, in line with the notion that TAZ functions as an oncogene in breast cancer. Tumours, as well as their corresponding in vitro cultured cells, were then subjected to gene expression profiling by RNA sequencing (RNA-seq). Interestingly, pathway analysis of the RNA-seq data indicated a TAZ-dependent enrichment of 'Inflammatory Response', a pathway correlated with TAZ expression levels also in human breast cancer tumours. Specifically, the RNA-seq analysis predicted a significant depletion of regulatory T cells (Tregs) in TAZ-deficient tumours, which was experimentally validated by the staining of tumour sections and by quantitative cytometry by time of flight (CyTOF). Strikingly, the differences in tumour size were completely abolished in immune-deficient mice, demonstrating that the immune-modulatory capacity of TAZ is critical for its oncogenic activity in this setting. Cytokine array analysis of conditioned medium from cultured cells revealed that TAZ increased the abundance of a small group of cytokines, including plasminogen activator inhibitor 1 (Serpin E1; also known as PAI-1), CCN family member 4 (CCN4; also known as WISP-1) and interleukin-23 (IL-23), suggesting a potential mechanistic explanation for its in vivo immunomodulatory effect. Together, our results imply that TAZ functions in a non-cell-autonomous manner to modify the tumour immune microenvironment and dampen the anti-tumour immune response, thereby facilitating tumour growth.
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Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional , Neoplasias de Mama Triplo Negativas , Animais , Feminino , Humanos , Camundongos , Via de Sinalização Hippo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Fatores de Transcrição/metabolismo , Neoplasias de Mama Triplo Negativas/genética , Neoplasias de Mama Triplo Negativas/patologia , Microambiente Tumoral , Proteínas com Motivo de Ligação a PDZ com Coativador Transcricional/metabolismoRESUMO
Breast cancer, the most frequent cancer in women, is generally classified into several distinct histological and molecular subtypes. However, single-cell technologies have revealed remarkable cellular and functional heterogeneity across subtypes and even within individual breast tumors. Much of this heterogeneity is attributable to dynamic alterations in the epigenetic landscape of the cancer cells, which promote phenotypic plasticity. Such plasticity, including transition from luminal to basal-like cell identity, can promote disease aggressiveness. We now report that the tumor suppressor LATS1, whose expression is often downregulated in human breast cancer, helps maintain luminal breast cancer cell identity by reducing the chromatin accessibility of genes that are characteristic of a "basal-like" state, preventing their spurious activation. This is achieved via interaction of LATS1 with the NCOR1 nuclear corepressor and recruitment of HDAC1, driving histone H3K27 deacetylation near NCOR1-repressed "basal-like" genes. Consequently, decreased expression of LATS1 elevates the expression of such genes and facilitates slippage towards a more basal-like phenotypic identity. We propose that by enforcing rigorous silencing of repressed genes, the LATS1-NCOR1 axis maintains luminal cell identity and restricts breast cancer progression.
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Neoplasias da Mama , Feminino , Humanos , Neoplasias da Mama/genética , Genes Reguladores , Proteínas Serina-Treonina Quinases/genética , Mama , Repressão Psicológica , Correpressor 1 de Receptor Nuclear/genéticaRESUMO
High-grade serous ovarian carcinoma (HGSOC) is the most common type of epithelial ovarian cancer. The majority of cases are diagnosed at advanced stages, when intraperitoneal (IP) spread has already occurred. Despite significant surgical and chemotherapeutic advances in HGSOC treatment over the past decades, survival rates with HGSOC have only modestly improved. Chimeric antigen receptor (CAR)-T cells enable T cells to directly bind to tumor-associated antigens in a major histocompatibility complex-independent manner, thereby inducing tumor rejection. While CAR-T cell therapy shows great promise in hematological malignancies, its use in solid tumors is limited. Therefore, innovative approaches are needed to increase the specificity of CAR-modified T cells against solid tumors. The aim of this study was to assess the efficacy and safety of intraperitoneal (IP) versus intravenous (IV) CAR-T cell therapy in the treatment of HGSOC. We constructed a CAR that targets the ErbB2/HER2 protein (ErbB2CAR), which is overexpressed in HGSOC, and evaluated the functionality of ErbB2CAR on ovarian cancer cell lines (OVCAR8, SKOV3, and NAR). Our findings show that an IP injection of ErbB2CAR-T cells to tumor-bearing mice led to disease remission and increased survival compared to the IV route. Moreover, we found that IP-injected ErbB2CART cells circulate to a lesser extent, making them safer for non-tumor tissues than IV-injected cells. Further supporting our findings, we show that the effect of ErbB2CAR-T cells on primary HGSOC tumors is correlated with ErbB2 expression. Together, these data demonstrate the advantages of an IP administration of CAR-T cells over IV administration, offering not only a safer strategy but also the potential for counteracting the effect of ErbB2CAR in HGSOC. Significance: IP-injected ErbB2CAR-T cells led to disease remission and increased survival compared to the IV route. These findings demonstrate the advantages of IP administration, offering a safe treatment strategy with the potential for counteracting the effect of ErbB2CAR in HGSOC.
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The TP53 gene is mutated in approximately 60% of all colorectal cancer (CRC) cases. Over 20% of all TP53-mutated CRC tumors carry missense mutations at position R175 or R273. Here we report that CRC tumors harboring R273 mutations are more prone to progress to metastatic disease, with decreased survival, than those with R175 mutations. We identify a distinct transcriptional signature orchestrated by p53R273H, implicating activation of oncogenic signaling pathways and predicting worse outcome. These features are shared also with the hotspot mutants p53R248Q and p53R248W. p53R273H selectively promotes rapid CRC cell spreading, migration, invasion and metastasis. The transcriptional output of p53R273H is associated with preferential binding to regulatory elements of R273 signature genes. Thus, different TP53 missense mutations contribute differently to cancer progression. Elucidation of the differential impact of distinct TP53 mutations on disease features may make TP53 mutational information more actionable, holding potential for better precision-based medicine.
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Neoplasias Colorretais , Proteína Supressora de Tumor p53 , Neoplasias Colorretais/genética , Genes p53 , Humanos , Mutação , Fenótipo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Translating preclinical studies to effective treatment protocols and identifying specific therapeutic responses in individuals with cancer is challenging. This may arise due to the complex genetic makeup of tumor cells and the impact of their multifaceted tumor microenvironment on drug response. To find new clinically relevant drug combinations for colorectal cancer (CRC), we prioritized the top five synergistic combinations from a large in vitro screen for ex vivo testing on 29 freshly resected human CRC tumors and found that only the combination of mitogen-activated protein kinase kinase (MEK) and proto-oncogene tyrosine-protein kinase Src (Src) inhibition was effective when tested ex vivo. Pretreatment phosphorylated Src (pSrc) was identified as a predictive biomarker for MEK and Src inhibition only in the absence of KRASG12 mutations. Overall, we demonstrate the potential of using ex vivo platforms to identify drug combinations and discover MEK and Src dual inhibition as an effective drug combination in a predefined subset of individuals with CRC.
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Neoplasias Colorretais , Quinases de Proteína Quinase Ativadas por Mitógeno , Linhagem Celular Tumoral , Proliferação de Células , Neoplasias Colorretais/tratamento farmacológico , Humanos , Mutação , Microambiente TumoralRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal cancer, which is refractory to all currently available treatments and bears dismal prognosis. About 70% of all PDAC cases harbor mutations in the TP53 tumor suppressor gene. Many of those are missense mutations, resulting in abundant production of mutant p53 (mutp53) protein in the cancer cells. Analysis of human PDAC patient data from The Cancer Genome Atlas (TCGA) revealed a negative association between the presence of missense mutp53 and infiltration of CD8+ T cells into the tumor. Moreover, CD8+ T cell infiltration was negatively correlated with the expression of fibrosis-associated genes. Importantly, silencing of endogenous mutp53 in KPC cells, derived from mouse PDAC tumors driven by mutant Kras and mutp53, down-regulated fibrosis and elevated CD8+ T cell infiltration in the tumors arising upon orthotopic injection of these cells into the pancreas of syngeneic mice. Moreover, the tumors generated by mutp53-silenced KPC cells were markedly smaller than those elicited by mutp53-proficient control KPC cells. Altogether, our findings suggest that missense p53 mutations may contribute to worse PDAC prognosis by promoting a more vigorous fibrotic tumor microenvironment and impeding the ability of the immune system to eliminate the cancer cells.
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Carcinoma Ductal Pancreático/genética , Fibrose , Mutação de Sentido Incorreto , Neoplasias Pancreáticas/genética , Proteína Supressora de Tumor p53/genética , Animais , Linfócitos T CD8-Positivos , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Linhagem Celular Tumoral , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Microambiente Tumoral/imunologia , Proteína Supressora de Tumor p53/metabolismo , Neoplasias PancreáticasRESUMO
BACKGROUND: Aim of this study was to evaluate the roles of inflammation and autophagy in obese patients with benign prostatic hyperplasia (BPH) and lower urinary tract symptoms (LUTS). METHODS: We analyzed 150 surgical specimens from patients underwent transurethral resection of the prostate (TURP) for LUTS/BPH (Median age 70.3±8.1 years, median BMI 25.7±4.0 kg/m2 and median PSA 6.0±5.4 ng/mL). All surgical specimens were investigated for the presence inflammatory infiltrates, according to the standardized classification of chronic prostatitis of the National Institute of Health. The inflammatory score (IS Score) was calculated. High IS score was defined as ≥7. Each sample was stained for anti-LC3B (cell signaling) and for anti-P62/SQSTM1 (MBL) according to manufacturer's suggestions and scored as follow: 0 (no dots); 1 (detectable dots in 5-25% of cells); 2 (readily detectable dots in 25-75% of cells); 3 (dots in >75% of cells). High percentage of p62 or LC3B was defined as >25%, whereas low percentage of p62 or LC3B was defined as <25% of cells with dots. RESULTS: Overall 74/150 (49.3%) patients were overweight or obese (BMI >25 kg/m2). Obese patients presented a higher inflammatory score. Obese/overweight patients presented a lower percentage of LC3B (58/74; 78.4%) and higher of p62 (49/74; 66.2%) compared to those of normal weight, which it means a deactivated autophagy (P<0.05). At multivariate analysis LC3B (OR=0.22; CI: 0.069-0.70; P=0.01) percentage and BMI (OR=1.118; CI: 1.001-1.250; P=0.04) were independent risk factors of prostatic inflammation (IS≥7). CONCLUSIONS: Here we confirm the association between obesity and prostatic inflammatory infiltrates and present the first evidence of autophagy deregulation in obese patients with LUTS/BPH. Further studies should better investigate this relationship and provide new possible therapeutic targets.
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Sintomas do Trato Urinário Inferior , Hiperplasia Prostática , Ressecção Transuretral da Próstata , Idoso , Autofagia , Humanos , Inflamação , Sintomas do Trato Urinário Inferior/etiologia , Masculino , Pessoa de Meia-Idade , Obesidade/complicações , Hiperplasia Prostática/complicaçõesRESUMO
Bacteria were first detected in human tumors more than 100 years ago, but the characterization of the tumor microbiome has remained challenging because of its low biomass. We undertook a comprehensive analysis of the tumor microbiome, studying 1526 tumors and their adjacent normal tissues across seven cancer types, including breast, lung, ovary, pancreas, melanoma, bone, and brain tumors. We found that each tumor type has a distinct microbiome composition and that breast cancer has a particularly rich and diverse microbiome. The intratumor bacteria are mostly intracellular and are present in both cancer and immune cells. We also noted correlations between intratumor bacteria or their predicted functions with tumor types and subtypes, patients' smoking status, and the response to immunotherapy.
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Bactérias/classificação , Microbiota , Neoplasias/microbiologia , Bactérias/genética , Bactérias/isolamento & purificação , Mama/microbiologia , Colo/microbiologia , Feminino , Humanos , Imunoterapia , Pulmão/microbiologia , Macrófagos/microbiologia , Masculino , Neoplasias/terapia , Ovário/microbiologia , RNA Ribossômico 16S/genéticaRESUMO
INTRODUCTION: Recently TRG, necrosis grade and the rate of viable cancer cells of colorectal liver metastases were correlated with the response to chemotherapy treatments, whereas K-RAS mutations and c-MET over-expression were correlated with the prognosis. METHODS: 58 resection specimens were assessed for regression grades. Patients undergone neo-adjuvant treatments were compared to patients who underwent therapy exclusively adjuvantly. We investigated the K-RAS mutational profile, the c-MET over-expression along with patients' survivals curves. RESULTS: Patients undergone neo-adjuvant treatment presented significant higher fibrosis rates and lower rates of viable cells. 36.7% of the patients had a K-RAS mutation and the 26.7% presented c-MET over-expression, but these features did not correlate with patients' clinical/pathological data. Survival analysis documented that K-RAS WT patients presenting c-MET over-expression had worse outcomes. CONCLUSION: Fibrosis and the rate of viable cells significantly correlate with the response to chemotherapy treatments. c-MET is a promising marker in K-RAS WT patients.
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Adenocarcinoma/secundário , Neoplasias Colorretais/patologia , Neoplasias Hepáticas/secundário , Proteínas Proto-Oncogênicas c-met/biossíntese , Proteínas Proto-Oncogênicas p21(ras)/genética , Adenocarcinoma/genética , Adenocarcinoma/terapia , Adulto , Idoso , Idoso de 80 Anos ou mais , Biomarcadores Tumorais/análise , Neoplasias Colorretais/genética , Neoplasias Colorretais/terapia , Análise Mutacional de DNA , Intervalo Livre de Doença , Feminino , Fibrose/patologia , Humanos , Estimativa de Kaplan-Meier , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/terapia , Masculino , Pessoa de Meia-Idade , Terapia NeoadjuvanteRESUMO
Hodgkin lymphoma (HL) associated with Warthin tumor (WT) is extremely rare, accounting for only 3 cases of classical HLs. Here, we report for the first time the occurrence of a nodular lymphocyte-predominant Hodgkin lymphoma (NLPHL) involving the lymphoid stroma of a WT of the parotid gland. Pathogenesis of WT is controversial, with both a nodal and a parenchymal possible origin. On the other hand, extranodal involvement by HLs is uncommon. In our case, the coexistence of a WT and of a NLPHL within its stroma and in cervical lymph node emphasizes the importance of a careful evaluation of the lymphoid tissue in WT in order to exclude the possibility of an associated lymphoid malignancy.
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Adenolinfoma/patologia , Doença de Hodgkin/patologia , Linfócitos/patologia , Neoplasias Primárias Múltiplas/patologia , Neoplasias Parotídeas/patologia , Idoso , Humanos , MasculinoRESUMO
Abstract Painful bladder syndrome/Interstitial cystitis (PBS/IC) is a chronic disorder characterized clinically by recurring episodes of pelvic pain and increased urination frequency, significantly impairing patients' quality of life. Despite this, there is an unmet medical need in terms of effective diagnostics and treatment. Animal models are crucial in this endeavor. Systemic chronic administration of cyclophosphamide (CYP) in mice has been proposed as a relevant preclinical model of chronic bladder pain. However, molecular mechanisms underlying the pathogenesis of this model are lacking. Here, we show that mice, subjected to repetitive systemic injections of CYP, developed mild inflammatory response in bladder tissue characterized by submucosal edema, moderate increase in proinflammatory cytokine gene expression, and mastocytosis. No signs of massive inflammatory infiltrate, tissue hemorrhages, mucosal ulcerations and urothelium loss were observed. Instead, CYP treatment induced urothelium hyperplasia, accompanied by activation of proliferative signaling cascades, and a decrease in the expression of urothelium-specific markers. Metabotropic glutamate (mGlu) receptors have been implicated in chronic pain disorders. CYP administration induced differential changes in mGlu receptors mRNA levels in bladder tissue, without affecting gene expression at spinal cord level, pointing to the potential link between peripheral mGlu receptors and inflammation-induced bladder malfunction and hyperalgesia. Taken together, these data indicate that chronic CYP treatment in mice is a model of PBS mostly relevant to the major, nonulcerative subtype of the syndrome, characterized by a relatively unaltered mucosa and a sparse inflammatory response. This model can help to elucidate the pathogenetic mechanisms of the disease.