Bladder cancer - the tenth most frequent cancer worldwide - has a heterogeneous natural history and clinical behaviour. The predominant histological subtype, urothelial bladder carcinoma, is ...characterized by high recurrence rates, progression and both primary and acquired resistance to platinum-based therapy, which impose a considerable economic burden on health-care systems and have substantial effects on the quality of life and the overall outcomes of patients with bladder cancer. The incidence of urothelial tumours is increasing owing to population growth and ageing, so novel therapeutic options are vital. Based on work by The Cancer Genome Atlas project, which has identified targetable vulnerabilities in bladder cancer, immune checkpoint inhibitors (ICIs) have arisen as an effective alternative for managing advanced disease. However, although ICIs have shown durable responses in a subset of patients with bladder cancer, the overall response rate is only ~15-25%, which increases the demand for biomarkers of response and therapeutic strategies that can overcome resistance to ICIs. In ICI non-responders, cancer cells use effective mechanisms to evade immune cell antitumour activity; the overlapping Warburg effect machinery of cancer and immune cells is a putative determinant of the immunosuppressive phenotype in bladder cancer. This energetic interplay between tumour and immune cells leads to metabolic competition in the tumour ecosystem, limiting nutrient availability and leading to microenvironmental acidosis, which hinders immune cell function. Thus, molecular hallmarks of cancer cell metabolism are potential therapeutic targets, not only to eliminate malignant cells but also to boost the efficacy of immunotherapy. In this sense, integrating the targeting of tumour metabolism into immunotherapy design seems a rational approach to improve the therapeutic efficacy of ICIs.
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FZAB, GEOZS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Tumour-associated macrophages have been implicated in pancreatic ductal adenocarcinoma (PDAC) therapy response and Extracellular vesicles (EVs) shed by macrophages might have a role in this process. ...Here, we demonstrated that large EVs released by anti-inflammatory human macrophages decreased PDAC cellular sensitivity to gemcitabine. Using proteomic analysis, chitinase 3-like-1 (CHI3L1) and fibronectin (FN1) were identified as two of the most abundant proteins in the cargo of macrophages-derived EVs. Overexpression of CHI3L1 and FN1, using recombinant human proteins, induced PDAC cellular resistance to gemcitabine through ERK (extracellular-signal-regulated kinase) activation. Inhibition of CHI3L1 and FN1 by pentoxifylline and pirfenidone, respectively, partially reverted gemcitabine resistance. In PDAC patient samples, CHI3L1 and FN1 were expressed in the stroma, associated with the high presence of macrophages. The Cancer Genome Atlas analysis revealed an association between CHI3L1 and FN1 gene expression, overall survival of PDAC patients, gemcitabine response, and macrophage infiltration. Altogether, our data identifies CHI3L1 and FN1 as potential targets for pharmacological inhibition in PDAC. Further pre-clinical in vivo work is warranted to study the possibility of repurposing pentoxifylline and pirfenidone as adjuvant therapies for PDAC treatment.
•CHI3L1 and FN1: the most abundant cargo proteins released by human macrophage EVs.•Both CHI3L1 and FN1 cause PDAC resistance to gemcitabine through the ERK signalling.•Pentoxifylline and Pirfenidone revert CHI3L1 and FN1 induced gemcitabine resistance.•CHI3L1 and FN1 are expressed in PDAC macrophage-enriched stroma.•The genes CHI3L1 and FN1 are associated with low overall survival of PDAC patients.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Muscle invasive bladder cancer (MIBC, stage ≥T2) is generally associated with poor prognosis, constituting the second most common cause of death among genitourinary tumours. Due to high molecular ...heterogeneity significant variations in the natural history and disease outcome have been observed. This has also delayed the introduction of personalized therapeutics, making advanced stage bladder cancer almost an orphan disease in terms of treatment. Altered protein glycosylation translated by the expression of the sialyl-Tn antigen (STn) and its precursor Tn as well as the activation of the PI3K/Akt/mTOR pathway are cancer-associated events that may hold potential for patient stratification and guided therapy. Therefore, a retrospective design, 96 bladder tumours of different stages (Ta, T1-T4) was screened for STn and phosphorylated forms of Akt (pAkt), mTOR (pmTOR), S6 (pS6) and PTEN, related with the activation of the PI3K/Akt/mTOR pathway. In our series the expression of Tn was residual and was not linked to stage or outcome, while STn was statically higher in MIBC when compared to non-muscle invasive tumours (p = 0.001) and associated decreased cancer-specific survival (log rank p = 0.024). Conversely, PI3K/Akt/mTOR pathway intermediates showed an equal distribution between non-muscle invasive bladder cancer (NMIBC) and MIBC and did not associate with cancer-specif survival (CSS) in any of these groups. However, the overexpression of pAKT, pmTOR and/or pS6 allowed discriminating STn-positive advanced stage bladder tumours facing worst CSS (p = 0.027). Furthermore, multivariate Cox regression analysis revealed that overexpression of PI3K/Akt/mTOR pathway proteins in STn+ MIBC was independently associated with approximately 6-fold risk of death by cancer (p = 0.039). Mice bearing advanced stage chemically-induced bladder tumours mimicking the histological and molecular nature of human tumours were then administrated with mTOR-pathway inhibitor sirolimus (rapamycin). This decreased the number of invasive lesions and, concomitantly, the expression of STn and also pS6, the downstream effector of the PI3K/Akt/mTOR pathway. In conclusion, STn was found to be marker of poor prognosis in bladder cancer and, in combination with PI3K/Akt/mTOR pathway evaluation, holds potential to improve the stratification of stage disease. Animal experiments suggest that mTOR pathway inhibition could be a potential therapeutic approach for this specific subtype of MIBC.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Circulating O-glycoproteins shed from cancer cells represent important serum biomarkers for diagnostic and prognostic purposes. We have recently shown that selective detection of cancer-associated ...aberrant glycoforms of circulating O-glycoprotein biomarkers can increase specificity of cancer biomarker assays. However, the current knowledge of secreted and circulating O-glycoproteins is limited. Here, we used the COSMC KO “SimpleCell” (SC) strategy to characterize the O-glycoproteome of two gastric cancer SimpleCell lines (AGS, MKN45) as well as a gastric cell line (KATO III) which naturally expresses at least partially truncated O-glycans. Overall, we identified 499 O-glycoproteins and 1236 O-glycosites in gastric cancer SimpleCells, and a total 47 O-glycoproteins and 73 O-glycosites in the KATO III cell line. We next modified the glycoproteomic strategy to apply it to pools of sera from gastric cancer and healthy individuals to identify circulating O-glycoproteins with the STn glycoform. We identified 37 O-glycoproteins in the pool of cancer sera, and only nine of these were also found in sera from healthy individuals. Two identified candidate O-glycoprotein biomarkers (CD44 and GalNAc-T5) circulating with the STn glycoform were further validated as being expressed in gastric cancer tissue. A proximity ligation assay was used to show that CD44 was expressed with the STn glycoform in gastric cancer tissues. The study provides a discovery strategy for aberrantly glycosylated O-glycoproteins and a set of O-glycoprotein candidates with biomarker potential in gastric cancer.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Pirfenidone, an antifibrotic drug, has antitumor potential against different types of cancers. Our work explored whether pirfenidone sensitizes non-small cell lung cancer (NSCLC) cell lines to ...chemotherapeutic treatments. The cytotoxic effect of paclitaxel in combination with pirfenidone against three NSCLC cell lines (A549, NCI-H322 and NCI-H460) was evaluated using the sulforhodamine B assay. The effects of this combination on cell viability (trypan blue exclusion assay), proliferation (BrdU incorporation assay), cell cycle (flow cytometry following PI staining) and cell death (Annexin V-FITC detection assay and Western blot) were analyzed on the most sensitive cell line (NCI-H460). The cytotoxic effect of this drug combination was also evaluated against two non-tumorigenic cell lines (MCF-10A and MCF-12A). Finally, the ability of pirfenidone to sensitize NCI-H460 cells to a combination of paclitaxel plus carboplatin was assessed. The results demonstrated that pirfenidone sensitized NCI-H460 cells to paclitaxel treatment, reducing cell growth, viability and proliferation, inducing alterations in the cell cycle profile and causing an increase in the % of cell death. Remarkably, this combination did not increase cytotoxicity in non-tumorigenic cells. Importantly, pirfenidone also sensitized NCI-H460 cells to paclitaxel plus carboplatin. This work highlights the possibility of repurposing pirfenidone in combination with chemotherapy for the treatment of NSCLC.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Estrogen receptors alpha (ERα) and beta (ERβ) and the cooperating protein GATA-binding factor 3 (GATA3) have been implicated in bladder carcinogenesis and tumour progression. GATA3 and ER have been ...functionally linked in the establishment of luminal fate in breast tissue, but to date their relationship in bladder cancer has not been established. This information will be useful to advance diagnostic and prognostic markers.
To determine the relationship between the expression of ERα, ERβ and GATA3 in bladder cancer, disclose their prognostic and diagnostic value and their association with clinicopathological characteristics.
A comprehensive literature search in PubMed database was performed for all immunohistochemical studies of ERα, ERβ and/or GATA3 in bladder cancer patients. We selected eligible studies in accordance with the PRISMA guidelines and evaluated methodological quality and risk of bias based on quality criteria from the reporting recommendations for tumour MARKer (REMARK) prognostic studies. Risk of bias assessment was performed using Review Manager 5. R software was used for all statistical analysis, the packages used were meta and dmetar for the standard meta-analysis, and netmeta for the network meta-analysis.
Thirteen studies were eligible for ERα, 5 for ERβ and 58 for GATA3 meta-analysis. Low grade tumours showed significantly lower ERα expression. GATA3 was widely expressed in bladder tumours, especially urothelial carcinomas, with higher expression of GATA3 in low grade and low stage tumours. Data was insufficient to determine the prognostic value of either ERα or ERβ, but GATA3-positivity was associated with higher recurrence free survival. A negative correlation between ERα or ERβ positivity and GATA3 expression was disclosed. Additionally, several sources of heterogeneity were identified, which can be used to improve future studies.
The clinicopathological value of ERα and ERβ was inconclusive due to low availability of studies using validated antibodies. Still, this meta-analysis supports GATA3 as good prognostic marker. On the contrary, ERα-positivity was associated to higher grade tumours; while ERα and ERβ were inversely correlated with GATA3 expression. Considering that it has previously been shown that bladder cancer cell lines have functional ERs, this suggests that ERα could be activated in less differentiated cells and independently of GATA3. Therefore, a comprehensive analysis of ERα and ERβ expression in BlaCa supported by complete patient clinical history is required for the identification of BlaCa subtypes and subgroups of patients expressing ERα, to investigate if they could benefit from treatment with hormonal therapy.
Prospero, CRD42021226836.
Monocarboxylate transporters (MCTs) are vital for intracellular pH homeostasis by extruding lactate from highly glycolytic cells. These molecules are key players of the metabolic reprogramming of ...cancer cells, and evidence indicates a potential contribution in urothelial bladder cancer (UBC) aggressiveness and chemoresistance. However, the specific role of MCTs in the metabolic compartmentalization within bladder tumors, namely their preponderance on the tumor stroma, remains to be elucidated. Thus, we evaluated the immunoexpression of MCTs in the different compartments of UBC tissue samples (n = 111), assessing the correlations among them and with the clinical and prognostic parameters. A significant decrease in positivity for MCT1 and MCT4 occurred from normoxic toward hypoxic regions. Significant associations were found between the expression of MCT4 in hypoxic tumor cells and in the tumor stroma. MCT1 staining in normoxic tumor areas, and MCT4 staining in hypoxic regions, in the tumor stroma and in the blood vessels were significantly associated with UBC aggressiveness. MCT4 concomitant positivity in hypoxic tumor cells and in the tumor stroma, as well as positivity in each of these regions concomitant with MCT1 positivity in normoxic tumor cells, was significantly associated with an unfavourable clinicopathological profile, and predicted lower overall survival rates among patients receiving platinum-based chemotherapy. Our results point to the existence of a multi-compartment metabolic model in UBC, providing evidence of a metabolic coupling between catabolic stromal and cancer cells' compartments, and the anabolic cancer cells. It is urgent to further explore the involvement of this metabolic coupling in UBC progression and chemoresistance.
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BFBNIB, GIS, IJS, KISLJ, NUK, PNG, UL, UM, UPUK
Abstract Purpose Pancreatic ductal adenocarcinoma (PDAC) is among the most aggressive malignancies. Our previous work revealed Chitinase 3-like 1 (CHI3L1) involvement in PDAC resistance to ...gemcitabine, identifying it as a promising therapeutic target. Here, we aimed to identify putative CHI3L1 inhibitors and to investigate their chemosensitizing potential in PDAC. Methods Docking analysis for CHI3L1 identified promising CHI3L1 inhibitors, including darifenacin (muscarinic receptor antagonist). PDAC cell lines (BxPC-3, PANC-1) and primary PDAC cells were used to evaluate darifenacin’s effects on cell growth (Sulforhodamine B, SRB), alone or in combination with gemcitabine or gemcitabine plus paclitaxel. Cytotoxicity against normal immortalized pancreatic ductal cells (HPNE) was assessed. Recombinant protein was used to confirm the impact of darifenacin on CHI3L1-induced PDAC cellular resistance to therapy (SRB assay). Darifenacin’s effect on Akt activation was analysed by ELISA. The association between cholinergic receptor muscarinic 3 (CHRM3) expression and therapeutic response was evaluated by immunohistochemistry of paraffin-embedded tissues from surgical resections of a 68 patients’ cohort. Results In silico screening revealed the ability of darifenacin to target CHI3L1 with high efficiency. Darifenacin inhibited PDAC cell growth, with a GI 50 of 26 and 13.6 µM in BxPC-3 and PANC-1 cells, respectively. These results were confirmed in primary PDAC-3 cells, while darifenacin showed no cytotoxicity against HPNE cells. Importantly, darifenacin sensitized PDAC cells to standard chemotherapies, reverted CHI3L1-induced PDAC cellular resistance to therapy, and decreased Akt phosphorylation. Additionally, high CHMR3 expression was associated with low therapeutic response to gemcitabine. Conclusion This work highlights the potential of darifenacin as a chemosensitizer for PDAC treatment.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
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•Low-field NMR associated with PLS were used to monitoring the quality of biodiesel.•A new methodology to determine kinematic viscosity, specific mass and refractive index in ...biodiesel is proposed.•The new methodology is exact, rapid, nondestructive and low-cost.
Because the methods specified by regulatory agencies for the determination of the physicochemical properties of biodiesel can be laborious and expensive, the development of alternative methodologies represents a major breakthrough. Thus, low-field nuclear magnetic resonance (NMR) is an advantageous option because it is nondestructive and reduces the cost and time consumption. In this study, the partial least squares (PLS) regression method was used to create models that correlated the decay curves of the Carr–Purcell–Meiboom–Gill (CPMG) signal, obtained from low-field NMR equipment (2.2 MHz for 1H), with the kinematic viscosity, specific mass and refractive index of biodiesel and their blends. Seventeen oilseeds diversified between edible and non-edible oils were utilized to synthesize the biodiesel and produce binary blends. Separately, multivariate calibration models were created only with biodiesel and blends with castor bean because these samples showed different tendencies from the others. The values of root mean square error of prediction (RMSEP) for the kinematic viscosity, specific mass and refractive index were equal to 0.1 mm2·s−1, 3.7 kg·m−3 and 0.002, respectively, for samples of biodiesel and blends without castor bean and 0.6 mm2·s−1, 2.0 kg·m−3 and 0.0005 for samples of biodiesel and blends with castor bean. The results reveal that the developed models are very satisfactory to predict the quality parameters of biodiesel and blends based on CPMG data with fairly good efficacy.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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