scholarly journals Tumor Microenvironment Alters Chemoresistance of Hepatocellular Carcinoma Through CYP3A4 Metabolic Activity

2021 ◽  
Author(s):  
Alican Özkan ◽  
Danielle L. Stolley ◽  
Erik N. K. Cressman ◽  
Matthew McMillin ◽  
Sharon DeMorrow ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Lines ◽  
Tumor Biology ◽  
Patient Specific ◽  
Type I ◽  
Activity Levels ◽  
Tissue Stiffness ◽  
Microenvironmental Factors ◽  
The Impact ◽  
Cyp3a4 Expression

AbstractVariations in tumor biology from patient to patient combined with the low overall survival rate of hepatocellular carcinoma (HCC) present significant clinical challenges. During the progression of chronic liver diseases from inflammation to the development of HCC, microenvironmental properties, including tissue stiffness and oxygen concentration, change over time. This can potentially impact drug metabolism and subsequent therapy response to commonly utilized therapeutics, such as doxorubicin, multi-kinase inhibitors (e.g., sorafenib), and other drugs, including immunotherapies. In this study, we utilized four common HCC cell lines embedded in 3D collagen type-I gels of varying stiffnesses to mimic normal and cirrhotic livers with environmental oxygen regulation to quantify the impact of these microenvironmental factors on HCC chemoresistance. In general, we found that HCC cells with higher baseline levels of cytochrome p450-3A4 (CYP3A4) enzyme expression, HepG2 and C3Asub28, exhibited a cirrhosis-dependent increase in doxorubicin chemoresistance. Under the same conditions, HCC cell lines with lower CYP3A4 expression, HuH-7 and Hep3B2, showed a decrease in doxorubicin chemoresistance in response to an increase in microenvironmental stiffness. This differential therapeutic response was correlated with the regulation of CYP3A4 expression levels under the influence of stiffness and oxygen variation. In all tested HCC cell lines, the addition of sorafenib lowered the required doxorubicin dose to induce significant levels of cell death, demonstrating its potential to help reduce systemic doxorubicin toxicity when used in combination. These results suggest that patient-specific tumor microenvironmental factors, including tissue stiffness, hypoxia, and CYP3A4 activity levels, may need to be considered for more effective use of chemotherapeutics in HCC patients.

scholarly journals Tumor Microenvironment Alters Chemoresistance of Hepatocellular Carcinoma Through CYP3A4 Metabolic Activity

2021 ◽  
Vol 11 ◽  
Author(s):  
Alican Özkan ◽  
Danielle L. Stolley ◽  
Erik N. K. Cressman ◽  
Matthew McMillin ◽  
Sharon DeMorrow ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Lines ◽  
Tumor Biology ◽  
Patient Specific ◽  
Type I ◽  
Activity Levels ◽  
Tissue Stiffness ◽  
Microenvironmental Factors ◽  
The Impact ◽  
Cyp3a4 Expression

Variations in tumor biology from patient to patient combined with the low overall survival rate of hepatocellular carcinoma (HCC) present significant clinical challenges. During the progression of chronic liver diseases from inflammation to the development of HCC, microenvironmental properties, including tissue stiffness and oxygen concentration, change over time. This can potentially impact drug metabolism and subsequent therapy response to commonly utilized therapeutics, such as doxorubicin, multi-kinase inhibitors (e.g., sorafenib), and other drugs, including immunotherapies. In this study, we utilized four common HCC cell lines embedded in 3D collagen type-I gels of varying stiffnesses to mimic normal and cirrhotic livers with environmental oxygen regulation to quantify the impact of these microenvironmental factors on HCC chemoresistance. In general, we found that HCC cells with higher baseline levels of cytochrome p450-3A4 (CYP3A4) enzyme expression, HepG2 and C3Asub28, exhibited a cirrhosis-dependent increase in doxorubicin chemoresistance. Under the same conditions, HCC cell lines with lower CYP3A4 expression, HuH-7 and Hep3B2, showed a decrease in doxorubicin chemoresistance in response to an increase in microenvironmental stiffness. This differential therapeutic response was correlated with the regulation of CYP3A4 expression levels under the influence of stiffness and oxygen variation. In all tested HCC cell lines, the addition of sorafenib lowered the required doxorubicin dose to induce significant levels of cell death, demonstrating its potential to help reduce systemic doxorubicin toxicity when used in combination. These results suggest that patient-specific tumor microenvironmental factors, including tissue stiffness, hypoxia, and CYP3A4 activity levels, may need to be considered for more effective use of chemotherapeutics in HCC patients.

scholarly journals ELIMINATOR: Essentiality anaLysIs using MultIsystem Networks And inTeger prOgRamming

2021 ◽  
Author(s):  
Asier Antoranz ◽  
Maria Ortiz ◽  
Jon Pey
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Cancer Cell ◽  
In Silico ◽  
Large Scale ◽  
Gene Knockout ◽  
Patient Specific ◽  
Gene Essentiality ◽  
Molecular Features ◽  
The Impact

A gene is considered as essential when it is indispensable for cells to grow and replicate under a certain environment. However, gene essentiality is not a structural property but rather a contextual one, which depends on the specific biological conditions affecting the cell. This circumstantial essentiality of genes is what brings the attention of scientist since we can identify genes essential for cancer cells but not essential for healthy cells. This same contextuality makes their identification extremely challenging. Huge experimental efforts such as Project Achilles where the essentiality of thousands of genes is measured in over one thousand cell lines together with a plethora of molecular data (transcriptomics, copy number, mutations, etc.) can shed light on the causality behind the essentiality of a gene in a given environment by associating the measured essentiality to molecular features of the cell line. Here, we present an in-silico method for the identification of patient-specific essential genes using constraint-based modelling (CBM). Our method expands the ideas behind traditional CBM to accommodate multisystem networks, that is a biological network that focuses on complex interactions within several biological systems. In essence, it first calculates the minimum number of non-expressed genes required to be active by the cell to sustain life as defined by a set of requirements; and second, it performs an exhaustive in-silico gene knockout to find those that lead to the need of activating extra non-expressed genes. We validated the proposed methodology using a set of 452 cancer cell lines derived from the Cancer Cell Line Encyclopedia where an exhaustive experimental large-scale gene knockout study using CRISPR (Achilles Project) evaluates the impact of each removal. We also show that the integration of different essentiality predictions per gene, what we called Essentiality Congruity Score, (derived from multiple pathways) reduces the number of false positives. Finally, we explored the gene essentiality predictions for a breast cancer patient dataset, and our results showed high concordance with previous publications. These findings suggest that identifying genes whose activity are fundamental to sustain cellular life in a patient-specific manner is feasible using in-silico methods. The patient-level gene essentiality predictions can pave the way for precision medicine by identifying potential drug targets whose deletion can induce death in tumour cells.

scholarly journals BTBD10 is a Prognostic Biomarker Correlated With Immune Infiltration in Hepatocellular Carcinoma

2022 ◽  
Vol 8 ◽  
Author(s):  
Jianhui Li ◽  
Xiaojuan Tian ◽  
Ye Nie ◽  
Ying He ◽  
Wenlong Wu ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Lines ◽  
Tumor Infiltrating Lymphocytes ◽  
Normal Liver ◽  
The Cancer Genome Atlas ◽  
Immune Checkpoints ◽  
Phosphatase 2A ◽  
Cancer Genome Atlas ◽  
Infiltrating Lymphocytes ◽  
The Impact

Background: BTBD10 serves as an activator of Akt family members through decreasing the protein phosphatase 2A-mediated dephosphorylation. The present study attempted to investigate the prognostic value of BTBD10 in hepatocellular carcinoma (HCC), specially, its relationship with tumor-infiltrating lymphocytes (TILs).Methods: BTBD10 expression was evaluated in HCC using The Cancer Genome Atlas (TCGA) and Xijing Hospital database, and verified in HCC cell lines. Cox analyses were performed to analyze independent prognostic risk factors for HCC. The optimal cut-off value of BTBD10 was calculated, by which all patients were divided into two groups to compare the overall survival (OS). The signaling pathways were predicted, by which BTBD10 may affect the progression of HCC. To investigate the impact of BTBD10 on HCC immunotherapy, correlations between BTBD10 and TILs, immune checkpoints, m6A methylation-related genes and ferroptosis-related genes were assessed. The distribution of half-maximal inhibitory concentration (IC50) of diverse targeted drugs was observed based on the differential expression of BTBD10.Results: BTBD10 expression was higher in HCC tissues and cell lines than that of normal liver tissues and cells. The patients with high expression of BTBD10 showed a worse OS, as compared to that of BTBD10 low-expressing group. Cox analyses indicated that BTBD10 was an independent prognostic risk factor for HCC. Several molecular pathways of immune responses were activated in HCC patients with high-expressing of BTBD10. Furthermore, BTBD10 expression was demonstrated to be positively correlated with tumor-infiltrating B cells, T cells, macrophages, neutrophils and dendritic cells. Meanwhile, the expression of BTBD10 was synchronized with that of several m6A methylation-related genes, ferroptosis-related genes and immune checkpoints. The IC50 scores of Sorafenib, Navitoclax, Veliparib, Luminespib, and Imatinib were found to be lower in BTBD10 high-expressing HCC group.Conclusion: BTBD10 negatively regulates tumor immunity in HCC and exhibits adverse effect on the prognosis of HCC, which could be a potential target for immunotherapy.

Targeting of SMAD5 Links MicroRNA-155 to the TGFβ Pathway and Lymphomagenesis.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 315-315
Author(s):  
Deepak Rai ◽  
Sang-Woo Kim ◽  
Morgan R McKeller ◽  
Ricardo C Aguiar
Keyword(s):  
Cell Lines ◽  
B Cell Lymphoma ◽  
Type I ◽  
Inhibitory Effects ◽  
Tgfβ Receptors ◽  
Growth Inhibitory ◽  
The Impact ◽  
Tgfβ Pathway

Abstract Abstract 315 The mechanisms by which microRNA (miRNA) dysfunction contribute to the pathogenesis of diffuse large B-cell lymphoma (DLBCL) are not well established. MiRNAs are small non-protein coding RNAs that function by regulating the expression of target transcripts. Thus, to capture the physiologic and pathologic impact of these small regulatory molecules it is necessary to identify the genes that they inhibit. MicroRNA-155 (miR-155) is overexpressed in aggressive subsets of DLBCL, and its enforced expression in a transgenic, Eμ-miR-155, mouse model is associated with the development of lymphoblastic leukemia/high grade lymphoma. However, although a series of bona-fide miR-155 targets have been identified, it is still unclear how overexpression of this miR-155 contributes to lymphomagenesis. Starting from unbiased genome-wide approaches in DLBCL, we discovered that miR-155 directly binds to two sites in the 3'UTR of the SMAD5 gene, inhibiting its expression. Surprisingly, although SMAD5 activity is classically associated with signals transduced by bone morphogenic protein (BMP) family of cytokines, we found that in DLBCLs a non-canonical signaling module linking TGFβ1 to SMAD5 is also active. Furthermore, using kinase inhibitors specific to BMP or TGFβ receptors, we found that the type I TGFβ receptors, in particular ALK5, were essential for the activation of SMAD5 by TGFβ1 in DLBCL. These data suggested that the impact of miR-155 on the TGFβ pathway could be broader than previously appreciated, and abrogate the growth inhibitory effects of both BMP and TGFβ1 in DLBCL. To test this hypothesis, we generated DLBCL cell lines (Ly1, Ly18 and Ly19) ectopically expressing miR-155 and found that they became resistant to the cytostatic effects of TGFβ1 and BMPs. This blockade in growth inhibition was associated with an impaired cell cycle arrest and defective induction of p21. Next, to firmly establish the role of SMAD5 targeting in the miR-155-mediated disruption of the TGFβ signals, we created DLBCL cell lines stably expressing two independent SMAD5-shRNA constructs. Indeed, knockdown of SMAD5 in DLBCL limited the growth inhibitory effects of BMP and TGFβ1 and abrogated p21 induction, largely recapitulating the effects of miR-155 expression. In addition, using a xenograft model of DLBCL with in vivo luminescent imaging capabilities, we found that miR-155 overexpression yielded larger and more widespread lymphomas. Confirming our in vitro data, DLBCLs with stable SMAD5 knockdown were also more aggressive, indicating that SMAD5 downregulation phenocopies the effects of miR-155 in vivo. To gain further insights into the functional consequences of the miR-155/SMAD5 interplay, we used real-time RT-PCR to measure the transcriptional activation of several SMAD5 direct targets. In DLBCL cell lines ectopically expressing miR-155 or SMAD5 shRNA constructs, we found that the expression of the SMAD5 targets, ID1, ID2, SMAD6, SMAD7, BMPR2 and BMP6, was significantly diminished. These data further stressed the relevance of miR-155-mediated SMAD5 downregulation, and started to unveil the downstream components of SMAD5's tumor suppressive activities. Finally, we expanded our observations to primary tumors, and used western blotting to show that miR-155 overexpressing DLBCLs had significantly lower levels of SMAD5, and exhibited an impaired expression of its transcriptional targets. Together, our data instructed on miR-155 mediated lymphomagenesis, highlighted a hitherto unappreciated role of SMAD5 as a lymphoma suppressor gene, and defined a novel mechanism used by cancer cells to escape TGFβ growth inhibitory effects. Disclosures: No relevant conflicts of interest to declare.

Inhibition of the Human Double Minute (HDM)-2 E3 Ubiquitin Ligase Activates Different Programmed Cell Death (PCD) Pathways in Models of Non-Hodgkin Lymphoma (NHL) with Wild Type (wt) and Mutant (mut) p53

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 3618-3618 ◽  
Author(s):  
Richard Julian Jones ◽  
Janine Arts ◽  
Robert Z Orlowski
Keyword(s):  
Cell Death ◽  
Cell Lines ◽  
Drug Exposure ◽  
Caspase 3 ◽  
Annexin V ◽  
Type I ◽  
Type Ii ◽  
Inhibition Of Proliferation ◽  
Acridine Orange Staining ◽  
The Impact

Abstract Background: The ubiquitin-proteasome pathway has been validated as a target for NHL with the recent approval of bortezomib for mantle cell lymphoma (MCL). In addition to anti-tumor activity, however, proteasome inhibitors have pleiotropic effects, including activation of anti-apoptotic heat shock proteins, and their use clinically is complicated by toxicities such as peripheral neuropathy. By targeting E3 ubiquitin ligases, which are involved in ubiquitination of only a small subset of cellular proteins, it may be possible to achieve more specific anti-tumor effects with a better therapeutic index. One attractive target is HDM-2, which is responsible for ubiquitination of the p53 tumor suppressor. Methods: To evaluate the therapeutic potential of agents targeting HDM-2, we studied the impact of the small molecule JNJ-26854165, an inhibitor of HDM-2-function, in both p53 wt and mut cell line models. Results: Treatment of wt p53 NHL cell lines with JNJ-26854165 induced a dose- and time-dependent inhibition of proliferation, with an IC50 in the 0.02–0.3 μM range. Cell death, which was typically seen within 48 hours of HDM-2 inhibition, occurred through induction of type I PCD, as judged by the appearance of increased staining with Annexin V and activation of caspase 3. While cell lines with mut p53 were generally less sensitive than their wt p53 counterparts, JNJ-26854165 remained potent, with an IC50 in the 0.05–0.6 μM range. The latter cell lines showed a longer kinetics of death, with PCD being seen within 72 hours of drug exposure. Notably, in these mut p53 cell lines, very little Annexin V staining or caspase 3 activation was seen, consistent with a minor role for type I PCD. Instead, mut p53 cell lines demonstrated an increased content of acidic vacuoles by acridine orange staining, increased expression of Beclin 1 and Sequestosome 1/p62, and conversion of microtubule-associated protein 1 light chain 3 form I to form II, consistent with activation of type II PCD, or autophagy. Also, electron microscopy showed an increased presence of autophagosomes and autolysosomes, further supporting the activation of this pathway. Combinations of JNJ-26854165 with other agents, including rapamycin, doxorubicin, and an inhibitor of Bcl 2, showed enhanced anti-proliferative effects in a sequence-dependent fashion, which were greatest when the chemotherapeutic preceded the HDM-2 inhibitor. Combination index analysis revealed that these interactions met criteria for synergy. Conclusions: Inhibition of the function of HDM-2 using JNJ-26854165 is a promising approach that is effective against both wt and mut p53 models by activating type I and type II PCD, respectively. The effectiveness of JNJ-26854165 was enhanced in combination with currently used chemotherapeutics in a sequence specific manner, providing a rationale for translation of this novel approach into the clinic.

scholarly journals NACC-1 regulates hepatocellular carcinoma cell malignancy and is targeted by miR-760

2020 ◽  
Vol 52 (3) ◽  
pp. 302-309
Author(s):  
Linan Yin ◽  
Tingting Sun ◽  
Ruibao Liu
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Lines ◽  
Bioinformatic Analysis ◽  
Cellular Growth ◽  
Luciferase Assay ◽  
Migration And Invasion ◽  
Cellular Processes ◽  
Therapeutic Benefits ◽  
The Impact

Abstract Hepatocellular carcinoma (HCC) is the most prominent form of presentation in liver cancer. It is also the fourth most common cause of cancer-associated deaths globally. The role of nucleus accumbens associated protein-1 (NACC-1) has been evaluated in several cancers. This protein is a transcriptional regulator that regulates a number of significant cellular processes. In the current study, we aimed to understand the role of NACC-1 in HCC. Primarily, we measured the expression of NACC-1 using quantitative real time polymerase chain reaction and western blot analysis. We knocked down the expression of NACC-1 in HCC cell lines Huh7 and HepG2 by transferring a commercially synthesized small interfering RNA and explored the impact of NACC-1 knockdown on cellular growth, migration, invasion, and chemoresistance to doxorubicin. Through bioinformatic analysis, we identified NACC-1 as a potential target of miR-760. Using a dual reporter luciferase assay, we confirmed the predicted target and assessed miR-760-mediated regulation of NACC-1 and rescue of tumorigenic phenotypes. We observed increased expression of NACC-1 in HCC. Furthermore, knockdown of NACC-1 resulted in reduced cell proliferation and invasion and increased susceptibility to doxorubicin-mediated chemosensitivity. Overexpression of miR-760 in HCC cell lines rescued NACC-1-mediated migration and invasion. We revealed that miR-760 regulated NACC-1 expression in HCC. Our data indicated that both miR-760 and NACC-1 could be used as prognostic markers, and miR-760 may have therapeutic benefits for HCC and other cancers.

Anti-Oxidant and Anticancerous Effect of Fomitopsis officinalis (Vill. ex Fr. Bond. et Sing) Mushroom on Hepatocellular Carcinoma Cells In Vitro through NF-kB Pathway

2021 ◽  
Vol 21 ◽  
Author(s):  
Nyamsambuu Altannavch ◽  
Xi Zhou ◽  
Md. Asaduzzaman Khan ◽  
Ashfaque Ahmed ◽  
Shinen Naranmandakh ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Cell Lines ◽  
Cancer Cell ◽  
Annexin V ◽  
Carcinoma Cells ◽  
Stress Disorders ◽  
Phase Cell ◽  
Hepatocellular Carcinoma Cells ◽  
The Impact

Background: Fomitopsis officinalis (Vill. ex Fr. Bond. et Sing) is a medicinal mushroom, commonly called ‘Agarikon’, traditionally used to treat cough and asthma in the Mongolian population. Objective: The objective of this study was to examine the significance of biological activity of F. officinalis, and evaluate the antioxidant and anticancer activity of six fractions of F. officinalis residues (Fo1-powder form dissolved in ethanol, Fo2-petroleum ether residue, Fo3-chloroformic, Fo4-ethylacetate, Fo5-buthanolic, and Fo6-water-ethanolic) against hepatocellular carcinoma cells. Methods: We performed in vitro studies of cell proliferation and viability assay, annexin V-FITC/Propidium Iodide assay, and NF-kB signaling pathway by immunoblot analysis. Results: Our findings revealed that all six fractions/extracts have antioxidant activity, and somehow, they exert anticancerous effects against cancer cells. In cancerous cell lines (HepG2 and LO2), Fo3 chloroformic extract promoted the cancer cell apoptosis, cell viability, activated G2/M-phase cell cycle, and selectively induced NF-kB proteins, revealing itself as a novel antitumor extract. Conclusion: This study reports that Fo3-chloroformic extract is rich in antitumor activity; it was previously not investigated in cancer. To study the impact of F. officinalis among natural products to treat/prevent oxidative stress disorders or cancers, further examinations are needed. However, this study assessed only one extract, Fo3-chloroformic, which has a significant impact on cancer cell lines.

scholarly journals Validation of Hepatocellular Carcinoma Experimental Models for TGF-β Promoting Tumor Progression

Cancers ◽  
2019 ◽  
Vol 11 (10) ◽  
pp. 1510 ◽  
Author(s):  
Serena Mancarella ◽  
Silke Krol ◽  
Alberto Crovace ◽  
Stefano Leporatti ◽  
Francesco Dituri ◽  
...  
Keyword(s):  
Hepatocellular Carcinoma ◽  
Tumor Progression ◽  
Cell Lines ◽  
Transforming Growth Factor Beta ◽  
Transforming Growth Factor ◽  
Tumor Stroma ◽  
Experimental Models ◽  
Tumor Promoter ◽  
Type I

Transforming growth factor beta (TGF-β) is a pleiotropic cytokine with dual role in hepatocellular carcinoma (HCC). It acts as tumor-suppressor and tumor-promoter in the early and late stage respectively. TGF-β influences the tumor-stroma cross-talk affecting the tumoral microenvironment. Therefore, inhibiting the TGF- β mediated pathway alone and/or in combination with chemotherapeutics represents an important therapeutic option. Experimental models to dissect the role of TGF-β in HCC tumor progression as well as the effectiveness of specific inhibitors are tricky. HCC cell lines respond to TGF-β according to their epithelial phenotype. However, the mesenchymal and more aggressive HCC cell lines in vitro, do not develop tumors when transplanted in vivo, thus hampering the understanding of molecular pathways that dictate outcome. In addition, in this model the native immune system is abolished, therefore the contribution of inflammation in hepatocarcinogenesis is unreliable. Different strategies have been set up to engineer HCC animal models, including genetically modified mice, chemically induced HCC, or hydrodynamic techniques. Patient-derived xenograft is currently probably the most fascinating model, keeping in mind that models cannot mirror all the reality. In this context, we discuss the different available HCC mouse models including our experimental model treated with inhibitor of TGF-β receptor Type I kinase (Galunisertib) and a potential role of exosomes in TGF-β moderated tumor progression of HCC. Unfortunately, no positive results were obtained in our treated orthotopic model because it does not reproduce the critical tumor-stroma interactions of the HCC.

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