AHR canonical pathway: in vivo findings to support novel antihypertensive strategies

2021 ◽  
pp. 105407
Author(s):  
Nuno R. Coelho ◽  
Clara Matos ◽  
António B. Pimpão ◽  
M. João Correia ◽  
Catarina O. Sequeira ◽  
...  
Keyword(s):  
Canonical Pathway

scholarly journals Panobinostat (LBH589) in Combination with the β-Catenin Inhibitor Tegavivint (BC2059) Exerts Significant Anti-Myeloma Activity Both in Vitro and In Vivo

Blood ◽  
2019 ◽  
Vol 134 (Supplement_1) ◽  
pp. 5540-5540
Author(s):  
Ioanna Savvidou ◽  
Tiffany Khong ◽  
Sophie Whish ◽  
Tara Sepehrizadeh ◽  
Sridurga Mithraprabhu ◽  
...  
Keyword(s):  
Research Funding ◽  
Volume Fraction ◽  
Aerobic Glycolysis ◽  
Canonical Pathway ◽  
Bone Homeostasis ◽  
Trabecular Thickness ◽  
Single Drug ◽  
Oncology Research

Introduction: Panobinostat (LBH589) is approved for the treatment of relapsed multiple myeloma (MM). The Wnt canonical pathway and its key player, β-catenin are dysregulated in advanced MM supporting the evaluation of β-catenin inhibition as a potential therapy. We evaluated the anti-MM effect of Tegavivint in combination with Panobinostat in vitro and in vivo. Results and Methods : In vitro combination of low doses (BC2059≤100nM; LBH589≤10nM) for 48h was synergistic against OCI-My1 and U266 MM cells, with combination indices from 0.569 to 0.883 (CI<1: synergism). The combination demonstrated synergistic killing of primary MM tumour cells in an autologous co-culture assay with synergism quotients from 1.2 to 2 (SQ>1: synergism). The combination rapidly (<24h) decreased the expression of downstream β-catenin targets myc, cyclin D1 and cyclin D2 as shown by immunoblotting. Furthermore, by 20h, the combination decreased both oxidative phosphorylation and aerobic glycolysis as measured by oxygen consumption rate (OCR) and extracellular acidification rate (ECAR), respectively (Seahorse XFe96 analyser). Basal, maximal and ATP coupled OCR were significantly reduced by the combination when compared to vehicle (for OCI-My1 from 203.93 to 56.20, 105.81 to 33.29 and 156.29 to 34.67 pmol/min/50K cells OCR respectively). Similarly, basal glycolysis and glycolytic capacity were both reduced by the combination (from 48.88 to 9.77 and 61.68 to 8.58 mpH/min/50K cells ECAR, respectively). In vivo, the combination was superior to either single drug treatment in a murine xenograft of U266-luciferase cells. Disease burden was reduced in the combination arm compared to single drug and vehicle arms as early as day 14 after inoculation (p=0.02). This difference increased further until the last bioluminescence imaging (day 49, p<0.001) and translated into significantly prolonged OS for the combination (p=0.006). Potential on-target toxicities with BC2059 are a concern as the Wnt canonical pathway is essential for stem cell maintenance in several organs and has a role in bone homeostasis, but the combination did not result in cytopenias nor body weight loss. After euthanasia, µCT demonstrated that neither BC2059 nor the combination negatively affected bone morphometric indices (bone volume fraction, trabecular thickness, connectivity density). Likewise, osteoblastic activity as measured by serum osteocalcin was unaffected, whereas osteoclastic activity (as indicated by serum CTX1) was reduced when compared to healthy mice (p=0.008 and p=0.013, respectively). In conclusion Tegavivint in combination with Panobinostat may be a useful therapeutic modality for MM patients with advanced/refractory disease and warrants further evaluation. Disclosures Khong: Novartis Oncology: Research Funding. Spencer:Celgene: Consultancy, Honoraria, Research Funding, Speakers Bureau; Amgen: Consultancy, Honoraria, Research Funding; Specialised Therapeutics Australia: Consultancy, Honoraria; Janssen: Consultancy, Honoraria, Research Funding, Speakers Bureau; Abbvie: Consultancy, Honoraria; Takeda: Consultancy, Honoraria, Research Funding, Speakers Bureau; Servier: Consultancy, Honoraria; Secura Bio: Consultancy, Honoraria.

scholarly journals ALIX- and ESCRT-III–dependent sorting of tetraspanins to exosomes

2020 ◽  
Vol 219 (3) ◽  
Author(s):  
Jorge Larios ◽  
Vincent Mercier ◽  
Aurélien Roux ◽  
Jean Gruenberg
Keyword(s):  
Intercellular Communication ◽  
Active Form ◽  
Canonical Pathway ◽  
Supported Bilayers ◽  
Membrane Deformation ◽  
Lysobisphosphatidic Acid ◽  
Signaling Receptors ◽  
Dependent Pathway

The intraluminal vesicles (ILVs) of endosomes mediate the delivery of activated signaling receptors and other proteins to lysosomes for degradation, but they also modulate intercellular communication when secreted as exosomes. The formation of ILVs requires four complexes, ESCRT-0, -I, -II, and -III, with ESCRT-0, -I, and -II presumably involved in cargo sorting and ESCRT-III in membrane deformation and fission. Here, we report that an active form of the ESCRT-associated protein ALIX efficiently recruits ESCRT-III proteins to endosomes. This recruitment occurs independently of other ESCRTs but requires lysobisphosphatidic acid (LBPA) in vivo, and can be reconstituted on supported bilayers in vitro. Our data indicate that this ALIX- and ESCRT-III–dependent pathway promotes the sorting and delivery of tetraspanins to exosomes. We conclude that ALIX provides an additional pathway of ILV formation, secondary to the canonical pathway, and that this pathway controls the targeting of exosomal proteins.

In Vitro and In Vivo Efficacy of a Novel Orally Bioavailable Beta-Catenin Inhibitor-BC2059- As Monotherapy or in Combination with Proteasome Inhibitors in Multiple Myeloma

Blood ◽  
2015 ◽  
Vol 126 (23) ◽  
pp. 1816-1816
Author(s):  
Ioanna Savvidou ◽  
Tiffany T. Khong ◽  
Stephen K. Horrigan ◽  
Andrew Spencer
Keyword(s):  
Multiple Myeloma ◽  
Cell Death ◽  
Single Agent ◽  
Proteasome Inhibitors ◽  
Canonical Pathway ◽  
Beta Catenin ◽  
Dependent Manner ◽  
Canonical Wnt

Abstract Background: The currently available treatment options are unlikely to be curative for the majority of Multiple Myeloma (MM) patients, emphasizing a continuing role for the introduction of investigational agents that overcome drug resistance. The canonical Wnt/β-catenin signalling pathway has been found to be dysregulated in MM, and its activation is associated with advanced stage MM, providing a rationale to evaluate the novel β-catenin inhibitor BC2059 in mono- and combination therapy with proteasome inhibitors in vitro and in vivo. Methods and Results: We evaluated the activation status of the canonical Wnt pathway in 12 genetically heterogeneous Human Myeloma Cell Lines (HMCL) by assessing the expression of β-catenin protein in the nuclear compartment (active form). This showed that nuclear β-catenin was present in all HMCL tested and absent in plasma cells derived from a healthy donor. Moreover, additional stimulation of the canonical pathway with rhWnt3a was shown to be pro-proliferative, in contrast, no proliferation was seen with activation of the non canonical pathway following treatment with rhWnt5a. BC2059 (50nM to 500nM) induced apoptosis of all 12 HMCL and was able to inhibit the proliferation of all HMCL tested in a dose and time dependent manner assessed by MTS assay and viable enumeration with trypan blue (IC50: 53nM to 247nM). Mimicking the bone marrow (BM) microenvironment by co-culturing HMCL with the immortalised human stromal cell line HS-5, BC2059 was able to overcome the protective effect of HS-5 (for example KMS18 at IC90=220nM had no stromal pro-survival effect). Similarly, BC2059 was able to abolish the pro-proliferative effect of rh-Wnt3a or conditioned media derived from MM patients' BM when used at doses >100nM or 50nM, respectively. BC2059 facilitated the degradation of β-catenin protein in the nuclear cellular compartment ( >50% decrease of nuclear β-catenin in KMS18 treated with 1.5xIC50 when compared with untreated cells), furthermore, using a reporter assay we showed that BC2059 inhibited TCF/LEF transcriptional activity in a dose-dependent manner and decreased the transcription of axin2, a down-stream target gene of β-catenin - 78% reduction in KMS18 cells treated with 1.5x IC50 when compared to untreated controls. BC2059-induced HMCL cell death was associated with activation of both the intrinsic and extrinsic caspase-dependent apoptotic pathways, as shown by the accumulation of the activated forms of caspases 8, 9 and 3 following BC2059 treatment. However, inhibition of the caspase-pathway by the addition of caspase inhibitors (pan-caspase inhibitor Z-VAD, and caspase-3 inhibitor Z-DEVD) could not abolish the pro-necrotic effect of BC2059 or BC2059 plus bortezomib, suggesting a possible role for autophagy-induced cell death. As β-catenin undergoes proteasome-mediated destruction and has been found to increase following bortezomib treatment, we evaluated the effect of combining BC2059 with Bortezomib. The combination was synergistic for 6/8 HMCL tested (e.g. for LP1 CI:0.64-0.55, where CI<1.1=synergism). We also evaluated the effect of the combination of BC2059 with next generation proteasome inhibitors (carfilzomib and marizomib) where it was shown to have synergistic and/or additive effects (e.g. for carfilzomib LP1 CI:0.33-0.99). Single agent BC2059 effectively killed primary MM tumour cells from relapsed/refractory MM patients (n=13) and the combination with bortezomib was synergistic (n=2) with no effect on healthy peripheral blood mononuclear cells (n=4). Finally, BC2059 (10mg/kg) prolonged survival of xenografted NSG mice compared to untreated controls with no major side effects in Wnt/β-catenin dependent tissues (GI track and haematopoiesis). Conclusion: We have demonstrated that BC2059 at nano-molar concentrations has a strong anti-MM effect both in vitro and in vivo and synergises with proteasome inhibitors. These data strongly support the clinical evaluation of BC2059 for the treatment of MM. Disclosures Horrigan: BetaCat Pharmaceuticals: Employment.

Fine Structural Changes in the Microvasculature of the Mouse Pinna: Aging and Radiation Injury

1974 ◽  
Vol 32 ◽  
pp. 54-55
Author(s):  
S. Phyllis Steamer ◽  
Rosemarie L. Devine
Keyword(s):  
Structural Changes ◽  
Radiation Treatment ◽  
Arterial Stenosis ◽  
Ultrastructural Changes ◽  
Vascular Effects ◽  
Microvascular Changes ◽  
Surrounding Connective Tissue ◽  
Fission Neutrons ◽  
Total Body

The importance of radiation damage to the skin and its vasculature was recognized by the early radiologists. In more recent studies, vascular effects were shown to involve the endothelium as well as the surrounding connective tissue. Microvascular changes in the mouse pinna were studied in vivo and recorded photographically over a period of 12-18 months. Radiation treatment at 110 days of age was total body exposure to either 240 rad fission neutrons or 855 rad 60Co gamma rays. After in vivo observations in control and irradiated mice, animals were sacrificed for examination of changes in vascular fine structure. Vessels were selected from regions of specific interest that had been identified on photomicrographs. Prominent ultrastructural changes can be attributed to aging as well as to radiation treatment. Of principal concern were determinations of ultrastructural changes associated with venous dilatations, segmental arterial stenosis and tortuosities of both veins and arteries, effects that had been identified on the basis of light microscopic observations. Tortuosities and irregularly dilated vein segments were related to both aging and radiation changes but arterial stenosis was observed only in irradiated animals.

Induction and Differentiation of Dental Epithelium in Vivo and in Vitro

1982 ◽  
Vol 40 ◽  
pp. 142-145
Author(s):  
E. J. Kollar
Keyword(s):  
Connective Tissue ◽  
Oral Mucosa ◽  
Basal Lamina ◽  
Functional Derivatives ◽  
Specific Source ◽  
Dental Epithelium ◽  
Connective Tissue Stroma

The differentiation and maintenance of many specialized epithelial structures are dependent on the underlying connective tissue stroma and on an intact basal lamina. These requirements are especially stringent in the development and maintenance of the skin and oral mucosa. The keratinization patterns of thin or thick cornified layers as well as the appearance of specialized functional derivatives such as hair and teeth can be correlated with the specific source of stroma which supports these differentiated expressions.

Ultrastructural Correlations between Clonal Human Tumor Colonies and in Vivo Neoplasms

1982 ◽  
Vol 40 ◽  
pp. 210-211
Author(s):  
M.J. Murphy ◽  
R.R. Price ◽  
J.C. Sloman
Keyword(s):  
Cultured Cells ◽  
Human Tumor ◽  
Cell Type ◽  
Tumor Mass ◽  
Initial Cell ◽  
Cloning Assay ◽  
Human Tumor Cloning ◽  
The Relationship

The in vitro human tumor cloning assay originally described by Salmon and Hamburger has been applied recently to the investigation of differential anti-tumor drug sensitivities over a broad range of human neoplasms. A major problem in the acceptance of this technique has been the question of the relationship between the cultured cells and the original patient tumor, i.e., whether the colonies that develop derive from the neoplasm or from some other cell type within the initial cell population. A study of the ultrastructural morphology of the cultured cells vs. patient tumor has therefore been undertaken to resolve this question. Direct correlation was assured by division of a common tumor mass at surgical resection, one biopsy being fixed for TEM studies, the second being rapidly transported to the laboratory for culture.

Structural analysis of the photosynthetic membrane from Ectothiorhodospira halochloris

1983 ◽  
Vol 41 ◽  
pp. 740-741
Author(s):  
H. Engelhardt ◽  
R. Guckenberger ◽  
W. Baumeister
Keyword(s):  
Lattice Structure ◽  
Bacterial Species ◽  
Hexagonal Lattice ◽  
Reaction Centre ◽  
Photosynthetic Membrane ◽  
Rhodopseudomonas Viridis ◽  
Photosynthetic Membranes ◽  
Ectothiorhodospira Halochloris ◽  
Main Components

Bacterial photosynthetic membranes contain, apart from lipids and electron transport components, reaction centre (RC) and light harvesting (LH) polypeptides as the main components. The RC-LH complexes in Rhodopseudomonas viridis membranes are known since quite seme time to form a hexagonal lattice structure in vivo; hence this membrane attracted the particular attention of electron microscopists. Contrary to previous claims in the literature we found, however, that 2-D periodically organized photosynthetic membranes are not a unique feature of Rhodopseudomonas viridis. At least five bacterial species, all bacteriophyll b - containing, possess membranes with the RC-LH complexes regularly arrayed. All these membranes appear to have a similar lattice structure and fine-morphology. The lattice spacings of the Ectothiorhodospira haloohloris, Ectothiorhodospira abdelmalekii and Rhodopseudomonas viridis membranes are close to 13 nm, those of Thiocapsa pfennigii and Rhodopseudomonas sulfoviridis are slightly smaller (∼12.5 nm).

The Infection of Cells by Bullet-Shaped Viruses

1969 ◽  
Vol 27 ◽  
pp. 372-373
Author(s):  
Frederick A. Murphy ◽  
Alyne K. Harrison ◽  
Sylvia G. Whitfield
Keyword(s):  
Electron Microscopy ◽  
Physical Properties ◽  
Host Cell ◽  
Thin Section ◽  
Cultured Cells ◽  
Cell Infection ◽  
Thin Section Electron Microscopy ◽  
Section Electron ◽  
Section Electron Microscopy

The bullet-shaped viruses are currently classified together on the basis of similarities in virion morphology and physical properties. Biologically and ecologically the member viruses are extremely diverse. In searching for further bases for making comparisons of these agents, the nature of host cell infection, both in vivo and in cultured cells, has been explored by thin-section electron microscopy.

Ultrastructure of melanocyte-keratinocyte interactions and pigment transfer in vitro

1978 ◽  
Vol 36 (2) ◽  
pp. 650-651
Author(s):  
Raul I. Garcia ◽  
Evelyn A. Flynn ◽  
George Szabo
Keyword(s):  
Direct Injection ◽  
Skin Pigmentation ◽  
Freeze Fracture ◽  
Pigment Granule ◽  
Time Lapse ◽  
Ultrastructural Level ◽  
Lanthanum Tracer ◽  
A Cell

Skin pigmentation in mammals involves the interaction of epidermal melanocytes and keratinocytes in the structural and functional unit known as the Epidermal Melanin Unit. Melanocytes(M) synthesize melanin within specialized membrane-bound organelles, the melanosome or pigment granule. These are subsequently transferred by way of M dendrites to keratinocytes(K) by a mechanism still to be clearly defined. Three different, though not necessarily mutually exclusive, mechanisms of melanosome transfer have been proposed: cytophagocytosis by K of M dendrite tips containing melanosomes, direct injection of melanosomes into the K cytoplasm through a cell-to-cell pore or communicating channel formed by localized fusion of M and K cell membranes, release of melanosomes into the extracellular space(ECS) by exocytosis followed by K uptake using conventional phagocytosis. Variability in methods of transfer has been noted both in vivo and in vitro and there is evidence in support of each transfer mechanism. We Have previously studied M-K interactions in vitro using time-lapse cinemicrography and in vivo at the ultrastructural level using lanthanum tracer and freeze-fracture.

Effect of Some Metabolic Inhibitors on Vinblastine-Induced Ribosomal-Granular Material Complexes

1971 ◽  
Vol 29 ◽  
pp. 548-549
Author(s):  
Awtar Krishan ◽  
Dora Hsu
Keyword(s):  
Granular Material ◽  
Rna Synthesis ◽  
Culture Medium ◽  
Actinomycin D ◽  
Metabolic Inhibitors ◽  
Protein And Rna Synthesis ◽  
Apparent Reduction ◽  
Plant Alkaloids ◽  
In Cells

Cells exposed to antitumor plant alkaloids, vinblastine and vincristine sulfate have large proteinacious crystals and complexes of ribosomes, helical polyribosomes and electron-dense granular material (ribosomal complexes) in their cytoplasm, Binding of H3-colchicine by the in vivo crystals shows that they contain microtubular proteins. Association of ribosomal complexes with the crystals suggests that these structures may be interrelated.In the present study cultured human leukemic lymphoblasts (CCRF-CEM), were incubated with protein and RNA-synthesis inhibitors, p. fluorophenylalanine, puromycin, cycloheximide or actinomycin-D before the addition of crystal-inducing doses of vinblastine to the culture medium. None of these compounds could completely prevent the formation of the ribosomal complexes or the crystals. However, in cells pre-incubated with puromycin, cycloheximide, or actinomycin-D, a reduction in the number and size of the ribosomal complexes was seen. Large helical polyribosomes were absent in the ribosomal complexes of cells treated with puromycin, while in cells exposed to cycloheximide, there was an apparent reduction in the number of ribosomes associated with the ribosomal complexes (Fig. 2).

Sign in / Sign up

Export Citation Format

Share Document