scholarly journals In-depth characterization of a new patient-derived xenograft model for metaplastic breast carcinoma to identify viable biologic targets and patterns of matrix evolution within rare tumor types

2021 ◽  
Author(s):  
M. D. Matossian ◽  
T. Chang ◽  
M. K. Wright ◽  
H. E. Burks ◽  
S. Elliott ◽  
...  
Keyword(s):  
Breast Carcinoma ◽  
Rapid Growth ◽  
Ex Vivo ◽  
Breast Cancer Subtype ◽  
Tumor Biology ◽  
Xenograft Model ◽  
Microscopy Imaging ◽  
Metaplastic Breast Carcinoma

AbstractMetaplastic breast carcinoma (MBC) is a rare breast cancer subtype with rapid growth, high rates of metastasis, recurrence and drug resistance, and diverse molecular and histological heterogeneity. Patient-derived xenografts (PDXs) provide a translational tool and physiologically relevant system to evaluate tumor biology of rare subtypes. Here, we provide an in-depth comprehensive characterization of a new PDX model for MBC, TU-BcX-4IC. TU-BcX-4IC is a clinically aggressive tumor exhibiting rapid growth in vivo, spontaneous metastases, and elevated levels of cell-free DNA and circulating tumor cell DNA. Relative chemosensitivity of primary cells derived from TU-BcX-4IC was performed using the National Cancer Institute (NCI) oncology drug set, crystal violet staining, and cytotoxic live/dead immunofluorescence stains in adherent and organoid culture conditions. We employed novel spheroid/organoid incubation methods (Pu·MA system) to demonstrate that TU-BcX-4IC is resistant to paclitaxel. An innovative physiologically relevant system using human adipose tissue was used to evaluate presence of cancer stem cell-like populations ex vivo. Tissue decellularization, cryogenic-scanning electron microscopy imaging and rheometry revealed consistent matrix architecture and stiffness were consistent despite serial transplantation. Matrix-associated gene pathways were essentially unchanged with serial passages, as determined by qPCR and RNA sequencing, suggesting utility of decellularized PDXs for in vitro screens. We determined type V collagen to be present throughout all serial passage of TU-BcX-4IC tumor, suggesting it is required for tumor maintenance and is a potential viable target for MBC. In this study we introduce an innovative and translational model system to study cell–matrix interactions in rare cancer types using higher passage PDX tissue.

scholarly journals Development and characterization of a novel human 3D model of bone metastasis from breast carcinoma in vitro cultured

2021 ◽  
Vol 4 (s1) ◽  
Author(s):  
Deyanira Contartese ◽  
Francesca Salamanna ◽  
Veronica Borsari ◽  
Stefania Pagani ◽  
Maria Sartori ◽  
...  
Keyword(s):  
Bone Metastasis ◽  
Breast Carcinoma ◽  
3D Model ◽  
Ex Vivo ◽  
Ex Vivo Model ◽  
Fresh Tissue ◽  
Vertebral Bone ◽  
Set Up

Breast cancer frequently metastasizes to the skeleton causing significant morbidity. Here, we set-up a novel and advanced ex vivo model by using fresh tissue from human vertebral bone metastasis from breast carcinoma patients able to retain the tumor microenvironment and tumor cells heterogeneity.

scholarly journals hPSC-Derived Enteric Ganglioids Model Human ENS Development and Function

2022 ◽  
Author(s):  
Homa Majd ◽  
Ryan M Samuel ◽  
Jonathan T Ramirez ◽  
Ali Kalantari ◽  
Kevin Barber ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Xenograft Model ◽  
Developmental Relationships ◽  
Drug Candidates ◽  
Effective Strategies ◽  
Wide Range ◽  
Functional Features ◽  
And Function

The enteric nervous system (ENS) plays a central role in gut physiology and mediating the crosstalk between the gastrointestinal (GI) tract and other organs. The human ENS has remained elusive, highlighting the need for an in vitro modeling and mapping blueprint. Here we map out the developmental and functional features of the human ENS, by establishing robust and scalable 2D ENS cultures and 3D enteric ganglioids from human pluripotent stem cells (hPSCs). These models recapitulate the remarkable neuronal and glial diversity found in primary tissue and enable comprehensive molecular analyses that uncover functional and developmental relationships within these lineages. As a salient example of the power of this system, we performed in-depth characterization of enteric nitrergic neurons (NO neurons) which are implicated in a wide range of GI motility disorders. We conducted an unbiased screen and identified drug candidates that modulate the activity of NO neurons and demonstrated their potential in promoting motility in mouse colonic tissue ex vivo. We established a high-throughput strategy to define the developmental programs involved in NO neuron specification and discovered that PDGFR inhibition boosts the induction of NO neurons in enteric ganglioids. Transplantation of these ganglioids in the colon of NO neuron-deficient mice results in extensive tissue engraftment, providing a xenograft model for the study of human ENS in vivo and the development of cell-based therapies for neurodegenerative GI disorders. These studies provide a framework for deciphering fundamental features of the human ENS and designing effective strategies to treat enteric neuropathies.  

scholarly journals In-Vitro, Ex-Vivo Characterization of Furosemide Bounded Pharmacosomes for Improvement of Solubility and Permeability

2014 ◽  
Vol 2 (5) ◽  
pp. 67-76
Author(s):  
Vivekanand K.Chatap ◽  
Prashant L. Patil ◽  
Savita D. Patil
Keyword(s):  
Ex Vivo

scholarly journals A novel BCMA/CD3 bispecific T-cell engager for the treatment of multiple myeloma induces selective lysis in vitro and in vivo

Leukemia ◽  
2016 ◽  
Vol 31 (8) ◽  
pp. 1743-1751 ◽  
Author(s):  
S Hipp ◽  
Y-T Tai ◽  
D Blanset ◽  
P Deegen ◽  
J Wahl ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Bone Marrow ◽  
T Cell ◽  
Plasma Cells ◽  
Ex Vivo ◽  
Xenograft Model ◽  
Selective Lysis ◽  
Bispecific T Cell Engager

Abstract B-cell maturation antigen (BCMA) is a highly plasma cell-selective protein that is expressed on malignant plasma cells of multiple myeloma (MM) patients and therefore is an ideal target for T-cell redirecting therapies. We developed a bispecific T-cell engager (BiTE) targeting BCMA and CD3ɛ (BI 836909) and studied its therapeutic impacts on MM. BI 836909 induced selective lysis of BCMA-positive MM cells, activation of T cells, release of cytokines and T-cell proliferation; whereas BCMA-negative cells were not affected. Activity of BI 836909 was not influenced by the presence of bone marrow stromal cells, soluble BCMA or a proliferation-inducing ligand (APRIL). In ex vivo assays, BI 836909 induced potent autologous MM cell lysis in both, newly diagnosed and relapsed/refractory patient samples. In mouse xenograft studies, BI 836909 induced tumor cell depletion in a subcutaneous NCI-H929 xenograft model and prolonged survival in an orthotopic L-363 xenograft model. In a cynomolgus monkey study, administration of BI 836909 led to depletion of BCMA-positive plasma cells in the bone marrow. Taken together, these results show that BI 836909 is a highly potent and efficacious approach to selectively deplete BCMA-positive MM cells and represents a novel immunotherapeutic for the treatment of MM.

scholarly journals Formulation and characterization of chlorhexidine HCl nanoemulsion as a promising antibacterial root canal irrigant: in-vitro and ex-vivo studies

2019 ◽  
Vol Volume 14 ◽  
pp. 4697-4708 ◽  
Author(s):  
Rehab Abdelmonem ◽  
Mona K. Younis ◽  
Doaa H Hassan ◽  
Mohamed Abd El-Gawad El-Sayed Ahmed ◽  
Ehab Hassanien ◽  
...  
Keyword(s):  
Root Canal ◽  
Ex Vivo ◽  
Root Canal Irrigant

Development of an in vitro model of calcified cerebral emboli in acute ischemic stroke for mechanical thrombectomy evaluation

2020 ◽  
Vol 12 (10) ◽  
pp. 1002-1007
Author(s):  
Sarah Johnson ◽  
Ray McCarthy ◽  
Brian Fahy ◽  
Oana Madalina Mereuta ◽  
Seán Fitzgerald ◽  
...  
Keyword(s):  
Ischemic Stroke ◽  
Acute Ischemic Stroke ◽  
In Vitro Model ◽  
Mechanical Thrombectomy ◽  
Ex Vivo ◽  
Guide Catheter ◽  
Vitro Model ◽  
Calcified Tissues

​BackgroundCalcified cerebral emboli (CCEs) are a rare cause of acute ischemic stroke (AIS) and are frequently associated with poor outcomes. The presence of dense calcified material enables reliable identification of CCEs using non-contrast CT. However, recanalization rates with the available mechanical thrombectomy (MT) devices remain low.ObjectiveTo recreate a large vessel occlusion involving a CCE using an in vitro silicone model of the intracranial vessels and to demonstrate the feasability of this model to test different endovascular strategies to recanalize an occlusion of the M1 segment of the middle cerebral artery (MCA).​MethodsAn in vitro model was developed to evaluate different endovascular treatment approaches using contemporary devices in the M1 segment of the MCA. The in vitro model consisted of a CCE analog placed in a silicone neurovascular model. Development of an appropriate CCE analog was based on characterization of human calcified tissues that represent likely sources of CCEs. Feasibility of the model was demonstrated in a small number of MT devices using four common procedural techniques.​ResultsCCE analogs were developed with similar mechanical behavior to that of ex vivo calcified material. The in vitro model was evaluated with various MT techniques and devices to show feasibility of the model. In this limited evaluation, the most successful retrieval approach was performed with a stent retriever combined with local aspiration through a distal access catheter, and importantly, with flow arrest and dual aspiration using a balloon guide catheter.​ConclusionCharacterization of calcified tissues, which are likely sources of CCEs, has shown that CCEs are considerably stiffer than thrombus. This highlights the need for a different in vitro AIS model for CCEs than those used for thromboemboli. Consequentially, an in vitro AIS model representative of a CCE occlusion in the M1 segment of the MCA has been developed.

scholarly journals Development and Characterization of Potential Ocular Mucoadhesive Nano Lipid Carriers Using Full Factorial Design

Pharmaceutics ◽  
2020 ◽  
Vol 12 (7) ◽  
pp. 682
Author(s):  
Eszter L. Kiss ◽  
Szilvia Berkó ◽  
Attila Gácsi ◽  
Anita Kovács ◽  
Gábor Katona ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Harmful Effect ◽  
Cell Junctions ◽  
Toxicity Tests ◽  
Eye Drops ◽  
Corneal Toxicity ◽  
Penetration Tests ◽  
Full Factorial

Generally, topically applied eye drops have low bioavailability due to short residence time and low penetration of the drug. The aim of the present study was to incorporate dexamethasone (DXM) into nano lipid carriers (NLC), which contain mucoadhesive polymer, in order to increase the bioavailability of the drug. A 23 factorial experimental design was applied, in which the three factors were the polymer, the DXM, and the emulsifier concentrations. The samples were analyzed for particle size, zeta potential, polydispersity index, and Span value. The significant factors were identified. The biocompatibility of the formulations was evaluated with human corneal toxicity tests and immunoassay analysis. The possible increase in bioavailability was analyzed by means of mucoadhesivity, in vitro drug diffusion, and different penetration tests, such as in vitro cornea PAMPA model, human corneal cell penetration, and ex vivo porcine corneal penetration using Raman mapping. The results indicated that DXM can be incorporated in stable mucoadhesive NLC systems, which are non-toxic and do not have any harmful effect on cell junctions. Mucoadhesive NLCs can create a depot on the surface of the cornea, which can predict improved bioavailability.

Construction and Characterization of a Novel Ribonuclease Immunotoxin Consisting of Two Ranpirnase (Rap) Molecules Fused to an Internalizing Anti-CD74 Humanized IgG4 Antibody.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 3289-3289
Author(s):  
Sailaja S. Vanama ◽  
Puja Sapra ◽  
Hans J. Hansen ◽  
Ivan D. Horak ◽  
David M. Goldenberg ◽  
...  
Keyword(s):  
Hodgkin’S Lymphoma ◽  
Xenograft Model ◽  
Hodgkin's Lymphoma ◽  
Light Chains ◽  
Non Hodgkin’S Lymphoma ◽  
Non Hodgkin's Lymphoma ◽  
Sds Page

Abstract Ranpirnase (Rap), isolated from frog (Rana pipiens) oocytes, is a monomeric ribonuclease (MW 11800) that kills cells by degrading t-RNA upon internalization. Previous studies indicated that the cytotoxicity of Rap could be enhanced more than 10,000-fold when the enzyme is chemically conjugated to an internalizing antibody. Here we describe the construction and characterization of 2L-Rap-hLL1-γ4P, composed of two Rap molecules fused to hLL1, an internalizing anti-CD74 humanized monoclonal antibody. To reduce unwanted cytotoxicity, the IgG1 constant region of hLL1 was replaced with an IgG4 that contains a proline mutation in the hinge region. The Rap gene was inserted at the N-terminus of the light chain in the expression vector of hLL1 and expressed in NS0 mouse myeloma cells. The fusion protein was characterized by a variety of techniques, including SE-HPLC, SDS-PAGE, in vitro transcription translation (IVTT) assay using luciferase reporter system, and competition ELISA to measure the binding affinity for CD74. The in vitro potency was determined in non-Hodgkin’s lymphoma (Daudi) and multiple myeloma (MC/CAR) cell lines by MTS tetrazolium dye reduction assay. In vivo pharmacokinetics and biodistribution of radiolabeled 2L-Rap-hLL1- γ4P was compared to radiolabeled hLL1 mAb in naïve mice and in vivo therapeutic efficacy of 2L-Rap-hLL1- γ4P was determined in a xenograft model of Burkitt’s non-Hodgkin’s lymphoma (Daudi). Purified 2L-Rap-hLL1- γ4P was shown to be a single peak by SE-HPLC and its MW determined by MALDI-TOF to be 177,150, which is in agreement with the MW of one IgG (150,000) plus two Rap molecules (24,000). Reducing-SDS-PAGE of 2L-Rap-hLL1- γ4P revealed the presence of 3 bands, one corresponding to the heavy chain and the other two appearing to be derived from the Rap-fused light chains (38,526 and 36,700 by MS). Occurrence of the 2 light chains was shown to be due to glycosylation of Rap at the N69 residue. The binding affinity of 2L-Rap-hLL1- γ4P for CD74 was indistinguishable from that of hLL1. Both 2L-Rap-hLL1- γ4P and hLL1 bound to CD74 with subnanomolar affinity. The EC50 of RNase activity, as measured by the IVTT assay, was 300 pM for 2L-Rap-hLL1- γ4P and 30 pM for recombinant Rap (expressed in E. coil). In in vitro cytotoxicity assays, 2L-Rap-hLL1- γ4P was significantly cytotoxic against Daudi (EC50 280 pM) and the myeloma cell line, MC/CAR (EC50 50 nM). In contrast, free Rap or naked hLL1 did not demonstrate significant cytotoxicity at the concentrations tested. In vivo, the pharmacokinetic profile of 2L-Rap-hLL1- γ4P was almost identical to that of naked hLL1. Both 2L-Rap-hLL1- γ4P and hLL1 showed biphasic clearance from the circulation; the α and β half-life (t1/2) of 2L-Rap-hLL1- γ4P were 5 h and 119 h, respectively, and those of hLL1 were 4 h and 125 h, respectively. In tissue biodistribution studies, no significant difference was observed between 2L-Rap-hLL1- γ4P and hLL1 with regards to normal tissue uptake. Early efficacy results in the Daudi Burkitt’s non-Hodgkin’s lymphoma xenograft model demonstrate that treatment with a single dose of 2L-Rap-hLL1- γ4P as low as 1 μg/mouse significantly improves survival in comparison to untreated control mice (P<0.0001).

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