scholarly journals Intratumoral Distribution and pH-Dependent Drug Release of High Molecular Weight HPMA Copolymer Drug Conjugates Strongly Depend on Specific Tumor Substructure and Microenvironment

2020 ◽  
Vol 21 (17) ◽  
pp. 6029
Author(s):  
Anne-Kathrin Noack ◽  
Henrike Lucas ◽  
Petr Chytil ◽  
Tomáš Etrych ◽  
Karsten Mäder ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Drug Release ◽  
High Molecular Weight ◽  
Cell Lines ◽  
Tumor Tissue ◽  
Tumor Type ◽  
Acidic Microenvironment ◽  
Drug Conjugates ◽  
Ph Dependent ◽  
Intratumoral Distribution

Stimulus-sensitive polymer drug conjugates based on high molecular weight N-(2-hydroxypropyl) methacrylamide (HPMA) copolymers carrying doxorubicin via a pH-dependent cleavable bond (pHPMA-Dox) were previously shown to be able to overcome multi-drug resistance. Nevertheless, a tumor type dependent differential response was observed. Although an improved and more selective tumor accumulation of pHPMA-Dox is generally achieved due to the enhanced permeability and retention (EPR) effect, little is known about the fate of these conjugates upon entering the tumor tissue, which could explain the different responses. In this study, we compared in vitro and in vivo accumulation and Dox-activation of pHPMA-Dox in three cancer cell line models (1411HP, A2780cis, HT29) and derived xenograft tumors using a near-infrared fluorescence-labeled pHPMA-Dox conjugate. Firstly, cytotoxicity assays using different pH conditions proved a stepwise, pH-dependent increase in cytotoxic activity and revealed comparable sensitivity among the cell lines. Using multispectral fluorescence microscopy, we were able to track the distribution of drug and polymeric carrier simultaneously on cellular and histological levels. Microscopic analyses of cell monolayers confirmed the assumed mechanism of cell internalization of the whole conjugate followed by intracellular cleavage and nuclear accumulation of Dox in all three cell lines. In contrast, intratumoral distribution and drug release in xenograft tumors were completely different and were associated with different tissue substructures and microenvironments analyzed by Azan- and Hypoxisense®-staining. In 1411HP tumors, large vessels and less hypoxic/acidic microenvironments were associated with a pattern resulting from consistent tissue distribution and cellular uptake as whole conjugate followed by intracellular drug release. In A2780cis tumors, an inconsistent pattern of distribution partly resulting from premature drug release was associated with a more hypoxic/acidic microenvironment, compacted tumor tissue with compressed vessels and specific pre-damaged tissue structures. A completely different distribution pattern was observed in HT29 tumors, resulting from high accumulation of polymer in abundant fibrotic structures, with small embedded vessels featuring this tumor type together with pronounced premature drug release due to the strongly hypoxic/acidic microenvironment. In conclusion, the pattern of intratumoral distribution and drug release strongly depends on the tumor substructure and microenvironment and may result in different degrees of therapeutic efficacy. This reflects the pronounced heterogeneity observed in the clinical application of nanomedicines and can be exploited for the future design of such conjugates.

scholarly journals MITOCHONDRIAL RNA FROM CULTURED ANIMAL CELLS

1970 ◽  
Vol 46 (2) ◽  
pp. 245-251 ◽  
Author(s):  
Bland S. Montenecourt ◽  
Margaret E. Langsam ◽  
Donald T. Dubin
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Cell Lines ◽  
Ribosomal Rna ◽  
Gel Electrophoresis ◽  
Base Composition ◽  
Size Class ◽  
Mitochondrial Rna ◽  
Animal Cells ◽  
Molecular Weights

Discrete RNA fractions sedimenting slightly slower than 18s ribosomal RNA have been found in mitochondrial preparations from both hamster (BHK-21) and mouse (L-929) cells. This RNA could be separated into two components, present in approximately equimolar amounts, by prolonged zonal centrifugation or acrylamide gel electrophoresis. The hamster components had sedimentation constants averaging 16.8 and 13.4, and molecular weights (estimated by gel electrophoresis) averaging 0.74 and 0.42 x 106 daltons. Mixed labeling experiments showed that the mouse components sedimented and electrophoresed 3–6% more slowly than the corresponding hamster components. The RNA from both cell lines resembled mitochondrial ribosomal RNA from yeast and Neurospora in being GC poor, and in addition the larger and smaller components resembled each other in base composition. These results, taken with those of other recent studies, are compatible with the idea that our high molecular weight mitochondrial RNA is ribosomal; such RNA would then constitute a uniquely small size-class of ribosomal RNA.

scholarly journals High-molecular-weight glycoproteins are the major carriers of the carbohydrate differentiation antigens I, i and SSEA-1 of mouse teratocarcinoma cells

1983 ◽  
Vol 215 (3) ◽  
pp. 491-503 ◽  
Author(s):  
R A Childs ◽  
J Pennington ◽  
K Uemura ◽  
P Scudder ◽  
P N Goodfellow ◽  
...  
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Cell Lines ◽  
Galactose Oxidase ◽  
Embryonal Carcinoma Cell ◽  
Teratocarcinoma Cell ◽  
Differentiation Antigens ◽  
Carcinoma Cell Lines ◽  
Teratocarcinoma Cells ◽  
Mouse Teratocarcinoma

The carriers of the carbohydrate differentiation antigens I, i and SSEA-1 were investigated in embryonal carcinoma cell lines of mouse and differentiated cell lines derived from them. Glycoproteins were studied by immunostaining (‘Western blotting’) of total cell lysates and immunoprecipitation from lysates of galactose oxidase/NaB3H4-labelled cells; glycolipids were investigated by immunostaining of thin layer chromatograms. The antigenic activities detected by immunofluorescence of cell smears were reflected in the antigenicities of high-molecular-weight glycoproteins. These were polydisperse and markedly susceptible to digestion with endo-beta-galactosidase. Only the I antigen was detected on minor glycolipids. These observations indicate that glycoproteins rather than glycolipids are the major carriers of carbohydrate differentiation antigens I, i and SSEA-1 in the teratocarcinoma cell lines.

scholarly journals Carbopol 71G-NF polymer –The next pillar of oral solid dosage form

2020 ◽  
Vol 1 (1) ◽  
pp. 010-017
Author(s):  
R. Priyanka ◽  
R. Senthil Prabhu
Keyword(s):  
Molecular Weight ◽  
Drug Release ◽  
Acrylic Acid ◽  
High Molecular Weight ◽  
Dosage Form ◽  
Carboxyl Groups ◽  
Dry Powders ◽  
Solid Dosage Form ◽  
Solid Dosage ◽  
Pharmaceutical Application

For pharmaceutical Oral Solid Dosage Form (OSDF) there are lots of excipients used and these excipients influence the drug release. In the recent decades there has been considerable interest in using carbopol polymers as excipients in a distinctive range of pharmaceutical application. Carbopol polymers are high molecular weight, cross linked, acrylic, acid-based polymers. Carbopol homopolymers are polymers of acrylic acid cross linked with ally sucrose or allylPentaerythritol. These polymers are offered as fluffy, white, dry powders. The carboxyl groups provided by the acrylic acid backbone of the polymer are responsible for many of the product benefits. This review work aims at guesstimate the characteristic of Carbopol 71G-NF polymer to be used as excipients in oral solid dosage form (OSDF).

FindingNemo in OneDay: Ultra-Long ONT Library Preparation from Cell to Flowcell in One Day v1

2021 ◽  
Author(s):  
Inswasti Cahyani ◽  
John Tyson ◽  
Nadine Holmes ◽  
Josh Quick ◽  
Nicholas Loman ◽  
...  
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Cell Lines ◽  
Human Cell ◽  
Human Cell Lines ◽  
Library Preparation ◽  
Dna Library ◽  
Oxford Nanopore ◽  
Working Day ◽  
Ultra High Molecular Weight

This protocol is focussed on the isolation of ultra-high molecular weight (UHMW) DNA, library preparation and clean up ready for sequencing on the Oxford Nanopore Technology (ONT) platforms, all within one working day. This protocol is optimised for human cell lines. Summary of the workflow:

scholarly journals Receptors for IgG: subclass specificity of receptors on different mouse cell types and the definition of two distinct receptors on a macrophage cell line.

1977 ◽  
Vol 145 (5) ◽  
pp. 1316-1327 ◽  
Author(s):  
C H Heusser ◽  
C L Anderson ◽  
H M Grey
Keyword(s):  
Molecular Weight ◽  
Cell Line ◽  
High Molecular Weight ◽  
Cell Lines ◽  
Gel Filtration ◽  
Aggregate Size ◽  
Cell Types ◽  
Mouse Cell ◽  
Receptor Sites ◽  
B Lymphoid

To evaluate subclass specificity and aggregate size requirements of IgG receptors on mouse cells, we measured binding of radiolabeled monomeric and BDB-aggregated mouse myeloma proteins fractionated into various sizes by means of gel filtration. Monomers, tetramers, and high molecular weight (approximately 10(7) daltons) aggregates were used. The various cells and cell lines studied could be segregated into three patterns of reactivity: (a) Macrophage and macrophage-like cell lines bound monomer IgG2a preferentially; high molecular weight IgG aggregates bound as follows: IgG1 = IgG2b = IgG2a. (b) Lymphoid lines D2N and S49 showed no capacity to bind monomer IgG2a; high molecular weight aggregates bound as follows: IgG1 = IgG2b less than IgG2a. (c) Other Thy-1-positive lymphoid cell lines (EL4 and L5178) and normal T and B cells showed no capacity to bind monomer IgG; high molecular weight IgG aggregates bound to a lesser extent than to cells of the first two categories in the following manner: IgG1 less than IgG2b greater than or equal to IgG2a. The variable pattern of reactivity of the macrophage-like cell lines with monomer and aggregated IgG suggested that two distinct receptors for IgG were present: one capable of binding IgG2a and another capable of binding all aggregates. Further evidence for this hypothesis was obtained by analysis of the inhibitory capacity of different IgG subclasses on the binding of aggregated IgG and monomer IgG2a to P388 cells. Inhibition of monomer IgG2a binding was effected only by monomer or aggregated IgG2a, whereas inhibition of binding of aggregated IgG1 or IgG2b was noted with aggregates of all three subclasses with some preferential inhibition by monomer IgG2b being observed. Furthermore, monomer IgG2b binding was preferentially inhibitable by monomer IgG2b. It is postulated from these data that two receptor sites are present on this macrophage-like cell line, one reactive with aggregates of all three subclasses as well as monomer IgG2b, and another receptor specific for monomer IgG2a which also binds aggregated IgG2a. Support of this concept was obtained by trypsinization experiments in which the binding of monomer IgG2a was markedly decreased by trypsin treatment of cells, whereas the binding of aggregated IgG2b was unaffected by this treatment.

Evidence of Mucin Secretion in Human Lung Adenocarcinoma Cell Lines NCIH650 and NCIH2077 and Effect of Select Secretagogues on Mucin Secretion

1999 ◽  
Vol 19 (5) ◽  
pp. 473-483 ◽  
Author(s):  
Prerna M. Sharma ◽  
Mangala G. Sarkar ◽  
Arvind K. Virmani ◽  
Adi F. Gazdar ◽  
Goverdhan P. Sachdev
Keyword(s):  
Molecular Weight ◽  
Lung Adenocarcinoma ◽  
High Molecular Weight ◽  
Cell Lines ◽  
In Vitro Model ◽  
Culture Conditions ◽  
Mucin Secretion ◽  
Adenocarcinoma Cell ◽  
Vitro Model

Mucins comprise an important class of tumor-associated antigens. The objectives of the present study were (a) to establish an in vitro model system using human non-small cell lung adenocarcinoma cell lines NCIH650 and NCIH2077 (b) provide evidence that these cell lines secrete mucin in culture conditions and (c) investigate the effects of select secretagogues on mucin secretion. The cell lines were established in ACL-4 medium containing several growth factors and retinoic acid and 5% fetal calf serum. The high molecular weight glycoconjugates secreted in the culture medium were purified by ammonium sulfate precipitation and Superose 6 and Superose 12 FPLC chromatography. The purified high molecular weight glycoconjugate fraction and the carcinoma cells were shown to have mucin by dot blot, Western blot and immunohistochemical analysis, respectively, using specific antibodies to purified major mucin, HTM-1. Also, incorporation experiments with mucin precursor 3H-glucosamine demonstrated that the cells indeed synthesize high molecular weight mucins. The effects of secretagogues such as, 8-bromocyclic AMP, ionomycin, phorbol-12-myristate-13-acetate and neutrophil elastase on mucin secretion were also investigated. Only 8-bromocyclic AMP and neutrophil elastase influenced mucin secretion. These studies provided strong evidence that the lung adenocarcinoma cell lines secrete high molecular weight mucins in culture conditions and only two of the four tested secretagogues significantly increased mucin secretion. Thus, this in vitro model system may be useful in determining alterations in mucin structure, if any, in lung adenocarcinomas as well as in studying the regulation of mucin gene expression.

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