Identification of madangamine A as a novel lysosomotropic agent to inhibit autophagy

To evaluate the internalization of cholecystokinin, monoiodinated imidoester of cholecystokinin octapeptide [125I-(IE)-CCK-8] was bound to dispersed pancreatic acinar cells, and surface-bound and internalized radioligand were differentiated by treating with an acidified glycine buffer. The amount of internalized radioligand was four- and sevenfold greater at 24 and 37 degrees C than at 4 degrees C between 5 and 60 min of association. Specific binding of radioligand to cell surface receptors was not significantly different at these temperatures. Chloroquine, a lysosomotropic agent that blocks intracellular proteolysis, significantly increased the amount of CCK-8 internalized by 18 and 16% at 30 and 60 min of binding, respectively, compared with control. Dithiothreitol (DTT), a sulfhydryl reducing agent, also augmented the amount of CCK-8 radioligand internalized by 25 and 29% at 30 and 60 min, respectively. The effect of chloroquine and DTT on the processing of internalized radioligand was also considered after an initial 60 min of binding of radioligand to acinar cells. After 180 min of processing, the amount of radioligand internalized was significantly greater in the presence of chloroquine compared with controls, whereas the amount of radioligand declined in acinar cells treated with DTT. Internalized and released radioactivity from acinar cells was rebound to pancreatic membrane homogenates to determine the amount of intact radioligand during intracellular processing. Chloroquine significantly increased the amount of intact 125I-(IE)-CCK-8 radioligand in released and internalized radioactivity while DTT increased the amount of intact radioligand only in internalized samples. This study shows that pancreatic acinar cells rapidly internalize large amounts of CCK-8 and that chloroquine and DTT inhibit intracellular degradation.

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Methylamine does not inhibit rates of endogenous lipolysis in isolated myocardial cells from rat heart

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y87-040 ◽

1987 ◽

Vol 65 (2) ◽

pp. 226-229 ◽

Cited By ~ 1

Author(s):

Albert Kryski Jr. ◽

Terje S. Larsen ◽

Ignasi Ramírez ◽

David L. Severson

Keyword(s):

Fatty Acids ◽

Lipase Activity ◽

Rat Heart ◽

Diabetic Rat ◽

Myocardial Cells ◽

Acid Lipase ◽

Ph Optimum ◽

Triacylglycerol Lipase ◽

Rat Hearts ◽

Lysosomotropic Agent

Triacylglycerol lipase activity with a pH optimum of 5 was present in homogenates of myocardial cells from rat heart. Acid lipase activity was inhibited by serum, heparin, and increased ionic strength. Methylamine, a lysosomotropic agent, did not inhibit the basal or isoproterenol-stimulated rate of endogenous lipolysis as measured by glycerol output from control myocytes. Similarly, accelerated rates of glycerol output that are a consequence of an elevation in the intracellular stores of triacylglycerols in myocytes from diabetic rat hearts and from myocytes prepared with free fatty acids in the isolation solutions were not reduced by methylamine. Therefore, the acid lysosomal triacylglycerol lipase must not be involved in the mobilization of endogenous triacylglycerols in myocardial cells from rat heart.

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Microencapsulated Multi-functionalized Graphene Oxide Equipped With Chloroquine for Efficient and Sustained Sirna Delivery

10.21203/rs.3.rs-1010499/v1 ◽

2021 ◽

Author(s):

Rana Imani ◽

Satya Prakash ◽

Hojatollah Vali ◽

John F. Presley ◽

Shahab Faghihi

Keyword(s):

Graphene Oxide ◽

Cellular Uptake ◽

Sirna Delivery ◽

Layer By Layer ◽

Functionalized Graphene ◽

Functionalized Graphene Oxide ◽

Lysosomal Ph ◽

Intracellular Targeting ◽

Lysosomotropic Agent ◽

Layer Coating

Abstract A multi-functionalized graphene oxide (GO)-based carrier with conjugation of aminated-polyethylenglycole (PEG-diamine), octaarginine (R8) and folic acid (FA), which also contains chloroquine (CQ), a lysosomotropic agent, is introduced. The cellular uptake mechanisms and intracellular targeting of FA functionalized nanocarriers are examined. The localized releases of CQ and siRNA intracellular delivery are evaluated. Microencapsulation of the nanocarrier complexed with genes in layer-by-layer coating of alginate micro-beads is also investigated. The covalently co-conjugated FA with PEG and R8 provides a stable formulation with increased cellular uptake compared to FA-free carrier. The CQ-equipped nanocarrier shows a 95% release of CQ at lysosomal pH. The localized release of the drug inside the lysosomes is verified which accelerates the cargo discharge into cytoplasm.

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“Amantadine disrupts lysosomal gene expression; potential therapy for COVID19”

10.1101/2020.04.05.026187 ◽

2020 ◽

Cited By ~ 3

Author(s):

Sandra P. Smieszek ◽

Bart P Przychodzen ◽

Mihael H Polymeropoulos

Keyword(s):

Gene Expression ◽

Gene Expression Analysis ◽

Cathepsin L ◽

Drug Screen ◽

Direct Inhibition ◽

The Us ◽

Lysosomotropic Agent ◽

A Cell ◽

Expression Potential ◽

Lysosomal Gene

AbstractSARS-coronavirus 2 is the causal agent of the COVID-19 outbreak. SARS-Cov-2 entry into a cell is dependent upon binding of the viral spike (S) protein to cellular receptor and on cleavage of the spike protein by the host cell proteases such as Cathepsin L and Cathepsin B. CTSL/B are crucial elements of lysosomal pathway and both enzymes are almost exclusively located in the lysosomes.CTSL disruption offers potential for CoVID-19 therapies. The mechanisms of disruption include: decreasing expression of CTSL, direct inhibition of CTSL activity and affecting the conditions of CTSL environment (increase pH in lysosomes).We have conducted a high throughput drug screen gene expression analysis to identify compounds that would downregulate the expression of CTSL/CTSB. One of the top significant results shown to downregulate the expression of the CTSL gene is Amantadine. Amantadine was approved by the US Food and Drug Administration in 1968 as a prophylactic agent for influenza and later for Parkinson’s disease. It is available as a generic drug..Amantadine in addition to downregulating CTSL appears to further disrupt lysosomal pathway, hence interfering with the capacity of the virus to replicate. It acts as a lysosomotropic agent altering the CTSL functional environment. We hypothesize that Amantadine could decrease the viral load in SARS-CoV-2 positive patients and as such it may serve as a potent therapeutic decreasing the replication and infectivity of the virus likely leading to better clinical outcomes. Clinical studies will be needed to examine the therapeutic utility of amantadine in COVID-19 infection.

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Eradicate Coronavirus by blocking replication, counteracting its defense system

10.31219/osf.io/psz35 ◽

2020 ◽

Author(s):

Kira Smith

Keyword(s):

Target Genes ◽

Defense Mechanism ◽

P53 Protein ◽

Sirtuin 1 ◽

Defense System ◽

Virus Growth ◽

Self Defense ◽

System P ◽

Lysosomotropic Agent ◽

Regulation Of Growth

SIRT1 inhibitors can reduce replication of many viruses with certain similar characteristics to those of Coronaviruses, while p53 protein is another important factor in down-regulation of growth. There are some molecules that inhibit Sirtuin 1 and 2, in addition to activate p53 protein, by means of regulation of the interactions used by Coronaviruses as self-defense mechanism, degradating it. Even mTOR signal will be regulate, as well as HIF-1α with the target genes and cytokines. Autophagy will be inhibited, being this compound like a lysosomotropic agent. By blocking virus growth and continuous replication, associating the already tested Antiviral medicines, Covid-19 could be eradicated.

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Regulation of Ca2+-dependent protein turnover in skeletal muscle by thyroxine

Biochemical Journal ◽

10.1042/bj2400269 ◽

1986 ◽

Vol 240 (1) ◽

pp. 269-272 ◽

Cited By ~ 16

Author(s):

R J Zeman ◽

P L Bernstein ◽

R Ludemann ◽

J D Etlinger

Keyword(s):

Skeletal Muscle ◽

Protein Turnover ◽

Proteinase Inhibitor ◽

Thyroid Status ◽

Protein Balance ◽

Lysosomotropic Agent ◽

Lysosomal Proteolysis ◽

Dependent Protein ◽

Protein Synthesis And Degradation ◽

Synthesis And Degradation

Dantrolene, an agent that inhibits Ca2+ mobilization, improved protein balance in skeletal muscle, as thyroid status was increased, by altering rates of protein synthesis and degradation. Thyroxine (T4) caused increases in protein degradation that were blocked by leupeptin, a proteinase inhibitor previously shown to inhibit Ca2+-dependent non-lysosomal proteolysis in these muscles. In addition, T4 abolished sensitivity to the lysosomotropic agent methylamine and the autophagy inhibitor 3-methyladenine, suggesting that T4 inhibits autophagic/lysosomal proteolysis.

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Bile protein secretion in the rat stimulated by taurocholate effect of chloroquine

Canadian Journal of Physiology and Pharmacology ◽

10.1139/y88-119 ◽

1988 ◽

Vol 66 (6) ◽

pp. 749-753 ◽

Cited By ~ 7

Author(s):

R. A. Marinelli ◽

C. E. Carnovale ◽

E. A. Rodríguez Garay

Keyword(s):

Acid Phosphatase ◽

Molecular Mass ◽

Protein Secretion ◽

Biliary Excretion ◽

Sodium Taurocholate ◽

Protein Excretion ◽

Chloroquine Treatment ◽

Lysosomotropic Agent ◽

The Individual ◽

Bile Protein

The biliary protein excretion during sodium taurocholate induced choleresis was studied in normal rats and in rats treated with the lysosomotropic agent, chloroquine. The analysis of the protein component in bile was made on SDS–polyacrilamide gel, and the individual polypeptides were quantitated by densitometry. The excretion of bile polypeptides was compared with that of lysosomal acid phosphatase. The biliary excretion of polypeptides of molecular mass lower than and equal to 54 kDa was markedly stimulated by taurocholate-induced choleresis. Chloroquine treatment of rats diminished the biliary excretion of such polypeptides and also inhibited their excretion induced by taurocholate. The behaviour of these polypeptides was well correlated to that of the lysosomal marker. The biliary excretion of polypeptide bands of a higher molecular mass (up to 140 kDa) did not show major changes during taurocholate-induced choleresis in any of the groups. The results indicate that biliary excretion of proteins in the rat may be either stimulated by taurocholate or may be independent of the bile salt. The former requires the functional integrity of chloroquine-sensitive hepatocyte compartments, which may involve the lysosomes.

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