Hyperactivity of cathepsin B and other lysosomal enzymes in fibroblasts exposed to azithromycin, a dicationic macrolide antibiotic with exceptional tissue accumulation

Gerbaux, Cécile; Bambeke, Françoise Van; Montenez, J P; Piret, Jocelyne; Morlighem, Grégory; Tulkens, Paul M.

doi:10.1016/0014-5793(96)00975-1

Cited by 19 publications

(10 citation statements)

References 23 publications

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“…First, AZM is a CAD and more than 50 marketed and experimental pharmaceuticals containing a CAD structure have been reported to elicit PLD (Sadrieh 2010). Second, AZM has been shown to promote the accumulation of phospholipids, cholesterol and lysosomal lamellar bodies in other cells (Gerbaux et al 1996; Munic et al 2011; Ribeiro et al 2009; Tyteca et al 2001; Van Bambeke et al 1998; Van Bambeke et al 1996). The hallmark feature, and indeed the gold standard, for identifying PLD is the demonstration of an intracellular accumulation of phospholipids and the concurrent development of lamellar bodies (Reasor and Kacew 2001; Sadrieh 2010).…”

Section: Discussionmentioning

confidence: 99%

One man's poison is another man's meat: Using azithromycin-induced phospholipidosis to promote ocular surface health

et al. 2014

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Drug-induced phospholipidosis (PLD) is a common adverse effect which has led to the termination of clinical trials for many candidate pharmaceuticals. However, this lipid-inducing effect may be beneficial in the treatment of meibomian gland dysfunction (MGD). MGD is the major cause of dry eye disease (DED), which affects 40 million people in the USA and has no cure. Azithromycin (AZM) is a PLD-inducing antibiotic that is used off-label to treat MGD, and is presumably effective because it suppresses the MGD-associated conjunctival inflammation (i.e. posterior blepharitis) and growth of lid bacteria. We hypothesize that AZM can act directly to promote the function of human meibomian gland epithelial cells by inducing PLD in these cells, characterized by the accumulation of lipids and lysosomes. We cultured immortalized human meibomian gland epithelial cells (HMGEC) were cultured with or without azithromycin for 5 days. Cells were evaluated for cholesterol (Filipin) and neutral lipid (LipidTox) staining, as well as the appearance of lysosomes (LysoTracker) and lamellar bodies (transmission electron microscopy, TEM). The lipid composition of cellular lysates was analyzed by high performance thin-layer chromatography. Our findings demonstrate that AZM stimulates the accumulation of free cholesterol, neutral lipids and lysosomes in HMGEC. This AZM-induced increase of neutral lipid content occurred predominantly within lysosomes. Many of these vesicles appeared to be lamellar bodies by TEM, which is the characteristic of PLD. Our findings also show that AZM promotes an accumulation of free and esterified cholesterol, as well as phospholipids in HMGEC. Our results support our hypothesis and confirm the beneficial effect of PLD induced by AZM on HMGEC. Our discovery reveals a new potential use of PLD-inducing drugs, and makes this adverse effect a beneficial effect.

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Section: Discussionmentioning

confidence: 99%

One man's poison is another man's meat: Using azithromycin-induced phospholipidosis to promote ocular surface health

et al. 2014

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“…3). Previous reports have shown that both nonselective, nonbasic cysteine cathepsin inhibitors (Kominami et al, 1987;Montenez et al, 1994) and lysosomotropic compounds, such as chloroquine (Gerard et al, 1988;Gerbaux et al, 1996), induce an increase of cathepsin and other lysosomal protein levels in both isolated cells and in vivo. The formation of a protein-ligand complex often results in protein stabilization toward thermal and chaotropic agent denaturation (Kleanthous et al, 1991), as well as protease susceptibility .…”

Section: Discussionmentioning

confidence: 99%

Effect of Cathepsin K Inhibitor Basicity on in Vivo Off-Target Activities

Desmarais

Black²,

Oballa³

et al. 2007

Mol Pharmacol

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Cathepsin K is a lysosomal cysteine protease that is a pharmacological target for the treatment of osteoporosis. Previous studies showed that basic, lipophilic cathepsin K inhibitors are lysosomotropic and have greater activities in cell-based assays against cathepsin K, as well as the physiologically important lysosomal cysteine cathepsins B, L, and S, than expected based on their potencies against these isolated enzymes. Long-term administration of the basic cathepsin K inhibitors N- (1-(((cyanomethyl) -006235) and balicatib to rats at a supratherapeutic dose of 500 mg/kg/day for 4 weeks resulted in increased tissue protein levels of cathepsin B and L but had no effect on cathepsin B and L message. This is attributed to the inhibitor engagement of these off-target enzymes and their stabilization to proteolytic degradation. No such increase in these tissue cathepsins was detected at the same dose of, a potent nonbasic cathepsin K inhibitor with a similar off-target profile, although all three inhibitors provided similar plasma exposures. Using an activity-based probe, 125 I-BIL-DMK, in vivo inhibition of cathepsins B, L, and S was detected in tissues of mice given a single oral dose of L-006235 and balicatib, but not in mice given L-873724. In each case, similar tissue levels were achieved by all three compounds, thereby demonstrating the in vivo cathepsin selectivity of L-873724. In conclusion, basic cathepsin K inhibitors demonstrate increased off-target cysteine cathepsin activities than their nonbasic analogs and potentially have a greater risk of adverse effects associated with inhibition of these cathepsins.The CA1 family of human papain-like cysteine proteases comprises 11 members. These enzymes are collectively known as cathepsins, a name which is derived from the Greek kathepsein, to digest. Being largely lysosomal enzymes, cathepsins have acidic pH activity and stability optima. These enzymes are synthesized as preproenzymes, the mature proteins sharing between 25 and 80% sequence identity (Lecaille et al., 2002;Turk et al., 2003). Cathepsin K (Cat K) is highly and somewhat specifically expressed in osteoclasts, the multinucleated giant cells of hematopoietic origin that are responsible for the normal physiological process of bone resorption. Cat K destroys the organic fraction of bone through its potent collagenase activity, this process taking place in the acidic pit between the osteoclast and the bone surface and also intracellularly within lysosomes of the osteoclast (Saftig et al., 1998). A large volume of genetic and pharmacological data points to a pivotal role for Cat K in bone resorption, and Cat K inhibitors are presently being evaluated in clinical trials as a treatment of osteoporosis, a disease characterized by an imbalance of bone resorption over bone formation (performed by osteoblasts) (Deaton and Tavares, 2005;Grabowskal et al., 2005;Yasuda et al., 2005;Boyce et al., 2006;Close et al., 2006). Both physiological and pathological roles have been identified for the remaining 10 ...

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“…The affinity for PL A2 and its competitive inhibition have been variable among different amphiphilic substances 3 . These amphiphilic drugs have modulated the activities of other lysosomal enzymes including cathepsin B protease 50 . Enhanced activities of the lysosomal hexosaminidase, acid sphingomyelinase and sulfatase B in cells of the kidney cortex are considered to be due to a compensatory increase in degrading enzymes due to PL inhibition 51 .…”

Section: Possible Mechanismsmentioning

confidence: 99%

Drug-induced Phospholipidosis -Pathological Aspects and Its Prediction

Nonoyama

Fukuda

2008

J Toxicol Pathol

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Drug-induced phospholipidosis (PLDsis) is an excessive accumulation of polar phospholipids in cells or tissues/organs caused by xenobiotics. Numerous drugs and chemicals are capable of inducing the storage disorder in animals and humans; however, despite their diverse pharmacological activities, each of these drugs shares common physicochemical properties: a hydrophobic aromatic ring structure on the molecule and a hydrophilic side chain with a charged cationic amine group and therefore are in the group of cationic amphiphilic drugs. In affected cells the appearance of membrane-bound inclusions, primarily lysosomal in origin, with a lamellar structure (lamellar bodies) is a definitive morphologic hallmark. Massive accumulations can occur in animal tissues such as the lung with little effect on organ function. The inducing drug also accumulates in association with the excess phospholipid. Although these alterations are generally reversible after cessation of drug treatment, PLDsis is of regulatory concern and an issue for drug safety for pharmaceutical companies. Thus, the assessment of potential target organ dysfunction and the identification of clinical biomarkers are important objectives for new drug development. Recent advances in biotechnology such as metabonomics and toxicogenomics have been providing novel tools to elucidate the mechanisms of PLDsis and to establish biomarkers for screening tests in the preclinical stages and for monitoring in the clinical phases in addition to conventional approaches such as morphology (lamellar bodies) and biochemical methods including assays of specific metabolites and phospholipase inhibition. (J Toxicol Pathol 2008; 21: 9-24)

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Hyperactivity of cathepsin B and other lysosomal enzymes in fibroblasts exposed to azithromycin, a dicationic macrolide antibiotic with exceptional tissue accumulation

Cited by 19 publications

References 23 publications

One man's poison is another man's meat: Using azithromycin-induced phospholipidosis to promote ocular surface health

One man's poison is another man's meat: Using azithromycin-induced phospholipidosis to promote ocular surface health

Effect of Cathepsin K Inhibitor Basicity on in Vivo Off-Target Activities

Drug-induced Phospholipidosis -Pathological Aspects and Its Prediction

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