FR3063640A1 - TREATMENT OF PULMONARY FIBROSIS USING A SELECTIVE INHIBITOR OF THE PRODUCTION OF REACTIVE SPECIES OF OXYGEN OF MITOCHONDRIAL ORIGIN - Google Patents

Abstract

The present invention relates to the prevention and / or treatment of diseases in which the reactive oxygen species (or ROS) of mitochondrial origin are involved. It relates more particularly to the use of a specific inhibitor of mitochondrial ROS production, in particular anethole trithione, for the treatment and prevention of pulmonary fibrosis.

Description

Holder (s): PETITJEAN OLIVIER, PETITJEAN GUILLAUME, PETITJEAN ELODIE, PETITJEAN GREGOIRE, CHILDS MARC, SAUZIERES JACQUES.

Extension request (s)

Agent (s): IP TRUST.

TREATMENT OF PULMONARY FIBROSIS USING A SELECTIVE INHIBITOR FOR THE PRODUCTION OF REACTIVE SPECIES OF OXYGEN OF MITOCHONDRIAL ORIGIN.

(5g) The present invention relates to the prevention and / or treatment of diseases in which reactive oxygen species (or ROS) are involved of mitochondrial origin. It relates more particularly to the use of a specific inhibitor of the production of mitochondrial ROS, in particular anethole trithione, for the treatment and prevention of pulmonary fibrosis.

FR 3 063 640 - A1

TREATMENT OF PULMONARY FIBROSIS USING A SELECTIVE INHIBITOR FOR THE PRODUCTION OF REACTIVE SPECIES OF MITOCHONDRIAL OXYGEN

The present invention relates to the prevention and / or treatment of diseases in which reactive oxygen species (or ROS) are involved of mitochondrial origin. It relates more particularly to the use of a specific inhibitor of the production of mitochondrial ROS, in particular anethole trithione, for the treatment of pulmonary fibrosis.

PRIOR ART

The mitochondria participate in the pathogenesis of almost all diseases associated with aging, including cardiovascular diseases, neurodegenerative diseases (Parkinson's disease, Alzheimer's disease, etc.), diabetes, as well as tissue dysfunctions of ischemic origin. . It is widely accepted that it plays a central role in the "theory of free radicals in aging". This theory asserts that the accumulation of damage caused by reactive oxygen species (ROS) affects many cellular functions, in particular mitochondrial functions, which are essential for energy supply and proper functioning. cellular. The mitochondria therefore appear to be the primary targets of ROS since optimal cell functioning is crucial to supply the energy that a cell needs to repair itself.

While mitochondria are the main source of ROS, they are also particularly sensitive to the damage caused by these ROS. Consequently, the mitochondria themselves generate ROS, which causes oxidative damage which they undergo and which contribute to their dysfunction and, subsequently, to cell death.

Numerous studies have been carried out to assess the ability of antioxidants to counteract the effect of ROS. Several antioxidant molecules have given satisfaction in preclinical studies, but their effectiveness has only been partially confirmed in most clinical trials (Orr et al., 2013, Free Radie Biol. Med, 65: 1047-59).

In addition, recent studies have shown that an excessive reduction in ROS has a deleterious effect on cells, suggesting that a balanced production of ROS contributes to good cellular functioning (Goodman et al., J. Notl. Cancer Inst. , 2004, 96: 1743-50; Bjelakovic G et al., JAMA. 2007, 297: 842-57).

TECHNICAL PROBLEM AND SOLUTION PROVIDED BY THE INVENTION

Studies relating to the role of oxidative stress in many pathologies have highlighted the advantage of having a selective inhibitor of ROS production of mitochondrial origin. The antioxidants available to date do not have such a specificity with, as a consequence, the risk of occurrence of side effects when the production of cytosolic ROS is affected, especially in case of prolonged treatment; these side effects are well described.

Several elements argue in favor of a deleterious effect of ROS on lung tissue. Indeed, it is known from the literature that any excess of ROS which will not be used for cell signaling, is toxic and potentially promoter of fibrosis. These observations are correlated by different publications (Faner R. et al. Am J Respir Crit Care Med, 2012, 186: 306-313; Kliment CR. Et al. Free Rad Biol Med, 2010, 49: 707-717; Liu RM ., Gaston Pravia KA. Free Rad Biol Med, 2010, 48: 1-15; Liu RM et al. Free Rad Biol Med, 2012.53: 554-563; Liu G. et al. Annu Rev Pathol: Mech Dis, 2013, 8: 161-187) who report that:

• oxidation products of lipids and proteins are found in excess in bronchoalveolar lavage (BAL) as well as in samples of fibrotic lung tissue;

• similarly, high levels of protein, DNA and lipid oxidation products are detected in models of bleomycin fibrosis;

• oxidized DNA is found in patients suffering from idiopathic pulmonary fibrosis as well as in subjects with asbestosis or silicosis and in animal models of asbestosis or silicosis;

• Finally, antioxidants and iron chelators reduce the severity of models of bleomycin fibrosis or of models of asbestosis.

In this context, the mitochondrial production of ROS appears to be pivotal in the constitution of pulmonary fibrosis. For example, Carter et al. have shown the key role of mitochondrial production of H ₂ O ₂ by alveolar macrophages in the constitution of asbestosis (Osborn-Herford HL et al., J Biol Chem, 2012, 287: 3301-3312; Murthy S. et al., J Biol Chem, 2010, 285: 25062-25073; Murthy S. et al., Am J Physiol Lung Cell Mol Physiol, 2009, 297: L846-L855; He C. et al., J Biol Chem, 2011, 286: 15595-15607).

It thus appears that any biological or environmental context which leads to an overproduction of ROS, whether it is mitochondrial or extramitochondrial, promotes the formation of fibrosis. There is also a close interrelation between the two ROS production pathways, one causing the stimulation of the other. The role of ROS in pulmonary fibrosis is consistent with the following observations:

1) The NOX pathway - NOX2 and NOX4 - has notably been implicated in the constitution of pulmonary fibrosis (Crestani B. et ai, Int J Biochem Cell Biol, 2011, 43: 1086-1089; Hecker L. et al., Cell Molec Life Sci, 2012, 69: 2365-2371). This NOX route is known to promote an overproduction of ROS. In this context, the inhibition of NOX4 also prevents the experimental induction of fibrosis by bleomycin and, at the same time, there is an overexpression of NOX4 in fibrosis with bleomycin as indeed in idiopathic fibrosis in l 'man. Likewise, the mouse made deficient in NOX2 does not develop bleomycin fibrosis.

2) The central role of mitochondrial ROS has also been highlighted through the phenomena of apoptosis and hypoxia. Indeed, apoptosis of alveolar epithelial cells appears as a facilitating and therefore major mechanism of the constitution of pulmonary fibrosis insofar as it will limit the normal repair of cells whose mitochondrial DNA has been affected to the point that the blocking this apoptosis alone prevents fibrosis (Lee CG. et al., J Exp Med, 2004, 200: 377-389; Budinger GR., Proc Nat Acad Sci USA, 2006, 103: 4604-4609).

3) The fibrotic process is linked to the hypoxia situation created by the loss of vascular endothelial cells (EMT for epithelio-mesenchymal transition) and the scarcity of capillaries, and which leads to the promotion of HIFla (Hypoxio Inducible Foctor), factor that activates ΙΈΜΤ as well as the production of NOX4, itself, responsible for the induction of an overproduction of ROS which maintains the system (Richter K. and Kietzmann T., Cell and Tissue Research, 2016, 365: 591 -605; Richter K. et al., Redox Biol, 2015, 6: 344-352).

4) Finally, the last piece of the puzzle is the TGFP (Tronsforming Growth Foctor) which is the most fibrogenic cytokine and which forms a vicious loop with NOX4 and ROS (each stimulating the production of the other) as shown in the figure below (where “mtROS” corresponds to mitochondrial ROS).

N0X4

This overproduction of TGFP has four contributing consequences in the promotion of pulmonary fibrosis (Cheresh P. et al., Biochim Biophys Acta, 2013, 1832: 1028-1040), namely:

- activation of fibroblast and collagen production;

- the induction of the epithelio-mesenchymal transition (EMT) and therefore the expansion of myofibroblasts (epithelial cells [or endothelial = EndoMT] then acquire a mesenchymal phenotype [fibroblast-like] and thereby invasive properties);

- promotion of apoptosis of epithelial cells;

- finally, the activation of the expression of the ECM gene (Extracellular Matrix) and the suppression of the degradation of the proteins of the cellular matrix which accumulate at the level of the interstitial tissue.

All these observations converge and advocate the determining role of ROS in pulmonary fibrosis, in particular mitochondrial ROS.

To date, pulmonary fibrosis does not benefit from any treatment allowing a cure.

Thus, in the present invention, the inventors propose to reduce the production of ROS, by acting at the mitochondrial level, to prevent the appearance of pulmonary fibrosis in any situation capable of promoting the appearance of such a pathology, or for treat already installed pulmonary fibrosis. This approach has the advantage of making it possible to envisage a benefit both in a preventive and curative framework, while avoiding the deleterious side effects associated with “classic” (non-selective) antioxidants.

The inventors have established, in a completely innovative manner, the interest of anethole trithione (ATT) as a specific inhibitor of ROS production of mitochondrial origin in the treatment and prevention of pulmonary fibrosis.

The ATT already benefits from a marketing authorization under the trade name of Sulfarlem® to increase the secretion of bile and saliva. It is used to treat difficult digestion and dry mouth. In the 70 years that this product has been marketed in France, no side effects have been reported to date relating to long-term use of this molecule.

Indeed, the mechanism of action of ATT can be deduced from the results of another experiment not published to date: the inventors have successfully treated a cat victim of acute poisoning with organochlorines. This result demonstrates that the target of ΙΆΤΤ is the complex I (or III) or, possibly, the mitochondrial l-lll supercomplex and this in connection with the results described in 2012 by Tebourbi (Tebourbi et al., Molecular mechanism of pesticide toxicity. In: STOYTCHEVA M. Ed. Pesticides in the modem world. Pests control and pesticides exposure and toxicity assessment. InTech publisher, 2012, chapter 15.). While knowing that as soon as ROS are overexpressed by the mitochondria, this induces a simultaneous overproduction of cytosolic ROS, whose production source is NADPH oxidase (or NOX, in general NOX4 and NOX2), some authors arrive at the conclusion that NOX are the first responsible when it is actually a mitochondrial production (Gosh R. et al., J. Clin. Diagn. Res., 2017, 11: BC09- BC12; Mangum LC et al ., Chem. Res. Toxicol, 2015, 28: 570-584).

In addition, ΓΑΤΤ is, unlike the majority of other antioxidants, a fat-soluble molecule which gives it excellent tissue and cell penetration (the Vd of ADT is, in rats, of the order of 2L / kg , or almost 10 times the volume of extracellular fluids [Yu HZ et al. J Pharm Sci, 2011, 100: 5048-5058]), explaining its access to the mitochondria. Thus, ΓΑΤΤ acts at concentrations of the order of a micromolar, 10 μΜ in the work of Pouzaud et al. (2004), concentration which is easy to reach at the usual doses; and this molecule seems to have persistent effects over time, certainly for several hours which should facilitate the method of administration of the product.

Finally, after administration in animals as in humans, ATT rapidly undergoes demethylation forming a derivative itself very likely to be active, 4-OHanetholtrithione (or ATX), carrying an alcohol function on the aromatic cycle. ; this function is esterifiable and can thus give access to water-soluble esters even though ATT and its metabolite ATX are insoluble in water.

The inventors have thus established that ATT, unlike conventional antioxidants, acts directly and selectively on the production of mitochondrial ROS, mainly at the level of complex I (or complex III) of the mitochondrial respiratory chain , sites which are both the main sites for the production of ROS and the main sites for mitochondrial dysfunctions (Tebourbi O., 2012, idem above).

ATT is therefore the first drug for human use authorized by the FDA and the EMA which prevents the mitochondria from producing ROS at the level of complexes I or III of the respiratory chain.

DETAILED DESCRIPTION OF THE INVENTION

A first object of the present invention relates to the use of a specific inhibitor of ROS production of mitochondrial origin to prevent and / or treat pulmonary fibrosis.

The term “specific inhibitor of ROS production of mitochondrial origin” means any compound capable of specifically inhibiting the production of ROS in the mitochondrial respiratory chain, without affecting the production of cellular ROS in the cytosol; this specificity is essential because it prevents the side effects associated with a defect in ROS in the cytosol, as can be observed in the event of excessive inhibition of ROS production by a non-selective antioxidant. In a preferred embodiment, this inhibitor is capable of inducing a specific inhibition of the production of ROS at the level of the complex I (or III) of the mitochondrial respiratory chain.

Such inhibitors are, for example, ATT, ΓΑΤΧ or NC-POBS, but any other compound having the same specificity of inhibition is suitable.

Within the meaning of the invention, the use of "an inhibitor" means the use of at least one specific inhibitor of the production of mitochondrial ROS; it can therefore be an inhibitor or a combination of several inhibitors, as described below.

ATT, for anethole trithione, is a 5- (4-methoxyphenyl) -3H-1,2-dithiole-3-thione. It is also known as ADT. Its formula is as follows:

gS

ATX corresponds to the phenolic form of ATT as it is metabolized by the liver, both in humans and in animals. This 4-OH-anethole trithione form has been described previously (Li et al., J Pharm Biomed Anal, 2008, 47: 612-617). The structure of ATT being conserved during this metabolization, there is every reason to believe that the antiROS activity carried by ATT is found in ATX, all the more so since after oral administration which is currently the marketed form, essential of the circulating product found is ΓΑΤΧ (Yu, 2011). In addition, ATX carries a phenol group in para which allows the formation of esters. In a particular embodiment, ATX is used in its esterified form, for example in the form of phosphate ester, ethylidenephosphate, sulfate, hemisuccinate, acetate, propionate, isobutyrate, d hexanoate, pivalate, ethoxycarbonate, nicotinate, or ester of amino acids such as glycine, diethylglycine or valine ester, and the list is not exhaustive.

NC-POBS corresponds to N-cyclohexyl-4- (4-nitrophenoxy) benzenesulfonamide, the only molecule described in the literature to date as a specific inhibitor of the production of mitochondrial ROS at site _Q of the respiratory chain ( Orr et al., 2013, Free Radie. Biol. Med., 65: 1047-1059).

In a preferred embodiment, the specific inhibitor is chosen from ΓΑΤΤ, ΙΆΤΧ or an ATX ester. In a particular embodiment, the treatment of pulmonary fibrosis is obtained by the association of at least two molecules among ΙΆΤΤ, ΙΆΤΧ and an ester of ATX.

By “treatment” is meant within the meaning of the invention both the treatment of an already established fibrosis and the prevention of the occurrence of such a pathology. Treatment also includes reducing symptoms and slowing the onset of signs associated with the condition. The subject to be treated can be a human being or an animal.

Pulmonary fibrosis, also called diffuse interstitial lung disease, corresponds to the modification of normal lung tissue into pathological fibrous tissue. This fibrosis is preferentially localized in the interalveolar spaces, where gaseous exchanges of oxygen and carbon dioxide take place, resulting in a drop in the oxygenation of the blood and respiratory insufficiency. The so-called "idiopathic" pulmonary fibrosis is a chronic, devastating and fatal disease, characterized by the progressive deterioration of pulmonary functions. Pulmonary fibrosis can be associated with asbestosis or silicosis.

In a particular embodiment, the present invention consists in treating a fibrosis associated with pulmonary edema.

In another particular embodiment, the present invention relates to a method of preventing pulmonary fibrosis associated with asbestosis or silicosis.

Asbestosis is a serious lung condition that affects people who have inhaled asbestos dust. After inhaling this dust, microscopic asbestos fibers settle in the lungs where they can cause permanent damage that can lead to pulmonary fibrosis.

Silicosis is a lung disease caused by the inhalation of particles of silica dust (crystalline silica) in mines, quarries, tunnel openings or construction sites, factories making "jeans", or even mills flour. It is an irreversible occupational disease.

"Prevention" means preventing any situation where the risk of developing pulmonary fibrosis is known and foreseeable, for example exposure to asbestos or silica, or the need for treatment with bleomycin .

The present invention also relates to a method of treating pulmonary fibrosis comprising administering a therapeutically effective dose of ATT to a patient in need thereof.

A second object of the present invention relates to the use of a pharmaceutical composition comprising ΓΑΤΤ for treating pulmonary fibrosis.

Such a composition is suitable for the therapeutic uses of the present invention and can be administered, for example, by the oral or intravenous route.

Claims (4)

1. Specific inhibitor of ROS production of mitochondrial origin chosen from anethole trithione (ATT), 4-OH-anethole trithione (ATX) or an ATX ester or a combination of at least two of these molecules , for use in the treatment of pulmonary fibrosis. 2. Pharmaceutical composition comprising a specific inhibitor of ROS production of mitochondrial origin chosen from anethole trithione (ATT), 4-OH-anethole trithione (ATX) or an ATX ester or a combination of at least two of these molecules and excipients for its use in the treatment of pulmonary fibrosis. 3. Inhibitor according to claim 1 or composition according to claim 2, for its use according to one of claims 1 or 2, in which the treatment consists in preventing the appearance of pulmonary fibrosis. 4. An inhibitor according to claim 1 or composition according to claim 2, for its use according to one of claims 1 or 2, in which the treatment consists in treating the symptoms of pulmonary fibrosis.