FR3144752A1 - USE OF PENTOSAN POLYSULFATE FOR THE TREATMENT OF DRY COUGH - Google Patents

Abstract

A composition comprising an effective amount of pentosan polysulfate for use as a medicament in the treatment of dry cough.

Description

USE OF PENTOSAN POLYSULFATE FOR THE TREATMENT OF DRY COUGH

The present invention relates to the use of a composition comprising pentosan polysulfate for the manufacture of a medicament for the treatment of dry cough.

Coughing is a normal reaction to irritation of the respiratory tract. Coughing is caused by irritation of the throat, trachea and bronchi. Coughing is not a disease but a symptom caused by irritation. Coughing is therefore generally an endogenous protective and cleansing mechanism.

Coughs can be grouped into two families: productive coughs and dry coughs.

A so-called productive cough is a productive or obstructed cough, intended to evacuate the airways of bodies obstructing these passages, due to the abundant production of phlegm and mucus.

The defense reflex due to bronchial or nasal secretions, from a wet cough, is reduced as soon as the bodies causing the irritation are expelled.

A so-called dry cough is a non-productive cough that is characterized by spasmodic, hard coughing fits in which no phlegm is expelled.

The defensive reflex due to the irritating sensations of a dry cough is difficult to alleviate.

The duration of a cough varies; it is considered an acute cough if a person has been coughing for less than three weeks, a subacute cough if it lasts for between three and eight weeks, and a chronic cough if the cough lasts for more than eight weeks.

A dry cough that does not go away on its own can be disabling or affect quality of life.

In general, medications to relieve dry cough are called antitussive drugs. They reduce the cough reflex by acting through the central nervous system. Antitussive drugs are divided into several categories, opiate antitussive drugs, antihistamine antitussive drugs and non-opiate and non-antihistamine antitussive drugs.

Opioid cough medicines have a sedative effect and should not be used in cases of respiratory failure or cough related to asthma. They can only be purchased with a prescription.

Antihistamine cough suppressants are recommended for dry, irritating coughs occurring at night, due to their sedative effect. They also have so-called atropinic effects, namely dry mouth, constipation, urinary blockage and visual disturbances.

Non-opiate and non-antihistamine cough suppressants are indicated for the short-term treatment of dry coughs. They are generally homeopathic medicines traditionally recommended for benign respiratory conditions.

Other alternatives have also been proposed, such as herbal supplements or the use of non-steroidal anti-inflammatory drugs.

Document EP 0 274 845 discloses the combination of ibuprofen as a non-steroidal anti-inflammatory drug with an opiate antitussive.

Document EP 2 106 258 describes the use of ibuprofen as a non-steroidal anti-inflammatory drug, as an antitussive, for the treatment of dry coughs of viral or bacterial origin.

WO 2014/161811 discloses the use of an astringent substance and a mucilaginous substance, such as Icelandic moss, for the treatment of irritations of the mucous membranes of the mouth and throat, as well as dry cough.

Document EP 3 086 799 describes the treatment of persistent coughs by a mechanism of physical protection of the mucous membranes of the upper airways, by the use of agents of plant origin.

The exact mechanism of a chronic cough is not clearly explained to date.

However, it is widely recognized that chronic cough is associated with airway inflammation.

Cough is generated by the cough reflex, a stimulus induced by activation of afferent, sensory nerves innervating the airway. These signals are transmitted centrally via the vagus nerve, which is connected to brainstem networks. Such networks coordinate the activation of motor output (e.g., phrenic, intercostal, and recurrent laryngeal nerves (RLNs) and the ultimate expression of cough) (Taylor-Clark TE. Peripheral neural circuitry in cough. Curr Opin Pharmacol. 2015 Jun;22:9-17.).

The majority of sensory nerve fibers innervating the airways are capsaicin-sensitive pulmonary vagal afferents (CSLV) (Agostoni, E., Chinnock, J. E., De Daly, M. B., and Murray, J. G. (1957). Functional and histological studies of the vagus nerve and its branches to the heart, lungs and abdominal viscera in the cat. J. Physiol. 135, 182–205.), the nociceptive sensory nerve endings.

These fibers are highly chemosensitive and can be activated by several endogenous mediators whose releases are elevated during pulmonary inflammation (Carr, M. J., and Undem, B. J. (2003). Bronchopulmonary afferent nerves. Respirology 8, 291–301.; Hsu, C. C., Lin, R. L., Lee, L. Y., and Lin, Y. S. (2013). Hydrogen sulfide induces hypersensitivity of rat capsaicin-sensitive lung vagal neurons: role of TRPA1 receptors. Am. J. Physiol. Regul. Integr. Comp. Physiol. 305, R769–R779.).

Anatomically, there are two types of neurons: myelinated Aδ fiber sensory neurons and C fiber sensory neurons. Myelinated Aδ fibers play a role in cough generation. Aδ fibers are responsible for the sudden, violent cough that occurs during aspiration (Lieu T, Undem BJ: Neuroplasticity in vagal afferent neurons involved in cough. Pulm Pharmacol Ther 2011, 24(3):276–279. ; Canning BJ, Mori N, Mazzone SB: Vagal afferent nerves regulating the cough reflex. Respir Physiol Neurobiol 2006, 152(3):223–242.).

In recent years, tremendous progress has been made in our understanding of the neural mechanism of itch. As an endogenous neurotransmitter, serotonin has emerged as a key player in both acute and chronic itch (X. Dong (2018): “Peripheral and central mechanisms of itch”. Neuron 98, 482–494.).

In our body, it appears that 95% of serotonin is synthesized and released by enterochromaffin cells in the intestine, in response to mechanical or chemical stimulation (Linan-Rico, A., et al. (2016), Front. Neurosci. 10:564.)

Recently, serotonin and its receptor 5HT7 have been identified as the key player in mediating acute and chronic skin itch sensation (Morita T, McClain SP, Batia LM, et al. 2015: HTR7 Mediates Serotonergic Acute and Chronic Itch. Neuron. 87:124–38).

Serotonin has been shown to produce strong and robust itch in mice and rats, as well as in humans (Akiyama et al., 2009: T. Akiyama, A.W. Merrill, M.I. Carstens, et al. Activation of superficial dorsal horn neurons in the mouse by a PAR-2 agonist and 5-HT: potential role in itch. J Neurosci, 29, pp. 6691-6699; Morita T, McClain Shannan P, Batia Lyn M, Pellegrino M, Wilson Sarah R, Kienzler Michael A, Lyman K, Olsen Anne Sofie B, Wong Justin F, Stucky Cheryl L, et al. (2015): “HTR7 Mediates Serotonergic Acute and Chronic Itch”. Neuron. ; 87:124–138.)

C fibers are also involved in cough generation. C fibers are thought to be important for dry coughs, which are used to get rid of an itchy feeling in the throat. This feeling is more typical of chronic coughs.

Serotonin and its receptors may also play a similar role in the lungs (Pecova T, Kocan I, Vysehradsky R, Pecova R. Itch and Cough - Similar Role of Sensory Nerves in Their Pathogenesis. Physiol Res. 2020 Mar 27;69(Suppl 1):S43-S54.).

Indeed, serotonin exerts a strong stimulatory effect on pulmonary C-fiber terminals via activation of the serotonin 5-HT3 receptor, which may contribute to airway hyperresponsiveness under pulmonary inflammation (Potenzieri C, Meeker S, Undem BJ. Activation of mouse bronchopulmonary C-fibers by serotonin and allergen-ovalbumin challenge. J Physiol. 2012 Nov 1;590(21):5449-59. ; Hsu CC, Ruan T, Lee LY, Lin YS. Stimulatory Effect of 5-Hydroxytryptamine (5-HT) on Rat Capsaicin-Sensitive Lung Vagal Sensory Neurons via Activation of 5-HT3 Receptors. Front Physiol. 2019 May 28;10:642.).

During tissue infections and inflammation, the number of myeloid-derived suppressor cells increases very significantly (Ost, M., Singh, A., Peschel, A., Mehling, R., Rieber, N., and Hartl, D. (2016). Myeloid Derived Suppressor Cells in Bacterial Infections. Front. Cell. Infect. Microbiol. 6).

In response to an inflammatory signal, myeloid cells produced by the bone marrow migrate into the blood and become functionally active, with the aim of restoring or replacing damaged peripheral populations (Panopoulos, A.D., and Watowich, S.S. (2008). Granulocyte colony-stimulating factor: molecular mechanisms of action during steady state and emergency hematopoiesis. Cytokine 42, 277–288.).

In addition to these conventional immune effector cells, hematopoietic stem cells have been shown to directly participate in the inflammatory process in respiratory diseases, such as asthma, allergies, etc. (Massberg S, Schaerli P, Knezevic-Maramica I, Köllnberger M, Tubo N, Moseman EA, Huff IV, Junt T, Wagers AJ, Mazo IB, von Andrian UH. (2007) “Immunosurveillance by hematopoietic progenitor cells trafficking through blood, lymph, and peripheral tissues.” Cell. 131(5):994-1008 ; Allakhverdi, Z., Comeau, M. R., Smith, D. E., Toy, D., Mfuna Endam, L., Desrosiers, M., Liu, Y-. J., Howie, K. J., Denburg, J. A., Gauvreau, G. M., Delespesse, G. (2009): “CD34+ hemopoietic progenitor cells are potent effectors of allergic inflammation”. Journal of Allergy and Clinical Immunology, Volume 123, Issue 2, Pages 472-478.e1; Blanchet, M. R., McNagny, K. M. (2009): “Stem cells, inflammation and allergy”. Allergy Asthma Clin Immunol. 5(1):13; Fischer, K. D., & Agrawal, D. K. (2013): “Hematopoietic stem and progenitor cells in inflammation and allergy”. Frontiers in immunology, 4, 428.).

These results are confirmed by the inventor's work: a large quantity of hematopoietic stem cells are visible on biopsies from patients suffering from inflammatory diseases. In addition, serotonin has been detected in these hematopoietic stem cells.

As mentioned previously, during inflammation, hematopoietic cells aggregate and produce serotonin, which is partly responsible for the itching, the stimulus for dry coughs.

The applicant discovered that pentosan polysulfate modified the morphology of hematopoietic cells. The modification of the morphology of these cells limits or even prevents their aggregation during an inflammation process, limiting the release of serotonin.

Pentosan polysulfate, also called pentosan polysulfate or pentosan sulfuric polyester, has been used in various therapeutic applications.

Pentosan polysulfate is a glucosaminoglycan analogue, derived from heparin, with anticoagulant, fibrinolytic and anti-inflammatory properties.

Document WO 93/23059 describes the use of compositions comprising pentosan polysulfate in the treatment of thrombocytopenia.

WO 2019/023761 discloses the use of pentosan polysulfate in the treatment of diseases or disorders associated with bone marrow pathologies.

Document EP 1 038 528 describes the use of pentosan polysulfate in the treatment of fibrosis.

Document EP 1 488 788 describes the use of compositions comprising pentosan polysulfate in the treatment of gastrointestinal disorders.

Document EP 2 164 501 discloses the use of pentosan polysulfate in the treatment of diseases such as asthma, allergic rhinitis and chronic obstructive pulmonary disease.

Pentosan polysulfate is also used as an active ingredient in creams aimed at combating inflammation. They are used in the local treatment of bruises and hematomas, and even muscle and joint pain.

Pentosan polysulfate is also indicated for the treatment of painful bladder syndrome characterized by glomerulations or Hunner's ulcers in adults with moderate to severe pain, urinary urgency, and pollakiuria. Its mechanism of action in the treatment of painful bladder syndrome is poorly understood; pentosan polysulfate appears to act by a local effect in the bladder through binding of glycosaminoglycans to the bladder mucosa and by an anti-inflammatory effect.

Excessive or chronic cough is a major problem that is poorly managed by currently available therapies.

Thus, there is a need for the development of new drugs.

The present invention provides for treating excessive or chronic dry coughs by the use of compositions comprising pentosan polysulfate.

The present invention therefore relates to a composition comprising an effective amount of pentosan polysulfate (PPS) for use as a medicament in the treatment of dry cough.

The implementation of the invention will be better understood with the aid of the detailed description which is set out below with regard to the appended figures in which:

Figures 1 to 3 illustrate images of lungs, pieces of lung parenchyma, and pieces of bronchial rings.

is an image representing normal lungs, uniform in color and smooth on the surface.

is an image representing a piece of lung parenchyma from a patient with pulmonary fibrosis.

is an image showing two pieces of bronchial rings, on the left, from an organ donor without respiratory disease; on the right, from a patient who received a lung transplant.

is a close-up image of blood congestions in the bronchial lumen of a patient with chronic lung disease.

Figures 5 and 6 illustrate optical microscope images of blood clot cells from an allergic donor, at x20 magnification.

is a contrast phase image.

is a fluorescent image showing a subset of blood clot cells expressing a high amount of serotonin represented by a red color in true color, and illustrated by light areas.

Figures 7 and 8 illustrate optical microscopic images of blood clot cells stained positive for the serotonin 5HT2C receptor by a specific monoclonal antibody, at ×20 magnification.

is an image of the contrast phase.

is a fluorescent image of 5-HT2C staining, colored green in true color, and illustrated by light areas.

Figures 9 to 11 illustrate optical microscope images of blood congestion cells.

illustrates a control image of blood congestion cells without treatment, magnification x20.

illustrates an image of blood clot cells after being exposed to an aqueous solution of 0.5% pentosan polysulfate for 24 hours, the blood clot cells became needle-shaped, magnified x20.

is an enlargement of the .

Thus the invention relates to the treatment of dry cough comprising the administration of a composition comprising an effective amount of pentosan polysulfate, for a duration and under conditions sufficient for the treatment of dry cough.

More specifically, the invention relates to the use of pentosan polysulfate in the manufacture of a medicament for the treatment of dry cough.

The term "effective amount" of pentosan polysulfate as used herein means a sufficient amount of pentosan polysulfate to provide the desired therapeutic or physiological effect.

The exact amount required varies from subject to subject, the age and general condition of the subject, the method of administration, etc.

An appropriate "effective amount" can be determined using routine experimentation. Effective amount in this context includes an amount required to treat dry cough.

These compositions are proposed for use as pharmaceutical compositions or formulations.

The compositions comprise advantageously aqueous and sterile solutions.

Sterile solutions are prepared by incorporating a required amount of pentosan polysulfate in a suitable solvent, possibly with other active ingredients, followed by sterilization or at least a process to reduce contaminating viruses, bacteria or other biological entities to acceptable levels.

Pentosan polysulfate (PPS) is preferably selected from the group consisting of: pentosan polysulfate sodium salt (NaPPS), pentosan polysulfate magnesium salt (MgPPS), pentosan polysulfate calcium salt (CaPPS), and pentosan polysulfate zinc salt (ZnPPS).

Preferably, the pentosan polysulfate (PPS) is sodium pentosan polysulfate (NaPPS).

In some embodiments, the medicament is administered by inhalation or as a nasal spray, delivered by an inhaler.

According to one characteristic, the mass concentration of pentosan polysulfate, expressed as a percentage, is between 0.5 and 3%.

According to one embodiment, the mass concentration of pentosan polysulfate, expressed as a percentage, is equal to 1%.

The solvent is preferably an aqueous saline solution, for example comprising a mass concentration, expressed as a percentage, of sodium chloride of 0.9%, buffered with 10mM HEPES (or 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid).

It is understood by other active ingredients, an antibiotic or antiviral molecule.

Healthy lungs are more or less uniform in color and shape, as shown in .

Lung parenchyma as well as bronchi of patients with chronic respiratory diseases, such as cystic fibrosis, idiopathic pulmonary fibrosis, sarcoidosis, show areas of aggregation of hematopoietic cells that are red in real color and illustrated by dark areas, as shown in Figures 2 to 4.

These areas of cellular aggregation are commonly referred to as blood congestion in the lungs.

These blood congestions thus directly participate in inflammatory responses and tissue repair.

By immunocytochemistry, the applicant discovered that the aggregation of hematopoietic cells in the lungs express a large amount of serotonin, as illustrated in .

These observations of the presence of serotonin were confirmed by two different antibodies in several independent experiments, with the same results, an example of such confirmation is illustrated in .

Thus, not only do the clusters of hematopoietic cells in the lungs express serotonin, but also a serotonin receptor, 5-HT2C ( ).

Based on these data, the applicant proposes the following scheme of chronic cough: during viral or bacterial infections, or tissue damage, pro-inflammatory mediators, including serotonin, are released in the affected area.

These mediators will recruit hematopoietic cells from surrounding lung tissues as well as from the bloodstream.

Serotonin would act on hematopoietic cells mainly via 5HT2C receptors in a paracrine and autocrine manner, amplifying the inflammatory response.

A large number of blood-forming cells will rapidly accumulate in the affected area, making the affected area red in color.

Blood cells release an increasing amount of serotonin which activates C fibers causing an itching sensation via the 5HT3 or 5HT7 receptor, activating the cough reflex.

As long as areas of significant aggregation of hematopoietic cells are present, for example as shown in Figures 2 and 3, the dry cough should not stop, due to the continued release of serotonin from the hematopoietic cell aggregates.

Additionally, dry cough can further increase serotonin secretion, creating a positive feedback loop.

The applicant has discovered that pentosan polysulfate has a direct effect on hematopoietic cells located in cell aggregates: as an anticoagulant.

Indeed, pentosan polysulfate significantly modifies the morphology of hematopoietic cells in vitro, as illustrated in .

Upon treatment with pentosan polysulfate, hematopoietic cells extracted from the aggregates changed morphology to a needle-like or nail-like shape, as illustrated in Figures 10 and 11.

To aggregate, cells must be disk-shaped, as shown in .

This morphological change induced by pentosan polysulfate would prevent hematopoietic cells from forming cell aggregates.

This hypothesis not only provides an explanation for how pentosan polysulfate works, but also a rationale for its use as a drug to break the vicious cycle of chronic inflammation in various inflammatory diseases, especially respiratory diseases, which are frequently associated with chronic cough.

Local and direct application of pentosan polysulfate by inhalation or as a nasal spray, to the respiratory tract, will have a double benefit, namely increased drug efficacy and reduced drug toxicity.

Moreover, by acting directly on hematopoietic cell aggregates, pentosan polysulfate does not alter the cough reflex, thus preserving the protective effects of coughing against harmful agents of the respiratory tract.

Materials and methods:

Lungs and lung biopsies:

The organ pieces shown in Figures 2–4 were obtained from patients who underwent lung transplantation or polypectomy. All experimental procedures were fully explained, and all subjects received informed consent. The study was conducted in accordance with the Declaration of Helsinki on Biomedical Research (Hong Kong Amendment, 1989), and received approval from the local ethics committee.

Isolation of hematopoietic stem cells from lung parenchyma:

Lung tissue samples were dissected into approximately 5 mm ³ pieces. Approximately 10 g of minced tissue was placed in 50 mL conical tubes filled with wash buffer (Dulbecco's Phosphate Buffered Saline (DPBS) each containing 100 U/mL penicillin and streptomycin). The tubes were inverted to wash the tissue and the supernatant removed, repeating these steps three times. As much liquid as possible was removed, minced and washed tissues were filtered through a layer of sterile gauze. Tissues on the gauze were distributed into 15 mL Falcon tubes, comprising approximately 1.5 g of tissue per tube. Filtrates were passed through a 10 μm cell strainer. Cells were collected by centrifugation.

Immunostaining, figures 5, 6 and 8:

Cells are grown on a coverslip and then fixed with a solution of 2% formaldehyde and 2.5% glutaraldehyde in 0.1 M sodium phosphate buffer for 30 minutes at room temperature. The fixed cells are then rinsed several times (3 X) with PBS (phosphate buffered saline). To facilitate antibody access, the fixed cells are then permeabilized with 0.25% Triton-X-100. Fixed cells were rinsed several times (3 X) with PBS. A solution of 100 µl of PBS supplemented with 10% serum (bovine, goat, or human, depending on the primary antibody used), was applied for 60 minutes at room temperature. 100 µl, or a volume sufficient to cover the cells, of primary antibody was applied and incubated for 30-60 minutes. Cells were rinsed with PBS 3 times for 2 minutes. 100 µl, or a volume sufficient to cover the cells was applied, of a solution of secondary antibody conjugated with a fluorophore (Alexa 488, Alexa 560). There The coverslip is then mounted on a glass slide for observation under a fluorescence microscope.

For the sample illustrated in the , the primary antibody is a specific antibody directed against serotonin (Sigma, S5545). This antibody was produced in rabbits (dilution 1 in 50 in PBS). The secondary antibody used is therefore an anti-rabbit IgG antibody, produced in goats, conjugated with a red fluorochrome [Alexa Fluor 568, Goat anti-rabbit IgG (H+L), dilution 1 in 500 in PBS], (Invitrogen, A11011). The photo was taken on a fluorescence microscope.

For the sample shown in the , the primary antibody is a specific antibody directed against the Serotonin receptor, 5HT2C (R&D Systems, Minneapolis, MAB5764). This antibody was produced in mice (dilution 1 in 50 in PBS). The secondary antibody used is therefore an anti-mouse IgG antibody, produced in goats, conjugated with a green fluorochrome [Alexa Fluor 488, Goat anti-rabbit IgG (H+L), dilution 1 in 500 in PBS], (Invitrogen, A11011). The photo was taken on a fluorescence microscope.

Claims (4)

A composition comprising an effective amount of pentosan polysulfate for use as a medicament in the treatment of dry cough. Composition according to claim 1, characterized in that the mass concentration, expressed as a percentage, of pentosan polysulfate is between 0.5 and 3%. Composition according to claim 1 or 2, characterized in that it comprises an antibiotic or antiviral molecule. Composition according to any one of claims 1 to 3, characterized in that it is capable of being packaged in an inhaler to be capable of being administered by inhalation or in the form of a nasal spray.