DESCRIPTION

FIELD OF THE INVENTION

The technical field relevant to the present invention is that of local (non-systemic) therapies for the treatment of candidiasis.

BACKGROUND ART

Vaginal candidiasis is a fairly common fungal infection. In 85% of cases, the fungus responsible for this disease is Candida albicans, which is normally present in the vaginal environment and in other hot-humid body districts, where it coexists with other microorganisms (fungi and bacteria) in its inactive, generally harmless form.

In normal physiological situations, the vaginal microbiota and the immune system participate in the maintenance of an equilibrium condition which prevents the proliferation of potentially harmful microorganisms. However, in particular situations these control systems fail and the Candida begins to replicate in an uncontrolled manner, giving rise to the condition known as Candida or candidiasis infection. Recurrent vulvovaginal candidiasis is a frustrating and disabling infection for the woman and for the intimacy of a couple.

Approximately 70-75% of women, particularly women of childbearing age, experience a vulvovaginal Candida infection at least once in their lifetime and 40-50% of these are recurring. Approximately 20% of acute infections evolves into a form of recurrent Candida vulvo-vaginitis, characterized by four or more episodes in a year (Murina F, Graziottin A, Felice R, Radici GL, Di Francesco S. The recurrent vulvovaginal candidiasis: proposal of a personalized therapeutic protocol. ISRN Obstet Gynecol. 2011; 2011: 806065). The diagnosis of recurrent candidiasis may not be easy, the symptomatology not blatant, the objectivity not conspicuous and the 20- 30% positivity rate of the culture examination forces the doctor to face a number of difficulties. Azole and triazole derivatives are the treatment of choice for Candida vulvovaginitis. Numerous topical azoles are available with different administration possibilities; these are safe and generally well tolerated. Candidiasis treatment occurs by means of drug therapy based on antifungal drugs. The most commonly used drugs in the presence of vaginal Candida are selected from: clotrimazole, econazole, fenticonazole nitrate, miconazole and nystatin, in the form of a vaginal cream or suppository; at the same time, the oral intake of one of fluconazole and itraconazole.

Problem of the prior art

However, given that the tendency of candidiasis to recur, even after 6 months of drug therapy, can reach 50% of treated women (Mendling W, Brasch J. German Society for Gynecology and Obstetrics. Guideline vulvovaginal candidosis (2010) of the German Society for Gynecology and Obstetrics, the Working Group for Infections and Infectimmunology in Gynecology and Obstetrics, the German Society of Dermatology, the Board of German Dermatologists and the German Speaking Mycological Society. Mycoses, 55, Suppl 3: 1-13, 2 12), it would be appropriate to develop new therapeutic protocols capable of replacing or flanking the pharmacological approach, increasing the effectiveness of the treatment in decreasing recurrences.

Therefore, the need remains to treat the symptoms associated with candidiasis with the use of associations of active ingredients with reduced or no side effects as opposed to conventional drug therapy and which, at the same time, improve patient compliance.

SUMMARY OF THE INVENTION

A first object of the present invention is an association comprising molecular oxygen and hyaluronic acid or salts thereof for use in the treatment of candidiasis, preferably for use in the treatment of recurrent candidiasis.

A further object of the present invention is a kit for use in the treatment of candidiasis, preferably of recurrent candidiasis.

Advantages of the invention

The association is particularly advantageous since it combines, at the tissue/genital level, the effects of two active ingredients, molecular oxygen and hyaluronic acid in an enhanced and/or synergistic way. This means that the molecular oxygen and hyaluronic acid comprised in the association of the invention support, promote and assist each other's action.

Although the use of an association of high concentration oxygen and hyaluronic acid for use in the topical treatment of vaginal disorders selected from vaginal dryness and vulvovaginal atrophy (AVV) is known in the prior art (European Patent Application No. 3645016), the Applicant is not aware of any disclosure related to the therapeutic use of such an association for the treatment of candidiasis.

The association for use for the purposes of the invention has proved to be a valid method for the treatment of candidiasis, in particular that of the recurrent type, as well as for the treatment of the symptoms associated with the candidiasis itself.

It is a totally painless therapy which shows excellent compliance from patients. It is a fast, non- invasive and repeatable treatment, with no side effects.

It is known that oxygen therapy increases the availability of oxygen to tissues, promotes the increase of tissue repair processes and increases collagen synthesis, allowing a normal hydroxylation of this protein. In fact, at tissue oxygen tensions which are lower than normal, collagen is not correctly synthesized, slowing the healing of ulcers and wounds (Hunt TK, Pai MP. The effect of varying ambient oxygen tensions on wound metabolism and collagen synthesis. Surg Gynecol Obstet. 1972;135(4):561- 567). Furthermore, oxygen induces a neo-angiogenetic stimulus by means of the release of factors such as Vascular Endothelial Growth Factor (VEGF) (Sheikh AY, Gibson JJ, Rollins MD, et al. Effect ofhyperoxia on vascular endothelial growth factor levels in a wound model. Arch. Surg. 2000; 135:1293-1297). This function is essential for the restoration of microcirculation in compromised vascular situations, restoring a vascular flow in hypoxic areas which ensures proper tissue regeneration (Tandara, A. & Mustoe, T. Oxygen in Wound Healing — More than a Nutrient. World J. Surg. (2004) 28: 294).

Hyaluronic acid is a natural polysaccharide which forms a fundamental part of the extracellular matrix of skin and cartilage. Hyaluronic acid exhibits remarkable adhesive, moisturising and repairing properties of the vaginal mucosa (Chen J et al. “Evaluation of the efficacy and safety of hyaluronic acid vaginal... ” J Sex Med. 2013; 10; 1575-84).

Therefore, the association for use which is the object of the invention promotes an improvement in the quality of life of people suffering from this disease. The Applicant has also demonstrated its effectiveness in the experimental test of Example 1.

DESCRIPTION OF THE FIGURES

Figure 1 - Depiction of the cannula-type dispensing device;

Figure 2 - Depiction of the spray gun-type dispensing device;

Figure 3 - Graphical depiction of the results after application of the administration protocol of Example 1.

DETAILED DESCRIPTION OF THE INVENTION

The Applicant describes the different objects of the invention in more detail below.

For the purposes of the invention, candidiasis (or Candida) means a fungal infection (or mycosis) caused by the fungus (or mycetes) of the genus Candida (or Candida sppi).

Preferably, the candidiasis is acute candidiasis (or of acute type). The symptoms associated with acute candidiasis are selected from the group consisting of: itching, preferably with rapid onset, redness, edema, preferably edema of the external genitalia, leukorrhea (or vaginal discharge, preferably white), burning and dyspareunia, preferably superficial.

Even more preferably, the candidiasis is recurrent candidiasis (invasive or cyclic). Recurrent candidiasis is preferably characterized by frequent recurrences, for example four or more episodes in a year. The symptomatology associated with recurrent candidiasis is preferably selected from the group consisting of: itching, preferably less intense than that attributable to acute candidiasis; burning; dyspareunia, preferably the latter two symptoms being more severe with respect to those associated with acute candidiasis; leukorrhea, preferably less abundant than in the acute form. Recurrent candidiasis is notoriously characterized by an excessive response, particularly of the immune-allergic type, to an amount of Candida antigens which would not otherwise cause any problems.

Symptomatology associated with candidiasis is preferably intended as that selected from the group consisting of: redness, burning, pain, preferably acute and/or chronic pain, recurrent infections, dyspareunia, itching, edema and combinations of the foregoing.

Association for use

As anticipated, the association which is the object of the invention comprises or consists of molecular oxygen (or gaseous oxygen) and hyaluronic acid or salts thereof.

The association is intended for the treatment of candidiasis, preferably recurrent candidiasis. More in particular, the association of the invention is intended for use in reducing the recurrent frequency of candidiasis.

According to a preferred embodiment, the association of the invention is also useful in the treatment of the symptomatology associated with candidiasis, where the symptomatology associated with candidiasis preferably means that selected from the group consisting of: redness, burning, pain, preferably acute and/or chronic pain, recurrent infections, dyspareunia, itching, edema and mixtures of the foregoing.

Preferably, the association which is the object of the invention is intended for topical vaginal use.

Still preferably, the association which is the object of the invention is intended for use as monotherapy in the treatment of candidiasis, preferably of recurrent candidiasis; or, is intended for use in the treatment of candidiasis, preferably of recurrent candidiasis, as an adjunct to traditional antifungal therapy.

It should be noted that traditional antifungal therapy means topical -genital and/or systemic therapy using one or more of the active substances selected from the group consisting of: clotrimazole, econazole, fenticonazole nitrate, miconazole, nystatin, fluconazole and itraconazole. The association according to the invention comprises molecular oxygen (or gaseous oxygen) with a degree of purity > 90% (v/v), preferably comprised between 90% and 99%, preferably comprised between 90% and 96% (v/v), preferably comprised between 92% and 95% (v/v), preferably equal to 95% (v/v).

The molecular (or gaseous) oxygen comprised in the association can be obtained by employing methodologies known to those skilled in the art; preferably, the molecular oxygen is obtained by virtue of an oxygen concentrator, which sucks external air and concentrates it in the oxygen fraction. The maximum delivery pressure by the oxygen concentrator is preferably comprised between 50 and 150 kPa, preferably it is 100 kPa.

The molecular oxygen dosage to be administered is preferably comprised between 5 and 130 L, preferably comprised between 5 and 120 L, preferably between 10 and 130 L, preferably comprised between 10 and 120 L, preferably between 20 and 120 L, preferably between 10 and 90 L, preferably between 10 and 70 L.

According to a preferred form of the invention, the molecular oxygen is delivered in a gas form at a flow preferably comprised between 0.5 and 7 L/min, preferably between 1 to 6 L/min, preferably between 1.5 and 4.5 L/min, preferably equal to 3 L/min.

The association according to the invention preferably comprises low molecular weight hyaluronic acid; preferably, the association according to the invention comprises hyaluronic acid with a molecular weight comprised between 200 kDa and 2 MDa.

According to a preferred embodiment, the hyaluronic acid is preferably dissolved in an aqueous solution at a concentration preferably comprised between 0.050% and 0.50% by weight, preferably comprised between 0.050% and 0.30% by weight, preferably comprised between 0.10% and 0.25% by weight, preferably equal to 0.20% by weight on the total volume of the aqueous solution. It should be noted that, preferably, the hyaluronic acid or salts thereof constitute the only active ingredient of the association which is dissolved in aqueous solution.

Preferably, the dosage of hyaluronic acid is comprised between 10 mg and 120 mg per dosage unit. Preferably, the dosage unit contains or consists of the aqueous solution of hyaluronic acid in a volume comprised between 5 and 15 ml, preferably equal to 10 ml. According to a preferred embodiment, the hyaluronic acid solution can be comprised in a closed container, preferably an ampoule, bottle, vial.

Preferably, the hyaluronic acid is in the form of a hyaluronate salt, preferably sodium hyaluronate.

According to a preferred embodiment, the association of the invention is dispensed directly at the level of the genital tissue outside or inside the vaginal cavity. It should be noted that the association of the invention is dispensed in the form of steam (i.e., vaporized or nebulized).

The association according to the invention can preferably be dispensed in a time comprised between 5 and 20 minutes (extremes included), preferably comprised between 5 and 10 minutes, preferably equal to 15 minutes, preferably equal to 10 minutes, preferably equal to 5 minutes.

Administration protocol

According to a preferred form of the invention, the association is dispensed following an administration protocol which comprises the following steps: i) dispensing the association to the external or vulvar tissue, ii) dispensing molecular oxygen inside the female genital tract or internal vaginal cavity, ii) dispensing the association inside the female genital tract or internal vaginal cavity, the association being that for use which is the object of the invention.

The anatomical organs and structures located inside the female genital tract are preferably: the vagina, the cervix, the uterus, the fallopian tubes and the ovaries; the organs and the anatomical structures located outside, on the other hand, are preferably: the mons pubis, the labia majora, the labia minora, Bartholin's glands and the clitoris.

For the dispensing of the association at the level of the genital tissue, external or internal, two types of dispensing devices can preferably be employed: a cannula or a spray gun. The cannula is preferably intended for the intravaginal dispensing of the oxygen, hyaluronic acid or salts thereof and/or the association of the invention; the spray gun is preferably employed for the external dispensing of the molecular (or gaseous) oxygen, the hyaluronic acid or salts thereof and/or the association of the invention.

The external application step of the association i) preferably has a duration comprised between 2 and 8 minutes, preferably equal to 5 minutes; the internal application step of molecular oxygen ii) preferably has a duration comprised between 7 and 13 minutes, preferably equal to 10 minutes; finally, the internal application step of the association iii) has a duration preferably comprised between 2 and 8 minutes, preferably equal to 5 minutes.

Preferably, the association according to the invention is to be administered weekly, preferably once weekly; in particular, the association is to be administered for a period of time comprised between 5 and 40 consecutive days, preferably comprised between 5 and 10 days, preferably equal to 7 days.

Kit

A further object of the present invention is a kit for use in the treatment of candidiasis, preferably of recurrent candidiasis, comprising:

- hyaluronic acid or salts thereof;

- molecular oxygen;

- instructions for the preparation of an association comprising molecular oxygen and hyaluronic acid or salts thereof;

- one or more dispensing devices suitable for nebulizing an association comprising molecular oxygen and hyaluronic acid or salts thereof, the association being that of the invention described above.

According to a preferred embodiment, the kit for use in the treatment of symptoms and/or disorders from (or associated with) candidiasis also comprises an oxygen concentrator. It should be noted that the molecular oxygen included in the kit can for example be in the form of pressurized, fillable or refillable cylinders. When the kit comprises the oxygen concentrator, the molecular oxygen is generated extemporaneously, without the need for storage.

According to a further preferred embodiment, the kit for use in the treatment of symptomatology and/or disorders from (or associated with) candidiasis further comprises a molecular oxygen transport cable.

The hyaluronic acid or salts thereof is preferably comprised inside a closed container, such as an ampoule, bottle, vial. The closed container forms a dosage unit preferably having a volume comprised between 5 and 15 ml, preferably equal to 10 ml; according to the latter embodiment, the number of closed containers comprising hyaluronic acid or salts thereof is equal to both the number of cannulas and the number of molecular oxygen transport cables.

Salt of hyaluronic acid is preferably intended as sodium hyaluronate.

The hyaluronic acid or salts thereof is preferably in the form of an aqueous solution of hyaluronic acid, preferably in a concentration comprised between 0.050% and 0.30% by weight, preferably comprised between 0.10% and 0.25% by weight, preferably equal to 0.20% by weight on the total volume of the aqueous solution. According to a preferred embodiment, the nebulization of the association comprising molecular oxygen and hyaluronic acid directly onto the areas affected by the infection is possible.

According to a preferred embodiment, the kit for use comprises a disposable dispensing device, preferably a vaginal cannula. It should be noted that the vaginal cannula forms a vaginal dispensing (or applicator) device.

A disposable dispensing device is a device which is used a single time or for a single treatment/protocol, then disposed of, i.e., not reusable a second time.

With reference to Figure 1, we describe the cannula device 100 below, configured for dispensing the association according to the invention.

The cannula 100 comprises a hollow container body 100a, extending between two ends, along a horizontal axis (X-X): a proximal end 101 and a distal end 102. The proximal end 101 comprises an opening (not shown in Figure 1), arranged on a flat surface in section with respect to the cannula container body. The opening comprises anchoring means for connecting the proximal end 101 to a molecular oxygen transport cable of an oxygen concentrator.

The container body 100a comprises an inner cavity, intended to receive the flow of molecular oxygen coming from the molecular oxygen transport cable and possibly an aqueous solution of hyaluronic acid; and a hole 100b which crosses the container body 100a, putting the inner cavity in fluid communication with the outside.

The distal end 102 comprises a head portion 102a, oval or rounded in shape, provided with at least two holes 102b for nebulizing the association of molecular oxygen and hyaluronic acid or salts thereof. Preferably, the head portion 102a is in a single piece with the container body 100a.

It should be noted that the head portion 102a is also hollow and in flow communication with the hollow container body 100a of the cannula.

According to a preferred embodiment, the head portion 102a comprises at least two shaped ribs 102c, which protrude from the outer surface of the head portion 102a, defining depressions 102d where the at least two holes 102b are located. The holes 102b cross the head portion 102a, putting the hollow container body 100a of the cannula in fluid communication with the outside.

Preferably, there are four shaped ribs 102c arranged crosswise on the head portion 102a. The ribs are preferably tapered, widening towards the distal end 102 and narrowing towards the proximal end 101, so as to give the head portion 102a a comfortable shape for intravaginal insertion.

According to a preferred embodiment, the holes 102b are located in pairs (equal to 2) on each of the depressions defined by the ribs 102c.

It should be noted that, advantageously, the ribs 102c thus shaped allow, at the time of use, to distance the mucosal tissues from the outer surface of the cannula, facilitating the exit of the molecular oxygen and/or the association comprising molecular oxygen and hyaluronic acid.

It should be noted that the molecular oxygen transport cable is the cable of an oxygen concentrator. Preferably, the molecular oxygen transport cable is of a material suitable for transporting molecular oxygen (or gaseous oxygen), preferably it is in polymeric material, preferably it is in polypropylene PP. The oxygen concentrator is an instrument known in the state of the art and commercially available, configured to suck in outdoor air and fdter it, to obtain air with an oxygen titre > 80%.

It should be noted that a generic oxygen concentrator preferably comprises:

- a compressed air generator or compressor configured to draw air from the external environment, filter and compress it.

- a machine body, comprising or equipped with molecular zeolite sieves. The machine body uses the principle of different absorption of gas molecules to let O2 pass and retain other gases in the air, such as nitrogen, argon, helium and hydrogen. The machine body transforms the outside air into 93-95% pure oxygen.

- oxygen transport cable, equipped with anchoring means suitable for coupling a dispensing device, said dispensing device preferably being chosen between vaginal cannula and spray gun.

According to a further embodiment, the kit for use according to the invention comprises a multi-purpose (or reusable) dispensing device, preferably a spray gun.

Preferably, the kit for use according to the invention comprises both a disposable dispensing device, preferably a vaginal cannula, and a multi-purpose (or reusable) dispensing device, preferably a spray gun.

With reference to Figure 2, a preferred embodiment of a spray gun suitable for the purposes of the invention is described below. The spray gun 200 comprises a hollow container body 200a, having an upper surface 201 and a lower surface 202; the hollow container body extends between two ends, along a horizontal axis Y-Y: a proximal end 203 and a distal end 204.

The container body comprises an inner cavity, suitable for receiving a pressurized flow of molecular oxygen.

The distal end 204 consists of a handle for use by a user.

The proximal end 203 comprises an opening (not shown in the figure), arranged on a flat surface in section with respect to the container body 200a of the spray gun, which puts the inner cavity in fluid communication with the outside. The opening comprises a nozzle 203a for the exit of the molecular oxygen or the association of molecular oxygen and hyaluronic acid.

The spray gun comprises a conduit 205, in fluid communication with the inner cavity of the container body, arranged between the proximal end 203 and the distal end 204, and protruding from one of the surfaces of the container body 200a.

The conduit 205 comprises anchoring means for the connection to a molecular oxygen transport cable of an oxygen concentrator.

The container body 200a further comprises a valve assembly 206 for regulating the flow of molecular oxygen passing through the inner cavity of the container body. The valve assembly 206 comprises a lever or a button 206a which adjusts the valve position to modulate the flow of oxygen. The lever or button is preferably located on the upper surface 201 of the spray gun, between the distal end 204 and the proximal end 203.

The spray gun comprises a tank 207, arranged on the upper surface 201 of the container body 200a, the tank 207 comprising an inner concavity 207a and a lid 207b. The inner concavity 207a comprises an opening in fluid communication with the inner cavity of the container body; such an inner concavity 207a, suitable for receiving a liquid, allows said liquid to flow towards the inner cavity of the container body.

Upon use, when the conduit 205 is anchored to the molecular oxygen transport cable of a pressure generator and the pressure generator is operated, the outflow of the liquid from the tank 207 to the container body 200a of the spray gun is induced by the formation of a pressure difference between the inner cavity of the container body and the inner concavity 207a of the tank.

The spray gun 200 is preferably made of a material suitable for withstanding sterilizations (such as autoclaving or the like), preferably made of steel.

The spray gun is preferably used for dispensing/nebulizing at the level of the external genitalia. In other words, the spray gun forms a vulvar dispensing device (or applicator).

EXAMPLES

The Applicant has reported examples of the invention below, for illustrative and nonlimiting purposes only.

Example 1 — Experimental test

1.1 Materials and methods

Five weekly oxygen therapy treatments were performed on 29 women diagnosed with recurrent candidiasis, for a total of five weeks.

The administration protocol comprises the following steps: i) dispensing oxygen and nebulized hyaluronic acid with a vulvar applicator for a period of five minutes (external application step of the association), ii) subsequently vaginal oxygen was applied for 10 minutes (internal application step of molecular oxygen (or gaseous oxygen)) and iii) dispensing oxygen and hyaluronic acid for an additional 5 minutes (internal application step).

The application of the above protocol (or treatment) consists of one session per week for a total five sessions. A post-treatment maintenance of one session per month was also recommended. Patients are taught behavioural norms for the home, necessary to contribute to pelvic health, including hygiene, good and healthy nutrition, indications for sexual intercourse.

For the treatment, an oxygen therapy device for gynaecological use was used which allows the topical administration of oxygen with a high degree of purity up to 93±3%, at a flow comprised between 1 and 6 L/min (litres/minute).

All the patients were evaluated for the presence of candidiasis at the end of the 5 treatment sessions, with follow-ups at 3 and 6 months after the end of treatment. The frequency of recurrent candidiasis in the previous 12 months was assessed for all patients.

1.2 Results

The 29 women had an average six-month relapse of 4.2±1.1.

At the end of the treatment (T5) all the detected infections had been resolved, demonstrating the effectiveness of the combined treatment of high concentration oxygen and hyaluronic acid (Figure 3).

Three months after the end of treatment, the percentage of patients who had had at least one recurrent episode was 6.8% (2/29), which increased to 55.2% (16/29) at six months.

The graph in figure 3 indicates the number of recurrent episodes for each patient in the six months before treatment and in the 2 follow-ups after treatment (3 and 6 months).

No treatment-associated side effects were reported by the patients.

Oxygen therapy combined with hyaluronic acid has proven to be a valid method for the treatment of symptoms associated with recurrent candidiasis, capable of resolving both the acute phase of the infection and of effectively preventing its recurrence. It is a totally painless therapy, with excellent compliance on the patients' part. It is a fast, non-invasive and repeatable treatment, with no side effects.