COMPOSITIONS FOR USE IN THE TREATMENT OR PREVENTION OF DYSBIOSIS

The present invention relates to compositions useful in the treatment or prevention of side effects of intestinal microbiota dysbiosis.

BACKGROUND OF THE INVENTION

When antibiotics are administered, either orally or parenterally, a significant fraction is not absorbed and reaches the gastro-intestinal tract. When those antibiotic residues reach the colon, they provoke a serious disruption of the intestinal microbiota: several bacterial populations are decimated whereas others (sometimes pathogenic and resistant to antibiotics) proliferate; this new state of the microbiota after the antibiotic-induced disruption is called dysbiosis. The intestinal microbiota balance is hence profoundly disrupted and may take weeks to months to fully recover, i.e. return to its original equilibrium. Other drugs are also known to disrupt the microbiota such as some anti-cancer chemotherapies.

Similarly to a damaged organ, a disrupted microbiota can no longer fulfil its physiological functions, leading to many adverse consequences such as altered immunity and immune response; colonization of the intestine by pathogenic bacteria such as Clostridioides difficile’, altered metabolism with increased risk of inflammation, metabolic syndrome or obesity; and colonization or emergence of antibiotic resistant bacteria or selection of genes of resistance to antibiotics, and their dissemination.

The medical community has well acknowledged today that preserving the microbiota balance and diversity during antibiotic treatments could prevent serious medical conditions such as C. difficile infections and graft-versus-host-disease. Maintaining a healthy intestinal microbiota could also prevent the selection and colonization of multi-resistant bacteria, and therefore limit the emergence and spread of antimicrobial resistance and prevent subsequent life-threatening infections. Finally, it is anticipated that maintaining the microbiota equilibrium is a driver for long-term health, and could favor better outcomes for cancer patients treated with cancer therapies, such as immune checkpoint inhibitors, and also patients with hematologic malignancies treated with hematopoietic stem cell transplants.

Compositions and methods were previously developed by the present Applicant to eliminate pharmaceutical agents that can induce intestinal dysbiosis, and to thus protect the intestinal microbiota. One approach to achieve this goal is to administer an adsorbent to eliminate such pharmaceutical agents, more specifically antibiotics, in the lower part of the intestine. More particularly, the adsorbent is released between the part of the intestine where such pharmaceutical agents are absorbed into the blood (e.g. duodenum and jejunum) and where their deleterious effect on the commensal bacteria occur (caecum and colon). These strategies were reported in WO2006122835, W02007132022 and WO2011104275.

The present invention provides further formulations useful to prevent intestinal dysbiosis.

SUMMARY OF THE INVENTION

In a first aspect the present invention relates to a solid dosage form comprising activated charcoal and a delivery system suitable to release the activated charcoal in a desired part of the intestine, wherein the activated charcoal is selected from activated charcoals having a R2B ratio higher or equal to 600 m ² /g: wherein:

S is the specific area of the activated charcoal expressed in m ² /g;

M is the total porous volume of the activated charcoal in cm ³ /g; and m is the microporous volume of the activated charcoal in cm ³ /g.

In a particular embodiment, the ratio of m over M is lower or equal to 50%.

In a further particular embodiment, S is higher or equal to 900 m ² /g.

In another particular embodiment, the activated charcoal is in admixture with at least one excipient. Representative suitable excipients include carrageenan. In another particular embodiment, the carrageenan is K-carrageenan.

In a further embodiment, the activated charcoal and the at least one excipient are in the form of a pellet. In yet another embodiment, the composition of activated charcoal and the at least one excipient comprises from 50 to 99% of activated charcoal, preferably from 75 to 95% of activated charcoal, by weight of the pellet.

In a particular embodiment, the dosage form comprises:

- a core comprising the activated charcoal; and

- a layer of an external coating formed around the core such as said core is released in the desired part of the intestine. In a particular embodiment, the core of the layered solid dosage form comprises the activated charcoal in admixture with K-carrageenan.

In a further embodiment, the desired part of the intestine into which the activated charcoal is intended to be released is the lower part of the intestine.

Moreover, according to another embodiment, the solid dosage form comprises an external coating comprising an anionic copolymer based on methyl acrylate, methyl methacrylate and methacrylic acid, such as poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) 7:3:1.

According to a second aspect, the solid dosage form according to the invention is for use in a method for the treatment of a side effect of an intestinal microbiota dysbiosis-inducing pharmaceutical agent.

DETAILED DESCRIPTION OF THE INVENTION

RZB ratio

The present invention relates to compositions useful in the treatment or prevention of side effects of intestinal microbiota dysbiosis. The invention is based on the identification of a criteria, a calculated ratio herein referred to as "R2B", useful to determine the optimal activated charcoal suitable for this purpose.

The R ₂ B ratio is calculated according to the following formula: wherein:

S is the specific area of the activated charcoal expressed in m ² /g;

M is the total porous volume of the activated charcoal in cm ³ /g; and m is the microporous volume of the activated charcoal in cm ³ /g.

According to IIIPAC nomenclature:

- micropores are pores with diameters below 2 nm;

- mesopores are pores with diameters ranging from 2 nm to 50 nm; and - macropores are pores with diameters above 50 nm

The total porous volume is the sum of the microporous, mesoporous and macroporous volumes.

Accordingly, the R ₂ B ratio is associated to the specific area available through the mesopores- macropores volume. Unexpectedly, the present inventors herein show that among different combinations of structural properties that were tested, this ratio is the best predictive parameter to select activated charcoals in order to have the best in vivo efficacy to adsorb antibiotics. Indeed, it was initially expected that activated charcoals with high specific area should lead to the best in vivo activity, not taking into account pore distribution. However, it was demonstrated that specific area only was absolutely not correlated to the ex vivo antibiotics adsorptions, and therefore could not predict the activated charcoal in vivo activity.

The specific area S and total porous volume M of an activated charcoal may be determined according to methods well known in the art. For example, one can use the Brunauer, Emmett and Teller (BET) method, which is based on the adsorption isotherm of nitrogen on the surface of a solid at the liquid nitrogen temperature: 77K. Details are available to those skilled in the art in the European Pharmacopeia, section 2.9.26: Specific surface area by gas adsorption (v. 04/2005:20926). In particular, one skilled in the art can use the method detailed in the experimental part of this application to determine S and M.

In a particular embodiment, S is determined as follows, using the BET method comprising the following steps:

- heating a known amount of an activated charcoal sample, for example a sample with a mass comprised between 70 and 150 mg, using a stream of nitrogen at 110 to 140°C during 20 to 60 minutes (for example 130°C during 30 minutes);

- vacuuming the sample with a pressure rate ranging from 200 to 300 mmHg/min during 10 to 15 minutes;

- cooling the sample until it reaches 15 to 30°C (room temperature);

- defining the adsorption isotherm on 56 to 70 points with a relative nitrogen pressure ranging from 0.0001 to 0.99;

- defining the desorption isotherms on 10 to 30 points, such as 22 points, with a relative nitrogen pressure ranging from 0.32 to 0.99; and

- determining the specific area of the activated charcoal sample with a suitable device, optionally performing a linearization of the BET isotherm curve on different points, such as from 10 to 30 points, in particular from 15 to 25 points, such as 19 points, with relative pressures of nitrogen (partial nitrogen pressure divided by saturation pressure) ranging from 0.01 to 0.15, such as from 0.01 to 0.10.

In a further particular embodiment, M is determined as follows, using the BET method comprising the following steps:

- heating a known amount of an activated charcoal sample, for example a sample with a mass comprised between 70 and 150 mg, using a stream of nitrogen at 110 to 140°C during 20 to 60 minutes (for example 130°C during 30 minutes);

- vacuuming the sample with a pressure rate ranging from 200 to 300 mmHg/min during 10 to 15 minutes;

- cooling the sample until it reaches 15 to 30°C (room temperature); and

- determining the total porous volume of the activated charcoal sample with a suitable device, by determination of nitrogen adsorption at its boiling temperature at a relative pressure of nitrogen of 0.960 to 0.999, for example 0.990.

In the context of the present invention, the volume of micropores with diameters between 1 and 2 nm is determined. The microporous volume m may be determined according to methods well known in the art. For example, one can use of density functional theory (DFT) method, such as the specific DFT method implemented in the experimental part below. One skilled in the art can also refer to Ravikovitch et al. https://doi.org/10.1016/S0001-8686(98)00047-5. This method is based on statistical mechanic calculations. In this model, pores are considered as independent to each other. Each pore size contributes proportionally to the adsorption isotherm depending on the sample surface it represents. Mathematically, this relation is expressed by:

With:

Q(p): the experimental quantity adsorbed at pressure p q(p,H): the quantity adsorbed per unit area at the same pressure p, in an ideal pore size of H f(H): the total area of pores of size H in the sample

As there is no analytic solution for this equation, the problem is best solved in a discrete form; the integral equation for pore size becomes a summation:

Based on this assumption, the experimental isotherm is compared to theoretical isotherm curves established according to the above equations and the volume of micropores in the sample is derived from the comparison.

As mentioned above, the R _2B ratio provided herewith is the best predictive parameter to select activated charcoals in order to have the best in vivo efficacy to adsorb antibiotics. In a particular embodiment, an activated charcoal having a R ₂ B ratio higher or equal to 600 m ² /g is selected. In another particular embodiment, an activated charcoal having a R ₂ B ratio higher or equal to 700 m ² /g is selected. In another particular embodiment, an activated charcoal having a R ₂ B ratio higher or equal to 800 m ² /g is selected. In a preferred embodiment, an activated charcoal having a R ₂ B ratio higher or equal to 850 m ² /g is selected. In another preferred embodiment, an activated charcoal having a R ₂ B ratio higher or equal to 1000 m ² /g is selected. Even more preferably an activated charcoal having a R ₂ B ratio higher or equal to 1100 m ² /g is selected.

Activated charcoal

Preferred activated charcoals for use in the invention are pharmaceutical grade activated charcoals, best suited for use in humans or animals for pharmaceutical or veterinary applications.

The activated charcoal may be of vegetal, mineral or synthetic origin, its surface being optionally modified by a physical or chemical treatment. In a particular embodiment, the activated charcoal is of vegetal origin. In a particular embodiment, the activated charcoal is derived from peat. In a particular embodiment, the activated charcoal is derived from coconut husks. In a particular embodiment, the activated charcoal is derived from different sources mixed together such as peat and coconut husks. In a particular embodiment, the activated charcoal is characterized by a European molasses number (of note the European molasses number is inversely related to the North American molasses number) which is preferably higher than 100, even more particularly greater than 200. In a particular embodiment, the activated charcoal has a phenazone number (measured according to the Ell Pharmacopeia) greater than 10 g/100 g, even more particularly greater than 20 g/100 g, even more particularly greater than 30 g/100 g, even more particularly greater than 40 g/100 g, even more particularly greater than 50 g/100 g, even more particularly greater than 60 g/100 g. In a particular embodiment, the activated charcoal is characterized by a density between 0.05 and 0.8, even more particularly between 0.1 and 0.6, even more particularly between 0.15 and 0.5, even more particularly between 0.2 and 0.4.

In a particular embodiment, the activated charcoal has a specific surface area higher or equal to 600 m ² /g, in particular higher or equal to 700 m ² /g, in particular higher or equal to 800 m ² /g, in particular higher or equal to 900 m ² /g , in particular higher or equal to 1000 m ² /g, in particular higher or equal to 1200 m ² /g, in particular higher or equal to 1400 m ² /g, in particular higher or equal to 1600 m ² /g, even more particularly higher or equal to 1800 m ² /g.

Solid dosage form

The activated charcoal is comprised in a solid dosage form further comprising a delivery system suitable to release the activated charcoal in a desired part of the intestine in particular in the lower part of the intestine, i.e. in the late ileum, the caecum or the colon.

In a particular embodiment, the activated charcoal is in admixture with at least one excipient. In yet another embodiment, said admixture of the activated charcoal and at least one excipient is in the form of a pellet, said pellet may serve as a core for the solid dosage form, said core being further coated with a layer of an external coating formed around the core such as said core is released in the desired part of the intestine. The external coating is in particular provided to minimize (preferably to totally prevent) the impact of the activated charcoal on the normal absorption process of a therapeutic agent (for example, an antibiotic) by the host organism when said therapeutic agent is administered orally along with the solid dosage form according to the invention. In addition or alternatively, the activated charcoal thus formulated is prevented from non-specifically adsorbing material present in the gastrointestinal tract all the way to the terminal part of the small intestine. This results in the release of a non-saturated activated charcoal, fully or almost fully efficient activated charcoal in the desired part of the intestine where its action is needed.

The amount of activated charcoal employed in the methods of the invention described below may vary depending upon the host/material being treated and the overall capacity, adsorption power and selectivity of the adsorbent. In a particular embodiment the amount of activated charcoal is an amount sufficient to prevent the deleterious impact of a substance, such as an antibiotic, on the intestinal microbiota known as “dysbiosis” or disruption of the gut microbiota. In a particular embodiment, the amount of activated charcoal is an amount sufficient to improve the efficacy of an immuno-oncology agent, or to improve the effectiveness of anticancer immunosurveillance in a subject. In a particular embodiment, the solid dosage form of the invention comprises from 50 to 99% of activated charcoal, by weight of the pellet comprising the activated charcoal in admixture with the at least one excipient(s). In a further embodiment, the composition comprises from 75 to 90% of activated charcoal, by weight of the pellet comprising the activated charcoal in admixture with the at least one excipient(s).

In a particular embodiment, excipients suitable to be mixed with activated charcoal, alone or in admixture, include, without limitation, carrageenan.

In one embodiment, the activated charcoal is mixed with carrageenan. Suitable carrageenan can be selected from kappa-, iota- and lambda-carrageenans, and mixtures thereof. In one aspect of this embodiment, the activated charcoal is in admixture with kappa-carrageenan. In a particular embodiment, the amount of carrageenan is between about 5% and about 25%, in particular between about 10% and about 20%, by weight of the mixture of the activated charcoal with the carrageenan. According to a specific embodiment of the invention, the amount of carrageenan is of about 15% by weight of said mixture. For example, the mixture may contain 85% of activated charcoal and 15% of carrageenan, by weight of the total mixture.

The core (or pellet) may be produced by any suitable means known to the skilled artisan. In particular, granulation techniques are adapted to produce said core. For example, the core may be obtained by mixing the adsorbent and the carrageenan in the ratio indicated above, adding a solvent such as water to proceed to wet granulation, followed by extrusion spheronization or one-pot pelletization. Any remaining water can be removed, for example, by drying using conventional techniques the resulting pellets.

Those skilled in the art will recognize that the core composition can further include conventional excipients such as antiadhesive agents, binders, fillers, diluents, flavours, colours, lubricants, glidants, preservatives, sorbents and sweeteners. The amounts of such excipients can vary, but will typically be in the range of 0.1 to 10% by weight of the pellet. Of course, the person skilled in the art will adapt these amounts so that the added excipient does not negatively impact on the advantageous properties of the mixture of the at least one excipient(s) described above in admixture with activated charcoal.

In one embodiment, the core, or pellet, of the invention has an average weight particle size in the range from 250 to 3000 pm, in particular 500 to 3000 pm. Several representative size ranges can be preferred. For example, the core size can be comprised between 500 and 1000 pm, or between 800 and 1600 pm. In the context of the present invention, the weight average particle size is determined by sieving different fractions in size, weighting the fractions and calculating the average particle size from the weights. The method is well known to a skilled person in the field of the invention.

The solid dosage form of the invention comprises a layer of an external coating formed around the core such that said core is released in the desired part of the intestine.

In a particular embodiment, the desired part of the intestine is the lower part of the intestine.

The external coating formed around the core is selected among coatings suitable to release the core in the desired part of the intestine.

Examples of suitable coatings include pH-dependent enterosoluble polymers, azopolymers, disulphide polymers, and polysaccharides, in particular amylose, pectin (e.g. pectin crosslinked with divalent cations such as calcium pectinate or zinc pectinate), chondroitin sulphate and guar gum. Representative pH-dependent enterosoluble polymers include cellulose acetate trimellitate (CAT), cellulose acetate phthalate (CAP), acrylic polymers, methacrylic polymers, anionic copolymers based on methylacrylate, methylmethacrylate and methacrylic acid, hydroxypropyl methylcellulose phthalate (HPMCP), hydroxypropylmethylcellulose acetate succinate (HPMCAS), methacrylic acid and ethyl acrylate copolymers, methacrylic acid and methyl methacrylate copolymers in a 1 : 1 molar ratio, methacrylic acid and methyl methacrylate copolymers in a 1 :2 molar ratio, polyvinyl acetate phthalate (PVAP) and shellac resins. Particularly preferred polymers include shellac, anionic copolymers based on methyl acrylate, methyl methacrylate and methacrylic acid, such as poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) in a 7:3:1 molar ratio, as well as methacrylic acid and methyl methacrylate copolymers in a 1 :2 molar ratio. Ideally, the polymer dissolves at a pH equal to 6.0 and above, preferably 6.5 and above. Suitable coatings may also be obtained by mixing the polymers and copolymers aforementioned. In another embodiment, suitable coatings are time-dependent coatings or based on time-dependent polymers such as mixture of ethylcellulose polymers with alginate sodium.

In a particular embodiment, the formulation comprises a further intermediate coating located between the core and the external pH-dependent layer. The intermediate coating can be formed from a variety of polymers, including pH-dependent polymers, pH-independent water soluble polymers, pH-independent insoluble polymers, and mixtures thereof. Examples of such pH-dependent polymers include shellac type polymers, anionic copolymers based on methylacrylate, methylmethacrylate and methacrylic acid, methacrylic acid and ethyl acrylate copolymers, hydroxypropyl methylcellulose phthalate (HPMCP), and hydroxypropylmethylcellulose acetate succinate (HPMCAS). Examples of pH-independent water soluble polymers include PVP or high molecular weight cellulose polymers such as hydroxypropylmethylcellulose (HPMC) or hydroxypropylcellulose (HPC). Examples of pH- independent insoluble polymers include ethylcellulose polymers or ethyl acrylate and methyl methacrylate copolymers.

In a particular embodiment, the invention uses a formulation comprising:

- a core comprising a pellet according to the invention,

- an intermediate coating selected in the group consisting of HPMC, ethylcellulose and a mixture of methacrylic acid and ethyl acrylate copolymer such as Eudragit® L30D-55, and ethyl acrylate and methyl methacrylate copolymer such as Eudragit® NE30D (for example in a mixture weight ratio of 1 :9 to 9:1 , preferably of 2:8 to 3:7), and

- an external layer of an anionic copolymer based on methyl acrylate, methyl methacrylate and methacrylic acid, such as poly(methyl acrylate-co-methyl methacrylate-co- methacrylic acid) 7:3:1 , e.g. Eudragit® FS30D.

In a specific embodiment, the formulation comprises a core, comprising a pellet according to the invention which comprises about 85% activated charcoal, and a coating with an anionic copolymer based on methyl acrylate, methyl methacrylate and methacrylic acid (such as poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) 7:3:1 , e.g. Eudragit® FS30D, Evonik, Darmstadt, Germany) or a mixture of methacrylic acid and ethyl acrylate copolymer (such as Eudragit® L30D55, Evonik, Darmstadt, Germany).

In a particular embodiment, the external coating comprises an anionic copolymer based on methyl acrylate, methyl methacrylate and methacrylic acid, such as poly(methyl acrylate-co- methyl methacrylate-co-methacrylic acid) 7:3:1.

In a particular embodiment, the solid dosage form of the invention comprises: a) a core composition in the form of a pellet comprising:

- from 50 to 99% of activated charcoal,

- from 1 to 50% of carrageenan; by weight of the core composition; and b) an external coating comprising poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) 7:3:1.

In a particular embodiment, the solid dosage form of the invention comprises: a) a core composition in the form of a pellet comprising: - from 50 to 99% of activated charcoal,

- from 1 to 50% of kappa-carrageenan; by weight of the core composition; and b) an external coating comprising poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) 7:3:1.

In a further particular embodiment, the solid dosage form of the invention comprises: a) a core composition in the form of a pellet comprising:

- from 75 to 90% of activated charcoal,

- from 10 to 25% of carrageenan; by weight of the core composition; and b) an external coating comprising poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) 7:3:1.

In yet another embodiment, the solid dosage form of the invention comprises: a) a core composition in the form of a pellet comprising:

- about 85 % of activated charcoal,

- about 15% of carrageenan; by weight of the core composition; and b) an external coating comprising poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) 7:3:1.

In yet another embodiment, the solid dosage form of the invention comprises: a) a core composition in the form of a pellet comprising:

- about 85 % of activated charcoal,

- about 15% of kappa-carrageenan; by weight of the core composition; and b) an external coating comprising poly(methyl acrylate-co-methyl methacrylate-co-methacrylic acid) 7:3:1.

Uses of the solid dosaqe forms of the invention

The solid dosage forms according to the invention can be used to treat conditions and disorders for which intestinal delivery of adsorbents is suitable.

Accordingly, the invention also relates to a solid dosage form as described above, for use as a medicament. In a particular embodiment, the solid dosage form of the invention is for use in a method for the treatment of a side effect of a dysbiosis-inducing pharmaceutical agent.

The solid dosage form according to the invention can be used to adsorb and therefore remove from the intestine any drug, metabolite or prodrug thereof, or toxin. This may be done after oral or parenteral administration of an active drug, which could be useful for limiting or decreasing adverse effects in the subject being treated the drug, metabolite or prodrug thereof reaches the lower intestine and/or colon.

As such, the present invention relates to the solid dosage form as described above, for use in a method for eliminating drugs in the intestinal tract before they reach the colon or as they reach the colon, preferably before they reach the caecum or as they reach the caecum and proximal colon.

The invention further provides a method for eliminating drugs in the intestinal tract before they reach the colon or as they reach the colon, preferably before they reach the caecum or as they reach the caecum and proximal colon, comprising administering to a patient in need thereof a formulation according to the invention.

Furthermore, the invention provides a formulation as described above, for use in a method for reducing or eliminating the side effect(s) of a drug in the intestinal tract, wherein the formulation eliminates the drug before it reaches the colon or as it reaches the colon, preferably before it reaches the caecum or as it reaches the caecum and proximal colon.

Furthermore, the invention provides a formulation as described above, for use in a method for reducing or eliminating the deleterious effect(s) of toxins in the intestinal tract, wherein the formulation eliminates the toxin in the colon.

The terms "substance", "drug", "therapeutic agent", "pharmaceutical agent", and terms derived therefrom, are herein used interchangeably and refer to a compound or composition that provides a desired biological or pharmacological effect when administered to a human or animal.

Treatment or prevention of conditions related to antibiotic administration The dysbiosis-inducing pharmaceutical agent may be an antibiotic, and the solid dosage form of the invention being used to treat a side effect of such an antibiotic.

Such side effects include, without limitation, the development of antibiotic resistance through the emergence of antibiotic-resistant bacteria, the dissemination of antibiotic-resistant bacteria, the development of an infection by Cl ostridi aides difficile or other pathogenic bacteria, a decrease in the efficacy of an anticancer agent in a subject in need thereof, a risk of developing or aggravating graft-versus-host disease in a subject and a risk of increasing the post-transplant mortality in subjects receiving or having received a hematopoietic stem cell transplant or the impaired immunity of the patient which can lead to immunological disorders such as auto-immune diseases, asthma, diabetes, obesity or other.

The adsorbent will adsorb residual antibiotics, and the solid dosage form according to the invention can be administered in a therapeutically effective amount to a patient who has been, is being, or will be administered an antibiotic. Any antibiotic that can be adsorbed into/onto the adsorbent can be inactivated and has no antibiotic activity once fully adsorbed.

The term “antibiotic” designates any compound that is active against bacteria. Antibiotics that may be eliminated thanks to the invention include but are not limited to:

- beta-lactams including:

- penicillins (such as penicillin G, penicillin V, ampicillin, amoxicillin, bacampicillin, carbenicillin, carbenicillin indanyl, ticarcillin, azlocillin, mezlocillin, piperacillin, and the like),

- penicillinase-resistant penicillins (such as methicillin, oxacillin, cioxacillin, dicloxacillin, nafcillin and the like),

- cephalosporins, such as: first generation cephalosporins (such as cefadroxil, cephalexin, cephradine, cephalothin, cephapirin, cefazolin, and the like) ; second generation cephalosporins (such as cefaclor, cefamandole, cefonicid, cefoxitin, cefotetan, cefuroxime, cefuroxime axetil, cefinetazole, cefprozil, loracarbef, ceforanide, and the like) ; third generation cephalosporins (such as cefepime, cefoperazone, cefotaxime, ceftizoxime, ceftriaxone, ceftazidime, cefixime, cefpodoxime, ceftibuten, and the like) ; fourth generation cephalosporins (such as cefclidine, cefepime, cefozopran, cefpirome, cefquionome and the like) ; fifth and further generation cephalosporins (such as ceftobiprole, ceftaroline, ceftolozane and the like),

- carbapenems (such as imipenem, meropenem, ertapenem, doripenem and the like)

- monobactams (such as aztreonam, and the like), - quinolones (such as nalidixic acid) and fluoroquinolones (such as cinoxacin, ciprofloxacin, moxifloxacin, levofloxacin, ofloxacin, gatifloxacin, gelifloxacin, norfloxacin and the like),

- sulfonamides (e.g., sulfanilamide, sulfadiazine, sulfamethoxazole, sulfisoxazole, sulfacetamide, sulfamethoxydiazine and the like),

- aminoglycosides (e.g., streptomycin, gentamicin, tobramycin, amikacin, netilmicin, kanamycin, neomycins B, C and E), spectinomycin, puromycin, gentamicin, and the like),

- tetracyclines (such as tetracycline, chlortetracycline, oxytetracycline, methacycline, doxycycline, minocycline, tigecycline, eravacycline and the like),

- macrolides (such as erythromycin, azithromycin, clarithromycin, fidaxomicin, telithromycin, josamycin, oleandomycin, spiramycin, tylosin, roxithromycin, cethromycin, solithromycin, and the like),

- glycopeptides (such as vancomycin, oritavancin, telavancin, teicoplanin, dalbavancin, ramoplanin and the like),

- oxazolidinones (such as linezolid, posizolid, tedizolid, radezolid, cycloserine and the like),

- phenicols (such a chloramphenicol, tiamphenicol and the like),

- lincosamides (such as clindamycin, lincomycin and the like),

- streptogramins (such as pristinamycin, quinupristin/dalfopristin, virginiamycin and the like)

- polymyxins (such as polymyxin A, B, C, D, E1 (colistin A), or E2, colistin B or C, and the like),

- diaminopyrimidines (such as trimethoprim, often used in conjunction with sulfamethoxazole, pyrazinamide, and the like),

- sulfones (such as dapsone, sulfoxone sodium, and the like),

- para-aminobenzoic acid,

- bacitracin,

- isoniazid,

- rifamycins (such as rifampicin, rifabutin, rifapentine, rifalasil, rimamixin, and the like)

- ethambutol,

- ethionamide,

- capreomycin,

- clofazimine, and

- any other antibacterial agent.

The term “antibiotic” also covers combinations of antibiotics.

The invention thus also relates to a solid dosage form as described above, for use in a method for eliminating residual antibiotics in the intestinal tract, preferably before they reach the colon or as they reach the colon. More preferably, the solid dosage form is used in a method for eliminating residual antibiotics in the intestinal tract, preferably before they reach the caecum or as they reach the caecum and proximal colon. According to the invention, the adsorbent is preferably delivered between the part of the intestine where the antibiotics are absorbed (duodenum and jejunum) and where their deleterious effect on the commensal bacteria occur (caecum and colon). The invention further relates to a method for eliminating residual antibiotics in the intestinal tract, preferably before they reach the colon or as they reach the colon, most preferably before they reach the caecum or as they reach the caecum and proximal colon comprising administering to a subject in need thereof an effective amount of the solid dosage form of the invention.

The invention further relates to a solid dosage form as described above, for use in a method for eliminating the adverse effects of antibiotic agents in the intestinal tract, in particular for eliminating the development of antibiotic resistance, antibiotic treatment-associated development of C. difficile (or other pathogenic bacteria), antibiotic treatment-associated fungal infections or antibiotic treatment-associated diarrhea. The invention further relates to a method for eliminating the adverse effects of antibiotic agents in the intestinal tract, comprising administering to a subject in need thereof an effective amount of the solid dosage form of the invention.

In another embodiment, the present invention provides a kit, comprising an antibiotic, and a solid dosage form of the invention. The kit may be a kit-of-parts, for simultaneous, separate or sequential use in the treatment of an infection against which the antibiotic is suitable.

Cancer treatment

The present invention relates to a solid dosage form as provided above, for use in a method for improving the therapeutic efficacy of an anticancer agent, such as an immuno-oncology agent. The invention also relates to a solid dosage form as provided above, for use in a method for treating or preventing cancer, in combination with an anticancer agent, such as an immuno- oncology agent. The invention further relates to a solid dosage form as provided above, for use in a method for treating or preventing cancer, in combination with an anticancer agent, such as an immuno-oncology agent, thereby improving the efficacy of said anticancer agent. The invention also relates to a solid dosage form as provided above, for use in a method for treating or preventing cancer, in combination with an anticancer agent, such as an immuno- oncology agent, thereby preserving the efficacy of said anticancer agent. The invention further relates to a solid dosage form as provided above, for use in a method for treating or preventing cancer, in combination with an anticancer agent, such as an immuno-oncology agent, thereby potentiating the efficacy of said anticancer agent. The solid dosage form may be administered at any point in the therapy, e.g. before, during and/or after the anticancer agent, such as an immuno-oncology agent. In particular, the solid dosage form may be administered as soon as the patient is diagnosed with a malignancy, even if the intent to administer an anticancer agent only constitutes a remote possibility.

Anticancer agents, also sometimes referred to as antineoplastic agents, are substances that act against cancer in a mammal, such as a human being. The term “anticancer agent” includes, without limitation, chemicals and biological agents that affect directly a cancer cell, or indirectly such as by affecting the vascularisation of the cancer cell. For example, anticancer agents include, without limitation, chemotherapeutic molecules such as cytostatic agents, cytotoxic agents and anti-angiogenesis agents, anticancer antibodies targeting cancer cells, anticancer peptides and anticancer viruses. Illustrative anticancer agents include, without limitation:

- tubulin poisons, taxanes, e.g. docetaxel, paclitaxel,

- platinum compounds, e.g. cisplatin, carboplatin, oxaliplatin,

- agents interfering with DNA replication such as DNA intercalating agents, for example anthracyclines,

- topoisomerase inhibitors such as etoposide,

- antimetabolites, e.g. methotrexate, cytarabine (ara-C), gemcitabine, 5-Fluorouracil,

- alkylators, e.g. mechlorethamine, melphalan, carmustine, ifosfamide, or cyclophosphamide,

- targeted agents, such as enzyme inhibitor, in particular kinase inhibitors, e.g. erlotinib, sorafenib, imatinib, or proteasome inhibitors such as bortezomib, Carfizomib, Ixazomib,

- monoclonal antibodies targeting the extracellular region of a growth factor receptor, such as trastuzumab, bevacizumab and cetuximab,

- immuno-oncology agents, and

- combinations thereof.

Anthracyclines include, without limitation, doxorubicin and daunorubicin.

Topoisomerase inhibitors further include, without limitation, camptothecin, irinotecan, topotecan, and derivatives thereof.

Anti metabolites further include, without limitation, capecitabine and pemetrexed.

In a particular embodiment, the anticancer agent is an immuno-oncology agent. Immuno- oncology agents (also known as immuno-targeted agents) act against tumors, at least in part, by involving the immune system, or by an immune system-related mode of action. An immuno- oncology may more particularly act by modulating the action of immune cells. Examples of immuno-oncology agents comprise agents that modulate immune checkpoints such as 2B4, 4-1 BB (CD137), AaR, B7-H3, B7-H4, BAFFR, BTLA, CD2, CD7, CD27, CD28, CD30, CD40, CD80, CD83 ligand, CD86, CD160, CD200, CDS, CEACAM, CTLA-4, GITR, HVEM, ICAM-1 , KIR, LAG-3, LAIR1 , LFA-1 (CD 11 a/CD 18), LIGHT, NKG2C, NKp80, 0X40, PD-1 , PD-L1 , PD-L2, SLAMF7, TGFRp, TIGIT, Tim3 and VISTA.

Immuno-oncology agents may be in the form of antibodies, peptides, small molecules or viruses. In a particular embodiment, the immuno-oncology agent is an antibody against PD-1 , PD-L1 or PD-L2.

In a particular embodiment, the immuno-oncology agent is an inhibitor of arginase, CTLA-4, indoleamine 2,3-dioxygenase, and/or PD-1/PD-L1. In certain embodiments, the immuno- oncology agent is abagovomab, adecatumumab, afutuzumab, alemtuzumab, anatumomab mafenatox, apolizumab, blinatumomab, BMS-936559, catumaxomab, durvalumab, epacadostat, epratuzumab, indoximod, inotuzumab, ozogamicin, intelumumab, ipilimumab, isatuximab, lambrolizumab, MED 14736, MPDL3280A, nivolumab, obinutuzumab, ocaratuzumab, ofatumumab, olatatumab, pembrolizumab, pidilizumab, rituximab, ticilimumab, samalizumab, or tremelimumab.

More generally, an immuno-oncology agent may be any agent that may be used in the treatment of malignant diseases and that acts, at least in part, by involving the immune system, or has an immune system-related mode of action. For example, the immuno-oncology agent may be selected from, without limitation:

- an immune checkpoint inhibitor such as a PD-1 inhibitor, e.g. nivolumab or pembrolizumab;

- an immune checkpoint inhibitor such as a PDL-1 inhibitor, e.g. atezolizumab, avelumab, or durvalumab; or a CTLA-4 inhibitor, e.g. ipilimumab,

- a cancer vaccine, e.g. sipuleucel-T;

- an immunomodulator such as thalidomide, lenalidomide, pomalidomide,

- a non-specific immunotherapy, e.g. interferons, or interleukins; and

- a chimeric antigen receptor (CAR)-T cell therapy, e.g. tisagenlecleucel, or axicabtagene ciloleucel, and

- combinations thereof.

In a particular embodiment, the anticancer agent is an anti-PD-1 antibody. In a further particular embodiment, the anti-PD-1 antibody is selected from nivolumab and pembrolizumab. In a particular embodiment of the invention, the anticancer agent is selected from Afatinib, Aflibercept, Alemtuzumab, Alitretinoin, Altretamine, Anagrelide, Arsenic trioxide, Asparaginase, Atezolizumab, Avelumab, Axitinib, Azacitidine, Bendamustine, Bevacizumab, Bexarotene, Bleomycin, Bortezomib, Bosutinib, Busulfan, Cabazitaxel, Capecitabine, Carboplatin, Carmofur, Carmustine, Cetuximab, Chlorambucil, Chlormethine, Cisplatin, Cladribine, Clofarabine, Crizotinib, Cyclophosphamide, Cytarabine, Dacarbazine, Dactinomycin, Dasatinib, Daunorubicin, Decitabine, Denileukin diftitox, Denosumab, Docetaxel, Doxorubicin, Durvalumab, Epirubicin, Erlotinib, Estramustine, Etoposide, Everolimus, Floxuridine, Fludarabine, Fluorouracil, Fotemustine, Gefitinib, Gemcitabine, Gemtuzumab ozogamicin, Hydroxycarbamide, Ibritumomab tiuxetan, Idarubicin, Ifosfamide, Imatinib, Ipilimumab, Irinotecan, Isotretinoin, Ixabepilone, Lapatinib, Lenalidomide, Lomustine, Melphalan, Mercaptopurine, Methotrexate, Mitomycin, Mitoxantrone, Nedaplatin, Nelarabine, Nilotinib, Nivolumab, Ofatumumab, Oxaliplatin, Paclitaxel, Panitumumab, Panobinostat, Pazopanib, Pembrolizumab, Pemetrexed, Pentostatin, Pertuzumab, Pomalidomide, Ponatinib, Procarbazine, Raltitrexed, Regorafenib, Rituximab, Romidepsin, Ruxolitinib, Sorafenib, Streptozotocin, Sunitinib, Tamibarotene, Tegafur, Temozolomide, Temsirolimus, Teniposide, Thalidomide, Tioguanine, Topotecan, Tositumomab, Trastuzumab, Tretinoin, Valproate, Valrubicin, Vandetanib, Vemurafenib, Vinblastine, Vincristine, Vindesine, Vinflunine, Vinorelbine and Vorinostat.

The solid dosage form of the invention and the anticancer agent may be used to treat or prevent a cancer or multiple cancers in a subject. In certain embodiments, the cancer may be one or a variant of a cancer selected from Acute Lymphoblastic Leukemia (ALL), Acute Myeloid Leukemia (AML), Adrenocortical Carcinoma, Anal Cancer, Appendix Cancer, Atypical Teratoid/Rhabdoid Tumor, Basal Cell Carcinoma, Bile Duct Cancer, Bladder Cancer, Bone Cancer, Brain Tumor, Astrocytoma, Brain and Spinal Cord Tumor, Brain Stem Glioma, Central Nervous System Atypical Teratoid/Rhabdoid Tumor, Central Nervous System Embryonal Tumors, Breast Cancer, Bronchial Tumors, Burkitt Lymphoma, Carcinoid Tumor, Carcinoma of Unknown Primary, Central Nervous System Cancer, Cervical Cancer, Childhood Cancers, Chordoma, Chronic Lymphocytic Leukemia (CLL), Chronic Myelogenous Leukemia (CML), Chronic Myeloproliferative Disorders, Colon Cancer, Colorectal Cancer, Craniopharyngioma, Cutaneous T-Cell Lymphoma Ductal Carcinoma In Situ (DCIS), Embryonal Tumors, Endometrial Cancer, Ependymoblastoma, Ependymoma, Esophageal Cancer, Esthesioneuroblastoma, Ewing Sarcoma, Extracranial Germ Cell Tumor, Extragonadal Germ Cell Tumor, Extrahepatic Bile Duct Cancer, Eye Cancer, Fibrous Histiocytoma of Bone, Gallbladder Cancer, Gastric Cancer, Gastrointestinal Carcinoid Tumor, Gastrointestinal Stromal Tumors (GIST), Germ Cell Tumor, Extracranial Germ Cell Tumor, Extragonadal Germ Cell Tumor, Ovarian Germ Cell Tumor, Gestational Trophoblastic Tumor, Glioma, Hairy Cell Leukemia, Head and Neck Cancer, Heart Cancer, Hepatocellular Cancer, Histiocytosis, Langerhans Cell Cancer, Hodgkin Lymphoma, Hypopharyngeal Cancer, Intraocular Melanoma, Islet Cell Tumors, Kaposi Sarcoma, Kidney Cancer, Langerhans Cell Histiocytosis, Laryngeal Cancer, Leukemia, Lip and Oral Cavity Cancer, Liver Cancer, Lobular Carcinoma In Situ (LCIS), Lung Cancer, Lymphoma, AIDS-Related Lymphoma, Macroglobulinemia, Male Breast Cancer, Medulloblastoma, Medulloepithelioma, Melanoma, Merkel Cell Carcinoma, Malignant Mesothelioma, Metastatic Squamous Neck Cancer with Occult Primary, Midline Tract Carcinoma Involving NUT Gene, Mouth Cancer, Multiple Endocrine Neoplasia Syndrome, Multiple Myeloma/Plasma Cell Neoplasm, Mycosis Fungoides, Myelodysplastic Syndrome, Myelodysplastic/Myeloproliferative Neoplasm, Chronic Myelogenous Leukemia (CML), Acute Myeloid Leukemia (AML), Myeloma, Multiple Myeloma, Chronic Myeloproliferative Disorder, Nasal Cavity Cancer, Paranasal Sinus Cancer, Nasopharyngeal Cancer, Neuroblastoma, Non-Hodgkin Lymphoma, Non-Small Cell Lung Cancer, Oral Cancer, Oral Cavity Cancer, Lip Cancer, Oropharyngeal Cancer, Osteosarcoma, Ovarian Cancer, Pancreatic Cancer, Papillomatosis, Paraganglioma, Paranasal Sinus Cancer, Nasal Cavity Cancer, Parathyroid Cancer, Penile Cancer, Pharyngeal Cancer, Pheochromocytoma, Pineal Parenchymal Tumors of Intermediate Differentiation, Pineoblastoma, Pituitary Tumor, Plasma Cell Neoplasm, Pleuropulmonary Blastoma, Breast Cancer, Primary Central Nervous System (CNS) Lymphoma, Prostate Cancer, Rectal Cancer, Renal Cell Cancer, Clear cell renal cell carcinoma, Renal Pelvis Cancer, Ureter Cancer, Transitional Cell Cancer, Retinoblastoma, Rhabdomyosarcoma, Salivary Gland Cancer, Sarcoma, Sezary Syndrome, Skin Cancer, Small Cell Lung Cancer, Small Intestine Cancer, Soft Tissue Sarcoma, Squamous Cell Carcinoma, Squamous Neck Cancer with Occult Primary (e.g., Metastatic), Squamous Cell Carcinoma of the Head and Neck (HNSCC), Stomach Cancer, Supratentorial Primitive Neuroectodermal Tumors, T- Cell Lymphoma, Testicular Cancer, Throat Cancer, Thymoma, Thymic Carcinoma, Thyroid Cancer, Transitional Cell Cancer of the Renal Pelvis and Ureter, Triple Negative Breast Cancer (T BC), Gestational Trophoblastic Tumor, Unknown Primary, Unusual Cancer of Childhood, Urethral Cancer, Uterine Cancer, Uterine Sarcoma, Waldenstrom Macroglobulinemia, and Wilms Tumor.

In particular, the cancer may be selected from:

- tumours of epithelial origin affecting organs such as breast (breast adenocarcinoma), skin (melanoma), lung (non-small cell lung cancer and small cell lung cancer), kidney (renal cell carcinoma), pancreas (pancreatic carcinoma), bladder,

- digestive tumours such as gastro-oesohagial adenocarcinomas,

- head and neck cancers (in particular squamous tumors), - squamous lung tumours,

- malignancies affecting blood of immune cells such as multiple myeloma, lymphoma (Hodgkin’s and non-Hodgkin’s of all types), leukemia among which lymphocytic leukemia (such as acute lymphoblastic leukemia (ALL), or chronic lymphocytic leukemia, (CLL)), myologenous leukemia (such as acute myolegenous leukemia (AML), and crhonic myelogenous leukemia (CML)), hairy cell leukemia, T-cell prolymphocytic leukemia, large granular lymphocytic leukemia, adut T-cell leukemia, adult T-cell lymphoma/leukemia.

In a particular embodiment, the cancer is selected from a cancer of the lung, a melanoma, a cancer of the pancreas, a cancer of the kidneys, refractory leukemia and lymphoma.

In certain embodiments, the method of the invention may further comprise administering one or more additional therapeutic agents conjointly with the anticancer agent. Representative therapeutic agents that may be conjointly administered with the anticancer agent include, without limitation: aminoglutethimide, amsacrine, anastrozole, asparaginase, AZD5363, Bacillus Calmette-Guerin vaccine (beg), bicalutamide, bleomycin, bortezomib, buserelin, busulfan, campothecin, capecitabine, carboplatin, carfilzomib, carmustine, chlorambucil, chloroquine, cisplatin, cladribine, clodronate, cobimetinib, colchicine, cyclophosphamide, cyproterone, cytarabine, dacarbazine, dactinomycin, daunorubicin, demethoxyviridin, dexamethasone, dichloroacetate, dienestrol, diethylstilbestrol, docetaxel, doxorubicin, epirubicin, erlotinib, estradiol, estramustine, etoposide, everolimus, exemestane, filgrastim, fludarabine, fludrocortisone, fluorouracil, fluoxymesterone, flutamide, gemcitabine, genistein, goserelin, hydroxyurea, idarubicin, ifosfamide, imatinib, interferon, irinotecan, lenalidomide, letrozole, leucovorin, leuprolide, levamisole, lomustine, lonidamine, mechlorethamine, medroxyprogesterone, megestrol, melphalan, mercaptopurine, mesna, metformin, methotrexate, miltefosine, mitomycin, mitotane, mitoxantrone, MK-2206, nilutamide, nocodazole, octreotide, olaparib, oxaliplatin, paclitaxel, pamidronate, pazopanib, pentostatin, perifosine, plicamycin, pomalidomide, porfimer, procarbazine, raltitrexed, rituximab, rucaparib, selumetinib, sorafenib, streptozocin, sunitinib, suramin, talazoparib, tamoxifen, temozolomide, temsirolimus, teniposide, testosterone, thalidomide, thioguanine, thiotepa, titanocene di chloride, topotecan, trametinib, trastuzumab, tretinoin, veliparib, vinblastine, vincristine, vindesine, and vinorelbine. Other representative therapeutic agents that may be conjointly administered with the anticancer agent include, without limitation, pemetrexed.

In a particular embodiment, anticancer therapy is a combination therapy with an immunooncology agent and one targeted therapy. For example, the patient may be administered with an immuno-oncology agent and at least one other anticancer agent selected from BRAF and MEK inhibitors.

In a particular embodiment, anticancer therapy is a combination therapy with an immuno- oncology agent and at least one other anticancer agent. For example, the patient may be administered with an immuno-oncology agent and at least one other anticancer agent selected from platinum salts (such as cisplatin, carboplatin and the like), pemetrexed and etoposide.

For example, the at least one other anticancer agent may be:

- pemetrexed,

- pemetrexed and platinum salts,

- etoposide, or

- etoposide and platinum salts.

In another embodiment, the present invention provides a kit, comprising an anticancer agent, and a solid dosage form of the invention. In certain embodiments, the kit may be for use in treating a condition or disease as described herein.

The present invention provides a method of treating or preventing cancer, comprising conjointly administering a solid dosage form according to the invention and an anticancer agent. Thanks to the invention, administering the anticancer agent and the solid dosage form according to the invention provides improved efficacy relative to individual administration of the anticancer agent.

In certain embodiments, the anticancer agent is administered within about 5 minutes to within about 7 hours after the solid dosage form according to the invention. In a particular embodiment, the solid dosage form according to the invention is administered multiple times before the anticancer agent is administered in order to ensure that the anticancer immunosurveillance system of the patient is improved. For example, the solid dosage form according to the invention may be administered daily, one or several times a day, for several days. For example, the solid dosage form according to the invention may be administered daily, one or several times a day, at least 2, at least 3, at least 4, at least 5, at least 6 or at least 7 days before administration of the anticancer agent.

In certain embodiments, the solid dosage form according to the invention is administered once daily, or multiple times daily such as twice daily or thrice daily, during the whole time of anticancer treatment and maintained between the different cycles of anticancer treatment. In certain aspects, the solid dosage form according to the invention is for use in a subject who has a cancer and who is administered, will be administered or has been administered with a substance, besides the anticancer agent, that may disturb the gut microbiota of said patient. Thanks to the invention, the deleterious impact of such substances may be prevented and thus the efficacy of the anticancer agent may be improved. Therefore, the invention relates to a method for mitigating the deleterious effects a substance may have on the gut microbiota of a subject suffering from cancer, said subject being the recipient of an anticancer agent therapy, comprising administering to said subject an effective amount of a solid dosage form according to the invention.

In certain embodiments, the substance is a pharmaceutical substance administered to treat a pathological condition in the patient. Indeed, certain pharmaceutical substances may be administered in order to treat a disease, but may have a deleterious effect on the gut microbiota when they reach the lower part of the intestine. The subject is still to receive the pharmaceutical substance for benefiting its desired effects but, on the other hand, solutions to avoid its secondary effects should be provided. Illustrative substances having this behaviour include antibiotics. Antibiotics may be administered to a subject in order to treat a bacterial infection. However, since antibiotics are, by design, able to affect bacterial growth or survival, they threaten the gut microbiota balance and may induce dysbiosis when they reach the lower part of the intestine. This induced dysbiosis may in turn result in a decrease in the efficacy of an anticancer drug administered to the subject. Other illustrative pharmaceutical substances that may induce dysbiosis (also referred to as “dysbiosis-inducing pharmaceutical substances”) include, without limitation: chemotherapy agents, such as taxanes (e.g. docetaxel, paclitaxel), a nth racy clines (e.g. doxorubicin), topoisomerase inhibitors (e.g. etoposide, irinotecan), antimetabolites (e.g. methotrexate, cytarabine, 5-fluorouracil, gemcitabine, pemetrexed), alkylating agents (e.g. melphalan), kinase inhibitors (e.g. erlotinib), antifungal agents, such as voroconazole, ambisome, posoconazole, antiviral agents, such as acyclovir, methisazone, anti-inflammatory agents, such as aspirin, ibuprofen; and proton pump inhibitors such as omeprazole, pantoprazole, esomeprazole.

Accordingly, in another aspect of the invention the solid dosage form of the invention is administered to a subject who has a cancer and who is treated, will be treated or has been administered with a dysbiosis-inducing pharmaceutical substance, such as an antibiotic. The solid dosage form of the invention may be administered to the subject even long before initial administration of the anticancer agent. For example, the subject may have been diagnosed with a malignancy but the treatment could not begin before several days, weeks, months or years. In this case, should the subject suffer, between these events, from a disease that would need a treatment with a dysbiosis-inducing pharmaceutical agent, such as an antibiotic, it would be advantageous to prevent gut microbiota dysbiosis by administering a solid dosage form of the invention as provided herein. Likewise, the solid dosage form of the invention may be administered to the subject even long before the start or after the end of administration of the anticancer agent. Firstly, it may unfortunately be that the subject’s cancer could relapse. In this case, halting the systematic administration of a solid dosage form of the invention when the subject receives a dysbiosis-inducing pharmaceutical substance, such as an antibiotic, could severely impair the efficacy of a future therapy with the same or another anticancer agent. Secondly, some therapies, such as gene therapies, may be efficient several years after administration, as long as the therapeutic gene is expressed. In that case, the administration of the solid dosage form of the invention would be beneficial for improving this kind of long-lasting anticancer therapies. Of course, the solid dosage form of the invention is preferably administered during the whole course of the anticancer agent therapy, when the subject is to receive a therapy with a dysbiosis-inducing pharmaceutical substance, such as an antibiotic.

In a particular embodiment, the invention relates to a solid dosage form of the invention for improving the efficacy of an anticancer agent in a subject in need of such an anticancer agent, wherein the subject is also administered with a dysbiosis-inducing pharmaceutical substance, such as an antibiotic.

The invention also relates to a solid dosage form of the invention for use in the prevention of the decrease of efficacy of an anticancer agent in a subject when said subject is administered with a dysbiosis-inducing pharmaceutical substance, such as an antibiotic.

The invention also relates to a solid dosage form of the invention for use to maintain the efficacy of an anticancer agent in a subject when said subject is administered with a dysbiosisinducing pharmaceutical substance, such as an antibiotic.

The invention further relates to a solid dosage form of the invention for use along with a dysbiosis-inducing pharmaceutical substance, such as an antibiotic, in a subject in need of an anticancer agent therapy. The invention further relates to a solid dosage form of the invention for use in combination with a dysbiosis-inducing pharmaceutical substance, such as an antibiotic, in a method for the treatment or prevention of a disease that may be treated or prevented with said dysbiosisinducing pharmaceutical substance, wherein the subject in need of said treatment is also in need of an anticancer therapy.

The invention further relates to a solid dosage form of the invention for use in a subject in need of an anticancer agent, for preventing the impact of a dysbiosis-inducing pharmaceutical substance, such as an antibiotic, on the efficacy of said anticancer agent.

The invention further relates to a solid dosage form of the invention for use in a subject in need of an anticancer agent, for preventing the decrease in efficacy of said anticancer agent potentially induced by a dysbiosis-inducing pharmaceutical substance, such as an antibiotic, administered to said subject to treat or prevent another pathological condition that may be treated or prevented with said dysbiosis-inducing pharmaceutical substance.

In a particular embodiment, the solid dosage form of the invention is administered to the subject almost simultaneously with a dysbiosis-inducing pharmaceutical substance, for example an antibiotic. By “almost simultaneously”, it is meant that the solid dosage form of the invention is administered shortly before, simultaneously, and/or shortly after administration of the dysbiosis-inducing pharmaceutical substance, in particular an antibiotic, preferably shortly before. In a particular embodiment, the solid dosage form of the invention is administered less than 30 minutes before or after the dysbiosis-inducing pharmaceutical substance, in particular an antibiotic, has been administered, in particular less than 20 minutes, less than 19, 18, 17, 16, 15, 14, 13, 12, 11 , 10, 9, 8, 7, 6, 5, 4, 3, 2 minutes, or less than one minute before or after the dysbiosis-inducing pharmaceutical substance, in particular an antibiotic, has been administered. In a further particular embodiment, the solid dosage form of the invention is administered at least once a day, in particular at least twice a day, more particularly three times a day or four times a day. In a further particular embodiment, the solid dosage form of the invention is administered during the whole course of the treatment with the dysbiosis-inducing pharmaceutical substance, in particular with an antibiotic. In a variant of this embodiment, the solid dosage form of the invention may be administered a longer time than the dysbiosisinducing pharmaceutical substance, in particular than an antibiotic, in order to ensure that any residual dysbiosis-inducing pharmaceutical substance, in particular any residual antibiotic, is eliminated. For example, the solid dosage form of the invention may still be administered at least one day after, such as two days after interruption of the administration of the dysbiosisinducing pharmaceutical substance, in particular after the administration of an antibiotic. In a particular embodiment, the invention relates to a solid dosage form of the invention for use in combination with an antibiotic, in particular almost simultaneously, to a subject who is in need of an anticancer agent. According to this embodiment, the solid dosage form of the invention prevents the adverse effects the antibiotic could have on the intestinal microbiota of the subject, and therefore may improve the therapeutic efficacy of the anticancer agent.

Thus, the invention thus also relates to a kit comprising a solid dosage form of the invention and a dysbiosis-inducing pharmaceutical substance, such as an antibiotic. The kit may be for use in the treatment or prevention of a pathological condition that may be treated or prevented with the dysbiosis-inducing pharmaceutical substance, such as an antibiotic. In a particular embodiment of the kit, the dysbiosis-inducing pharmaceutical substance is an antibiotic. The kit may further comprise instructions to implement the methods of the present invention, aiming at preventing the decrease in the efficacy of an anticancer agent. The components of the kit may be administered simultaneously, separately or sequentially. As provided above, the solid dosage form of the invention may, in particular, be administered before, during, or after the administration of the dysbiosis-inducing pharmaceutical agent, such as an antibiotic, in particular shortly before or shortly after, more particularly shortly before.

Graft versus host disease (GVHD)

The present invention also to the treatment, prevention or delaying GVHD or reduction of the severity of GVHD based on the use of a solid dosage form of the invention.

In particular, the present invention can be used to prevent the disruption of the microbiota in patients receiving an allogeneic hematopoietic stem cells transplant and prevent or delay the occurrence of or reduce the severity of GVHD.

In certain aspects, the solid dosage form of the invention according to the invention is for use in a subject who is administered, will be administered or has been administered with an agent that may disturb the gut microbiota of said subject. Thanks to the invention, the deleterious impact of such agents may be prevented. Therefore, the invention relates to a method for mitigating the deleterious effects a pharmaceutical agent may have on the gut microbiota of a subject who is or could be a recipient of an immuno-competent transplant, comprising administering to said subject an effective amount of a solid dosage form of the invention, suitable for inactivating a dysbiosis-inducing pharmaceutical agent. The dysbiosis-inducing pharmaceutical agent may be a pharmaceutical agent administered to treat a pathological condition in the subject as described above.

The solid dosage form of the invention may be administered to the subject even long before transplantation. For example, the subject may have been selected as a transplant recipient but the treatment could not begin before several days, weeks, months or years. In this case, should the subject suffer, between these events, from a disease that would need a treatment with a dysbiosis-inducing pharmaceutical agent, such as an antibiotic, it would be advantageous to prevent gut microbiota dysbiosis by administering a solid dosage form as provided herein. Likewise, the solid dosage form of the invention may be administered to the subject even long after the day of transplantation. In particular, it may unfortunately be that the subject’s transplant be rejected by the host. In this case, halting the systematic administration of a solid dosage form of the invention when the subject receives a dysbiosis-inducing pharmaceutical substance, such as an antibiotic, could severely impair the efficacy of a future transplantation.

In a particular embodiment, the solid dosage form of the invention is administered to the subject almost simultaneously with a dysbiosis-inducing pharmaceutical agent, for example an antibiotic, as defined above in the section relating to cancer treatment.

In a particular embodiment, the invention relates to a solid dosage form of the invention for use in combination with an antibiotic, in particular almost simultaneously, to a subject who is in need of a transplant. According to this embodiment, the solid dosage form of the invention prevents the adverse effects the antibiotic could have on the intestinal microbiota of the subject, and therefore may treat or prevent GVHD.

In a specific embodiment, the invention can be used appropriately in patients at risk of GVHD such as patients taking antibiotics waiting for a hematopoietic stem cell transplant (HSCT) procedure, to prevent GVHD occurrence or reduce the severity of a GVHD episode should one episode occur despite the initial treatment with the invention.

In particular, the invention can be used in patients in wait of, or during the course of a HSCT procedure when they receive antibiotics, in particular during the neutropenia phase. The invention can also be used in these patients when they receive antibiotics before the neutropenia phase in order to maintain an optimal microbiota equilibrium. The invention can also be used in patients diagnosed with a cancer of the blood or bone-marrow when they receive antibiotics in order to maintain the microbiota in the best possible state for the longest possible time and improve the outcome of a HSCT if this procedure is deemed necessary to cure the patient.

The invention can also be used in patients having received a HSCT procedure when they receive antibiotics in order to prevent the occurrence of the GVHD syndrome or avoid the worsening of acute or chronic GVHD if the patient already suffers from the disease.

In particular embodiments, the invention can be used every time the subject takes antibiotics. The invention may also be used after the subject has received a fecal microbial transplant or a treatment with probiotics to restore his or her microbiota diversity and is at risk of GVHD.

In a particular embodiment, the subject was administered with an immunosuppressive agent, such as methotrexate, tacrolimus, everolimus, sirolimus, mycophenolate mofetil or cyclosporine A. In another particular embodiment, the subject was administered with an antiinflammatory drug such as with a corticosteroid.

In a further particular embodiment, the subject has fever. In particular, the antibiotic to be eliminated from the intestine of the subject has been prescribed because of said fever.

In a further particular embodiment, the solid dosage form of the invention is for use in a method for preventing the alteration of the microbiota in a subject who has received, receives or will received an allogeneic transplantation.

The invention can further be used in subjects at high risk of GVHD such as subjects who had a previous episode of GVHD in the years prior to a novel antibiotic cure, a novel hospitalization or a novel immune-suppressive cure.

Thus, the invention also relates to a kit comprising a solid dosage form of the invention and a dysbiosis-inducing pharmaceutical agent, such as an antibiotic, or to a kit comprising a solid dosage form of the invention and an antibiotic. The kit may be for use in the treatment or prevention of a pathological condition that may be treated or prevented with the dysbiosisinducing pharmaceutical agent, such as an antibiotic. In a particular embodiment of the kit, the dysbiosis-inducing pharmaceutical agent is an antibiotic. The kit may further comprise instructions to implement the methods of the present invention, aiming at treating or preventing GVHD. The components of the kit may be administered simultaneously, separately or sequentially. As provided above, the solid dosage form of the invention may, in particular, be administered before, during, or after the administration of the dysbiosis-inducing pharmaceutical agent, such as an antibiotic, in particular shortly before or shortly after, more particularly shortly before.

LEGEND OF THE FIGURE

Figure 1 : Survival results obtained with hamsters treated with clindamycin (CLI) in combination with (i) a placebo ; (ii) activated charcoal “D” at a dose of 600 mg/kg bid ; (iii) activated charcoal “I” at a dose of 600 mg/kg bid ; (iv) activated charcoal “D” at a dose of 1350 mg/kg bid ; or (v) activated charcoal “I” at a dose of 1350 mg/kg bid.

EXAMPLES

Example 1 : Determination of specific area of activated charcoal

Specific surface of activated charcoal is determined with the Brunauer, Emmett and Teller (BET) method, which is based on the adsorption isotherm of nitrogen on the surface of the solid (at the liquid nitrogen temperature: 77K). The isotherm curve reports the quantity of adsorbed nitrogen according to the relative pressure of nitrogen (partial nitrogen pressure divided by saturation pressure). On a certain range of relative pressures, the isotherm can be linearized. From the linear fit equation, the number of nitrogen molecules required to form a monolayer on the surface of 1 g of sample can be determined. Knowing the surface occupied per one nitrogen molecule (16,21 A ² ), the specific area of the solid can be calculated.

BET analysis was performed on samples with a mass comprised between 70 and 150 mg. First, the activated charcoal samples were heated using a stream of nitrogen at 130°C during 30 minutes. Then, they were vacuumed under a pressure rate of 200 mmHg/min during 10 minutes, using a Micromeritics Flowprep 060 device. Finally, they were cooled until they reach ambient temperature (between 15 and 30 °C). After this pre-processing, the samples were analysed with a Micromeritics Gemini VII 2390 analyser. Specific area is a standard property that the Gemini VII management software is able to determine based on the range of relative pressures parameterized in the analysis methods. The characterization was performed by the nitrogen adsorption at its boiling temperature.

The nitrogen adsorption isotherm was defined on 56 points, with relative nitrogen pressure ranging from 0,01 to 0,99. The nitrogen desorption isotherm was defined on 22 points, with relative nitrogen pressure ranging from 0,32 to 0,99. The equilibration time was set to 5 seconds. The linearization of the BET isotherm curve was performed on 19 points with relative pressures of nitrogen ranging from 0.01 to 0.10.

The specific areas of 8 different activated charcoals samples were measured. The results are presented in table 1 :

Table 1 : Specific area results of 8 different activated charcoal samples

Example 2: Determination of porous volume of activated charcoal

Porous volume of activated charcoal is determined with the Brunauer, Emmett and Teller (BET) method, which is based on the adsorption isotherm of nitrogen on the surface of the solid (at the liquid nitrogen temperature: 77K). The isotherm curve reports the quantity of adsorbed nitrogen according to the relative pressure of nitrogen (partial nitrogen pressure divided by saturation pressure). The total porous volume is calculated based on the quantity of adsorbed nitrogen gas at high relative pressure.

BET analysis was performed on samples with a mass comprised between 70 and 150 mg. First, the activated charcoal samples were heated using a stream of nitrogen at 130°C during 30 minutes. Then, they were vacuumed under a pressure rate of 200 mmHg/min during 10 minutes, using a Micromeritics Flowprep 060 device. Finally, they were cooled until they reach ambient temperature (between 15 and 30 °C). After this pre-processing, the samples were analysed with a Micromeritics Gemini VII 2390 analyser. Total porous volume is a standard property that the Gemini VII management software is able to determine based on the range of relative pressures parameterized in the analysis methods. The porous volume determination was performed by the nitrogen adsorption at its boiling temperature and at a relative pressure of 0.99 (enabling to consider all the pores with diameters up to 300 pm) and was calculated by the software depending on the adsorbed quantity of nitrogen in these conditions.

The porous volumes of the 8 activated charcoals samples of example 1 were measured. The results are presented in table 2:

Table 2: Porous volume results of 8 different activated charcoal samples

Example 3: Determination of microporous volume of activated charcoal

The DFT (density functional theory) method is applied to determine the microporous volume of the sample. This method is based on statistical mechanic calculations as laid out in Ravikovitch et al. https://doi.org/10.1016/S0001-8686(98)00047-5.

In this model, pores are considered as independent to each other. Each pore size contributes proportionally to the adsorption isotherm depending on the sample surface it represents. Mathematically, this relation is expressed by: With:

Q(p): the experimental quantity adsorbed at pressure p q(p,H): the quantity adsorbed per unit area at the same pressure p, in an ideal pore size of H f(H): the total area of pores of size H in the sample

As there is no analytic solution for this equation, the problem is best solved in a discrete form; the integral equation for pore size becomes a summation:

The experimental isotherm is compared to theoretical isotherm curves and the pore size distribution is derived from the comparison. The available reference theoretical isotherms are based on different models depending on the adsorbed gas or on the pore geometry (cylindrical or in slit). In this case, nitrogen gas was used and the in slit geometry was selected, as it was considered as the most relevant model to characterize activated charcoals.

The microporous volumes of the 8 activated charcoals samples A-P were measured. The results are presented in table 3:

Table 3: Microporous volume results of 8 different activated charcoal samp es

Example 4: Determination of the in vitro adsorption of antibiotics by activated charcoal 0.2 mL of a suspension of activated charcoal to be tested (10 mg/mL) was mixed with 7.8 mL of a defined buffer (50 nM sodium phosphate + 80 mM sodium chloride, pH =6.0). 2 mL of a solution of antibiotic (MXF: moxifloxacin; CM: clindamycin; CRO: ceftriaxone) at 1 mg/mL was then added to the previous mix. The solution was then sampled at different timepoints (t=0, t=30min, t=1h and t=2h). The adsorption capacities were determined by measuring the residual antibiotic concentration at the end of the test by spectrophotometry.

The in vitro adsorption capacities of 7 activated charcoals samples from different grades and different suppliers were measured. The results are presented in table 4:

Example 5: Determination of the ex vivo adsorption of antibiotics by activated charcoal

The antibiotic of interest and the activated charcoal sample were preincubated separately in pig caecal medium during 2 hours. Different quantities of activated charcoals were preincubated in order to test different ratios of [activated charcoal: anti biotic] (10:1 ; 25:1 ; 50:1 ; 75:1 and 100:1). Indeed, the ex vivo adsorption result can be determined only if:

• the activated carbon is saturated with the antibiotic, and

• the antibiotic concentration variation is significant over the test Accordingly, the antibiotic solution was then added to the activated charcoals suspensions with the different ratios. The adsorption kinetic is monitored over 5 hours with sampling points at t+30min, t+60min, t+180min, t+300min (to is sampled only for controls runs: antibiotic in buffer and antibiotic in caecal medium). The residual quantities of antibiotic were measured by microbiological dosage.

The ex vivo adsorption capacities of 8 different activated charcoals samples. The results are presented in table 5:

Example 6: Correlation of in vitro data with ex vivo data

The ex vivo method is considered as the most representative method to predict the in vivo efficacy of activated charcoals. However, the performance of the ex vivo method is costly, timeconsuming and variable. The prediction of the in vivo activity of activated charcoals from their in vitro adsorption properties would be extremely helpful.

The adsorption capacities of 7 samples of activated charcoals were assessed:

• In vitro, as described in example 4, with the following antibiotics: MXF, CRO and CM

• Ex vivo, as described in example 5, with the following antibiotics: MXF, CRO and CM The mean adsorption score was defined for each method as the mean of the three adsorption values obtained with the three antibiotics. Table 6 summarizes the in vitro and ex vivo adsorption scores of the seven activated charcoals samples used in the study:

The correlation between the results was then studied for each antibiotic separately and for the adsorption score by using a linear regression to fit the 2 variables. The resulting coefficients of determination are presented in table 7:

Table 7: Correlation results between ex vivo and in vitro adsorption assessment methods We can conclude that the in vitro adsorption is not a strong predictive tool of the ex vivo adsorption of activated charcoal and cannot reasonably be used to anticipate which activated charcoals could lead to the best ex-vivo adsorption properties.

Example 7: Correlation of specific area with ex vivo adsorption

As the in vitro adsorption method is not a strong predictive tool of the ex vivo adsorption of activated charcoals, a correlation between their structural properties and the ex vivo adsorption was assessed. The correlation was considered as good if the coefficient of determination R ² obtained by linear regression was above 0.4. To consider a parameter as predictive, the correlation must be good for all the antibiotics taken individually and for the adsorption score. Eight activated charcoals were tested for specific area and for their ex vivo adsorption capacities (cf previous examples): samples A, D, E, F, G, H, I and O. The use of linear regression to fit the 2 variables lead to the following coefficients of determination:

Table 8: Correlation results between ex vivo adsorption results and specific area

We can conclude that specific area is not predictive of the ex vivo and in vivo adsorption of activated charcoal and cannot reasonably be used to select the most appropriate activated charcoals.

Example 8: Correlation of microporous volume with ex vivo adsorption As the in vitro adsorption method is not a strong predictive tool of the ex vivo adsorption of activated charcoals, a correlation between their structural properties and the ex vivo adsorption was assessed. The correlation was considered as good if coefficient of determination R ² obtained by linear regression was above 0.4. To consider a parameter as predictive, the correlation must be good for all the antibiotics taken individually and for the adsorption score.

Eight activated charcoals sampled were tested for microporous volume and for their ex vivo adsorption capacities (samples A, D, E, F, G, H, I and O). The use of linear regression to fit the 2 variables lead to the following coefficients of determination:

Table 9: Correlation results between ex vivo adsorption results and microporous volume

We can conclude that microporous volume is not predictive of the ex vivo and in vivo adsorption of activated charcoal and cannot reasonably be used to select the most appropriate activated charcoals.

Example 9: Correlation of the proportion of micropores with ex vivo adsorption

As the in vitro adsorption method is not a strong predictive tool of the ex vivo adsorption of activated charcoals, a correlation between their structural properties and the ex vivo adsorption was assessed. The correlation was considered as good if coefficient of determination R ² obtained by linear regression was above 0.4. To consider a parameter as predictive, the correlation must be good for all the antibiotics taken individually and for the adsorption score. The proportion of micropores is defined as follows:

[microporous volume]

R _2A = [proportion micropore] = — - - - —

[porous volume]

Eight activated charcoals were tested for microporous volume, porous volume and for their ex vivo adsorption capacities (samples A, D, E, F, G, H, I and O). The proportion of micropores has been calculated for those samples and correlated to the ex vivo adsorption results. The use of linear regression to fit the 2 variables lead to the following coefficients of determination:

Table 10: Correlation results between ex vivo adsorption results and proportion of micropores

We can conclude that the proportion of micropores is not sufficiently predictive of the ex vivo and in vivo adsorption of activated charcoal and cannot reasonably be used to select the most appropriate activated charcoals.

Example 10: Correlation of the macroporous/mesoporous volume with ex vivo adsorption

As the in vitro adsorption method is not a strong predictive tool of the ex vivo adsorption of activated charcoals, a correlation between their structural properties and the ex vivo adsorption was assessed. The correlation was considered as good if coefficient of determination R ² obtained by linear regression was above 0.4. To consider a parameter as predictive, the correlation must be good for all the antibiotics taken individually and for the adsorption score.

The macroporous/mesoporous volume is defined as follow: R1 = [macroporous/mesoporous volume] = [porous volume] — [microporous volume]

Eight activated charcoals were tested for microporous volume, porous volume and for their ex vivo adsorption capacities (samples A, D, E, F, G, H, I and O). The macroporous/mesoporous volume has been calculated forthose samples and correlated to the ex vivo adsorption results. The use of linear regression to fit the 2 variables lead to the following coefficients of determination:

Table 11: Correlation results between ex vivo adsorption results and macroporous/mesoporous volume

We can conclude that the macroporous/mesoporous volume is not fully predictive of the ex vivo and in vivo adsorption of activated charcoal and cannot reasonably be used to select the most appropriate activated charcoals.

Example 11 : Correlation of the specific area available through mesopores-macropores with ex vivo adsorption

As the in vitro adsorption method is not a strong predictive tool of the ex vivo adsorption of activated charcoals, a correlation between their structural properties and the ex vivo adsorption was assessed. The correlation was considered as good if coefficient of determination R ² obtained by linear regression was above 0.4. To consider a parameter as predictive, the correlation must be good for all the antibiotics taken individually and for the adsorption score. The specific area available through mesopores-macropores volume is defined as follow:

Eight activated charcoals were tested for microporous volume, porous volume, specific area and for their ex vivo adsorption capacities (samples A, D, E, F, G, H, I and O). The specific area available through mesopores-macropores has been calculated for those samples and correlated to the ex vivo adsorption results. The use of linear regression to fit the 2 variables lead to the following coefficients of determination:

Table 12: Correlation results between ex vivo adsorption results and specific area available through macropores/mesopores

The correlations are considered as good (R ² >0.4) for all the antibiotics and the ex vivo adsorption score.

Unlike the other combinations of structural properties and unexpectedly, the observation of the specific area available through mesopores-macropores seems well correlated with all ex vivo adsorptions results. R2B ratio is thus considered as the best predictive parameter to select activated charcoals in order to have the best in vivo efficacy.

Based on the experimental data, a R2B ratio of 600 m ² /g was identified as a threshold value to identify activated charcoals associated with efficient ex vivo adsorption capacities. Thus, activated charcoals having a R2B ratio higher or equal to 600 m ² /g are expected to show good ex vivo and in vivo performances. Example 12: Confirmation that the R2B ratio is a reliable parameter to predict ex vivo adsorption capacities

Activated charcoal “P”:

An additional activated charcoal sample (named “P”), which was initially not taken into account to establish previous correlations, was tested for ex vivo adsorption, specific area, microporous volume and total porous volume.

The structure characteristics of activated charcoal P are presented hereafter:

• Specific area: 1975.4 m ² /g

• Total porous volume: 1.707 cm ³ /g

• Microporous volume: 0.319 cm ³ /g

R2B ratio was calculated at 1606 m ² /g for activated charcoal P. This value is above the defined lower limit of 600 m ² /g for R2B and is significantly higher than all R2B values obtained on activated charcoals taken into account to establish correlation between R2B ratio and ex vivo adsorption performances. Therefore, good ex vivo adsorption performances are expected for activated charcoal P.

Table 13: ex vivo adsorption results for activated charcoal P

Excellent ex vivo adsorption performances were obtained with activated charcoal P: this activated charcoal presented ex vivo adsorption values significantly higher than those obtained with activated charcoals taken into account to establish previous correlation between R2B ratio and ex vivo adsorption performances. Thus, R2B ratio is confirmed as a relevant indicator to predict ex vivo adsorption performances.

Activated charcoal “Q”:

An additional activated charcoal sample (named “Q”) was tested for ex vivo adsorption, specific area, microporous volume and total porous volume.

The structure results are presented hereafter:

• Specific area: 1623.2 m ² /g

• Total porous volume: 0.701 cm ³ /g

• Microporous volume: 0.478 cm ³ /g

R2B ratio was calculated at 516 m ² /g for activated charcoal Q. This value is below the defined lower limit of 600 m ² /g for R2B and is in the lower range of R2B values obtained on activated charcoals taken into account to establish correlation between R2B ratio and ex vivo adsorption performances. Therefore, poor ex vivo adsorption performances are expected for activated charcoal Q.

Table 14: ex vivo adsorption results for activated charcoal Q

Very poor ex vivo adsorption performances were obtained with activated charcoal Q: this activated charcoal presented ex vivo adsorption values mainly lower than those obtained with activated charcoals taken into account to establish previous correlation between R ₂ B ratio and ex vivo adsorption performances. Thus, R2B ratio is confirmed as a relevant indicator to predict ex vivo adsorption performances. Example 13: in vivo performance of activated charcoal samples presenting high R2B ratios

The activated charcoal sample I (see previous examples) is herein tested in vivo to assess its efficacy in protecting the gut microbiome in animal models and avoiding the clinical manifestations of gut dysbiosis such as C. difficile infection.

The R _2B ratio for activated charcoal I was assessed to be 857 m ² /g: it is above the threshold of 600 m ² /g which predicts good ex vivo and in vivo performances. In Example 6, the good ex vivo adsorption performances of activated charcoal I were demonstrated.

The in vivo performance of activated charcoal I to prevent a side effect of antibiotics, namely the development of an infection by Clostridioides difficile possibly resulting in death, was then evaluated in a hamster model and compared with activated charcoal D characterized by a R _2B ratio of 1113 m ² /g.

A hamster model of Clostridioides difficile infection (GDI) induced by clindamycin was selected. Each hamster was treated concomitantly with an antibiotic (Clindamycin) and :

- Activated charcoal I, at 600 mg/kg bid or 1350 mg/kg bid; or

- Activated charcoal D, at 600 mg/kg bid or 1350 mg/kg bid; or a placebo.

The antibiotic was administered once a day during 8 days. Activated charcoal (D or I) was administered twice a day during 8 days, which is also the antibiotic treatment duration (administered once a day). Clostridioides difficile strain inoculation (Clostridioides difficile UNT103-1 (VA-11) REA J strain) was performed on Day 3. Hamsters were observed twice a day until D20.

As shown in Figure 1 , treatment with activated charcoal D protected 100% of hamsters from mortality, either at 600 or 1350 mg/kg. Treatment with activated charcoal I protected 100% of hamsters at 1350 mg/kg, but none of the hamster at 600 mg/kg

The activated charcoal I (characterized by a R _2B ratio of 857 m ² /g) demonstrated good in vivo performances, with suitable protection against GDI and subsequent death. It was shown to be less efficient than activated charcoal D which is characterized by a higher R ₂ B ratio (1113 m ² /g). Indeed, activated charcoal D protected all the hamsters from death at the two doses tested, whereas activated charcoal I protected all of them only at the highest dose.

Those results support the predictivity of R2B ratio to select activated charcoals presenting good in vivo efficacy in antibiotic adsorption and protection against antibiotic side effects, such as the development of GDI, possibly resulting in death.