Prodrugs intended for treatment of bacterial infections resistant to antibiotics of beta-lactam family

Abstract

The present invention relates to the field of antibacterial treatments intended to combat antibiotic resistant infections. The present invention more specifically relates to a prodrug that enables the topical and specific inhibition of bacterial infections due to bacteria that are resistant to antibiotics of the beta-lactam family. The prodrug comprises an antibacterial ingredient from a novel therapeutic class bound to a bait molecule comprising a functional group that can be cleaved by beta-lactamase as defined in the classification of Ambler (1980) and Bush and Jacoby (2010). The antibacterial ingredient is masked and thus inactive in the prodrug state and restores its natural structure and antibacterial properties once the prodrug is cleaved in situ by the pathogenic bacteria.

Description

[0001] The present invention relates to the field of antibacterial therapy aimed at combating antibiotic-resistant infections. The invention more particularly relates to a prodrug that enables local and specific inhibition of bacterial infections caused by bacteria resistant to antibiotics of the β-lactam family. The prodrug comprises an antibacterial component from a new therapeutic class bound to a decoy molecule comprising a functional group cleavable by β-lactamases as defined in the classifications of Ambler (1980) and Bush and Jacoby (2010). The antibacterial component is derived from trans-cinnamaldehyde; it is masked and therefore inactive in the prodrug state and restores its native structure and antibacterial properties once the prodrug is cleaved in situ by the pathogen. Technical Field

[0002] Fighting bacterial infections is a worrying public health issue when it comes to dealing with multi-drug resistant bacteria. In fact, many bacteria have developed resistance mechanisms to conventional antibiotics, especially those in the β-lactam family. To treat these infections, second generation antibiotics are administered, but these antibiotics have side effects. Moreover, their systemic use (i.e., non-local and non-specific) contributes to the emergence of new resistances against these new molecules.

[0003] Strategies have been proposed to target antibacterial molecules to infection sites of antibiotic-resistant bacteria, especially by targeting treatment by exploiting resistance mechanisms deployed by bacteria. Evans et al. (J.Med.Chem 2019, 62, 4411-4425) describe such methods; they consist of prodrugs activated by β-lactamases produced by resistant bacteria. In this way, the conventional antibiotic molecule ciprofloxacin is chemically conjugated to a β-lactam motif recognized by bacteria and cleaved by the latter to release the antibiotic molecule. However, the specificity of the prodrug may be limited by the fact that the molecular structure of the antibiotic agent is in its "active" form within the prodrug. This means that the prodrug is expected to exert antibiotic activity, albeit to a lesser extent, even before it is cleaved by β-lactamase. This limits the specificity of the prodrug's action on multidrug-resistant bacteria, but this aspect is necessary to avoid systemic exposure and give bacteria that are not yet multidrug-resistant the opportunity to develop resistance mechanisms. Although this system provides a local antibacterial effect at the site of infection, it does not solve the problem of antibiotic resistance.

[0004] There is a clear need for new therapeutic strategies to treat infections caused by antibiotic-resistant bacteria. Solutions to address this need must include (1) novel therapeutic targets and (2) specific local action to avoid systemic exposure and the development of resistance. Summary of the invention

[0005] The inventors have proposed a new therapeutic strategy to combat antibiotic-resistant bacteria. The method is based on the antibacterial action of small molecules targeting new molecular targets whose mechanism of action is not offset by conventional defense or resistance systems developed by bacteria, such as certain molecules derived from essential oils. However, since these molecules are unstable and their bioavailability is low, it is necessary to associate an innovative delivery system with them so that the therapeutic solution according to the present invention takes the form of a prodrug. The method consists of a prodrug that enables the active molecule to be delivered only locally to the target site, where the infection site contains resistant bacteria, due to an enzymatic reaction specific to multidrug-resistant bacteria that can cleave the prodrug. This prevents any harmful systemic effects of the active ingredient.

[0006] Therefore, a first object of the present invention relates to a prodrug intended for the treatment of bacterial infections resistant to antibiotics of the β-lactam family, the prodrug comprising an antibacterial component linked to a decoy molecule containing a functional group capable of being cleaved by β-lactamase, characterized in that it corresponds to formula (I):

[0007] [Chemical formula 1]

[0008]

[0009] in

[0010] X is either absent or is a spacer group selected from phenylene, alkenylene or alkynylene,

[0011] R1 is a group selected from alkyl, alkylsulfonylethyl or alkylaminoethyl, for example methyl, methylsulfonylethyl or dimethylaminoethyl,

[0012] R2 is a group selected from methyl, 2,2,2-trifluoroethyl,

[0013] R3 is a masked antimicrobial ingredient, the masked active ingredient corresponding to the structure of trans-cinnamaldehyde without its aldehyde function or one of the variants of this structure having retained antimicrobial activity,

[0014] R4 is a β-lactam group.

[0015] A second object of the present invention relates to a pharmaceutical composition comprising a prodrug according to the present invention and at least one pharmaceutically acceptable excipient.

[0016] A third object of the invention relates to the use of the prodrug according to the invention in the treatment of bacterial infections resistant to antibiotics of the β-lactam family.

[0017] Advantages of the invention

[0018] The present invention differs from existing solutions by the choice of the antibacterial active ingredient and the fact that this active ingredient is masked within the prodrug (and is therefore rendered inactive in prodrug form). In fact, the inventors have developed a method to exploit the antibacterial properties of small molecules whose instability and low bioavailability have hitherto been an obstacle to their therapeutic use, as is the case with those derived from essential oils (EO). The latter are known to have antibacterial activity and a novel mechanism of action which is less likely to induce bacterial resistance. The object of the present invention is therefore to obtain a new class of antibacterial drugs (prodrugs) which are effective and safe (do not induce new resistances) for the treatment of infections caused by beta-lactam-resistant bacteria.

[0019] The drug in question is in the form of a prodrug, comprising an antibacterial component derived from a small molecule carrying an aldehyde functional group; this functionality is exposed to a variety of mechanisms that are rapidly altered by the patient's metabolism, resulting in poor bioavailability of the small molecule. In order to ensure that they reach the site of infection, they need to be incorporated into a molecular construct with suitable stability, water solubility and bioavailability. Therefore, the active ingredient is modified by conjugation with a decoy molecule. Another advantage of this system is that it masks the activity of the active ingredient, but ensures its release in the presence of bacteria resistant to antibiotics of the β-lactam family. Once released, the molecule restores its active form and antibacterial properties. Therefore, the delivery of the active ingredient is targeted to the site of infection, thereby avoiding the side effects and resistance development risks associated with the systemic distribution of antibacterial agents.

[0020] The release of the active ingredient depends on the action of resistant bacteria. In fact, the prodrug includes a group recognized by the β-lactamase secreted by the resistant pathogen, which catalyzes the cleavage of the decoy molecule unit (β-lactam unit) of the prodrug. The disintegration of this unit releases the antibacterial ingredient and delivers the active molecule to the site of infection. Therefore, the prodrug acts as a Trojan horse because the bacteria induce the release of antibacterial molecules that destroy and / or inhibit the bacteria by activating their defense system - the secretion of β-lactamase type enzymes.

[0021] In more detail, between the β group and the active ingredient (R3 group), the prodrug comprises an N / O-acetal-oxycarbonyl group connected to the X spacer group. It is the N / O-acetal-oxycarbonyl group that ensures the incorporation and masking of the active ingredient, as well as its release after the prodrug disintegrates due to the action of the target enzyme. This is the original feature of the prodrug structure, and the innovative characteristics of the present invention are based on this original feature, which is related to similar prodrug molecules in the prior art: the release is effective, but the prodrug is stable to spontaneous hydrolysis degradation in the absence of the target enzyme. This characteristic also ensures that the prodrug cannot exert any inherent antibiotic effect, because the structure of the active ingredient cannot be restored without cleavage. These two characteristics make it possible to provide the clinical use of the prodrug according to the present invention.

[0022] The system according to the invention therefore relies on a local, immediate and specific action of an antimicrobial molecule from a new therapeutic class that is less likely to induce new resistances. An example of such a molecule is trans-cinnamaldehyde, the active ingredient in cinnamon essential oil. DETAILED DESCRIPTION

[0023] The first object of the present invention relates to a prodrug intended for the treatment of bacterial infections resistant to antibiotics of the β-lactam family, the prodrug comprising an antibacterial active ingredient linked to a decoy molecule comprising a functional group capable of being cleaved by β-lactamase, characterized in that it corresponds to formula (I):

[0024] [Chemical formula 1]

[0025]

[0026] in:

[0027] X is either absent or is a spacer group selected from phenylene, alkenylene or alkynylene,

[0028] R1 is a group selected from alkyl, alkylsulfonylethyl or alkylaminoethyl, for example methyl, methylsulfonylethyl or dimethylaminoethyl,

[0029] R2 is a group selected from methyl, 2,2,2-trifluoroethyl,

[0030] R3 is a masked antimicrobial ingredient, the masked active ingredient corresponding to the structure of trans-cinnamaldehyde without its aldehyde function or one of the variants of this structure having retained antimicrobial activity,

[0031] R4 is a β-lactam group.

[0032] The term "prodrug" refers to a drug precursor. This is an inactive / less active form of a molecule that is metabolized in vivo to an active metabolite. In the present invention, the prodrug consists of a masked antibacterial component and a decoy molecule; therefore, the antibacterial component is a metabolite.

[0033] The active ingredient is a small molecule drug with antibacterial properties. It is masked in the prodrug, i.e., present in an inactive form. Such active ingredients may be derived from various types of plants, including plants or lichens, and may in particular be derived from essential oils, or from animals, in particular marine animals.

[0034] In a preferred embodiment of the invention, the antimicrobial active ingredient is derived from EO with known antimicrobial properties, trans-cinnamaldehyde as shown below, or one of its variants. "Trans-cinnamaldehyde variant" refers to a molecule having the same structure as cinnamaldehyde, exhibiting similar, identical or superior antimicrobial activity and / or conferring enhanced stability to the prodrug. Such variants may be substituted, in particular, on the phenyl unit, for example by replacing H with methoxy, ethoxy, alkylamino (such as dimethylamino) or acylamino.

[0035] [Chemical formula 2]

[0036]

[0037] In the prodrug, the active ingredient is inactivated by masking its aldehyde function. This modification has two functions: it enables binding to the decoy molecule and it masks (temporarily inhibits) the antibacterial activity of the active ingredient in prodrug form.

[0038] In the context of the present invention, masking is achieved by converting the aldehyde function into an N / O acetal, the nitrogen atom of which is acylated (more precisely, "oxy-carbonylated") to give the acetal a higher stability against spontaneous hydrolysis (degradation) and to enable the release of the oxy-carbonylated N / O acetal as a carbamate (a good leaving group) upon activation of the decoy moiety of the prodrug by the target β-lactamase. Thus, "R3" represents a phenyl-vinyl unit.

[0039] "Decoy molecules" in the sense of the present invention are groups that are recognized by bacteria and are able to induce the activation of their defense functions against antibiotics of the β-lactam family. These defense functions consist in the secretion of enzymes capable of cleaving the β-lactam nucleus present in this family of antibiotics.

[0040] The bait molecule according to the present invention comprises an R4 group recognized by bacteria resistant to beta-lactams. The R4 group is a beta-lactam group that simulates the presence of antibiotics. As explained above, the recognition of this group can serve as a signal that triggers the secretion of beta-lactamase by bacteria, which plays a role in defense. The bait molecule also comprises a central group (N / O-acetal-oxycarbonyl group) of formula (I) and an optional spacer group X.

[0041] A particularly innovative aspect of the present invention lies in the structure of the cleavable functional group - which corresponds to the central N / O-acetal-oxycarbonyl group of formula (I), which fulfils three functions: (i) binding of the active ingredient to the β-lactam group, (ii) masking the antibacterial activity of the active ingredient by binding the active ingredient to the N / O-acetal-oxycarbonyl unit, and (iii) activation of the antibacterial activity by cleavage of the β-lactam group after secretion of β-lactamase by the bacteria, followed by self-disintegration of the structure, resulting in the release of the active ingredient in a restored active form with novel antibiotic properties. In fact, it is noteworthy that the cleavable group has the following properties, which allow the active ingredient that has been modified to synthesize a prodrug to restore its natural form and antibacterial properties after being released. Here, the active ingredient carries an aldehyde functional group connected to the bait molecule at R3, as shown in formula (I). The aldehyde functional group of the active ingredient is reformed in situ. In [ Figure 3 ] shows the mechanism of action.

[0042] In a preferred embodiment of the invention, the β-lactam group (R4 group) is derived from a cephalosporin, a penicillin or a carbapenem. This group is capable of being cleaved by a β-lactamase, which may be a carbapenemase.

[0043] Many R4 groups are described in the literature and those skilled in the art will be able to recognize them. A specific example is cephalothin.

[0044] [Chemical formula 3]

[0045]

[0046] The R4 group can be more generally represented by the following formula (II):

[0047] [Chemical formula 4]

[0048]

[0049] Wherein R7 is selected from alkyl, benzyl, substituted or unsubstituted aryl.

[0050] Examples of specific R7 groups are shown below:

[0051]

[0052] The spacer group X modulates the properties of the prodrug depending on the choice of group X and the active ingredient associated with the prodrug. Its presence is optional. In the configuration in which the X group is absent, the R4 group is directly attached to the N / O-acetal-oxycarbonyl group.

[0053] Examples of prodrugs according to the invention are the molecules OSTA-1, OSTA1b, OSTA-5, OSTA-6 and OSTA-7 shown below:

[0054] [Chemical formula 5]

[0055]

[0056] [Chemical formula 6]

[0057]

[0058] [Chemical formula 7]

[0059]

[0060] [Chemical formula 8]

[0061]

[0062] [Chemical formula 9]

[0063]

[0064] A second object of the present invention relates to a pharmaceutical composition comprising the prodrug as described above and at least one acceptable excipient.

[0065] A third object of the present invention relates to the use of the prodrug as previously described in the treatment of bacterial infections resistant to antibiotics of the β-lactam family.

[0066] Bacterial infections resistant to β-lactam antibiotics are mainly caused by Gram-negative bacteria that produce β-lactamases, but some Gram-positive bacteria (such as MRSA) can also produce β-lactamases.

[0067] The prodrugs according to the invention are particularly directed to:

[0068] - Gram-negative bacteria belonging to one of the following families: Moraxellaceae, Legionella, Enterobacteriaceae,

[0069] Pseudomonadaceae, Alcaligenesaceae, Sphingomonasaceae, Pasteurellaceae, Neisseriaceae, Pasteurellaceae, Flavobacteriaceae, Bacteroidetes, Fusobacteriaceae, Burkholderia cepacea, Cronobacter

[0070] malonaticus, Cronobacter malonaticus, Enterobacter sakazakii, Helicobacter pylori, Micrococcus luteus, Prevotella, Proteus mirabilis, Proteus vulgaris G, Salmonella Enteritidis G-, Salmonella typhimurium, Salmonella typhimuriu, Salmonella paratyphi A, Vibrio fluviatilis (MDR), Vibrio parahaemolyticus, Acinetobacter baumannii,

[0071] - Gram-negative bacteria belonging to one of the following genera: Klebsiella, Serratia, Citrobacter, Salmonella, Shigella, Proteus, Haemophilus, Campylobacter, Legionella, Sphingomonas, Moraxella, Kingella, Pasteurella, Capnophaga, Neisseria, Bacteroides, Enterobacter, Pseudomonas, Fusobacterium, Achromobacter, Escherichia and Acinetobacter,

[0072] - Gram-negative bacteria belonging to one of the following species: Klebsiella pneumoniae, Klebsiella oxytoca, Acinetobacter baumannii, Acinetobacter calcoaceticus, Serratia marcescens, Citrobacter freundii, Citrobacter cohnii, Haemophilus influenzae, Haemophilus parainfluenzae, Campylobacter jejuni, Campylobacter coli, Legionella pneumophila, Legionella longbeach, Legionella Legionella micda, Enterobacter cloacae, Escherichia coli, Escherichia herschii, Pseudomonas aeruginosa, Pseudomonas fluorescens, Pseudomonas putida, Pseudomonas stutzeri, Achromobacter xyloxydans, Achromobacter denitrificans, Sphingomonas paucimobilis, Moraxella catarrhalis, Kingella kingella, Pasteurella multocida, Capnocytophaga canimorsus, Neisseria gonorrhoeae, Neisseria actamica, and Bacteroides fragilis,

[0073] - Particularly Pseudomonas aeruginosa and Acinetobacter baumannii (particularly A. baumannii 5377 or A. baumannii myxoides).

[0074] - The following Gram-positive bacteria:

[0075] Families: Staphylococcus, Micrococcaceae, Streptococcus, Enterococcaceae, Bacillaceae, Clostridiaceae, Listeriaceae, Corynebacteraceae,

[0076] Genus: Staphylococcus, Micrococcus, Streptococcus, Enterococcus, Bacillus, Clostridium, Listeria, Corynebacterium,

[0077] Species (examples): Staphylococcus aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus, Micrococcus luteus, Streptococcus pyogenes, Streptococcus agalactiae, Streptococcus pneumoniae, Streptococcus viridans, Enterococcus faecalis, Enterococcus faecium, Bacillus anthracis, Clostridium difficile, Listeria monocytogenes, Corynebacterium diphtheriae,

[0078] -Other non-Gram-stainable bacteria:

[0079] Family: Mycobacteriaceae;

[0080] Genus: Mycobacterium;

[0081] Species: Mycobacterium tuberculosis, Mycobacterium leprae, Mycobacterium abscessus, Mycobacterium ulcerans.

[0082] These infections may particularly affect the digestive tract, urinary tract, lung tract, or skin and mucous membranes, and may involve specific organs or be systemic. The prodrugs according to the invention can be used to treat all these forms of infection.

[0083] Examples of infections (with or without concomitant diseases, such as genetic factors, immunosuppression, etc.) for which the molecules according to the invention are particularly attractive are, respiratory tract infections (with or without concomitant diseases): pneumonia, HAP, VAP, tuberculosis, sinusitis, cystic fibrosis; blood or blood-borne infections: bacteremia, sepsis; urinary tract infections: cystitis, pyelonephritis; skin and soft tissue infections: abscesses, furuncle, impetigo, non-healing wounds, necrotizing infections; surgical site infections; infections associated with prostheses, medical devices (catheters, tubes, etc.); gastrointestinal infections; sexually transmitted infections; infections of the central or peripheral nervous system, heart, eyes, ears, mouth, bones and joints.

[0084] The present invention will be better understood by reading the following examples, which are provided by way of illustration and are in no way to be considered as limiting the scope of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0085] [ Figure 1 ][ Figure 1 ]: Representation of the OSTA-1 prodrug synthesis method.

[0086] [ Figure 2 ][ Figure 2 ]: UV absorption spectrum of OSTA-1 molecule at 285nm in vitro.

[0087] [ Figure 3 ][ Figure 3 ]: Activation mechanism of active ingredients after disintegration of OSTA-1 prodrug.

[0088] Example

[0089] Example 1: Synthesis of Compound A (OSTA-1)

[0090] The synthesis scheme of OST1 prodrug molecule includes five steps and two auxiliary steps. Figure 1 ]middle.

[0091] Example 2: Functional analysis of OSTA-1 molecules

[0092] In vitro cleavage of A-OSTA-1 molecules

[0093] Cleavage of the OSTA-1 prodrug was tested in vitro by exposure to β-lactamase in saline aqueous medium.

[0094] Experimental conditions :

[0095] The cleavage test was performed in physiological saline buffer (phosphate buffered saline, PBS) at pH 7.0. OSTA-1 molecules were added at a concentration of 100 μM, and commercial β-lactamase was added at a concentration of 1 unit / mL. Incubation was performed at 37°C. The appearance of free trans-cinnamaldehyde in the culture medium was then monitored by UV absorption at 285 nm.

[0096] The results are in [ Figure 2 ] is shown in ].

[0097] The release of the active ingredient (in this case, trans-cinnamaldehyde) was monitored by UV absorption at 285 nm. The release progress of the active ingredient (100 μM) of the prodrug molecule was observed in the presence and absence of β-lactamase. It is clear that in the presence of β-lactamase, a large amount of the active ingredient is released after 2 hours of reaction. In the absence of β-lactamase, no spontaneous release of the active ingredient was observed.

[0098] Conclusion: The OSTA-1 prodrug molecule functions as expected: it can be cleaved by β-lactamase and releases the active ingredient.

[0099] B-Activation of the active ingredient after prodrug disintegration

[0100] The activation mode of the active ingredient is Figure 3 ] which shows the mechanism of disintegration of OSTA-1 prodrug and release of cinnamaldehyde in its active native form.

Claims

1. A prodrug intended for use in the treatment of bacterial infections resistant to antibiotics of the β-lactam family, the prodrug comprising an antibacterial active ingredient linked to a decoy molecule containing a functional group capable of being cleaved by β-lactamase, characterized in that: It corresponds to formula (I): [Chemical formula 1] in X is either absent or is a spacer group selected from phenylene, alkenylene or alkynylene, R1 is a group selected from alkyl, alkylsulfonylethyl or alkylaminoethyl, for example methyl, dimethyl, methylsulfonylethyl or dimethylaminoethyl, R2 is a group selected from methyl, 2,2,2-trifluoroethyl, R3 is a masked antimicrobial ingredient, said masked active ingredient corresponding to the structure of trans-cinnamaldehyde without its aldehyde function or one of the variants of said structure having retained antimicrobial activity, R4 is a β-lactam group.

2. The prodrug of claim 1, wherein the R4 group is a cephalosporin, a penicillin or a carbapenem derivative.

3. The prodrug according to one of the preceding claims, wherein the group X is absent.

4. Prodrug according to one of the preceding claims, which is selected from the molecules OSTA-1, OSTA-1b, OSTA-5, OSTA-6 or OSTA-7.

5. A pharmaceutical composition comprising the prodrug according to one of claims 1 to 4 and at least one acceptable excipient.

6. Use of a prodrug according to one of claims 1 to 4 for the treatment of bacterial infections resistant to β-lactam antibiotics.

Images