EP2866819A1

EP2866819A1 - Dosing instructions for endotoxin-binding lipopeptides

Info

Publication number: EP2866819A1
Application number: EP13732900.9A
Authority: EP
Inventors: Dieter Falkenhagen; Stephan Harm; Jens Hartmann
Original assignee: Zentrum fur Biomedizinische Technologie Der Donau- Universitat Krems
Current assignee: Zentrum fur Biomedizinische Technologie Der Donau- Universitat Krems
Priority date: 2012-06-29
Filing date: 2013-06-27
Publication date: 2015-05-06
Also published as: EP2679236A1; WO2014001444A1; US20150190461A1

Abstract

The invention relates to an endotoxin-binding lipopeptide selected from the group consisting of polymyxins, polymyxin derivatives, polymyxin analogues, prodrugs thereof and pharmaceutically acceptable salts thereof, and also relates to a preparation for parenteral administration comprising a lipopeptide of this type, for prophylaxis or for treatment of diseases and conditions caused by endotoxemia, by i) parenterally administering a bolus of the lipopeptide to achieve a lipopeptide serum concentration of 0.01 µg/ml to 0.8 µg/ml and ii) maintaining this lipopeptide serum concentration by parenterally administering the lipopeptide over a specifiable period of time.

Description

DOSAGE INSTRUCTIONS FOR ENDOTOXIN BINDING LIPOPEPTIDES

The invention relates to an endotoxin-binding lipopeptide selected from the group consisting of polymyxins, polymyxin derivatives, polymyxin analogues, their prodrugs and their pharmaceutically acceptable salts for the prophylaxis or treatment of diseases and conditions caused by endotoxemia. The invention further relates to a preparation for parenteral administration comprising at least one such endotoxin-binding lipopeptide as the active ingredient and at least one pharmaceutically acceptable carrier and / or adjuvant for the prophylaxis or treatment of diseases and conditions caused by endotoxemia.

Endotoxins are lipopolysaccharides (LPS) in the cell wall of Gram-negative bacteria and are released by cell lysis and cell division. In fact, lipopolysaccharides are the most abundant lipid component of the outer cell membrane of Gram-negative bacteria. Endotoxins are pyrogenic substances and the affected individual reacts with a strong inflammatory reaction and fever when endotoxins enter the body, for example in the course of a microbial intoxication, and as key mediators cause an uncontrolled activation of the mononuclear-phagocytic system. An accumulation of endotoxins in the bloodstream as a result of endotoxemia leads to an uncontrolled activation of the immune cells and an imbalance of the coagulation system. This can lead to sepsis, which is characterized among other things by high fever, low blood pressure and, in severe cases, by multi-organ failure. Sepsis is a very serious condition; the lethality of individuals with severe sepsis or septic shock is about 30-60%, depending on the severity of the disease. Endotoxemia due to Gram-negative bacterial infection is one of the most common causes of Systemic Inflammatory Response Syndrome (SIRS), sepsis, severe sepsis or septic shock, and the resulting serious complications Patients with impaired immune responses, such as liver patients or patients undergoing chemotherapy, are susceptible to bacterial infections, presenting symptoms of endotoxin poisoning, and acute liver failure or acute decompensation in chronic liver failure can also result in endotoxemia, which causes biochemically very similar to sepsis. For example, acute decompensation may occur in patients with chronic liver failure. In this condition, endotoxins derived from normal gut flora can overcome the intestinal barrier and stimulate the release of inflammatory mediators in the body, causing a sepsis-like condition.

The structure of a lipopolysaccharide molecule is in three parts: a lipid A forms the region of the molecule which faces the bacterial cell; Lipid A anchors the molecule in the outer membrane of the Gram-negative bacterium. The LPS molecule also has a central, highly conserved core region bound to the lipid A. The third and outermost region is formed by an O-specific polysaccharide (O-antigen), the structure of which can vary widely within the various Gram-negative bacteria. The toxic effect is due to the lipid A, which is released only during cell lysis.

Polymyxins are antibiotic substances originally derived from the bacterium Bacillus polymyxa and have been used for a long time to treat infections with Gram-negative bacteria in humans and animals. Polymyxins interfere with the cell wall structure by increasing the permeability of the cell membrane, which causes cell lysis. Polymyxins bind not only phospholipids but also lipopolysaccharides (endotoxins) with high affinity. The antibacterial mechanism of polymyxins is described, for example, in a publication by Tony Velkov et al. (Velkov et al., 2010 Journal of Medicinal Chemistry: 53 (5): 1898-1916).

Due to the neurotoxic and nephrotoxic effects of polymyxins, only polymyxin B and polymyxin E (colistin) have gained certain therapeutic importance as antibiotics. To date, these two polymyxins are the only therapeutically approved representatives of their class of substances. Polymyxin B and colistin are FDA approved for parenteral infusion in the US. Polymyxin B and Colistin have been used for decades for oral or topical therapies. For parenteral, systemic treatment of diseases and conditions due to infection with Gram-negative bacteria, however, they come only as a last resort as an antibiotic for therapeutic use due to their neuro-and nephrotoxic side effects. Colistin seems to be less nephrotoxic than polymyxin B, by However, the required higher dosage is at least partially compensated, so that can be expected in the clinical routine about the same extent with nephrotoxic reactions. However, sufficient data on the nephrotoxicity of the two antibiotics are not available from today's perspective. New York-based infectiologists describe renal failure in 14% of 60 patients treated with polymyxin B. Doctors in Greece describe marked nephrotoxicity in the majority of patients with renal insufficiency already at baseline. In contrast, no significant changes were found in patients with normal kidney function. A detailed review of the toxicity of polymyxins can be found in a publication by Falagas and Kasiakou (Falagas and Kasiakou, 2006, Critical Care 10: R27). The dosage of polymyxins thus plays a central role in the prevention or minimization of toxic side effects, in particular nephrotoxic side effects.

Due to the frequent occurrence of severe disease progression as a result of infections with multidrug-resistant pathogenic strains observed in recent years, for example in acute infections with strains of the bacterium Pseudomonas aeruginosa, despite their toxicity, polymyxins are again necessarily administered parenterally as an antibiotic. A reference source for polymyxin B in the form of the sulfate salt of Polymxin Bl and B2 for parenteral administration is currently available from Bedford Laboratories ("Polymyxin B for Injection 500,000 Units", manufacturer: Bedford Laboratories) According to the manufacturer's information, parenteral administration is intravenous, intramuscular or in the case of meningitis intrathecal, wherein the maximum daily dose reported is typically 2.5 mg / kg body weight per day divided into two to three infusions Typically, the serum concentration of polymyxin after administration is in the range of 1 to 6 μg / ml In severe cases, it may also be higher in the range of 6 to 50 g / ml Colistin is administered predominantly in the form of colistin-methanesulfonate with a serum concentration in the range of approximately 1 to 3 μg / ml. Polymyxin E) is used in the same way as Polymyxin B, usually in higher doses.

Resistance to polymyxin B is rather unusual but may develop if the antibiotic fails to reach the cytoplasmic membrane due to changes in the outer membrane. Polymyxins are effective against many Gram-negative pathogens, such as E. coli, Enterobacter, Klebsiella spp. and also against P. aeruginosa. Proteus arias and S. marcescens, which are normally resistant; the sensitivity of ß. fragilis is variable. The minimum inhibitory concentrations for E. coli are in the range of 0.04 - 3.7 mg / 1 and for P. aeruginosa between 1.2 and 33.3 mg / 1 (Garidel and Brandenburg, 2009, Anti-Infective Agents in Medicinal Chemistry, 8: 367-385).

Since the doses of polymyxin B and colistin used in clinical practice to induce nephro- and neurotoxic side effects when administered parenterally, new treatment strategies and therapeutic approaches have been developed in the past in connection with the use of endotoxin-binding lipopeptides such as polymyxin.

As a commonly used alternative to the administration of polymyxins in the form of a medicament, extracorporeal blood and / or blood plasma purification methods (therapeutic apheresis) have been established using suitable adsorption materials. Apheresis methods and adsorber materials for eliminating toxic and / or harmful substances from blood and blood plasma are well known in the art. Known adsorber materials comprise porous or fibrous carrier materials, on the surfaces of which polymyxin B is immobilized, for example, covalently or by means of hydrophobic interaction. To date, no neurotoxic and nephrotoxic side effects have been reported with such adsorbent materials, which are widely used in the treatment of septic conditions. Adsorbent materials functionalized with polymyxin B are known, for example, from EP 0 110 409 A1, WO 2010/083545 and WO 2011/160149. However, apheresis methods using suitable adsorbers have the disadvantage that they can not be applied across the board on account of the high technical complexity, the limited availability of the therapy sites and the significantly higher manufacturing and therapy costs compared to the medicamentous treatment and therefore only those for intensive care Supply necessary needs can be met.

Although the lethality of patients with endotoxemia-induced diseases, in particular sepsis, has been reduced by the clinical use of the abovementioned polymyxin-based adsorber materials, the mortality of patients with severe sepsis and septic shock is still very high despite maximal therapy. For this reason, as well as due to the ever-increasing problem of multi-resistance of In the presence of bacteria against antibiotics and the associated increasing incidence of serious illnesses, there is still a high demand for low-cost, nationwide and low-side-effects forms of therapy. Consequently, in recent years, they have again turned to the drug therapy approaches. For example, by chemically modifying the Dab side chains, the cyclic peptide ring or the fatty acid chain of the polymyxin molecular structure, a variety of synthetic polymyxin analogs / derivatives have been developed to provide drugs with lower toxic side effects and improved endotoxin binding efficiency, respectively. However, most of these modifications resulted in inactive compounds. Furthermore, no toxicological data are known for many of the analogs / derivatives described. A detailed review of polymyxin-based antibiotics, analogs and derivatives is described in the publication by Velkov et al. (Velkov et al., 2010, Journal of Medicinal Chemistry, 53 (5): 1898-1916).

EP 2 332 965 describes synthetic peptides derived from naturally occurring polymyxins and octapeptins having antibacterial properties for use as an antibiotic to treat individuals with a bacterial infection, and to methods for producing these peptides.

WO 2010/075416 discloses polymyxin, in particular polymyxin B-derived chemical compounds with antibacterial properties as antibiotic against a variety of Gram-negative bacteria. The compounds described therein have a reduced toxicity compared to polymyxin B.

Despite the numerous attempts to produce peptide compounds to provide a drug with polymyxin B or colistin comparable endotoxin binding efficiency and antibacterial activity, and with markedly reduced toxicity, none of these compounds has heretofore been approved for clinical use.

It is therefore an object of the invention to eliminate or minimize the drawbacks known from the prior art and to provide a new dosage regimen for drug-administered endotoxin binding lipopeptides such as polymyxins, polymyxin analogs or polymyxin prodrugs and their pharmaceutically acceptable salts such that prophylaxis or treatment of diseases and conditions caused by endotoxemia. It is in particular a task of Invention to provide a new dosage regimen for naturally occurring and clinically approved polymyxins such as polymyxin B and colistin. The new dosage regimen is expected to bring significant improvements in toxic side effects while maintaining high efficacy and to be a much less expensive treatment option for patients with endotoxemia than the adsorbent materials used to date for these purposes.

This object is achieved by a new dosage regimen which is characterized by i) parenteral administration of a bolus of the lipopeptide to reach a lipopeptide serum concentration of 0.01 ^ g / ml to 0.8 μg / ml and

ii) maintenance of this lipopeptide serum concentration by parenteral administration of the lipopeptide over a predetermined period of time.

Thanks to the invention, it is possible for the first time not to use polymyxins or polymyxin derivatives and polymyxin analogues in the conventional sense as antibiotics, ie to eliminate gram-negative bacteria, but in significantly lower concentrations (4- to 100-fold lower) for inactivating the endotoxins in patients with endotoxemia. Inactivation of the endotoxins means that their biological activity, in particular on the release of inflammatory mediators such as cytokines, is inhibited or blocked. As a result, the proinflammatory phase, e.g. in sepsis or in SIRS caused by gram-negative endotoxins, is suppressed or reduced.

The novel dosing regimen allows parenteral administration of polymyxins and their analogues, derivatives and prodrugs while circumventing nephro- and neurotoxic side-effects. The serum lipopeptide concentration in the dosage regimen according to the invention is thus at a factor of 4 to 100 below the concentration achieved when the approved antibiotic polymyxin B preparations are administered parenterally.

The surprising fact has been found that even very low serum concentrations of polymyxin B of 0.0 ^ g / ml to 0.8 g / ml are sufficient to inhibit endotoxins in their activity, while neuro and nephrotoxic effects are excluded. The effect of this novel dosing instruction was demonstrated with the aid of endotoxins from E. coli and Pseudomonas aeruginosa, in which experiments an endotoxin Initial concentration of 0.5 ng / ml was chosen, ie a concentration that already represents a maximum value in a clinically relevant sepsis.

The dosing instructions according to the invention are based on the stated surprising fact that in the investigation of various polymyxin-functionalized adsorbent materials, endotoxin binding by polymxin occurs exclusively via a very small amount of desorbed and transferred into the blood or blood plasma polymyxin molecules. These surprising and unpredictable results are based on the fact that after targeted washing of the adsorber materials in which desorbable polymyxin molecules were removed from the adsorber surface, no endotoxin adsorption could be detected by the polymyxin molecules still immobilized on the adsorber material surface. It has been found that even with covalent binding of polymyxin to the adsorber surface, a certain amount of polymyxin molecules is bound by nonspecific forces. These nonspecifically bound poly-myxin molecules can desorb from the adsorber surface during therapeutic application and in the released state render harmless endotoxins present in the blood or blood plasma of the patient. Thus, it can be summarized that the very good Endotoxinadsorption by adsorbent materials based on porous or fibrous support materials, on the surfaces of polymyxin B, for example, covalently or immobilized by hydrophobic interaction, exclusively on a very small material desorbed from the carrier and released into the blood or blood plasma Amount of free polymyxin molecules is due. The binding between polymyxin and endotoxins apparently occurs only when both the hydrophobic and the positively charged amino groups of polymyxin B are accessible to the endotoxins.

The novel dosage initiation described in this disclosure is based on these surprising results and defines significantly lower serum concentrations of endotoxin-binding lipopeptides such as polymyxin compared to those established in the standard polymyxin B and colistin therapies that have been practiced for many decades.

The terms "polymyxin" and "polymyxins" as used herein refer to known, naturally occurring chemical compounds which are originally derived from the bacterium Bacillus polymyxa (polymyxin B) and Bacillus colistinus (polymyxin E). The polymyxins can either be isolated from bacteria or made synthetically. The bacterium-derived polymyxin B is composed of 6 derivatives called polymyxin Bl, polymyxin B2, polymyxin B3, polymyxin B4, polymyxin B5 and polymyxin B6. By comparison, the polymyxin approved by the FDA for parenteral infusion is composed only of polymyxin Bl to B4. As previously mentioned, only polymyxin B and polymyxin E (colistin) are clinically relevant.

The term "polymyxin derivative" refers to a polymyxin-derived compound obtainable by modification of naturally occurring polymyxins, for example, by chemical modification of the dab side chains, the cyclic peptide ring or the fatty acid chain of the polymyxin molecular structure A detailed review of Polymy - xin-based antibiotics, analogs and derivatives are described in the publication by Velkov et al., (Velkov et al., 2010, Journal of Medicinal Chemistry, 53 (5): 1898-1916) A representative example of a polymyxin derivative is polymyxin nonapeptide, a polymyxin B derivative lacking the hydrophobic moiety and an amino acid.

The term "polymyxin analog" as used herein refers to a chemical lipopeptide compound structurally similar or similar to a polymyxin (polymyxin-like lipopeptide) and having the same endotoxin binding activity as polymyxins or polymyxins has comparable endotoxin binding activity. A representative example of such a polymyxin analogue can be found in the disclosure WO 2008/006125 Al.

The term "prodrug" as used herein refers to a precursor compound of the endotoxin-binding lipopeptide as defined wherein the precursor compound is converted to the active endotoxin-binding lipopeptide in vivo As representative examples, the prodrugs are colistin methanesulfonate and polymyxin B methanesulfonate sodium call.

The term "endotoxinemia" is used herein for all conditions in which clinically relevant amounts of endotoxins are present in the blood of the patient which subsequently result in the induction of cytokines, preferably in conditions such as sepsis and SIRS. The dosing instructions are suitable both for the treatment and the prophylaxis of diseases and conditions caused by endotoxemia. By "prophylactic" is meant administration of the endotoxin-binding lipopeptide when endotoxemia is present but no clinical symptoms are present Prophylactic therapy may be particularly indicated in those patients who are expected to develop endotoxemia due to their disease For example, in patients suffering from acute liver failure or acute decompensation in chronic liver failure, such that in the course of monitoring these patients when endotoxemia is present and even before the onset of clinical symptoms, the endotoxin-binding lipopeptides are administered according to the dosage regimen of the present invention.

In addition, it is known that the use of antibiotics in an infection with Gram-negative bacteria and by the induced by the antibiotic cell lysis leads to increased endotoxin secretion or can come. The invention can therefore be used advantageously as an additional therapeutic or prophylactic measure in the context of a conventional treatment of bacterial infectious diseases by means of antibiotics to intercept the induced by antibiotics endotoxin secretion, which subsequently leads to the induction of cytokines.

The term "parenteral administration" as used herein refers to routes of administration other than enteral and topical administration, and more particularly relates to an infusion, the infusion preferably including, but not limited to, intravenous, subcutaneous, Parenteral administration has the advantage that the lipopeptide serum concentration to be achieved can be rapidly adjusted and maintained both during inital bolus administration and while maintaining serum concentration for a predeterminable period of time Form of intravenous infusion or by infusion into the blood carried in extracorporeal blood circulation as described in detail below.

The invention can be used for both the human and the veterinary medical field. The term "patient" as used herein thus refers to humans and animals. Because of the increased incidence of multiresistant infections, tente strains and the associated need for new forms of therapy, the invention is particularly for the human medical area of high relevance.

Since naturally occurring polymyxins, originally derived from the bacterium Bacillus polymyxa, are among the best-studied peptide antibiotics and have been used for decades in the treatment of diseases and conditions due to endotoxemia, it is preferred that the endotoxin-binding lipoprotein be used. tid is a polymyxin. More preferably, the lipopeptide is selected from the group consisting of the only so far approved for clinical use polymyxins polymyxin B and colistin (polymyxin E). However, most preferred is polymyxin B, as it has been found to be most effective for human medical use. Polymyxin B is preferably used in the form of polymyxin B sulfate.

A further subject of the invention is further a preparation for parenteral administration comprising at least one endotoxin-binding lipopeptide as defined in this disclosure as the active ingredient and optionally at least one pharmaceutically acceptable carrier and / or adjuvant. The preparation may comprise only one type of endotoxin-binding lipopeptide or a mixture of two or more endotoxin-binding lipopeptides, for example a mixture of polymyxin Bl, B2, B3 and B4.

A "pharmaceutically acceptable carrier and / or adjuvant" may be any substance known for the preparation of parenteral dosage forms such as injections, infusion solutions, etc. Formulations suitable for the invention for injection and infusion solutions are given in Example 5 below.

Preferably, the preparation for parenteral administration is in the form of an injection preparation or an infusion preparation. The endotoxin-binding lipopeptide is preferably present in the form of a lyophilized powder for producing a sterile aqueous injection preparation or infusion preparation, wherein the powder can be dissolved, for example, in sterile water, 5% dextrose solution, a Ringer's solution or a physiological sodium chloride solution.

The lipopeptide is in the preparation in dissolved form in step i) for the parenteral administration of a bolus, preferably in a concentration of 5 mg / 1 to 200 mg / l, and in step ii) for maintaining serum concentration preferably at a concentration of 0.04 mg / 1 to 13 mg / 1, more preferably from 0.1 mg / 1 to 7 mg / 1, most preferably 0.5 mg / 1 to 4 mg / 1 before.

Preferably, the serum lipopeptide concentration is in a range of 0.1 μg / ml to 0.6 μg / ml, preferably 0.1 μg / ml to 0.4 μg / ml, most preferably between 0.1 μg / ml to 0.25 μg / ml, since at these serum concentrations an efficient therapy without neuro-and nephrotoxic side effects is feasible even in serious disease processes such as sepsis, severe sepsis or septic shock.

The lipopeptide serum concentration according to the invention in a first step i) as defined in the claims achievable by a single bolus dose of the lipopeptide to obtain the desired serum concentration. In the context of this disclosure, the term "bolus" is thus understood as a single parenteral administration of the endotoxin-binding lipopeptide in the form of a preparation, preferably in the form of an injection or infusion preparation, wherein the formulation for bolus administration preferably has a higher concentration of lipopeptide than that preparation used in step ii) for the subsequent maintenance of lipopeptide serum concentration.

The bolus delivery in step i) is preferably over a period of at least 10 minutes, more preferably at least 60 minutes, most preferably at least 120 minutes, preferably by parenteral injection or infusion, ideally not during a dialysis treatment. To determine the amount of polymyxin required, the patient's blood volume should be considered. The bolus administration (step i)) is preferably by injection, for example, administering a once-off bolus of 10 to 250 ml of the prepared solution for injection at the start of treatment. The bolus administration may also be by infusion.

In a second step ii), this serum concentration of lipopeptides, which is adjusted very rapidly by means of bolus administration, is maintained for a predeterminable time, wherein the maintenance of the serum concentration is preferably carried out by infusion, which preferably takes place continuously. The infusion rate is dependent on the serum half-life for the lipopeptide in the patient. For example, the serum half-life for polymyxin B in patients with normal renal function is typically 13 hours, that is for Colistin depending on the literature at 6 to 7.4 hours. For simultaneous treatment by means of an extracorporeal blood purification method, the clearance of the particular filter used and / or the clearance of an adsorber system for the administered lipopeptide must be taken into consideration, as described in detail below.

Preferably, in step ii), the period of maintaining the lipopeptide serum concentration is the total duration of therapy, that is, as long as there is endotoxemia. This period can last from a few hours to 2 weeks, or even longer, during which time the unwanted induction of cytokines is reduced or suppressed.

A detailed description of how the desired serum concentration can be achieved can be found below in the Examples.

In a variant of the invention, which can be used particularly cost-effectively and nationwide, the maintenance of the lipopeptide serum concentration by intravenous administration takes place in step ii). In this variant, the endotoxin-binding lipopeptide is administered intravenously via a venous access, preferably by means of a metering pump.

In a further variant, in step ii) the maintenance of the lipopeptide serum concentration by infusion into the blood of a patient guided in an extracorporeal blood circulation of an extracorporeal perfusion system takes place at a position downstream of a dialyzer (dialysis filter) assigned to the extracorporeal blood circulation. This variant is indicated as an additional therapeutic measure especially in severe or life-threatening conditions of the patient (eg sepsis), which require intensive care measures in the form of extracorporeal blood and / or plasma cleaning (therapeutic apheresis). The infusion of the lipopeptide takes place via a supply line into the blood which circulates in the extracorporeal blood circulation, at a point downstream of the dialyzer, ie immediately before the blood is returned to the patient. Although direct bolus administration is preferred for the bolus administration administered in step i), bolus delivery downstream of the dialyzer may also be injected into the extracorporeal bloodstream. The bolus dose should preferably be taken before dialysis treatment, continuous infusion simultaneously with dialysis and at a position preferably downstream of the dialyzer.

Since in this variant, the endotoxin-binding lipopeptide is not only degraded by the body, but is also removed via the dialyzer from the blood, it is useful in the dosage of the infused lipopeptide in the blood guided in the extracorporeal blood circulation, the lipopeptide clearance of the body and the Lipopeptide clearance of the dialyzer to be considered.

If a depletion device, for example in the form of an adsorber cartridge or a plasma circuit with adsorbent particles suspended therein, is also associated with the extracorporeal perfusion system, it is favorable if the lipopeptide clearance of a depletion device assigned to the extracorporeal perfusion system is additionally taken into account when metering the infused lipopeptide. For example, it is known that adsorbers of a polystyrene-divinylbenzene copolymer adsorb lipopeptides such as polymyxins in addition to physiologically relevant components such as cytokines, so that the consideration of the lipopeptide clearance of the adsorber is advantageous for the dosage of the infused lipopeptide.

The invention is advantageously used for the treatment of an infection with gram-negative bacteria, in particular for the prophylaxis or for the treatment of a systemic inflammatory reaction (SIRS), sepsis, severe sepsis or septic shock. Representative examples of conditions that can be treated according to the present disclosure are those that occur as a result of infection with Gram-negative bacteria and that can subsequently result in SIRS, sepsis, severe sepsis with multiple organ failure, or septic shock.

Representative examples of Gram-negative bacteria include Escherichia spp, Haemophilus influenzae, Pseudomonas aeruginosa, Pasteurella, Enterobacter spp., Salmonella spp., Shigella spp. The invention is particularly advantageous in gram-negative bacteria, for which an increased occurrence of multidrug-resistant strains has been observed, in which case Pseudomonas aeruginosa should be emphasized as a particularly relevant representative. As already stated above, when antibiotics are used in the case of an infection with Gram-negative bacteria and the cell lysis induced by the administration of antibiotics leads to increased endotoxin release. An increased endotoxin release has been described, for example, for antibiotics which preferentially bind to the PBP-3 ("penicillin-binding protein-3"), for example the frequently used antibiotics of the cephalosporin group such as ceftazidimine therapeutic or prophylactic measure in the context of a conventional treatment of bacterial infectious diseases by means of antibiotics used to intercept the antibiotic-induced endotoxin secretion, which subsequently leads to the induction of cytokines.

In a further aspect, the invention advantageously has the prophylaxis or treatment of an inflammatory response due to acute liver failure or acute decompensation in chronic liver failure, in particular a systemic inflammatory response (SIRS), sepsis, severe sepsis with multiple organ failure or septic shock. In patients with intact liver functions, the endotoxins that pass from the intestine to the bloodstream are eliminated by the reticuloendothelial system (RES) or the Kupffer cells by endocytosis. Patients with chronic liver failure may experience acute decompensation. In this case, endotoxins of the normal intestinal flora can overcome the intestinal barrier and therefore pass unhindered through the liver leading to a systemic inflammatory response (SIRS), sepsis, severe sepsis with multiple organ failure, or septic shock.

The invention further relates to a method for the prophylaxis or treatment of diseases and conditions caused by endotoxemia by administering an endotoxin binding lipopeptide selected from the group consisting of polymyxins, polymyxin derivatives, polymyxin analogs, their prodrugs and their pharmaceutically acceptable Salting, by i) parenterally administering a bolus of the lipopeptide to reach a serum lipopeptide concentration of from 0.01 μg / ml to 0.8 μg / ml and ii) maintaining that lipopeptide serum concentration by parenteral administration of the lipopeptide for a preselected period of time. The above definitions and developments are equally applicable to the method. The invention is explained in more detail below by way of non-limiting examples.

Example V. Endotoxin (LPS) inactivation as a function of polymyxin concentration using endotoxins from E. coli and Pseudomonas aeruginosa.

1.1. aim

The aim of this experiment is to determine the plasma polymyxin B (PMB) concentration-dependent endotoxin activation in the plasma (batch test I). In addition, it will be investigated to what extent this endotoxin inactivation results in an inhibition of cytokine release (batch test II).

1.2. blood donation

9 donor blood collection tubes (9 ml each) are spiked with 5 IU heparin. The plasma is centrifuged off and the cell pellet is incubated on a roller mixer. The plasma is spiked with LPS and used for the batch test I:

1.3. LPS-spike, polymyxin B solutions and batch test I

LPS: Pseudomonas aeruginosa (L-7018 Fa. Sigma Lot: 128K4115, -70 ° C, a 100 μΐ 10-3 g / ml (1 mg / ml))

LPS: E. coli (L-4130 Fa. Sigma Lot: 110M4086M, -70 ° C, a 100 μΐ 10-3 g / ml (1 mg / ml))

The LPS is used in batch with a final concentration of 0.5 ng / ml. The batches are carried out in 3 ml of pyrogen-free glass vials. In batch test I, different PMB concentrations (from Sigma, P-1004) are added in a two-fold mixture and incubated for 60 min at 37 ° C. on the overhead shaker (see Table 2).

In batch test I, PMB concentrations of 0 (without PMB), 10, 100, 250, 500 and 1000 ng / ml are used. For this purpose, sterile PMB solutions (pyrogen-free autoclaved at 121 ° C., 90 min) are prepared with the following concentrations (Table 1): Table 1:

1.4. Endotoxin analysis

Charles River's Limulus Amebocyte Lysate test (LAL) measures endotoxins as EU / ml.

1.5. Cytokine Batch (Bat test II)

The plasma spiked with LPS and PMB is returned after the Bat test I to the cell concentrate obtained from the blood donor in a ratio of 1: 1 (see Table 2). For the cytokine batch the samples from batch test I were used with a PMB concentration of 0 (without PMB), 250, 500 and 1000 ng / ml. As a control, a sample without LPS and with 1000 ng / ml PMB was carried. After the incubation times of 4 h and 12 h at 37 ° C. on the roller mixer (5 revolutions / min), samples are taken, centrifuged off and 50 μΐ plasma frozen at -80 ° C. for later cytokine quantification. The experimental data for the cytokine batch are listed in Table 2.

1.6. Results

Endotoxin Batch (batch test I):

1 shows the inhibition of LPS from E. coli in plasma (original LPS concentration: 0.5 ng / ml) as a function of the PMB concentration (n = 2) after an incubation time of 60 min.

FIG. 2 shows the inhibition of LPS from Pseudomonas aeruginosa in plasma (original LPS concentration: 0.5 ng / ml) as a function of the PMB concentration (n = 2) after an incubation time of 60 min.

The results clearly show that even at a very low PMB concentration in the plasma, i. in a range of 50 to 300 ng / ml (0.05 to 0.3 μg / ml), a strong inhibition of LPS from E. coli and Pseudomonas aeruginosa takes place, with increasing PMB concentration, the LPS inhibition no longer increases significantly , Consequently, even very low levels of PMB are sufficient to inhibit LPS (endotoxins) activity. At these low concentrations, neuro- or nephrotoxic side effects are ruled out.

Cytokine Batch (Batch Test II):

The release of the cytokines TNF-alpha (Figure 3), IL-1beta (Figure 4), IL-6 (Figure 5) and IL-8 (Figure 6) by the blood cells as a function of PMB concentration (without PMB, 250 ng / ml, 500 ng / ml and 1000 ng / ml; 1000 ng / ml control without LPS) in LPS (E.coh ^' ) spiked plasma after 4 hours of incubation is shown in Figs. The results of batch test II clearly show that even at very low PMB concentrations not only a strong inhibition of LPS (see batch test I), but also a strong inhibition of cytokine release takes place. This is particularly pronounced in the inhibition of the key mediator TNF-alpha (FIG. 3). 2. Example 2: Dosage instructions for polymyxin B in direct intravenous administration and examples of formulations for preparations for parenteral administration of polymyxin B (PMB)

2.1. Injection solutions (for bolus administration):

2.1.1. Bolus dose for a PMB serum concentration of 100 ng / ml plasma

Assumption: Patient with 70 kg body weight and 60% of the body weight are distribution volume for PMB -> 42000 ml volume of distribution.

The aim is a PMB serum concentration of 100 ng PMB / ml plasma -> a total of 4.2 mg PMB is required.

Injection solution for a bolus dose over a period of 60 min: 4.2 mg PMB in 100 ml physiological saline = finished solution for bolus administration over a period of 60 min.

2.1.2. Bolus dose for a PMB serum concentration of 250 ng / ml plasma

The target is a PMB serum concentration of 250 ng PMB / ml plasma -> a total of 10.5 mg PMB is required.

Injection solution for a bolus dose over a period of 120 minutes: 10.5 mg PMB in 100 ml physiological saline = ready-to-use solution for bolus administration over a period of 120 min.

2.2. Infusion solutions (for maintaining serum concentration):

Assumption: patient with 70 kg -> distribution volume for PMB (60% of body mass) 42000 ml body fluid with 100 ng PMB / ml -> 4.2 mg PMB in the volume of distribution (see under 2.1.1.). Infusion solution for a 24 h infusion at a serum half-life of 6 h: Assumed half-life for PMB in serum 6 h: 2.1 mg PMB per 6 h or 8.4 mg PMB / day are reduced -> 8.4 mg PMB in 1 L physiological saline = infusion solution for 24 h infusion.

Infusion solution for a 24 h infusion with a serum half-life of 14 h: Half-life for PMB in serum 14 hours: 4.2 mg PMB / 14 h or 7.2 mg PMB / day are degraded -> 7.2 mg PMB in 1 L physiological saline = infusion solution for 24 h infusion.

3. Example 3: Dosing instructions for polymyxin B (PMB) during extracorporeal blood purification (dialysis and adsorption treatment) to maintain an existing PMB serum concentration

The blood purification device according to a known type comprises an extracorporeal blood circulation in which the blood of the patient is guided and a dialyzer (dialysis filter) arranged in the extracorporeal blood circulation. Furthermore, in the illustrated calculation example it is assumed that the blood purification device is assigned an adsorber system, for example in the form of an adsorption cartridge, the adsorber system being located on the blood side in the extracorporeal blood circulation (hemoperfusion) or in a plasma circulation connected to the extracorporeal blood circulation via a plasma filter (plasma or fractionated plasma adsorption ) can be connected.

The following calculation example assumes an already existing serum PMB concentration. This is done by bolus administration before starting the dialysis and adsorption treatment. This can be done under Example 2 / 2.1. described injection solutions are used.

To calculate the dosage, the PMB clearance of the patient's body, the dialyzer and the adsorber system are taken into account:

- The PMB dialysis clearance (CDial) can be determined experimentally and depends on the plasma flow and the type of dialysis filter used. In the example given, this is 60 ml / min. - The PMB clearance of the adsorber (Cads) depends on the adsorber material used as well as on the filtrate flow or, in the case of hemoperfusion, on the blood flow. In the example given, this is 45 ml / min.

- The PMB patient clearance was determined in the example given from the half-life for PMB of 13.6 and is 36 ml / min.

From the individual PMB clearance rates, the PMB total clearance (C total) is obtained by addition. The resulting decrease in PMB is shown in FIG. The negative increase in PMB decrease (Ctotal) at a given time, as seen in Figure 7, corresponds to the necessary PMB infusion to maintain the PMB serum concentration at that time.

For the example given, the following infusion rates result:

- 0.84 mg PMB / h during treatment with dialysis and adsorption

- 0.21 mg PMB / h without dialysis treatment and adsorption

This results in the following amounts of PMB to be infused during 24 hours with extracorporeal treatment over 6 hours:

6 hours treatment with dialysis and adsorption:

18 hours only PMB infusion (without dialysis and adsorption)

Total amount of PMB infused during 24 hours: 9.0 mg

Claims

An endotoxin-binding lipopeptide selected from the group consisting of polymyxins, polymyxin derivatives, polymyxin analogues, their prodrugs and their pharmaceutically acceptable salts for the prophylaxis or treatment of diseases and conditions caused by endotoxemia

i) parenteral administration of a bolus of the lipopeptide to achieve a serum lipopeptide concentration of 0.01 μg / ml to 0.8 μg / ml and

The endotoxin-binding lipopeptide of claim 1, wherein the lipopeptide is a polymyxin.

The endotoxin-binding lipopeptide of claim 2, wherein the lipopeptide is selected from the group consisting of polymyxin B and colistin.

The endotoxin-binding lipopeptide of claim 3, wherein the lipopeptide is polymyxin B.

A preparation for parenteral administration comprising at least one endotoxin-binding lipopeptide according to any one of claims 1 to 4 as an effective ingredient and optionally at least one pharmaceutically acceptable carrier and / or adjuvant for the prophylaxis or treatment of diseases and conditions caused by endotoxemia, by

i) parenteral administration of a bolus of the lipopeptide to achieve a lipopeptide serum concentration of 0.01 ^ g / ml to 0.8 μg / ml and

6. A preparation for parenteral administration according to claim 5 in the form of an injection preparation or an infusion preparation.

7. A preparation for parenteral administration according to claim 5 or 6, wherein the lipopeptide is present in the preparation in dissolved form in i) for the parenteral administration of the bolus in a concentration of 5 mg / 1 to 200 mg / 1 and in ii) for the Maintenance of the serum concentration in a concentration of 0.04 mg / 1 to 13 mg / 1, preferably from 0.1 mg / 1 to 7 mg / 1, most preferably from 0.5 mg / 1 to 4 mg / 1.

8. The endotoxin-binding lipopeptide according to any one of claims 1 to 4 or formulation for parenteral administration according to any one of claims 5 to 7, wherein in i) the period for parenteral administration of the bolus is at least 10 minutes, preferably at least 60 minutes, most preferably at least 120 min.

The endotoxin-binding lipopeptide according to any one of claims 1 to 4 and 8 or preparation for parenteral administration according to any one of claims 5 to 8, wherein the serum lipopeptide concentration is in a range from 0.1 μg / ml to 0.6 μg / ml, preferably 0.1 μg / ml to 0.4 μg / ml, most preferably between 0.1 μg / ml to 0.25 μg / ml.

10. Endotoxin-binding lipopeptide according to any one of claims 1 to 4 and 8 to 9 or preparation for parenteral administration according to any one of claims 5 to 9, wherein in ii) the maintenance of the lipopeptide serum concentration by intravenous administration.

11. The endotoxin-binding lipopeptide according to any one of claims 1 to 4 and 8 to 9 or preparation for parenteral administration according to any one of claims 5 to 9, wherein in ii) the maintenance of the lipopeptide serum concentration by infusion into that in an extracorporeal blood circulation of an extracorporeal perfusion system guided blood of a patient is carried out at a position downstream of a dialyzer associated with the extracorporeal blood circulation.

12. Endotoxin-binding lipopeptide according to claim 11 or preparation for parenteral administration according to claim 11, wherein in the dosage of the infused lipopeptide into the blood carried in the extracorporeal blood circulation, the lipopeptide clearance of the body and the lipopeptide clearance of the dialyzer are taken into account.

13. The endotoxin-binding lipopeptide according to claim 12 or preparation for parenteral administration according to claim 12, wherein the dosage of the infused lipopeptide additionally takes into account the lipopeptide clearance of a depletion device associated with the extracorporeal perfusion system.

14. Endotoxinbindendes lipopeptide according to any one of claims 1 to 4 and 8 to 13 or preparation for parenteral administration according to any one of claims 5 to 13, for the treatment of an infection with gram-negative bacteria, in particular for the prophylaxis or for the treatment of a systemic inflammatory reaction ( SIRS), sepsis, severe sepsis or septic shock.

15. An endotoxin-binding lipopeptide according to any one of claims 1 to 4 and 8 to 13 or preparation for parenteral administration according to any one of claims 5 to 13, for the prophylaxis or treatment of an inflammatory response due to acute liver failure or acute decompensation in chronic liver failure, especially one systemic inflammatory response (SIRS), sepsis, severe sepsis or septic shock.