Treatment of sepsis and septic shock

Abstract

The present invention relates to a composition comprising a mixture of empty liposomes, wherein the mixture of empty liposomes comprises: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin, the composition being useful for treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably sustained hypotension, in animals, preferably humans, for treating hypotension, preferably such sustained hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury in animals, preferably humans, or for treating toxic shock syndrome in animals, preferably humans.

Description

[0001] The present invention relates to a composition comprising a mixture of empty liposomes, wherein the mixture of empty liposomes comprises: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin, the composition being useful for treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, in an animal, preferably a human, for treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury in an animal, preferably a human, or for treating toxic shock syndrome in an animal, preferably a human. In particular, the present invention relates to a composition for treating sepsis or septic shock in an animal, preferably a human. Background Art

[0002] Sepsis is a potentially life-threatening organ dysfunction caused by a dysregulated host response to infection. It is characterized by physiological, pathological, and biochemical abnormalities that lead to organ dysfunction (Levy M et al. Intensive Care Med 2003; 29: 530-38; Singer M et al. JAMA 2016; 315(8): 801-810). Sepsis can lead to tissue damage, multisystem organ failure, and subsequent death (Cohen, 2002, Nature 420, 885-891). Despite maximal care, approximately 30% to 50% of patients with sepsis die. The mechanisms underlying this dysregulated systemic inflammatory response are complex and may involve multiple pathways. Therefore, sepsis cannot be effectively treated with antimicrobial agents alone. In fact, despite the availability of antibiotics, current therapies for the treatment of sepsis have been shown to be ineffective. Neutralization of specific inflammatory cytokines was also ineffective, highlighting the need for new therapies (Wenzel and Edmond, 2012, N Engl J Med 366, 2122-2124).

[0003] Septic shock occurs in a subset of patients with sepsis and is associated with increased mortality. The pathophysiology of septic shock is not precisely understood; it includes underlying circulatory and cellular / metabolic abnormalities severe enough to significantly increase mortality. Patients with septic shock can be identified by a clinical construct of sepsis characterized by: persistent hypotension requiring vasopressors to maintain a mean arterial pressure of 65 mm Hg or greater, despite adequate volume resuscitation; organ hypoperfusion; and serum lactate levels greater than 2 mmol / L (18 mg / dL) (Singer M et al. JAMA 2016; 315(8):801-10).

[0004] Sepsis and septic shock have lasting effects on patients. For example, chronic tissue hypoperfusion can lead to long-term neurological and cognitive sequelae.

[0005] In addition to bacterial infections, viral infections also predispose patients to infections that may cause sepsis or septic shock by increasing susceptibility to bacterial co-infections. For example, influenza patients often show increased susceptibility to co-infection with Streptococcus pneumoniae, and sepsis has been reported to be a common cause of death during influenza pandemics. The detailed mechanisms by which viral infections predispose patients to bacterial infections and subsequently to sepsis are not fully understood, and there is a need for therapies for treating and preventing sepsis and septic shock in patients with viral infections.

[0006] In recent years, customized empty liposomes, such as those composed of cholesterol and / or sphingomyelin, and their use in the treatment of bacterial infections have been described as acting as traps for virulence factors such as bacterial toxins, enzymes, and toxic appendages (WO 2013 / 186286; Henry BD et al., Nat Biotechnol 2015;33(1):81-88; Azeredo da Silveira, S and Perez, A, Expert Rev. Anti Infect. Ther. 2015;13(5):531-533; Azeredo da Silveira, S. and Perez, A. Expert Rev. Anti Infect. Ther. 2017;15:973-975). These tailored empty liposomes have also been described to exhibit antiviral activity and therefore serve as a therapy against viral infections, in particular as a therapy for neutralizing enveloped viruses such as influenza virus (WO 2017 / 216282). Summary of the Invention

[0007] In the first human study conducted on patients with severe pneumonia, the preferred composition of the present invention showed surprising positive results that improved clinical signs and symptoms. In addition, and importantly, it was surprisingly found that the preferred composition of the present invention improved hemodynamic parameters, prevented hemodynamic deterioration and accelerated the resolution of septic shock. Therefore, as revealed by accelerating the normalization of hemodynamic instability, thereby curing patients and accelerating their discharge from the intensive care unit (ICU), the composition of the present invention is effective in treating sepsis and septic shock. Based on this promising efficacy data and its credible mechanism of action, the efficacy in the envisioned further studies of patients is reasonable, and the patients are suspected or diagnosed with infection, regardless of their pathogen, and show signs of complications or severity development, especially those with community-acquired pneumonia, hospital-acquired pneumonia, ventilation-associated pneumonia, intra-abdominal infection, skin and soft tissue infection, urinary tract infection or bacteremia. Further, based on this promising efficacy data and its plausible mechanism of action, the compositions of the present invention are particularly believed to be very beneficial for the treatment and prevention of sepsis or septic shock caused by or associated with bacterial or viral pathogens that use specific lipid microdomains to attack the host and thereby attack animals, preferably human patients.

[0008] Thus, in a first aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) first empty liposomes comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) second empty liposomes comprising sphingomyelin; said composition being for use in treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, in an animal, preferably a human, for treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury in an animal, preferably a human, or for treating toxic shock syndrome in an animal, preferably a human.

[0009] In another aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; the composition is for use in treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, in an animal, preferably a human.

[0010] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition being useful in treating septic shock in an animal, preferably a human.

[0011] Further aspects and embodiments of the invention will become apparent as this description proceeds. BRIEF DESCRIPTION OF THE DRAWINGS

[0012] Figure 1: Evolution of the cardiovascular SOFA score from baseline (before dosing) to day 8 in the entire study population presented as absolute values ​​in the placebo (diamonds), CAL02 low-dose (triangles) and CAL02 high-dose (squares) groups ( Figure 1A ), and the evolution of the cardiovascular SOFA score from baseline (before dosing) to day 8 in the entire study population excluding patients without any hypotensive events (three patients in the CAL02 high-dose group) presented as the difference in score from baseline ( Figure 1B ). *p<0.05

[0013] Figure 2 : Evolution of cardiovascular SOFA scores from baseline (before dosing) to day 8 in patients already in septic shock at baseline, presented as absolute values, in the placebo (diamonds), CAL02 low-dose (triangles), and CAL02 high-dose (squares) groups. DETAILED DESCRIPTION

[0014] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0015] As used herein, the term "about" shall have a meaning of + / - 5%. For example, about 50% shall mean 47.5% to 52.5%. Preferably, as used herein, the term "about" shall have a meaning of + / - 3%. For example, about 50% shall mean 48.5% to 51.5%.

[0016] Unless otherwise indicated, when the term "a" or "an" is used herein, it means "at least one / a kind". Specifically, the term "a" or "an" used in conjunction with the single empty liposome, the first empty liposome, and the second empty liposome to describe the empty liposomes and the mixture of empty liposomes according to the present invention should typically and preferably refer to the single empty liposome and the mixture of empty liposomes including the first empty liposome and the second empty liposome.

[0017] All ranges of values ​​disclosed herein should refer to all values ​​within the range, including the values ​​defining the range. For example, as an illustration, a value of 12 to 13 should refer to the value 12 or 13, or all values ​​between 12 and 13.

[0018] As used herein, the term "empty liposome" refers to a liposome, preferably an artificial liposome, having an average diameter of 20 nm to 10 μm, preferably 20 nm to 500 nm, and further preferably having an average diameter of 20 nm to 400 nm, again further preferably 40 nm to 400 nm or 20 nm to 200 nm, and consisting of one or more phospholipid bilayers, and typically and preferably unilamellar vesicles and multilamellar vesicles, more preferably small unilamellar vesicles (SUVs). In a preferred embodiment, the term "empty liposome" as used herein typically and preferably refers to a liposome that is not incorporated with any drug, typically and preferably refers to a liposome that is not incorporated with any pharmaceutical drug. As used herein and when referring to the empty liposomes of the present invention, "incorporated / Incorporating" typically and preferably means encapsulated in the cavity of the liposome, within the potential bilayer of the liposome, or as part of the membrane layer of the liposome. In another preferred embodiment, the term "empty liposome," as used herein, typically and preferably refers to a liposome according to the invention consisting of sphingomyelin and cholesterol, or consisting of sphingomyelin, and further comprising only water-soluble inorganic compounds and / or water-soluble organic molecules, wherein typically and preferably, the water-soluble inorganic compounds and / or water-soluble organic molecules are derived from the synthesis of the empty liposomes of the invention, and wherein typically and preferably, the water-soluble inorganic compounds are inorganic salts, preferably selected from the group consisting of NaCl, KCl, and MgCl2, and wherein the water-soluble organic molecules are buffers, wherein preferably, the water-soluble organic molecules are selected from the group consisting of glucose and HEPES. Typically and preferably, the water-soluble inorganic compounds and / or water-soluble organic molecules are incorporated into the empty liposomes of the invention due to their presence during the production of the empty liposomes of the invention. In another preferred embodiment, the term "empty liposome," as used herein, typically and preferably refers to a liposome according to the invention consisting of sphingomyelin and cholesterol, or consisting of sphingomyelin, and wherein the empty liposomes do not include an antioxidant.In a further preferred embodiment, the term "empty liposome" as used herein typically and preferably refers to a liposome consisting of sphingomyelin and cholesterol or consisting of sphingomyelin according to the present invention, and further comprises only water-soluble inorganic compounds and / or water-soluble organic molecules, wherein typically and preferably, the water-soluble inorganic compound and / or water-soluble organic molecule are derived from the synthesis of the empty liposomes of the invention, and wherein typically and preferably, the water-soluble inorganic compound is an inorganic salt, which is preferably selected from NaCl, KCl and MgCl2, and wherein the water-soluble organic molecule is a buffer, wherein preferably the water-soluble organic molecule is selected from glucose and HEPES, and wherein the empty liposome consisting of sphingomyelin and cholesterol or consisting of sphingomyelin according to the present invention does not comprise an antioxidant.

[0019] Sepsis or septic shock: Sepsis is a disease with an infectious cause and manifests as the pathology of systemic inflammatory response syndrome (SIRS); it is defined as life-threatening organ dysfunction caused by a dysregulated host response to infection (Levy M et al. Intensive Care Med. 2003; 29: 530-38; Singer M et al. JAMA 2016; 315(8): 801-810). The initial symptoms include chills, sweats, fever, and hypotension. When various inflammatory mediators and coagulation factors increase throughout the body, microcirculation becomes disturbed, leading to a worsening of the pathological state. Septic shock includes abnormal organ perfusion, uncontrolled hypotension, and multi-organ dysfunction, which may lead to death. The clinical structure of sepsis is characterized by: persistent hypotension requiring vasopressors to maintain a MAP ≥ 65 mm Hg despite adequate volume resuscitation; and serum lactate levels > 2 mmol / L (18 mg / dL). Therefore, the term "sepsis" as used herein shall refer to life-threatening organ dysfunction caused by a dysregulated host response to infection, as defined and suggested in Singer M et al. JAMA. 2016;315(8):801-810 (recommendations; Box 3). Organ dysfunction can be identified as an acute change in the total SOFA (Sequential [Sepsis-related] Organ Failure Assessment) score ≥2 points after infection. For patients known not to have pre-existing organ dysfunction, the baseline SOFA score can be assumed to be zero. A SOFA score ≥2 reflects an overall mortality risk of approximately 10% in a general hospital population with suspected infection. Even patients presenting with moderate dysfunction may deteriorate further, highlighting the severity of this condition and the need for timely and appropriate intervention, if such intervention has not already been implemented. Patients with suspected infection who are at risk of prolonged ICU stay or death in the hospital can be rapidly identified as more likely to have adverse outcomes typical of sepsis if they have at least two of the following clinical criteria that collectively constitute a new bedside clinical score, called quick SOFA (qSOFA): respiratory rate of 22 / min or higher; altered mental activity, i.e., altered mental status; or systolic blood pressure of 100 mm Hg or less. As used herein and as defined and suggested by Singer M et al. JAMA. 2016;315(8):801-810 (recommendations; Box 3), the term "septic shock" is a subtype of sepsis in which the underlying circulatory and cellular / metabolic abnormalities are severe enough to significantly increase mortality, and thus, the risk of death associated with septic shock is higher than that associated with sepsis alone.Patients with septic shock can be identified using the clinical construct of sepsis with the following characteristics: persistent hypotension requiring vasopressors to maintain a mean arterial pressure (MAP) ≥ 65 mm Hg, despite adequate volume resuscitation (and therefore in the absence of hypovolemia); and serum lactate levels > 2 mmol / L (18 mg / dL). In the presence of these criteria, hospital mortality exceeds 40%.

[0020] Animal: As used herein, the term "animal" refers to a living, multicellular vertebrate organism, a class that includes, for example, mammals and birds. The term mammal includes humans and non-human mammals. Similarly, the term "subject" includes humans and livestock subjects.

[0021] As used herein, the terms "treating," "treatment," or "therapy" refer to means for achieving a desired physiological effect. An effect can be therapeutic in terms of partially or completely curing a disease or condition and / or symptoms attributable to the disease or condition, including ameliorating signs or symptoms of a disease, such as sepsis or septic shock, or a pathological condition associated with the disease, such as reducing fever in a subject with septic shock or stabilizing the subject's blood pressure or improving symptoms such as, but not limited to, chills, sweating, or increasing organ function. As used herein in its broadest sense, the term "treatment" shall include and refer to "prevention" of a disease. "Preventing" or "prevention" of a disease refers to inhibiting a disease or condition, such as sepsis or septic shock, i.e., arresting the partial or complete development of a disease, such as sepsis or septic shock, in a person suffering from or at risk of a bacterial infection. Thus, as used herein, the term "treatment," for purposes of defining and characterizing preferred aspects and embodiments of the present invention, shall exclude and shall not refer to "prevention" of a disease in its preferred sense. Furthermore, in other embodiments and aspects of the present invention for preventing sepsis or septic shock, particularly in animals, preferably humans at risk of sepsis or septic shock, the compositions used and the methods of the present invention are then used to delay or prevent the development of sepsis or septic shock. Thus, the methods comprise: selecting a human patient at risk of sepsis or septic shock, typically and preferably caused by an infection; and administering to the human patient one or more of the compositions disclosed herein. The human patient can be, for example, intubated, i.e., a person under invasive mechanical ventilation, or a person who has been exposed to specific bacteria such as Streptococcus pneumoniae or Staphylococcus aureus.

[0022] Singer et al., in JAMA 2016, well described patients with possible sepsis: Any two of three clinical variables—Glasgow Coma Scale score of 13 or less; systolic blood pressure of 100 mm Hg or less; and respiratory rate of 22 / min or more—provided predictive validity similar to that of the full SOFA score outside the ICU. Based on external US and non-US datasets, this model was shown to be robust to multiple sensitivity analyses that included a simpler assessment of altered mental status (Glasgow Coma Scale score <15) performed in out-of-hospital, emergency department, and ward settings. For patients suspected of infection in the ICU, the SOFA score had superior predictive validity to this model, which may reflect the moderating effects of interventions (eg, vasopressors, sedatives, mechanical ventilation).

[0023] A "therapeutically effective amount" is the amount of a composition that achieves the desired effect in the subject being treated. For example, this may be the amount necessary to inhibit septic shock, reduce fever, or prevent multiple organ failure in an animal, preferably a human patient, such as a patient with pneumonia and / or a patient infected with Streptococcus pneumoniae. When administered to an animal, preferably a human patient, a dosage that will achieve an effective target tissue concentration will generally be used.

[0024] As used herein, "therapeutic dose" refers to a dose known to those skilled in the art to have a therapeutic effect.

[0025] The term "pneumonia" as used herein shall encompass "community-acquired pneumonia" (CAP), "hospital-acquired pneumonia" (HAP), or "ventilator-associated pneumonia" (VAP).

[0026] The term "community-acquired pneumonia" or "CAP" is known to those skilled in the art, see, for example, IDSA / ATS Guidelines for CAP in Adults (CID 2007:44(Suppl 2)S27). Specifically, the term refers to pneumonia acquired outside a hospital.

[0027] The term "hospital-acquired pneumonia (HAP)" refers to pneumonia acquired during or after hospitalization for another illness or surgery, where the onset occurs at least 48 to 72 hours after admission.

[0028] As defined herein, "ventilator-associated pneumonia (VAP)" is pneumonia that develops 48 hours or more after mechanical ventilation and is characterized by microbial invasion of the lower respiratory tract and lung parenchyma. VAP is a potentially serious medical condition.

[0029] Pneumonia is caused by infection with a variety of microorganisms, including bacteria such as Streptococcus pneumoniae, Haemophilus influenzae, Legionella pneumophilia, Staphylococcus aureus, and Pseudomonas aeruginosa for CAP, and Gram-negative bacilli such as Pseudomonas aeruginosa and Serratia marcescens, and Staphylococcus aureus, Klebsiella pneumoniae, Escherichia coli, Stenotrophomonas maltophilia, Acinetobacter species, and Haemophilus influenzae for HAP (see Cilloniz et al., Thorax. 2011 Apr;66(4):340-6 and Jones RN. Clin Infect. Dis. 2010 Aug;51(Suppl 1):S81-7).

[0030] The term "severe community-acquired pneumonia" or "sCAP" is known to those skilled in the art. Specifically, the term "severe community-acquired pneumonia" or "sCAP" refers to a subgroup of patients with community-acquired pneumonia who require intensive care. The Infectious Diseases Society of America (IDSA) and the American Thoracic Society (ATS) have published guidelines on the management of CAP, including a definition of sCAP (see Mandell et al., 2007, Infectious Diseases Society of America / American Thoracic Society Consensus Guidelines on the Management of Community-Acquired Pneumonia in Adults, Clin. Inf. Dis. 2007: 44: S27-72 (Suppl. 2), Table 4). According to the IDSA / ATS guidelines, sCAP is defined as CAP requiring intensive care. If a patient with CAP exhibits one or two of the two major criteria from the list described in Example 1 and implemented in the presented study, or if there are three minor criteria from the list, admission to an intensive care unit is recommended.

[0031] As used herein, the term "for" used in "a composition for treating a disease" shall also disclose the corresponding method of treatment and the corresponding use for preparing a formulation for preparing a medicament for treating a disease.

[0032] In one aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition being useful for treating septic shock in an animal, preferably a human.

[0033] In another aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition is for treating hypotension, preferably sustained hypotension, in an animal, preferably a human, wherein the hypotension, preferably the sustained hypotension is associated with septic shock.

[0034] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; the composition is for use in treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, in an animal, preferably a human, for treating hypotension, preferably persistent hypotension in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury in an animal, preferably a human, or for treating toxic shock syndrome in an animal, preferably a human.

[0035] In another aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; the composition is for use in treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, in an animal, preferably a human.

[0036] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition being useful for treating sepsis or severe sepsis in an animal, preferably a human.

[0037] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition being useful in treating septic shock in an animal, preferably a human.

[0038] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition is for use in treating long-term and severe hypotension, preferably persistent hypotension, in an animal, preferably a human.

[0039] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition being useful for treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury in an animal, preferably a human.

[0040] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) first empty liposomes comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) second empty liposomes comprising sphingomyelin; said composition being for use in treating persistent hypotension in septic shock in an animal, preferably a human.

[0041] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition being for use in treating hypotension, preferably sustained hypotension, in septic shock in an animal, preferably a human.

[0042] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) first empty liposomes comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) second empty liposomes comprising sphingomyelin; said composition being for use in treating persistent hypotension in septic shock in a human.

[0043] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) first empty liposomes comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) second empty liposomes comprising sphingomyelin; said composition being for use in treating hypotension, preferably persistent hypotension, in sepsis or severe sepsis in an animal, preferably a human.

[0044] In a further aspect, the present invention provides a composition comprising, preferably consisting of, a mixture of empty liposomes, wherein the mixture of empty liposomes comprises, preferably consists of: (a) a first empty liposome comprising cholesterol, wherein the amount of cholesterol is at least 30% (weight / weight); and (b) a second empty liposome comprising sphingomyelin; said composition being useful for treating toxic shock syndrome in an animal, preferably a human.

[0045] In a further aspect and very preferred embodiment, the present invention provides a composition of the invention for use in treating hypotension, preferably persistent hypotension, in said animal, preferably said human, suffering from sepsis. In a further aspect and very preferred embodiment, the present invention provides a composition of the invention for use in treating hypotension, preferably persistent hypotension, in said human, suffering from sepsis. In a further aspect and very preferred embodiment, the present invention provides a composition of the invention for use in treating persistent hypotension in said human, suffering from sepsis.

[0046] In a further aspect and very preferred embodiment, the present invention provides a composition of the invention for use in treating hypotension, preferably persistent hypotension, in the animal, preferably the human, suffering from septic shock. In a further aspect and very preferred embodiment, the present invention provides a composition of the invention for use in treating hypotension, preferably persistent hypotension, in the human, suffering from septic shock. In a further aspect and very preferred embodiment, the present invention provides a composition of the invention for use in treating persistent hypotension in the human, suffering from septic shock.

[0047] In another aspect, the present invention provides a method of treating sepsis or septic shock, preferably septic shock, in an animal, preferably a human, in need thereof, the method comprising administering a therapeutically effective amount of a composition as defined in the appended claims. Preferably, the human patient suffers from pneumococcal pneumonia, or the sepsis or septic shock, preferably septic shock, is caused by pneumococcal pneumonia.

[0048] For all aspects and embodiments disclosed herein, a therapeutically effective amount of the composition of the invention is typically and preferably used in the disclosed treatment.

[0049] Furthermore, all embodiments and preferred embodiments disclosed herein are to be understood as embodiments and preferred embodiments of any and all aspects of the present invention.

[0050] The first study compared a preferred composition of the present invention plus standard antibiotic therapy with placebo plus standard antibiotic therapy in adult patients admitted to the intensive care unit (ICU) for severe community-acquired pneumococcal pneumonia. Two different doses of the preferred composition of the present invention were compared: a low dose (4 mg / kg - low dose) and a high dose (16 mg / kg - high dose). The results showed a synergistic effect of the preferred composition of the present invention (designated CAL02) with antibiotic treatment.

[0051] As a result, the preferred compositions of the present invention thus capture and neutralize toxins released from a variety of bacteria associated with serious infections in a manner that synergizes with antibiotic treatment in human patients. Preferred compositions of the present invention work regardless of the resistance profile of the target pathogen and do not induce the emergence of resistance.

[0052] In a preferred embodiment, the second empty liposome (b) comprises the sphingomyelin as the only lipid component. In a preferred embodiment, the second empty liposome (b) consists of sphingomyelin.

[0053] In a preferred embodiment, the amount of cholesterol in the first empty liposome (a) is 30% to 70% (w / w), and preferably the amount of cholesterol in the first empty liposome (a) is 35% to 60% (w / w).

[0054] In a preferred embodiment, the amount of cholesterol in the empty liposomes (a) is 45% to 55% (w / w), and preferably the amount of cholesterol in the empty liposomes (a) is about 50% (w / w).

[0055] In a preferred embodiment, the first empty liposome (a) consists of cholesterol and sphingomyelin, and wherein preferably the amount of cholesterol in the empty liposome (a) is 45% to 55% (weight / weight), and wherein further preferably the amount of cholesterol in the empty liposome (a) is about 50% (weight / weight).

[0056] In a preferred embodiment, the first empty liposome (a) comprises the cholesterol and the sphingomyelin as the only lipid components.

[0057] In a preferred embodiment, the amount of cholesterol in the empty liposomes (a) is 45% to 55% (weight / weight), and wherein preferably the amount of cholesterol in the empty liposomes (a) is about 50% (weight / weight), and wherein the second empty liposomes (b) are composed of sphingomyelin.

[0058] In a preferred embodiment, the first empty liposome (a) consists of cholesterol and sphingomyelin, and wherein the amount of cholesterol in the empty liposome (a) is 45% to 55% (weight / weight), and wherein preferably the amount of cholesterol in the empty liposome (a) is about 50% (weight / weight), and wherein the second empty liposome (b) consists of sphingomyelin.

[0059] In a preferred embodiment, the first empty liposome (a) consists of cholesterol and sphingomyelin, and wherein the amount of cholesterol of the empty liposome (a) is 45% to 55% (weight / weight), and wherein preferably the amount of cholesterol of the empty liposome (a) is about 50% (weight / weight), and wherein the first empty liposome (a) comprises said cholesterol and said sphingomyelin as the only lipid components, and wherein the second empty liposome (b) consists of sphingomyelin.

[0060] In a preferred embodiment, the mixture of empty liposomes comprises at least 20% (weight / weight) of the first empty liposomes (a) and the second empty liposomes (b), and wherein preferably the mixture of empty liposomes comprises at least 30% (weight / weight) of the first empty liposomes (a) and the second empty liposomes (b).

[0061] In a preferred embodiment, the mixture of empty liposomes comprises at least 40% (weight / weight) of the first empty liposomes (a) and the second empty liposomes (b).

[0062] In a preferred embodiment, the first empty liposomes (a) consist of cholesterol and sphingomyelin, and wherein further preferably the amount of cholesterol in the empty liposomes (a) is about 50% (w / w), and wherein the mixture of empty liposomes comprises at least 40%, preferably at least 45% (w / w) of the first empty liposomes (a) and the second empty liposomes (b).

[0063] In a preferred embodiment, the first empty liposomes (a) consist of a 1:1 (weight / weight - w / w) mixture of the first empty liposomes and the second liposomes, wherein the first empty liposomes are composed of cholesterol and sphingomyelin in a 1:1 weight ratio (1:1 w / w; molar ratio 35:65) and the second empty liposomes are composed only of sphingomyelin.

[0064] In a preferred embodiment, the first empty liposomes (a) consist of a 1:1 (weight / weight - w / w) mixture of the first empty liposomes and the second empty liposomes, wherein the first empty liposomes are composed of cholesterol and sphingomyelin in a 1:1 weight ratio (1:1 w / w; molar ratio 35:65), and the second empty liposomes are composed only of sphingomyelin, and wherein the first empty liposomes (a) include the cholesterol and the sphingomyelin as the only lipid components, and the second empty liposomes (b) include the sphingomyelin as the only lipid component.

[0065] In a preferred embodiment, the average diameter of the first empty liposomes is about 130 nm, and the average diameter of the second empty liposomes is about 90 nm.

[0066] In a preferred embodiment, the composition is for use in treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury, preferably for treating hypotension, preferably persistent hypotension, in septic shock, wherein the hypotension, preferably the persistent hypotension, is associated with a systolic blood pressure <90 mm Hg or a mean arterial pressure <70 mm Hg.

[0067] In a preferred embodiment, the composition is for use in treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury, preferably for treating hypotension, preferably persistent hypotension, in septic shock, wherein the hypotension, preferably the persistent hypotension, is associated with a systolic blood pressure < 90 mm Hg.

[0068] In a preferred embodiment, the composition is for use in treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury, preferably for treating hypotension, preferably persistent hypotension, in septic shock, wherein the hypotension, preferably persistent hypotension, is associated with a mean arterial pressure <70 mm Hg.

[0069] In a preferred embodiment, the composition is for use in treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury, preferably for treating hypotension, preferably persistent hypotension, in septic shock, wherein the hypotension, preferably persistent hypotension, is associated with a systolic blood pressure <90 mm Hg or a mean arterial pressure <70 mm Hg, and wherein the hypotension, preferably persistent hypotension, is pretreated with a vasopressor for at least 2 hours.

[0070] In a preferred embodiment, the composition is for use in treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury, preferably for treating hypotension, preferably persistent hypotension, in septic shock, wherein the hypotension is associated with a systolic blood pressure <90 mm Hg or a mean arterial pressure <70 mm Hg, and wherein the hypotension, preferably the persistent hypotension, is pretreated with vasopressors for at least 2 hours after fluid resuscitation.

[0071] In a preferred embodiment, the composition is for use in treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury, preferably for treating hypotension, preferably said persistent hypotension, in septic shock, wherein the hypotension, preferably the persistent hypotension is associated with a systolic blood pressure <90 mm Hg or a mean arterial pressure <70 mm Hg, and wherein the hypotension, preferably the persistent hypotension is pretreated with at least one, preferably one, therapeutic dose of vasopressor for at least 2 hours.

[0072] In a preferred embodiment, the composition is for treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury, preferably for treating hypotension, preferably persistent hypotension, in septic shock, wherein the hypotension, preferably the persistent hypotension is associated with a systolic blood pressure <90 mm Hg or a mean arterial pressure <70 mm Hg, and wherein the hypotension, preferably the persistent hypotension is pretreated with at least one, preferably one therapeutic dose of a vasopressor for at least 2 hours, wherein the vasopressor is selected from dopamine, epinephrine, norepinephrine, phenylephrine or vasopressin.

[0073] In a preferred embodiment, the composition is for treating hypotension, preferably persistent hypotension, in septic shock, sepsis, severe sepsis, acute respiratory distress syndrome or acute lung injury, preferably for treating hypotension, preferably persistent hypotension in septic shock, wherein the hypotension, preferably the persistent hypotension is associated with a systolic blood pressure <90 mm Hg or a mean arterial pressure <70 mm Hg, and wherein the hypotension, preferably the persistent hypotension is pretreated with at least one, preferably one, therapeutic dose of a vasopressor for at least 2 hours, wherein the therapeutic dose of the vasopressor is dopamine >5 mg / kg / min or a dose corresponding thereto of the vasopressor, preferably a dose corresponding thereto of epinephrine, norepinephrine, phenylephrine or vasopressin.

[0074] In a preferred embodiment, the composition is used to treat and prevent septic shock. In a preferred embodiment, the composition is used to treat hypotension, preferably persistent hypotension. In a preferred embodiment, the hypotension, preferably persistent hypotension, is associated with septic shock. In a preferred embodiment, the composition is used to treat persistent hypotension. In a preferred embodiment, the persistent hypotension is associated with septic shock.

[0075] In preferred embodiments, the hypotension, preferably sustained hypotension, is associated with a systolic blood pressure <90 mm Hg (or a mean arterial pressure <70 mm Hg). In preferred embodiments, the hypotension, preferably sustained hypotension, is associated with a systolic blood pressure <90 mm Hg (or a mean arterial pressure <70 mm Hg) despite treatment with therapeutic doses of vasopressors (i.e., dopamine >5 mg / kg / min or any dose of epinephrine, norepinephrine, phenylephrine, or vasopressin) for at least 2 hours after adequate fluid resuscitation.

[0076] In a preferred embodiment, the treatment is an adjunct to antibiotic therapy, preferably an adjunct to standard antibiotic therapy. In a preferred embodiment, the antibiotic therapy, preferably the antibiotic of the standard antibiotic therapy, is selected from the group consisting of: ceftriaxone, spiramycin, amoxicillin, amoxicillin / clavulanic acid, gentamicin, piperacillin / tazobactam, cefuroxime, penicillin, azithromycin, clarithromycin, erythromycin, doxycycline, cefotaxime, ampicillin, ertapenem, cefepime, imipenem, meropenem, ciprofloxacin, levofloxacin, vancomycin, linezolid, moxifloxacin and gemifloxacin, and wherein preferably the antibiotic therapy, preferably the antibiotic of the standard antibiotic therapy, is selected from the group consisting of: ceftriaxone, spiramycin, amoxicillin, gentamicin, levofloxacin, piperacillin / tazobactam, amoxicillin / clavulanic acid, cefuroxime and penicillin. In preferred embodiments, the antibiotic therapy, preferably standard antibiotic therapy, is intravenous (IV) or oral antibiotic therapy, preferably standard antibiotic therapy.

[0077] In a preferred embodiment, the human patient suffers from pneumonia, wherein preferably the pneumonia is selected from community-acquired pneumonia (CAP), hospital-acquired pneumonia (HAP) and ventilator-associated pneumonia (VAP). In a preferred embodiment, the human patient suffers from pneumonia, wherein the pneumonia is community-acquired pneumonia (CAP). In a preferred embodiment, the human patient suffers from pneumonia, wherein the pneumonia is hospital-acquired pneumonia (HAP). In a preferred embodiment, the human patient suffers from pneumonia, wherein the pneumonia is ventilator-associated pneumonia (VAP). In a preferred embodiment, the human patient suffers from pneumonia, wherein the pneumonia is severe pneumonia, preferably severe community-acquired pneumonia (sCAP) or severe community-acquired pneumococcal pneumonia (sCAPP). In a preferred embodiment, the human patient suffers from pneumonia, wherein the pneumonia is community-acquired pneumonia (CAP) or community-acquired pneumococcal pneumonia (CAPP).

[0078] In preferred embodiments, the human patient has pneumonia, and wherein the pneumonia is caused by Streptococcus pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Enterococcus faecium, Legionella pneumophila, Haemophilus influenzae, Klebsiella pneumoniae, Escherichia coli, Acinetobacter baumanii, Bordetella pertussis, Serratia marcescens, Stenotrophomonas maltophilia, Moraxella catarrhalis, or Mycobacterium tuberculosis. In a preferred embodiment, the human patient suffers from pneumonia, and wherein the pneumonia is a severe pneumonia caused by Streptococcus pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Enterococcus faecalis, Legionella pneumophila, Haemophilus influenzae, Klebsiella pneumoniae, Escherichia coli, Acinetobacter baumannii, Bordetella pertussis, Serratia marcescens, Stenotrophomonas maltophilia, Moraxella catarrhalis or Mycobacterium tuberculosis, and wherein preferably the pneumonia, preferably the severe pneumonia is caused by Streptococcus pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Enterococcus faecalis, Haemophilus influenzae, Klebsiella pneumoniae, Escherichia coli, Acinetobacter baumannii, Bordetella pertussis, Serratia marcescens or Mycobacterium tuberculosis, and wherein further preferably the pneumonia, preferably the severe pneumonia is caused by Streptococcus pneumoniae, Staphylococcus aureus, Klebsiella pneumoniae or Pseudomonas aeruginosa, and wherein again further preferably the pneumonia or the severe pneumonia is caused by Streptococcus pneumoniae.

[0079] In a preferred embodiment, the composition is in the form of a solution for intravenous administration, preferably intravenous infusion, comprising between 10 and 40 grams of the mixture of empty liposomes per liter of solution, preferably between 10 and 20 grams of the mixture of empty liposomes per liter of solution.

[0080] In a preferred embodiment, the infusion time of said intravenous administration is up to 3 hours, and wherein preferably the infusion time of said intravenous administration is from 10 minutes to 2 hours.

[0081] In a preferred embodiment, the composition is administered in at least 2 doses. In a preferred embodiment, the composition is administered in at least 2 doses: a first dose and a second dose, and wherein the interval between the first dose and the second dose is 6 hours to 96 hours, preferably 12 hours to 72 hours, further preferably 24 hours to 48 hours, and even further preferably 24 hours or 48 hours.

[0082] In a preferred embodiment, the composition is administered in 2 to 4 doses, preferably in two doses within 12 to 72 hours, further preferably in two doses within 24 to 48 hours, preferably with an interval of 24 hours or 48 hours.

[0083] In a preferred embodiment, the composition is administered to the human patient in at least 2 doses: a first dose and a second dose, and wherein the interval between the first dose and the second dose is 20 hours to 28 hours, preferably 24 hours.

[0084] In a preferred embodiment, the composition is administered to the human patient in two doses: a first dose and a second dose, wherein the interval between the administration of the first dose and the administration of the second dose is 20 hours to 28 hours, preferably 24 hours.

[0085] In a preferred embodiment, each of the doses is 1 mg / kg to 64 mg / kg, preferably 2 mg / kg to 32 mg / kg, further preferably 3 mg / kg to 25 mg / kg, and even further preferably 4 mg / kg to 16 mg / kg.

[0086] In a preferred embodiment, each of the doses is 2 mg / kg to 8 mg / kg, preferably 2 mg / kg to 6 mg / kg, further preferably 3 mg / kg to 5 mg / kg, and even further preferably 4 mg / kg.

[0087] In a preferred embodiment, each of the doses is 10 mg / kg to 22 mg / kg, preferably 12 mg / kg to 20 mg / kg, further preferably 14 mg / kg to 18 mg / kg, and even further preferably 16 mg / kg.

[0088] In a preferred embodiment, the composition is administered in the form of a solution for intravenous administration.

[0089] In a preferred embodiment, said composition is administered to said human patient in preferably at least 2 doses: a first dose and a second dose, wherein the interval between said administration of said first dose and said administration of said second dose is 20 hours to 48 hours, preferably 24 hours, and wherein said composition is in the form of a solution for intravenous administration.

[0090] In a preferred embodiment, the composition is for treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, and wherein preferably the composition is for treating septic shock in an animal, preferably a human, and wherein the sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, preferably septic shock requires hospitalization.

[0091] In a preferred embodiment, the composition is used to treat sepsis, septic shock or hypotension, preferably persistent hypotension, in an animal, preferably a human.

[0092] In a preferred embodiment, the composition is used to treat sepsis, septic shock, or hypotension, preferably persistent hypotension, in a human. In a very preferred embodiment, the composition is used to treat sepsis in a human. In a very preferred embodiment, the composition is used to treat septic shock in a human. In a very preferred embodiment, the composition is used to treat hypotension, preferably persistent hypotension, in a human. In a preferred embodiment, the sepsis, septic shock, or hypotension, preferably persistent hypotension, requires hospitalization of the human, preferably in an intensive care unit (ICU) of a hospital.

[0093] In a preferred embodiment, the composition is for treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, and wherein preferably the composition is for treating septic shock in an animal, preferably a human, and wherein the sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, preferably septic shock requires admission to an intensive care unit (ICU), preferably to an intensive care unit (ICU) in a hospital.

[0094] In a preferred embodiment, the composition is for use in the treatment of sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, and wherein preferably the composition is for use in the treatment of septic shock in an animal, preferably a human, and wherein the sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, preferably septic shock requires hospitalization, and wherein the treatment reduces the length of stay in the hospital compared to the length of stay in the hospital when no such treatment is administered.

[0095] In a preferred embodiment, the reduction in hospital stay due to the treatment is at least one day, preferably two days, more preferably three days, even more preferably four days, even more preferably five days, even more preferably six days, even more preferably seven days, even more preferably eight days, even more preferably nine days. In a preferred embodiment, the hospital stay is at most 18 days.

[0096] In a preferred embodiment, the composition is for use in the treatment of sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, and wherein preferably the composition is for use in the treatment of septic shock in an animal, preferably a human, and wherein the sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, preferably septic shock requires admission to an intensive care unit (ICU), preferably to an intensive care unit (ICU) of a hospital, and wherein the treatment reduces the length of stay in the intensive care unit (ICU) compared to the length of stay in the intensive care unit (ICU) when such treatment is not administered.

[0097] In a preferred embodiment, the reduction in the length of stay in the intensive care unit (ICU) is at least one day, preferably two days, more preferably three days, even more preferably four days, even more preferably five days, even more preferably six days, even more preferably seven days. In a preferred embodiment, the length of stay in the intensive care unit (ICU) is at most 18 days.

[0098] In a preferred embodiment, the composition is for use in treating sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, and wherein preferably the composition is for use in treating septic shock in an animal, preferably a human, and wherein the sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, preferably septic shock is cured in less time than when no such treatment is administered.

[0099] In a preferred embodiment, the healing time is less than at least one day, preferably two days or further preferably at least three days, further preferably at least four days, further preferably at least five days, further preferably at least six days, further preferably at least seven days.

[0100] In a preferred embodiment, the composition is for use in the treatment of sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, and wherein preferably the composition is for use in the treatment of septic shock in an animal, preferably a human, and wherein the treatment reduces the cardiovascular SOFA score compared to the cardiovascular SOFA score in the absence of such treatment.

[0101] In a preferred embodiment, the composition is for use in the treatment of sepsis, severe sepsis, septic shock or prolonged and severe hypotension, preferably persistent hypotension, and wherein preferably the composition is for use in the treatment of septic shock in an animal, preferably a human, and wherein the treatment reduces the cardiovascular SOFA score compared to the cardiovascular SOFA score in the absence of such treatment, and wherein the reduction is at least 50%, preferably at least 60%, further preferably at least 70%, yet further preferably at least 80% 7 days after the start of the treatment, preferably 6 days after the start of the treatment, further preferably after 5 days.

[0102] Examples

[0103] Liposomes:

[0104] Egg yolk sphingomyelin (CAS No. 85187-10-6) was purchased from Sigma (S0756), Avanti Polar Lipids (860061), or Lipoid GmbH. Cholesterol from sheep lanolin (CAS No. 57-88-5) was purchased from Sigma (C-8667), Avanti Polar Lipids (70000), or Dishman Netherlands B.V. According to the present invention, the sphingomyelin and cholesterol included in or consisting of the mixture of the empty liposomes of the present invention can be obtained from natural sources as described above, or alternatively, can be obtained by chemical synthesis.

[0105] Liposome preparation:

[0106] Sphingomyelin monolayers: Cholesterol (35:65 molar ratio) and sphingomyelin only (100%) liposomes were prepared using sonication or microfluidization (eg, high pressure homogenization) or according to a hydration, extrusion, and diafiltration protocol.

[0107] Ultrasound:

[0108] Lipids were dissolved in chloroform at a concentration of 1 mg / ml and stored at -20°C. To prepare liposomes, the chloroform solutions of each lipid were mixed in proportion as needed to routinely produce a final solution of 50 μl to 500 μl. Chloroform was completely evaporated at 60°C for 20 to 50 minutes. 50 μl or 100 μl of Tyrode's buffer (140 mM NaCl, 5 mM KCl, 1 mM MgCl2, 10 mM glucose, 10 mM HEPES; pH = 7.4) containing 2.5 mM CaCl2 was added to the tube containing the dried lipid film and vortexed vigorously. The lipid suspension was incubated at 45°C for 20 to 30 minutes under vigorous shaking in an Eppendorf thermomixer. To produce liposomes, the final lipid suspension was sonicated 3 × 5 seconds at 6°C in a Bandelin Sonopuls sonicator at 70% power. Liposome preparations were kept at 6°C for at least 1 hour before use in experiments.

[0109] Hydration, extrusion and filtration process procedures:

[0110] In an alternative method, each liposome formulation is prepared by an ethanol hydration and extrusion method. The lipids are dissolved in ethanol and tert-butanol, respectively, while mixing at high temperature (~55°C). The lipid solution is then added to a PBS buffer solution (sodium chloride, monosodium phosphate dihydrate, and disodium phosphate dihydrate dissolved in water for injection while mixing, adjusted to a pH of 7.0 to 7.4 with hydrochloric acid (HCl) or sodium hydroxide (NaOH) as needed, and filtered through a 0.2 μm filter) while mixing at high temperature (~65°C) for approximately 30 minutes. The resulting process fluid is then repeatedly extruded through a series of polycarbonate track-etched membranes at high pressure and temperature (~65°C) until the desired particle size is reached as measured by dynamic light scattering (Example of an extruder: Extruder). The resulting process fluid is then concentrated approximately 2-fold using a hollow fiber filter cartridge with a molecular weight cutoff of 100,000 and then diafiltered against approximately 10 volume exchanges of PBS buffer to remove ethanol and n-butanol. At the end of the diafiltration, the process fluid is concentrated by approximately 30% to allow subsequent dilution to the target lipid concentration. Prior to dilution, the process fluid is filtered through a 0.2 μm sterilizing grade filter to remove any larger liposomes that may clog the filter during sterile filtration. The process fluid is then diluted with PBS buffer to a target of 40 mg / mL total lipid. The final formulation is sterile filtered through two 0.2 μm sterilizing grade filters in series and aseptically filled into glass vials.

[0111] The concentrations of individual lipids in the liposomes are always given as weight / weight ratios. In liposomes containing sphingomyelin and cholesterol, a 1:1 (weight / weight) ratio corresponds to a 50% (weight / weight) or 35:65 molar ratio. The specifications are described in Table 1.

[0112] Table 1: Liposome specifications

[0113] Average diameter (nm) Polydispersity index Zeta potential (mV) Osmolarity (mmol / kg) pH 40 to 400 <0.45 -25 to +2 250-400 6.5-8.0

[0114] CURB-65 Rating:

[0115] The CURB-65 (also known as the CURB criteria) is a clinical prediction rule well known to those skilled in the art that has been validated to predict mortality in community-acquired pneumonia (Lim WS et al. (2003), Thorax 58(5):377–82). The British Thoracic Society recommends the CURB-65 for use in assessing the severity of pneumonia (British Thoracic Society Standards of Care Committee (2001). "BTS Guidelines for the Management of Community Acquired Pneumonia in Adults". Thorax. 56. Suppl 4: IV1–64).

[0116] Score is an acronym for each of the risk factors measured. Each risk factor is scored one point, with a maximum score of 5:

[0117] -New onset confusion (defined as 8 or fewer AMTS)

[0118] -Blood urea nitrogen greater than 7 mmol / L (19 mg / dL)

[0119] - A respiratory rate of 30 breaths per minute or more

[0120] - Blood pressure less than 90 mm Hg, with systolic or diastolic blood pressure of 60 mm Hg or less

[0121] - Aged 65 or above.

[0122] APACHE II:

[0123] APACHE II is the acronym for Acute Physiology and Chronic Health Evaluation and is calculated according to Table 2 (Knaus WA et al. APACHE II: a severity of disease classification system. Crit Care Med. 1985 Oct;13(10):818-29):

[0124] Table 2: APACHE II score

[0125]

[0126]

[0127]

[0128]

[0129] The APACHE II score is intended to provide an approximation of the probability of death for a patient or group of patients based on the total score obtained. The mortality interpretation of the score is described in Table 3:

[0130] Table 3: Apache II Score / Approximate Mortality Rate Interpretation

[0131]

[0132] SOFA:

[0133] SOFA score is an acronym for Sequential Organ Failure Assessment and is calculated according to Table 4 (S. Vosylius, J. Sipylaite, and J. Ivaskevicius, Croatia Med J, 45 (2004), 715-20):

[0134] Table 4: SOFA score

[0135]

[0136] Cardiovascular hypotension:

[0137] Cardiovascular hypotension is a cardinal feature of septic shock. It may be due to low cardiac output or low systemic vascular resistance. Its severity can be assessed using the cardiovascular SOFA (Sequential Organ Failure Assessment) score, which is defined by mean arterial pressure or the need for vasopressor administration, as shown in Table 5 (S. Vosylius, J. Sipylaite, and J. Ivaskevicius, Croatian Journal of Medicine, 45 (2004), 715-20):

[0138] Table 5: Cardiovascular SOFA score

[0139] Mean arterial pressure or need for vasopressors score No hypotension 0 MAP<70mm / Hg 1 <![CDATA[ Dopamine ≤5 μg / kg / min or Dobutamine (Any dose)]]> 2 <![CDATA[dopamine > 5 μg / kg / min or adrenaline ≤ 0.1 μg / kg / min or Norepinephrine ≤ 0.1 mcg / kg / min > 3 Dopamine > 15 mcg / kg / min, epinephrine > 0.1 mcg / kg / min, or norepinephrine > 0.1 mcg / kg / min 4

[0140] Example 1

[0141] Treatment of sepsis and septic shock in patients with severe infection

[0142] In a study reported subsequently, a highly preferred mixture of empty liposomes of the present invention (designated CAL02) was administered intravenously (IV) to patients admitted to the intensive care unit (ICU) with severe community-acquired pneumonia (CAP) caused by Streptococcus pneumoniae as an adjunct to standard antibiotic therapy. The highly preferred mixture of empty liposomes of the present invention (CAL02) consisted of a 1:1 (weight / weight - w / w) mixture of the first empty liposomes and the second empty liposomes, wherein the first empty liposomes consisted of sphingomyelin and cholesterol in a 1:1 weight ratio (1:1 w / w; molar ratio 35:65) and the second empty liposomes consisted solely of sphingomyelin.

[0143] In addition to standard-of-care antibiotic therapy (Mandel et al., CID 2007:44(Suppl 2), S27-S72), each patient received two infusions of CAL02 or placebo (physiological 0.9% NaCl solution) (with an interval of 24 or 48 hours between these administrations). The first administration was performed shortly after the severity was diagnosed. The severity was diagnosed based on at least one of the following severity criteria:

[0144] i. Invasive mechanical ventilation support

[0145] ii. After adequate fluid resuscitation, treatment with therapeutic doses of vasopressors (i.e., dopamine >5 mg / kg / min or any dose of epinephrine, norepinephrine, phenylephrine, or vasopressin) for at least 2 hours to maintain or attempt to maintain systolic blood pressure >90 mm Hg (or mean arterial pressure >70 mm Hg)

[0146] Or based on at least three of the following minor severity criteria:

[0147] i. Respiratory rate ≥ 30 breaths / minute

[0148] ii. PaO2 / FiO2 ratio ≤ 250 mm Hg

[0149] iii. Multi-leaf infiltration

[0150] iv. Confusion / disorientation (must be documented before the use of sedatives or other new psychotropic medications)

[0151] v. Urea >7 mM (>40 mg / dL)

[0152] vi. Leukopenia (white blood cell count <4,000 cells / mm 3 )

[0153] vii. Thrombocytopenia (platelet count < 100,000 cells / mm 3 )

[0154] viii. Hypothermia (core temperature <36°C)

[0155] ix. Systolic blood pressure <90 mm Hg or mean arterial pressure <70 mm Hg and receiving ≥40 mL / kg of fluid resuscitation for at least 2 hours.

[0156] Two dose levels of CAL02 were tested: 4 mg / kg (low dose) and 16 mg / kg (high dose). Five patients were randomized to the placebo group, 11 patients to the CAL02 high dose group, and 3 patients to the CAL02 low dose group.

[0157] At the time of treatment, 56% of the patients had developed septic shock: two patients in the placebo group, five patients in the high-dose CAL02 group, and all three patients in the low-dose CAL02 group.

[0158] Study Results

[0159] Table 6: Patient characteristics at baseline

[0160]

[0161] When considering the entire study population and when only patients presenting with septic shock at baseline were assessed, a more rapid decline in the cardiovascular SOFA score was observed in the CAL02 group compared with the placebo group: in the CAL02 group, the SOFA score had already decreased by 100% within 6 days, whereas at the same time point, the reduction in the placebo group had not reached 40% ( Figures 1 and 2 ).

[0162] Among patients presenting with septic shock, hypotension and septic shock had complete resolution in all patients in the CAL02 high-dose group (5 / 5, 100%) and 66% (2 / 3) of patients in the CAL02 low-dose group by day 8 after treatment, compared to no patients in the placebo group (0 / 2, 0%).

[0163] Of the three patients in the placebo group who were not in septic shock at baseline, one patient was hypotensive at baseline and developed septic shock on day 4, and two patients who were not hypotensive at baseline developed hypotension during the first eight days. In contrast, in the CAL02 group, three patients who were not hypotensive at baseline did not develop any hypotension, and all other patients improved to a resolved state by day 6. This suggests that CAL02 prevented hypotension and hemodynamic instability, and prevented the development of septic shock.

[0164] Resolution of septic shock was accompanied by a substantial reduction in mean ICU length of stay, from 32 days for patients in the placebo group to 5.4 days and 15 days for patients in the CAL02 high-dose group and the CAL02 low-dose group, respectively. Furthermore, the CAL02 group had a lower incidence of death (20% and 33% in the CAL02 high-dose and CAL02 low-dose groups, respectively) compared to the placebo group (50%) (Table 7).

[0165] Table 7: Results

[0166]

[0167] The effects of CAL02 treatment on vital signs (including heart rate, systolic and diastolic blood pressure, and core body temperature) and lactate levels were also assessed.

Claims

1. Use of a composition for the preparation of a medicament for treating hypotension in sepsis in a human, wherein the treatment is an adjunct to antibiotic therapy, the composition comprising a mixture of empty liposomes, wherein the mixture of empty liposomes comprises: (a) a first empty liposome comprising cholesterol and sphingomyelin, wherein the amount of cholesterol is at least 30% weight / weight; and (b) a second empty liposome comprising sphingomyelin, wherein the second empty liposome (b) comprises the sphingomyelin as the only lipid component.

2. The method according to claim 1, wherein the hypotension is persistent hypotension.

3. Use according to any one of the preceding claims, wherein the amount of cholesterol of the empty liposomes (a) is 45% to 55% w / w, and wherein the second empty liposomes (b) consist of sphingomyelin.

4. The use according to claim 1, wherein the first empty liposome (a) consists of cholesterol and sphingomyelin, and wherein the amount of cholesterol in the empty liposome (a) is 47.5-52.5% weight / weight, and wherein the mixture of empty liposomes comprises at least 40% weight / weight of the first empty liposome (a) and the second empty liposome (b).

5. The use according to claim 1 , wherein the mixture of empty liposomes consists of a 1:1 weight / weight mixture of the first empty liposomes and the second empty liposomes, wherein the first empty liposomes are composed of cholesterol and sphingomyelin in a 1:1 weight ratio, and the second empty liposomes are composed only of sphingomyelin, and wherein the first empty liposomes (a) comprise the cholesterol and the sphingomyelin as the sole lipid components, and the second empty liposomes (b) comprise the sphingomyelin as the sole lipid component.

6. Use according to claim 1, wherein the hypotension is associated with a mean arterial pressure < 70 mm Hg.

7. The use according to claim 6, wherein the hypotension is pretreated with vasopressors for at least 2 hours.

8. The use according to claim 1, wherein the antibiotic therapy is intravenous (IV) or oral antibiotic therapy.

9. The use according to claim 1, wherein the human suffers from pneumonia, wherein the pneumonia is selected from community-acquired pneumonia (CAP), hospital-acquired pneumonia (HAP) and ventilator-associated pneumonia (VAP).

10. The use according to claim 9, wherein the pneumonia is severe pneumonia.

11. The method according to claim 9, wherein the pneumonia is caused by Streptococcus pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Enterococcus faecalis, Legionella pneumophila, Haemophilus influenzae, Klebsiella pneumoniae, Escherichia coli, Acinetobacter baumannii, Bordetella pertussis, Serratia marcescens, Stenotrophomonas maltophilia, Moraxella catarrhalis or Mycobacterium tuberculosis.

12. The use according to claim 1, wherein the composition is in the form of a solution for intravenous administration.

13. The use according to claim 1, wherein the composition is administered to the human in at least 2 doses: a first dose and a second dose, and wherein the interval between the first dose and the second dose is 6 hours to 96 hours.

14. The use according to claim 1, wherein the hypotension requires hospitalization.

15. Use according to claim 14, wherein said treatment reduces the length of stay in said hospital compared to the length of stay in said hospital in the absence of such treatment.

16. Use according to claim 15, wherein the reduction in hospital stay due to said treatment is at least one day.

17. The use according to claim 1, wherein the time to resolution of the hypotension is less than in the absence of such treatment.

18. The use according to claim 17, wherein the healing time that is less is at least one day less in time.

19. The use of claim 1, wherein the treatment reduces the cardiovascular SOFA score compared to the cardiovascular SOFA score in the absence of such treatment.

20. The use according to claim 19, wherein said reduction is at least 50% 7 days after the start of said treatment.

21. The use according to claim 4, wherein the mixture of empty liposomes comprises at least 45% weight / weight of the first empty liposomes (a) and the second empty liposomes (b).

22. The use according to claim 9, wherein the pneumonia is severe community-acquired pneumonia (sCAP).

23. The use according to claim 9, wherein the pneumonia is severe community-acquired pneumococcal pneumonia (sCAPP).

24. The use according to claim 9, wherein the pneumonia is caused by Streptococcus pneumoniae.

25. The use according to claim 13, wherein the interval between the first dose and the second dose is 12 hours to 72 hours.

26. The use according to claim 13, wherein the interval between the first dose and the second dose is 24 hours to 48 hours.

27. The use according to claim 13, wherein the interval between the first dose and the second dose is 24 hours or 48 hours.

28. The use of claim 14, wherein the hypotension requires admission to a hospital intensive care unit (ICU).

29. The method of claim 16, wherein the reduction in hospital stay due to the treatment is two days, or three days, or four days, or five days, or six days, or seven days, or eight days, or nine days.

30. The use of claim 18, wherein the healing time is less than two days in time.

31. The use of claim 18, wherein the healing time is less than at least three days less in time.

32. The use of claim 18, wherein the healing time that is less is at least four days less in time.

33. The use of claim 18, wherein the healing time that is less is at least five days less in time.

34. The use of claim 18, wherein the healing time that is less is at least six days less in time.

35. The use of claim 18, wherein the healing time that is less is at least seven days less in time.

36. The use of claim 20, wherein the reduction is at least 50% 6 days after the start of the treatment.

37. The use of claim 20, wherein the reduction is at least 50% 5 days after the start of the treatment.

38. The use of claim 20, wherein the reduction is at least 60% 7 days after the start of the treatment.

39. The use of claim 20, wherein the reduction is at least 60% 6 days after the start of the treatment.

40. The use of claim 20, wherein the reduction is at least 60% 5 days after the start of the treatment.

41. The use of claim 20, wherein the reduction is at least 70% 7 days after the start of the treatment.

42. The use of claim 20, wherein the reduction is at least 70% 6 days after the start of the treatment.

43. The use of claim 20, wherein the reduction is at least 70% 5 days after the start of the treatment.

44. The use of claim 20, wherein the reduction is at least 80% 7 days after the start of the treatment.

45. The use of claim 20, wherein the reduction is at least 80% 6 days after the start of the treatment.

46. ​​The use of claim 20, wherein the reduction is at least 80% 5 days after the start of the treatment.

47. The method of claim 10, wherein the severe pneumonia is caused by Streptococcus pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Enterococcus faecalis, Legionella pneumophila, Haemophilus influenzae, Klebsiella pneumoniae, Escherichia coli, Acinetobacter baumannii, Bordetella pertussis, Serratia marcescens, Stenotrophomonas maltophilia, Moraxella catarrhalis, or Mycobacterium tuberculosis.

48. The use according to claim 10, wherein the severe pneumonia is caused by Streptococcus pneumoniae.

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