Methods of treating excessive inflammatory disorders using lipid binding protein-based complexes

Abstract

Methods of treating excessive inflammatory conditions, such as hemophagocytic lymphocyte hyperplasia (HLH), dengue hemorrhagic fever, and dengue shock syndrome, using lipid binding protein based complexes.

Description

[0001] 1. Cross - reference to related applications

[0002] This application claims priority to U.S. Application No. 63 / 351,129, filed on June 10, 2022, the content of which is incorporated herein by reference in its entirety.

[0003] 2. Sequence Listing

[0004] This application contains a sequence listing that has been electronically submitted in XML format and is hereby incorporated by reference in its entirety. The XML sequence listing was created on June 1, 2023, named CRN - 050WO_SL.xml, and is 3,275 bytes in size. Background Art

[0005] Multiple studies have reported that the levels of ApoA - I and HDL are lower in COVID - 19 patients, especially in the most severe cases, and that the HDL in COVID - 19 patients has a weaker protective effect on endothelial cells triggered by inflammation and cannot protect them from apoptosis. A low level of ApoA - I in serum may increase the risk of contracting COVID - 19 and the risk of severe forms of COVID - 19. This decrease in ApoA - I or HDL is a common phenomenon in cytokine storms and has also been observed in virus - induced hemophagocytic lymphohistiocytosis and familial hemophagocytic lymphohistiocytosis (HLH) as well as dengue shock syndrome.

[0006] Current treatments for such hyper - inflammatory conditions are generally insufficient or ineffective. Therefore, new therapies for hyper - inflammatory conditions (such as those caused by viruses) are needed.

[0007] 4. Summary of the Invention

[0008] The present disclosure provides methods for treating a subject having or at risk of having an inflammatory disorder such as hemophagocytic lymphohistiocytosis (HLH), dengue hemorrhagic fever, and dengue shock syndrome. In some embodiments, the subject has hyper - inflammation characterized by severe inflammation accompanied by a cytokine storm.

[0009] In some embodiments, the subject is treated with a high dose of the lipocalin complex. The high dose is generally higher than the dose used to treat chronic diseases (such as familial hypercholesterolemia). The high dose is generally administered over a relatively short period of time, such as over a period of one day to two weeks, and generally includes multiple administrations of the lipocalin complex, such as two to ten separate doses. Each separate dose may be administered less than one day apart (e.g., twice a day) or one day or more apart (e.g., once a day).

[0010] In some embodiments of the methods of the present disclosure, the lipocalin-based complex comprises sphingomyelin and / or negatively charged lipids, such as CER-001. CER-001 is a negatively charged lipoprotein complex that comprises recombinant human ApoA-I, sphingomyelin (SM), and 1,2-dipalmitoyl-sn-glycero-3-phosphate-(1'-rac-glycerol) (dipalmitoyl phosphatidylglycerol; DPPG). It mimics native, nascent discoidal pre-βHDL, which is the form that HDL particles adopt before acquiring cholesterol. Without being bound by theory, it is believed that CER-001 therapy can reduce the levels of inflammatory cytokines (such as IL-6) in the serum, thereby providing clinical benefit to subjects suffering from the inflammatory disorders described herein (such as subjects suffering from or at risk of a virus-induced inflammatory state).

[0011] In one aspect, the present disclosure provides a method of treating a subject suffering from or at risk of HLH, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0012] In one aspect, the present disclosure provides a method of treating a subject suffering from or at risk of familial HLH, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0013] In one aspect, the present disclosure provides a method of treating a subject suffering from HLH secondary to a malignant disease (e.g., acute leukemia or lymphoma) or a non-malignant disease (e.g., an autoimmune disease or an infection) or at risk of suffering from HLH secondary to a malignant disease (e.g., acute leukemia or lymphoma) or a non-malignant disease (e.g., an autoimmune disease or an infection), comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0014] In one aspect, the present disclosure provides a method of treating a subject suffering from or at risk of virus-induced HLH, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0015] In another aspect, the present disclosure provides a method of treating a subject suffering from a dengue infection, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0016] In another aspect, the present disclosure provides a method of treating a subject suffering from or at risk of dengue hemorrhagic fever, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0017] In another aspect, the present disclosure provides a method of treating a subject having or at risk of having dengue shock syndrome, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0018] In another aspect, the present disclosure provides a method of treating a subject having a herpes simplex infection, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0019] In some aspects, the present disclosure provides a dosing regimen for the lipocalin-based treatment (e.g., CER-001 treatment) of the subjects described herein.

[0020] The dosing regimen of the present disclosure generally requires multiple administrations of CER-001 to the subject (e.g., once or twice a day). The CER-001 therapy can be continued for a predetermined period of time, e.g., for one week or less (e.g., one, two, three, four, five, six, or seven days) or for a period of time longer than one week (e.g., two weeks). Alternatively, CER-001 can be continuously administered to the subject until one or more symptoms of the disorder (e.g., acute inflammation or cytokine release syndrome (CRS)) are alleviated, or until the serum levels of one or more inflammatory markers are reduced, e.g., to normal levels or reduced relative to baseline measurements taken prior to initiation of CER-001 therapy. For a subject having an infection (e.g., viral infection), in some embodiments, the treatment can be continued until the subject has recovered from the infection.

[0021] The dosing regimen of the present disclosure can comprise administering to the subject a lipocalin-based complex (e.g., CER-001) according to an initial “induction” regimen, followed optionally by administering to the subject a lipocalin-based complex according to a “consolidation” regimen.

[0022] The induction regimen generally comprises administering to the subject multiple doses of a lipocalin-based complex (e.g., CER-001), e.g., six doses over three days.

[0023] Consolidation regimens generally include administering to a subject one or more doses of a lipocalin-based complex (e.g., CER-001) after the final dose of an induction regimen (e.g., one or more days after the final dose of an induction regimen). In some embodiments, the first dose of the consolidation regimen is administered on the third day after the final dose of the induction regimen. For example, the dosing regimen may include administering a lipocalin-based complex (e.g., CER-001) to the subject on days 1, 2, and 3 according to an induction regimen, and administering a lipocalin-based complex to the subject on day 6 according to a consolidation regimen. In some embodiments, the consolidation regimen includes two doses of a lipocalin-based complex.

[0024] In certain embodiments, the present disclosure provides a method of treating a subject having HLH (e.g., virus-induced HLH, familial HLH, or HLH secondary to acute leukemia or lymphoma) or at risk of having HLH (e.g., virus-induced HLH, familial HLH, or HLH secondary to acute leukemia or lymphoma), having dengue infection, having dengue hemorrhagic fever or at risk of having dengue hemorrhagic fever, having dengue shock syndrome or at risk of having dengue shock syndrome, or having herpes simplex infection, using a lipocalin-based complex (e.g., CER-001) according to a dosing regimen comprising:

[0025] - On days 1, 2, and 3, take 2 doses per day (induction regimen), followed optionally by

[0026] - Inject 2 subsequent doses on day 4 or later (consolidation regimen).

[0027] In some embodiments, the regimen comprises:

[0028] - On days 1, 2, and 3, take 2 doses per day (induction regimen), then

[0029] - Take 2 doses on day 6 (consolidation regimen).

[0030] In certain aspects, the lipocalin-based complex (e.g., CER-001) is co-administered with a standard of care therapy for the subject's disease or condition.

[0031] In certain aspects, an antihistamine (e.g., dextrochlorpheniramine, hydroxyzine, diphenhydramine, cetirizine, fexofenadine, or loratadine) can be administered before administering the lipocalin-based complex (e.g., CER-001). The antihistamine can reduce the likelihood of an allergic reaction. 5. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figures 1A - 1DShows the schedule of four subjects with COVID-19 treated with CER-001 (Example 1). Figure 1A Subject 1; Figure 1B Subject 2; Figure 1C Subject 3; Figure 1D Subject 4. DXM = dexamethasone; TCZ = tocilizumab.

[0034] Figures 2A - 2D Shows the apolipoprotein A-I (ApoA-I) levels of Subjects 1-4 in Example 1. Figure 2A Subject 1; Figure 2B Subject 2; Figure 2C Subject 3; Figure 2D Subject 4. The X-axis shows the number of days measured from the first administration of CER-001.

[0035] Figures 3A - 3D Shows the HDL levels of Subjects 1-4 in Example 1. Figure 3A Subject 1; Figure 3B Subject 2; Figure 3C Subject 3; Figure 3D Subject 4. The X-axis shows the number of days measured from the first administration of CER-001.

[0036] Figures 4A - 4D Shows the ferritin levels of Subjects 1-4 in Example 1. Figure 4A Subject 1; Figure 4B Subject 2; Figure 4C Subject 3; Figure 4D Subject 4. The X-axis shows the number of days measured from the first administration of CER-001.

[0037] Figures 5A - 5D Shows the interleukin 8 (IL-8) levels of Subjects 1-4 in Example 1. Figure 5A Subject 1; Figure 5B Subject 2; Figure 5C Subject 3; Figure 5D Subject 4. The X-axis shows the number of days measured from the first administration of CER-001.

[0038] Figures 6A - 6D Shows the tumor necrosis factor α (TNF-α) levels of Subjects 1-4 in Example 1. Figure 6A Subject 1; Figure 6B Subject 2; Figure 6C Subject 3; Figure 6D Subject 4. The X-axis shows the number of days measured from the first administration of CER-001.

[0039] Figures 7A - 7D Shows the platelet counts of Subjects 1 - 4 in Example 1. Figure 7A , Subject 1; Figure 7B , Subject 2; Figure 7C , Subject 3; Figure 7D , Subject 4. The X-axis shows the number of days measured starting from the first administration of CER-001. 6. DETAILED DESCRIPTION OF THE INVENTION

[0041] The present disclosure provides methods of treating a subject having or at risk of having an inflammatory disorder (such as hemophagocytic lymphohistiocytosis (HLH), dengue hemorrhagic fever, and dengue shock syndrome) with a lipocalin-based complex.

[0042] In some embodiments, the methods comprise administering a high dose of a lipocalin-based complex.

[0043] In one aspect, the present disclosure provides methods of treating a subject having or at risk of having HLH, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0044] In one aspect, the present disclosure provides methods of treating a subject having or at risk of having familial HLH, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0045] In one aspect, the present disclosure provides methods of treating a subject having or at risk of having HLH secondary to a malignant disease (such as acute leukemia or lymphoma) or a non-malignant disease (such as an autoimmune disease or an infection), comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0046] In one aspect, the present disclosure provides methods of treating a subject having or at risk of having virus-induced HLH, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0047] In another aspect, the present disclosure provides methods of treating a subject having a dengue infection (such as a subject having dengue, dengue hemorrhagic fever, or dengue shock syndrome), comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0048] In another aspect, the present disclosure provides a method of treating a subject having or at risk of having dengue hemorrhagic fever, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0049] In another aspect, the present disclosure provides a method of treating a subject having or at risk of having dengue shock syndrome, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0050] In another aspect, the present disclosure provides a method of treating a subject having a herpes simplex infection, comprising administering to the subject a lipocalin-based complex (e.g., CER-001).

[0051] In some embodiments, the lipocalin-based complex is an Apomer, a Cargomer, an HDL-based complex, or an HDL mimetic-based complex. In certain embodiments, the lipocalin-based complex is CER-001.

[0052] Exemplary characteristics of lipocalin-based complexes useful in the methods and compositions of the present disclosure are described in Section 6.1. Exemplary characteristics of lipocalin-based complexes useful in the methods and compositions of the present disclosure are described in Section 6.2.

[0053] In some embodiments, the methods of the present disclosure comprise administering the lipocalin-based complex (e.g., CER-001) to the subject in two phases. First, the lipocalin-based complex (e.g., CER-001) is administered in an initial, intense "induction" regimen. The induction regimen is followed by a less intense "consolidation" regimen. Alternatively, the lipocalin-based complex (e.g., CER-001) can be administered to the subject in a single phase, e.g., according to a dosing regimen corresponding to the doses and dosing frequencies of the induction or consolidation regimens described herein.

[0054] The induction regimen useful in the methods of the present disclosure is described in Section 6.3, and the consolidation regimen useful in the methods of the present disclosure is described in Section 6.3.2. The dosing regimens of the present disclosure comprise administering the lipocalin-based complex (e.g., CER-001) as a monotherapy or as part of a combination therapy with one or more drugs, e.g., in combination with the standard of care therapy for the subject's disease or condition. Combination therapies are described in Section 6.4.

[0055] 6.1. Lipocalin-based complexes

[0056] 6.1.1. HDL-based and HDL mimetic-based complexes

[0057] In one aspect, the lipocalin-based complex includes an HDL-based or HDL mimetic-based complex. For example, the complex can include a lipoprotein complex as described in U.S. Patent No. 8,206,750, PCT Publication WO 2012 / 109162, PCT Publication WO 2015 / 173633 A2 (e.g., CER-001), or US 2004 / 0229794 A1, the content of each patent being incorporated herein by reference in its entirety. The terms "lipoprotein" and "apolipoprotein" are used interchangeably herein, and unless the context requires otherwise, the term "lipoprotein" encompasses lipoprotein mimetics. The terms "lipocalin" and "lipocalin polypeptide" are also used interchangeably herein, and unless the context requires otherwise, the term does not denote an amino acid sequence of a specific length.

[0058] The lipoprotein complex can comprise a protein moiety (e.g., an apolipoprotein moiety) and a lipid moiety (e.g., a phospholipid moiety). The protein moiety includes one or more lipocalin molecules, such as apolipoproteins, peptides, or apolipoprotein peptide analogs or mimetics, such as one or more of the lipocalin molecules described in Section 6.1.2.

[0059] The lipid moiety generally includes one or more phospholipids, which can be neutral, negatively charged, positively charged, or a combination thereof. Exemplary phospholipids and other amphiphilic molecules that can be included in the lipid moiety are described in Section 6.1.3.

[0060] In certain embodiments, the lipid moiety contains at least one neutral phospholipid (e.g., sphingomyelin (SM)) and optionally one or more negatively charged phospholipids. In a lipoprotein complex comprising neutral and negatively charged phospholipids, the neutral and negatively charged phospholipids can have fatty acid chains of the same or different carbon numbers and the same or different degrees of saturation. In some cases, the neutral and negatively charged phospholipids will have the same acyl tails, such as C16:0 or palmitoyl acyl chains. In a particular embodiment, especially in an embodiment using egg SM as the neutral lipid, the weight ratio of the apolipoprotein moiety:lipid moiety ranges from about 1:2.7 to about 1:3 (e.g., 1:2.7).

[0061] Any phospholipid that is at least partially negatively charged at physiological pH can be used as the negatively charged phospholipid. Non-limiting examples include negatively charged forms such as salts of phosphatidylinositol, phosphatidylserine, phosphatidylglycerol, and phosphatidic acid. In a specific embodiment, the negatively charged phospholipid is 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] or DPPG, i.e., phosphatidylglycerol. Preferred salts include potassium salts and sodium salts.

[0062] In some embodiments, the lipoprotein complex used in the methods of the present disclosure is the lipoprotein complex described in U.S. Patent No. 8,206,750 or WO 2012 / 109162 (and its U.S. counterpart US 2012 / 0232005), the respective contents of which are incorporated herein by reference in their entirety. In certain embodiments, the protein component of the lipoprotein complex is as described in Section 6.1 of WO 2012 / 109162 (and US 2012 / 0232005), preferably as described in Section 6.1.1, and the lipid component is as described in Section 6.2 of WO 2012 / 109162 (and US 2012 / 0232005), which may optionally be complexed together in the amounts described in Section 6.3 of WO 2012 / 109162 (and US 2012 / 0232005). The content of each of these sections is incorporated herein by reference. In certain aspects, the lipoprotein complexes of the present disclosure are in a population of complexes that is at least 85%, at least 90%, at least 95%, at least 97% or at least 99% homogeneous, as described in Section 6.4 of WO 2012 / 109162 (and US 2012 / 0232005), the content of which is incorporated herein by reference.

[0063] In a specific embodiment, the lipoprotein complex useful in the methods of the present disclosure comprises 2 - 4 ApoA-I equivalents, 2 charged phospholipid molecules, 50 - 80 lecithin molecules, and 20 - 50 SM molecules.

[0064] In another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure comprises 2 - 4 ApoA-I equivalents, 2 charged phospholipid molecules, 50 lecithin molecules, and 50 SM molecules.

[0065] In yet another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure comprises 2 - 4 ApoA-I equivalents, 2 charged phospholipid molecules, 80 lecithin molecules, and 20 SM molecules.

[0066] In yet another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure comprises 2 - 4 ApoA-I equivalents, 2 charged phospholipid molecules, 70 lecithin molecules, and 30 SM molecules.

[0067] In yet another specific embodiment, the lipoprotein complex useful in the methods of the present disclosure comprises 2 - 4 ApoA-I equivalents, 2 charged phospholipid molecules, 60 lecithin molecules, and 40 SM molecules.

[0068] In a specific embodiment, the lipoprotein complex that can be used in the method of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 50-80 lecithin molecules, and 20-50 SM molecules.

[0069] In another specific embodiment, the lipoprotein complex that can be used in the method of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 50 lecithin molecules, and 50 SM molecules.

[0070] In yet another specific embodiment, the lipoprotein complex that can be used in the method of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 80 lecithin molecules, and 20 SM molecules.

[0071] In yet another specific embodiment, the lipoprotein complex that can be used in the method of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 70 lecithin molecules, and 30 SM molecules.

[0072] In yet another specific embodiment, the lipoprotein complex that can be used in the method of the present disclosure consists essentially of 2-4 ApoA-I equivalents, 2 charged phospholipid molecules, 60 lecithin molecules, and 40 SM molecules.

[0073] In a specific embodiment, the lipoprotein complex that can be used in the method of the present disclosure comprises a lipid component, and the lipid component comprises about 90 to 99.8 wt% SM and about 0.2 to 10 wt% negatively charged phospholipids, such as about 0.2-1 wt%, 0.2-2 wt%, 0.2-3 wt%, 0.2-4 wt%, 0.2-5 wt%, 0.2-6 wt%, 0.2-7 wt%, 0.2-8 wt%, 0.2-9 wt%, or 0.2-10 wt% of the total negatively charged phospholipids. In another specific embodiment, the lipoprotein complex that can be used in the method of the present disclosure comprises about 90 to 99.8 wt% lecithin and about 0.2 to 10 wt% negatively charged phospholipids, such as about 0.2-1 wt%, 0.2-2 wt%, 0.2-3 wt%, 0.2-4 wt%, 0.2-5 wt%, 0.2-6 wt%, 0.2-7 wt%, 0.2-8 wt%, 0.2-9 wt%, or 0.2-10 wt% of the total negatively charged phospholipids.

[0074] In a specific embodiment, the lipoprotein complex that can be used in the methods of the present disclosure comprises a lipid component consisting essentially of about 90 to 99.8 wt% SM and about 0.2 to 10 wt% negatively charged phospholipids, such as a total of about 0.2 - 1 wt%, 0.2 - 2 wt%, 0.2 - 3 wt%, 0.2 - 4 wt%, 0.2 - 5 wt%, 0.2 - 6 wt%, 0.2 - 7 wt%, 0.2 - 8 wt%, 0.2 - 9 wt% or 0.2 - 10 wt% of negatively charged phospholipids. In another specific embodiment, the lipoprotein complex that can be used in the methods of the present disclosure consists essentially of about 90 to 99.8 wt% lecithin and about 0.2 to 10 wt% negatively charged phospholipids, such as a total of about 0.2 - 1 wt%, 0.2 - 2 wt%, 0.2 - 3 wt%, 0.2 - 4 wt%, 0.2 - 5 wt%, 0.2 - 6 wt%, 0.2 - 7 wt%, 0.2 - 8 wt%, 0.2 - 9 wt% or 0.2 - 10 wt% of negatively charged phospholipids.

[0075] In yet another specific embodiment, the lipoprotein complex that can be used in the methods of the present disclosure comprises a lipid fraction that contains about 9.8 to 90 wt% SM, about 9.8 to 90 wt% lecithin, and about 0.2 - 10 wt% negatively charged phospholipids, such as a total of about 0.2 - 1 wt%, 0.2 - 2 wt%, 0.2 - 3 wt%, 0.2 - 4 wt%, 0.2 - 5 wt%, 0.2 - 6 wt%, 0.2 - 7 wt%, 0.2 - 8 wt%, 0.2 - 9 wt% to 0.2 - 10 wt% of negatively charged phospholipids.

[0076] In yet another specific embodiment, the lipoprotein complex that can be used in the methods of the present disclosure comprises a lipid fraction consisting essentially of about 9.8 to 90 wt% SM, about 9.8 to 90 wt% lecithin, and about 0.2 - 10 wt% negatively charged phospholipids, such as a total of about 0.2 - 1 wt%, 0.2 - 2 wt%, 0.2 - 3 wt%, 0.2 - 4 wt%, 0.2 - 5 wt%, 0.2 - 6 wt%, 0.2 - 7 wt%, 0.2 - 8 wt%, 0.2 - 9 wt% to 0.2 - 10 wt% of negatively charged phospholipids.

[0077] In another specific embodiment, the lipoprotein complex that can be used in the methods of the present disclosure includes apoA-I apolipoprotein and a lipid fraction, wherein the lipid fraction includes sphingomyelin and about 3 wt% of negatively charged phospholipids, wherein the molar ratio of the lipid fraction to the apoA-I apolipoprotein is about 2:1 to 200:1, and wherein the complex is a small or large discoidal particle containing 2 - 4 apoA-I equivalents.

[0078] In another specific embodiment, the lipoprotein complex that can be used in the methods of the present disclosure includes ApoA-I apolipoprotein and a lipid fraction, wherein the lipid fraction consists mainly of sphingomyelin and about 3 wt% of negatively charged phospholipids, wherein the molar ratio of the lipid fraction to ApoA-I apolipoprotein is about 2:1 to 200:1, and wherein the complex is a small or large discoidal particle containing 2-4 ApoA-I equivalents.

[0079] HDL-based or HDL-mimetic complexes can include a single type of lipid-binding protein, or a mixture of two or more different lipid-binding proteins, which can be from the same or different species. Although not essential, the complex preferably contains a lipid-binding protein from the animal species being treated or corresponding in amino acid sequence to the animal species being treated to avoid inducing an immune response to the treatment. Thus, for the treatment of human patients, lipid-binding proteins of human origin are preferably used. The use of peptide-mimicking apolipoproteins can also reduce or avoid immune responses.

[0080] In some embodiments, the lipid component includes two types of phospholipids: sphingomyelin (SM) and negatively charged phospholipids. Exemplary SM and negatively charged lipids are described in Section 6.1.3.1.

[0081] The lipid component including SM can optionally include a small amount of additional lipids. Virtually any type of lipid can be used, including but not limited to lysophospholipids, galactocerebrosides, gangliosides, cerebrosides, glycerolipids, triglycerides, and cholesterol and its derivatives.

[0082] When included, such optional lipids generally account for less than about 15 wt% of the lipid portion, although more optional lipids can be included in certain cases. In some embodiments, the optional lipids account for less than about 10 wt%, less than about 5 wt%, or less than about 2 wt%. In some embodiments, the lipid portion does not include optional lipids.

[0083] In one specific embodiment, the phospholipid fraction contains egg SM or palmitoyl SM or phytosphingomyelin and DPPG, and the weight ratio (SM: negatively charged phospholipid) is 90:10 to 99:1, more preferably 95:5 to 98:2. In one embodiment, the weight ratio is 97:3.

[0084] The molar ratio of the lipid component to the protein component of the disclosed complex can vary and will depend on factors such as the characteristics of the apolipoprotein that makes up the protein component, the characteristics and amount of the lipids that make up the lipid component, and the desired size of the complex. Since the biological activity of apolipoproteins (such as ApoA-I) is thought to be mediated by the amphipathic helices that make up the apolipoprotein, it is convenient to use ApoA-I protein equivalents to represent the apolipoprotein portion of the lipid:apolipoprotein molar ratio. ApoA-I is generally thought to contain 6 - 10 amphipathic helices, depending on the method used to calculate the helices. Other apolipoproteins can be expressed in ApoA-I equivalents based on the number of amphipathic helices they contain. For example, ApoA-I M , which is usually present in the form of a disulfide-bridged dimer, can be represented as 2 ApoA-I equivalents because each ApoA-I M molecule contains twice the number of amphipathic helices as an ApoA-I molecule. In contrast, a peptide apolipoprotein containing a single amphipathic helix can be represented as 1 / 10 - 1 / 6 ApoA-I equivalents because each molecule contains 1 / 10 - 1 / 6 the number of amphipathic helices as an ApoA-I molecule. Generally, the lipid:ApoA-I equivalent molar ratio (defined herein as "Ri") of the lipoprotein complex ranges from about 105:1 to 110:1. In some embodiments, Ri is about 108:1. A weight ratio can be obtained using an MW of about 650 - 800 for the phospholipid.

[0085] In some embodiments, the molar ratio of lipid:ApoA-I equivalents ("RSM") ranges from about 80:1 to about 110:1, such as from about 80:1 to about 100:1. In a particular instance, the RSM of the complex can be about 82:1.

[0086] In some embodiments, the lipoprotein complex used in the disclosed method is a negatively charged complex that contains a protein portion that is preferably mature full-length ApoA-I, and a lipid portion that contains neutral phospholipids, sphingomyelin (SM), and negatively charged phospholipids.

[0087] In one specific embodiment, the lipid component contains SM (e.g., egg SM, palmitoyl SM, phytoSM (plant SM), or a combination thereof) and a negatively charged phospholipid (e.g., DPPG), and the weight ratio (SM:negatively charged phospholipid) is from 90:10 to 99:1, more preferably from 95:5 to 98:2, such as 97:3.

[0088] In a specific embodiment, the ratio range of the protein component to the lipid component can be from about 1:2.7 to about 1:3, preferably 1:2.7. This corresponds to a molar ratio range of ApoA-I protein to lipid from about 1:90 to 1:140. In some embodiments, the molar ratio of protein to lipid in the complex is from about 1:90 to about 1:120, from about 1:100 to about 1:140, or from about 1:95 to about 1:125.

[0089] In a particular embodiment, the complex comprises CER-001, CSL-111, CSL-112, CER-522 or ETC-216. In a preferred embodiment, the complex is CER-001.

[0090] CER-001 as used in the literature and in the examples below refers to the complex described in Example 4 of WO 2012 / 109162. WO 2012 / 109162 refers to CER-001 as a complex having a lipoprotein weight:total phospholipid weight ratio of 1:2.7 and an SM:DPPG weight:weight ratio of 97:3. Example 4 of WO 2012 / 109162 also describes its manufacturing method.

[0091] When used in the context of the methods and / or CER-001 dosing regimens of the present disclosure, CER-001 refers to a lipoprotein complex, the individual components of which may differ from CER-001 as described in Example 4 of WO 2012 / 109162 by up to 20%. In certain embodiments, the components of the lipoprotein complex differ from CER-001 as described in Example 4 of WO 2012 / 109162 by up to 10%. Preferably, the components of the lipoprotein complex are the components described in Example 4 of WO 2012 / 109162 (plus / minus acceptable manufacturing tolerance variations). The SM in CER-001 can be natural or synthetic. In some embodiments, the SM is natural SM, such as the natural SM described in WO 2012 / 109162, such as egg SM. In some embodiments, the SM is synthetic SM, such as the synthetic SM described in WO 2012 / 109162, such as synthetic palmitoylsphingomyelin, as described in WO 2012 / 109162. Methods for synthesizing palmitoylsphingomyelin are known in the art, such as those described in WO 2014 / 140787. The lipoprotein in CER-001, apolipoprotein AI (ApoA-I), preferably has an amino acid sequence corresponding to amino acids 25 to 267 of SEQ ID NO:2 (previously published as SEQ ID NO:1 of WO 2012 / 109162). ApoA-I can be purified or recombinantly produced from animal sources (particularly human sources). In a preferred embodiment, the ApoA-I in CER-001 is recombinant ApoA-I. The CER-001 used in the dosing regimens of the present disclosure is preferably highly homogeneous, such as at least 80%, at least 85%, at least 90%, at least 95%, at least 97%, at least 98% or at least 99% homogeneous, as reflected by a single peak in gel permeation chromatography. See, for example, Section 6.4 of WO 2012 / 109162.

[0092] In certain embodiments, the ApoA-I in CER-001 is recombinant ApoA-I produced by a mammalian host cell. The host cell can be from any mammalian cell line. The polynucleotide encoding ApoA-I can be codon optimized for expression in the recombinant host cell. Preferred host cells are mammalian host cells, including but not limited to Chinese hamster ovary cells (e.g., CHO-K1; ATCC No. CCL 61; CHO-S (GIBCO Life Technologies Inc., Rockville, MD, Catalog #11619012)), VERO cells, BHK (ATCC No. CRL 1632), BHK 570 (ATCC No. CRL 10314), HeLa cells, COS-1 (ATCC No. CRL 1650), COS-7 (ATCC No. CRL 1651), MDCK cells, 293 cells (ATCC No. CRL 1573; Graham et al., J. Gen. Virol. 36:59-72, 1977), 3T3 cells, myeloma cells (especially murine), PC12 cells, and W138 cells. In some embodiments, the mammalian cells such as CHO-S cells (Invitrogen™, Carlsbad CA) are adapted to grow in serum-free media. Additional suitable cell lines are known in the art and are available from public repositories such as the American Type Culture Collection, Manassas, Va.

[0093] In a preferred embodiment, the recombinant ApoA-I is produced by CHO cells. The recombinant ApoA-I expressed by a mammalian host cell (e.g., CHO cells) may undergo post-translational processing (e.g., glycosylation, etc.). The resulting recombinant ApoA-I may have one or more structural features (e.g., different glycosylation patterns) that are different from ApoA-I purified from human plasma.

[0094] For the recombinant expression of ApoA-I, a polynucleotide encoding ApoA-I is operably linked to one or more control sequences such as a promoter or terminator, which regulate the expression of ApoA-I in a host cell of interest. The one or more control sequences can be native or foreign to the ApoA-I-encoding sequence and also native or foreign to the host cell in which ApoA-I is expressed. Control sequences include, but are not limited to, a promoter, ribosome binding site, leader sequence, polyadenylation sequence, propeptide sequence, signal peptide sequence, and transcription terminator. In some embodiments, the control sequences include a promoter, ribosome binding site, and transcription and translation termination signals. The control sequences can also include one or more adaptors for the purpose of introducing specific restriction sites, which facilitate the ligation of the control sequences to the coding region of the nucleotide sequence encoding ApoA-I.

[0095] The promoter driving the recombinant expression of ApoA-I can be a constitutive promoter, a regulatable promoter, or an inducible promoter. Suitable promoter sequences can be obtained from genes encoding extracellular or intracellular polypeptides, which are endogenous or heterologous to the host cell. Methods for the isolation, identification, and manipulation of promoters of variable length are available in the art or can be readily modified from the art. See, e.g., Nevoigt et al. (2006) Appl. Environ. Microbiol. 72:5266-5273, the contents of which are incorporated herein by reference.

[0096] One or more control sequences can be derived from a viral source. For example, in certain aspects, the promoter is derived from the polyoma or adenovirus major late promoter. In other aspects, the promoter is derived from simian virus 40 (SV40), which can be obtained as a fragment that also contains the SV40 viral origin of replication (Fiers et al., 1978, Nature, 273:113-120) or from cytomegalovirus such as the simian cytomegalovirus immediate early promoter (see U.S. Patent 4,956,288). Other suitable promoters include those from the metallothionein gene (see U.S. Patents 4,579,821 and 4,601,978).

[0097] The present application also provides a recombinant ApoA-I expression vector. The recombinant expression vector can be any vector that can be processed by recombinant DNA technology to facilitate the expression of heterologous ApoA-I in recombinant host cells, such as a plasmid or a virus. The expression vector can be integrated into the chromosome of the recombinant host cell and contains one or more heterologous genes operably linked to one or more control sequences for producing ApoA-I. In other embodiments, the expression vector is an extrachromosomal replicating DNA molecule, such as a linear or closed circular plasmid, which is found to have a low copy number (e.g., about 1 to about 10 copies / genome equivalent) or a high copy number (e.g., more than about 10 copies / genome equivalent). In multiple embodiments, the expression vector includes selectable markers, such as genes that confer antibiotic resistance (e.g., ampicillin, kanamycin, chloramphenicol, or tetracycline resistance) to the recombinant host organism containing the vector. In a specific aspect, the DNA construct, vector, and polynucleotide are suitable for the expression of ApoA-I in mammalian cells. The vector for the expression of ApoA-I in mammalian cells can contain an origin of replication compatible with the host cell system, a promoter, and any necessary ribosome binding sites, RNA splicing sites, polyadenylation sites, and transcription termination sequences. In certain aspects, the origin of replication is heterologous to the host cell, for example, it is of viral origin (e.g., SV40, Polyoma, Adeno, VSV, BPV). In other aspects, the origin of replication is provided by the host cell chromosomal replication mechanism.

[0098] Methods, reagents, and tools for introducing foreign DNA into mammalian host cells are known in the art and include, but are not limited to, calcium phosphate-mediated transfection (Wigler et al., 1978, Cell 14:725; Corsaro et al., 1981, Somatic Cell Genetics 7:603; Graham et al., 1973, Virology 52:456), electroporation (Neumann et al., 1982, EMBO J. 1:841-5), DEAE-dextran-mediated transfection (Ausubel et al. (eds.), Short Protocols in Molecular Biology, 3rd Edition (John Wiley & Sons 1995)), and liposome-mediated transfection (Hawley-Nelson et al., 1993, Focus 15:73; Ciccarone et al., 1993, Focus 15:80).

[0099] For high-yield production, stable expression of ApoA-I is preferred. For example, after introducing exogenous DNA into a host cell, the host cell can be grown in enriched medium for 1-2 days and then switched to selective medium. Instead of using an expression vector containing a viral replication origin, the host cell can be transformed with a vector having a nucleotide sequence containing the ApoA-I-encoding sequence, which is controlled by appropriate expression control elements and optional markers. The optional markers in the vector confer resistance to selection and allow the cells to stably integrate the vector into their chromosomes and grow to form foci, which can then be cloned and expanded into cell lines. A number of selection systems can be used, including but not limited to those that can be used in tk - , hgprt - or aprt - cells, such as the herpes simplex virus thymidine kinase (Wigler et al., 1977, Cell 11: 223), hypoxanthine guanine phosphoribosyltransferase (Szybalska & Szybalski, 1962, Proc. Natl. Acad. Sci. USA 48: 2026), and adenine phosphoribosyltransferase (Lowy et al., 1980, Cell 22: 817) genes. Similarly, antimetabolite drug resistance can be used as a basis for selection by using, for example, dhfr, which confers resistance to methotrexate (Wigler et al., 1980, Natl. Acad. Sci. USA 77: 3567; O’Hare et al., 1981, Proc. Natl. Acad. Sci. USA 78: 1527); gpt, which confers resistance to mycophenolic acid (Mulligan & Berg, 1981, Proc. Natl. Acad. Sci. USA 78: 2072); neo, which confers resistance to the aminoglycoside G-418 (Colberre-Garapin et al., 1981, J. Mol. Biol. 150: 1); and / or hyg, which confers resistance to hygromycin (Santerre et al., 1984, Gene 30: 147).

[0100] Stable high-yield expression can also be achieved by using retroviral vectors integrated into the host cell genome (see, for example, U.S. Patent Publications 2008 / 0286779 and 2004 / 0235173). Alternatively, stable high-yield expression of ApoA-I can be achieved by gene activation methods that cause activation and amplification of the endogenous ApoA-I gene in the genomic DNA of selected mammalian cells, as described, for example, in WO 1994 / 012650. Increasing the copy number of the ApoA-I gene (containing the ApoA-I coding sequence and one or more control elements) can promote high-yield expression of ApoA-I. Preferably, the mammalian host cells in which ApoA-I is expressed have an ApoA-I gene copy index of at least 2, at least 3, at least 4, or at least 5. In some specific embodiments, the mammalian host cells in which ApoA-I is expressed have an ApoA-I gene copy index of at least 6, at least 7, at least 8, at least 9, or at least 10.

[0101] In certain embodiments, the mammalian cells are adapted to produce at least 0.5 g / L, at least 1 g / L, at least 1.5 g / L, at least 2 g / L, at least 2.5 g / L, at least 3 g / L, at least 3.5 g / L of ApoA-I, and optionally produce up to 4 g / L, up to 4.5 g / L, up to 5 g / L, up to 5.5 g / L, or up to 6 g / L of ApoA-I. The mammalian host cells preferably are capable of producing at least about 0.5, 1, 2, or 3 g / L ApoA-I in culture and / or producing up to about 20 g / L ApoA-I in culture, such as producing up to 4, 5, 6, 7, 8, 9, 10, 12, or 15 g / L ApoA-I in culture.

[0102] In certain embodiments, the mammalian cells are adapted to grow in serum-free medium. In these embodiments, the ApoA-I is secreted by the cells. In other embodiments, the ApoA-I is not secreted by the cells.

[0103] The mammalian host cells provided by the present application can be used to produce ApoA-I. Generally, the method includes culturing the mammalian host cells described in the present application under conditions for expressing ApoA-I. In addition, the method can include recovering and optionally purifying mature ApoA-I from the supernatant of the mammalian cell culture.

[0104] The culture conditions (including medium, temperature, pH) can be suitable for the mammalian host cells to be cultured and the selected culture mode (shake flask, bioreactor, roller bottle, etc.). The mammalian cells can be grown in large-scale batch cultures, continuous cultures, or semi-continuous cultures.

[0105] The present application also provides a mammalian cell culture comprising a plurality of the ApoA-I-producing mammalian host cells described herein. In some embodiments, the mammalian cell culture comprises at least 0.5 g / L, at least 1 g / L, at least 1.5 g / L, at least 2 g / L, at least 2.5 g / L, at least 3 g / L, at least 3.5 g / L, and optionally comprises up to 4 g / L, up to 4.5 g / L, up to 5 g / L, up to 5.5 g / L or up to 6 g / L of ApoA-I. The culture can be of any scale, ranging from about 150 mL to about 500 mL, 1 L, 10 L, 15 L, 50 L, 100 L, 200 L, 250 L, 300 L, 350 L, 400 L, 500 L, 750 L, 1000 L, 1500 L, 2000 L, 2500 L, 3000 L, 5000 L, 7500 L, 10000 L, 15000 L, 20000 L, 25000 L, 50000 L or more. In some cases, the culture is a large-scale culture, such as 15 L, 20 L, 25 L, 30 L, 50 L, 100 L, 200 L, 300 L, 500 L, 1000 L, 5000 L, 10000 L, 15000 L, 20000 L, 25000 L, up to 50000 L or more.

[0106] The mammalian host cells of the present disclosure can grow in culture. Accordingly, the present disclosure also provides a mammalian cell culture comprising a plurality of mammalian host cells as described above. The cell culture may comprise one or more of the following features: (a) the culture (optionally a large-scale batch culture of at least 10 liters, at least 20 liters, at least 30 liters, at least 50 liters, at least 100 liters, 300 L, 500 L, 1000 L, 5000 L, 10,000 L, 15,000 L, 20,000 L, 25,000 L, up to 50,000 L or a continuous culture of at least 10 liters, at least 20 liters, at least 30 liters, at least 50 liters, at least 100 liters, 300 L, 500 L, 1000 L, 5000 L or up to 10,000 L) contains at least about 0.5, 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0 g / L or more of mature ApoA-I protein, which comprises or consists of the amino acid sequence corresponding to amino acids 25 to 267 of SEQ ID NO: 2; (b) at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98% or at least 99% of the proteins in the culture medium are ApoA-I proteins lacking a signal sequence; (c) at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98% or at least 99% of the proteins in the culture medium are mature ApoA-I proteins lacking a signal sequence and a propeptide sequence; and (d) at least 75%, at least 80%, at least 85%, at least 90%, at least 95% of the mature ApoA-I is not truncated, oxidized or deamidated.

[0107] CSL-111 is a recombinant human ApoA-I purified from plasma complexed with soybean phosphatidylcholine (SBPC) (Tardif et al., 2007, JAMA 297:1675-1682).

[0108] CSL-112 is an ApoA-I preparation purified from plasma and reconstituted to form HDL suitable for intravenous infusion (Diditchenko et al., 2013, DOI 10.1161 / ATVBAHA.113.301981).

[0109] ETC-216 (also known as MDCO-216) is delipidated HDL containing recombinant ApoA-I Milano See Nicholls et al., 2011, Expert Opin Biol Ther. 11(3):387-94. doi: 10.1517 / 14712598.2011.557061.

[0110] In another embodiment, the complex that can be used in the methods of the present disclosure is CER-522. CER-522 is a lipoprotein complex comprising a combination of three phospholipids and a 22-amino acid peptide CT80522:

[0111]

[0112] CT80522

[0113] The phospholipid component of CER-522 consists of egg sphingomyelin, 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (dipalmitoyl phosphatidylcholine, DPPC), and 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (dipalmitoyl phosphatidylglycerol, DPPG) in a weight ratio of 48.5:48.5:3. The ratio of peptide to total phospholipids in the CER-522 complex is 1:2.5 (w / w).

[0114] In some embodiments, the lipoprotein complex is delipidated HDL. Most HDL in plasma is cholesterol-rich. The lipids in HDL can be depleted, for example, partially and / or selectively, for example, to reduce its cholesterol content. In some embodiments, delipidated HDL can be similar to small α, pre-β-1, and other pre-β forms of HDL. Methods for selectively depleting HDL are described in Sacks et al., 2009, J Lipid Res. 50(5): 894–907.

[0115] In certain embodiments, the lipoprotein complex comprises bioactive agent delivery particles as described in US 2004 / 0229794.

[0116] The bioactive agent delivery particles can comprise a lipid-binding polypeptide (e.g., an apolipoprotein as previously described in this section or in Section 6.1.2), a lipid bilayer (e.g., comprising one or more phospholipids as previously described in this section or in Section 6.1.3.1), and a bioactive agent (e.g., an anti-cancer agent), wherein the interior of the lipid bilayer comprises a hydrophobic region and wherein the bioactive agent is associated with the hydrophobic region of the lipid bilayer. In some embodiments, the bioactive agent delivery particles are as described in US 2004 / 0229794.

[0117] In some embodiments, the bioactive agent delivery particles do not comprise a hydrophilic core.

[0118] In some embodiments, the bioactive agent delivery particles are discoidal (e.g., having a diameter of about 7 to about 29 nm).

[0119] The bioactive agent delivery particles include bilayer-forming lipids, such as phospholipids (e.g., as previously described in this section or in Section 6.1.3.1). In some embodiments, the bioactive agent delivery particles include bilayer-forming lipids and non-bilayer-forming lipids. In some embodiments, the lipid bilayer of the bioactive agent delivery particles includes phospholipids. In one embodiment, the phospholipids incorporated into the delivery particles include dimyristoylphosphatidylcholine (DMPC) and dimyristoylphosphatidylglycerol (DMPG). In one embodiment, the lipid bilayer includes DMPC and DMPG in a molar ratio of 7:3.

[0120] In some embodiments, the lipid-binding polypeptide is an apolipoprotein (e.g., as previously described in this section or in Section 6.1.2). The major interaction between the lipid-binding polypeptide (e.g., an apolipoprotein molecule) and the lipid bilayer is generally a hydrophobic interaction between residues on the hydrophobic surface of the amphipathic structure, e.g., the α-helix of the lipid-binding polypeptide and the fatty acyl chains of the lipids on the outer surface of the particle periphery. The bioactive agent delivery particles can include exchangeable and / or non-exchangeable apolipoproteins. In one embodiment, the lipid-binding polypeptide is ApoA-I.

[0121] In some embodiments, the bioactive agent delivery particles include lipid-binding polypeptide molecules, such as apolipoprotein molecules, that have been modified to increase the stability of the particles. In one embodiment, the modification includes introducing cysteine residues to form intramolecular and / or intermolecular disulfide bonds.

[0122] In another embodiment, the bioactive agent delivery particles include chimeric lipid-binding polypeptide molecules, such as chimeric apolipoprotein molecules, that have one or more binding functional moieties, such as one or more targeting moieties and / or one or more moieties having a desired biological activity (e.g., antibacterial activity), which can enhance or act synergistically with the activity of the bioactive agent incorporated into the delivery particles.

[0123] 6.1.2. Lipid-Binding Protein Molecules

[0124] Lipid-binding protein molecules that can be used in the complexes described herein include apolipoproteins (apolipoproteins as described in Section 6.1.2.1) and apolipoprotein mimetic peptides (apolipoprotein mimetic peptides as described in Section 6.1.2.2). In some embodiments, the complex contains a mixture of lipid-binding protein molecules. In some embodiments, the complex contains a mixture of one or more lipid-binding protein molecules and one or more apolipoprotein mimetic peptides.

[0125] In some embodiments, the complex comprises 1 to 8 ApoA-I equivalents (e.g., 1, 2, 3, 4, 5, 6, 7, 8, 1 to 7, 1 to 6, 1 to 5, 1 to 4, 1 to 3, 1 to 2, 2 to 8, 2 to 6, 2 to 4, 4 to 6, or 4 to 8 ApoA-I equivalents). Lipocalins can be expressed in ApoA-I equivalents based on the number of amphipathic helices they contain. For example, ApoA-I, which typically exists as a disulfide-bridged dimer, M can be represented as 2 ApoA-I equivalents because each ApoA-I M molecule contains twice as many amphipathic helices as an ApoA-I molecule. In contrast, a peptide mimetic containing a single amphipathic helix can be represented as 1 / 10 - 1 / 6 ApoA-I equivalents because each molecule contains as many amphipathic helices as 1 / 10 - 1 / 6 of an ApoA-I molecule.

[0126] 6.1.2.1. Apolipoproteins

[0127] Suitable apolipoproteins that can be included in the lipocalin-based complex include apolipoprotein ApoA-I, ApoA-II, ApoA-IV, ApoA-V, ApoB, ApoC-I, ApoC-II, ApoC-III, ApoD, ApoE, ApoJ, ApoH, and any combination of two or more of the foregoing. Polymorphic forms, subtypes, variants, and mutants, as well as truncated forms of the foregoing apolipoproteins can also be used, among which the most common ones are apolipoprotein A-IMilano (ApoA-IM), apolipoprotein A-IParis (ApoA-IP), and apolipoprotein A-IZaragoza (ApoA-IZ). Apolipoprotein mutants containing cysteine residues are also known and can also be used (see, for example, US Publication No. 2003 / 0181372). The apolipoprotein can be in the form of a monomer or a dimer, which can be a homodimer or a heterodimer. For example, homodimers or heterodimers of ApoA-I (Duverger et al., 1996, Arterioscler. Thromb. Vasc. Biol. 16(12):1424-29), ApoA-IM (Franceschini et al., 1985, J. Biol. Chem. 260:1632-35), ApoA-IP (Daum et al., 1999, J. Mol. Med. 77:614-22), ApoA-II (Shelness et al., 1985, J. Biol. Chem. 260(14):8637-46; Shelness et al., 1984, J. Biol. Chem. 259(15):9929-35), ApoA-IV (Duverger et al., 1991, Euro. J. Biochem. 201(2):373-83), ApoE (McLean et al., 1983, J. Biol. Chem. 258(14):8993-9000), ApoJ, and ApoH (when feasible) can all be used.

[0128] The primary sequence of the apolipoprotein can be modified to render it less susceptible to oxidation, for example, as described in U.S. Publication Nos. 2008 / 0234192 and 2013 / 0137628 and U.S. Patent Nos. 8,143,224 and 8,541,236. The apolipoprotein may include residues corresponding to elements that facilitate its isolation, such as His tags, or other elements designed for other purposes. Preferably, the apolipoprotein in the complex is soluble in a biological fluid (e.g., lymph, cerebrospinal fluid, vitreous humor, aqueous humor, blood, or a blood fraction (e.g., serum or plasma)).

[0129] In some embodiments, the complex comprises covalently bound lipocalin monomers, such as dimeric apolipoprotein A-IMilano, which is a cysteine-containing mutant form of ApoA-I. The cysteine allows for the formation of disulfide bonds, which can result in the formation of homodimers or heterodimers (e.g., ApoA-I Milano-ApoA-II).

[0130] In some embodiments, the apolipoprotein molecule comprises an ApoA-I, ApoA-II, ApoA-IV, ApoA-V, ApoB, ApoC-I, ApoC-II, ApoC-III, ApoD, ApoE, ApoJ, or ApoH molecule or a combination thereof.

[0131] In some embodiments, the apolipoprotein molecule comprises an ApoA-I molecule or consists of an ApoA-I molecule. In some embodiments, the ApoA-I molecule is a human ApoA-I molecule. In some embodiments, the ApoA-I molecule is recombinant. In some embodiments, the ApoA-I molecule is not ApoA-IMilano.

[0132] In some embodiments, the ApoA-I molecule is an apolipoprotein A-IMilano (ApoA-IM), apolipoprotein A-IParis (ApoA-IP), or apolipoprotein A-IZaragoza (ApoA-IZ) molecule.

[0133] Apolipoproteins can be purified or recombinantly produced from animal sources (especially human sources), as is well known in the art, see, for example, Chung et al., 1980, J. Lipid Res. 21(3):284-91; Cheung et al., 1987, J. Lipid Res. 28(8):913-29. See also U.S. Patent Nos. 5,059,528, 5,128,318, 6,617,134; U.S. Publication Nos. 2002 / 0156007, 2004 / 0067873, 2004 / 0077541 and 2004 / 0266660; and PCT Publication Nos. WO2008 / 104890 and WO 2007 / 023476. Other purification methods can also be employed, such as the method described in PCT Publication No. WO 2012 / 109162, the disclosure of which is incorporated herein by reference in its entirety.

[0134] The apolipoprotein can be in the prepro-form, pro-form or mature form. For example, the complex can comprise ApoA-I (e.g., human ApoA-I), where ApoA-I is prepro-ApoA-I, pro-ApoA-I or mature ApoA-I. In some embodiments, the complex comprises ApoA-I having at least 90% sequence identity with SEQ ID NO:1:

[0135] In other embodiments, the complex comprises ApoA-I having at least 95% sequence identity with SEQ ID NO:1. In other embodiments, the complex comprises ApoA-I having at least 98% sequence identity with SEQ ID NO:1. In other embodiments, the complex comprises ApoA-I having at least 99% sequence identity with SEQ ID NO:1. In other embodiments, the complex comprises ApoA-I having 100% sequence identity with SEQ ID NO:1.

[0136] In some embodiments, the complex comprises ApoA-I having at least 90% sequence identity with amino acids 25 to 267 of SEQ ID NO:2:

[0137]

[0138] In other embodiments, the complex comprises ApoA-I having at least 95% sequence identity with amino acids 25 to 267 of SEQ ID NO:2. In other embodiments, the complex comprises ApoA-I having at least 98% sequence identity with amino acids 25 to 267 of SEQ ID NO:2. In other embodiments, the complex comprises ApoA-I having at least 99% sequence identity with amino acids 25 to 267 of SEQ ID NO:2. In other embodiments, the complex comprises ApoA-I having 100% sequence identity with amino acids 25 to 267 of SEQ ID NO:2.

[0139] In some embodiments, the complex comprises 1 to 8 apolipoprotein molecules (e.g., 1 to 6, 1 to 4, 1 to 2, 2 to 8, 2 to 6, 2 to 4, 4 to 8, 4 to 6, or 6 to 8 apolipoprotein molecules). In some embodiments, the complex comprises 1 apolipoprotein molecule. In some embodiments, the complex comprises 2 apolipoprotein molecules. In some embodiments, the complex comprises 3 apolipoprotein molecules. In some embodiments, the complex comprises 4 apolipoprotein molecules. In some embodiments, the complex comprises 5 apolipoprotein molecules. In some embodiments, the complex comprises 6 apolipoprotein molecules. In some embodiments, the complex comprises 7 apolipoprotein molecules. In some embodiments, the complex comprises 8 apolipoprotein molecules.

[0140] The apolipoprotein molecule may comprise a chimeric apolipoprotein that comprises an apolipoprotein and one or more attached functional moieties, e.g., one or more CRN-001 complexes, one or more targeting moieties, a moiety having a desired biological activity, an affinity tag for facilitating purification, and / or a reporter molecule for characterization or localization studies. The attached moiety having biological activity may have an activity capable of enhancing and / or synergizing with the biological activity of a compound bound to the complex of the present disclosure. For example, the moiety having biological activity may have antimicrobial (e.g., antifungal, antibacterial, antiprotozoal, bacteriostatic, fungistatic, or antiviral) activity. In one embodiment, the attached functional moiety of the chimeric apolipoprotein does not contact the hydrophobic surface of the complex. In another embodiment, the attached functional moiety contacts the hydrophobic surface of the complex. In some embodiments, the functional moiety of the chimeric apolipoprotein may be inherent to the native protein. In some embodiments, the chimeric apolipoprotein comprises a ligand or sequence that is recognized by or capable of interacting with a cell surface receptor or other cell surface moiety.

[0141] In one embodiment, the chimeric apolipoprotein comprises a targeting moiety not possessed by a native apolipoprotein, e.g., the Saccharomyces cerevisiae α-mating factor peptide, folic acid, transferrin, or lactoferrin. In another embodiment, the chimeric apolipoprotein comprises a moiety having a desired biological activity that enhances and / or synergizes with the activity of a compound bound to the complex of the present disclosure. In one embodiment, the chimeric apolipoprotein may comprise a functional moiety inherent to the apolipoprotein. An example of an apolipoprotein-inherent functional moiety is an inherent targeting moiety formed by approximately amino acids 130-150 of human ApoE, which contains a receptor-binding region recognized by members of the low-density lipoprotein receptor family. Other examples of apolipoprotein-inherent functional moieties include the ApoB-100 region that interacts with the low-density lipoprotein receptor and the ApoA-I region that interacts with the class B type I scavenger receptor. In other embodiments, functional moieties may be synthetically added or recombinantly added to produce chimeric apolipoproteins. Another example is an apolipoprotein having a prosequence or a proregion from another proapolipoprotein (e.g., the prosequence from proapolipoprotein A-II replaces the prosequence of proapolipoprotein A-I). Another example is an apolipoprotein in which some amphipathic sequence segments have been replaced by other amphipathic sequence segments from another apolipoprotein.

[0142] As used herein, "chimeric" refers to two or more molecules that can exist separately and combine together to form a single molecule having the desired functions of all of its constituent molecules. The constituent molecules of the chimeric molecule can be synthetically linked by chemical bonding, or, when the constituent molecules are all polypeptides or analogs thereof, the polynucleotides encoding the polypeptides can be recombinantly fused together such that a single continuous polypeptide is expressed. Such a chimeric molecule is referred to as a fusion protein. A "fusion protein" is a chimeric molecule in which the constituent molecules are all polypeptides and are linked (fused) to each other such that the chimeric molecule forms a continuous single chain. The various components can be directly linked together or coupled through one or more linkers. For example, one or more fragments of the various components can be inserted into the sequence of the apolipoprotein, or, as another example, added to the N-terminus or C-terminus of the apolipoprotein sequence. For example, the fusion protein can comprise an antibody light chain, an antibody fragment, a heavy chain antibody, or a single-domain antibody.

[0143] In some embodiments, chimeric apolipoproteins are prepared by chemically conjugating an apolipoprotein and a functional moiety to be linked. Methods for chemically conjugating molecules are well known to those skilled in the art. Such methods will vary depending on the structure of the moiety to be linked, but are readily determinable by those skilled in the art. Polypeptides typically contain various functional groups, such as carboxylic acid (--COOH), free amino (--NH2), or sulfhydryl (--SH) groups, which can react with suitable functional groups on the functional moiety or linker to attach the moiety thereto. The functional moiety can be linked to a functional group on the N-terminus, C-terminus, or internal residue (i.e., a residue at an intermediate position between the N-terminus and C-terminus) of the apolipoprotein molecule. Alternatively, the apolipoprotein and / or the moiety to be labeled can be derivatized to expose or attach additional reactive functional groups.

[0144] In some embodiments, a fusion protein comprising a polypeptide functional moiety is synthesized using a recombinant expression system. Generally, this involves creating a nucleic acid (e.g., DNA) sequence encoding the apolipoprotein and the functional moiety such that the two polypeptides are in-frame upon expression, placing the DNA under the control of a promoter, expressing the protein in a host cell, and isolating the expressed protein.

[0145] The nucleic acid encoding the chimeric apolipoprotein can be incorporated into a recombinant expression vector in a form suitable for expression in a host cell. As used herein, an "expression vector" is a nucleic acid that can be transcribed and translated into a polypeptide when introduced into a suitable host cell. The vector can also include regulatory sequences, such as promoters, enhancers, or other expression control elements (e.g., polyadenylation signals). Such regulatory sequences are known to those skilled in the art (see, e.g., Goeddel, 1990, Gene Expression Technology: Meth. Enzymol. 185, Academic Press, San Diego, Calif.; Berger and Kimmel, Guide to Molecular Cloning Techniques, Methods in Enzymology 152 Academic Press, Inc., San Diego, Calif.; Sambrook et al., 1989, Molecular Cloning--A Laboratory Manual (2nd ed.) Vol. 1-3, Cold Spring Harbor Laboratory, Cold Spring Harbor Press, NY, etc.).

[0146] In some embodiments, the apolipoprotein has been modified such that when incorporated into the complexes of the present disclosure, the modification will increase the stability of the complex, confer targeting ability, or increase capacity. In one embodiment, the modification includes introducing cysteine residues into the apolipoprotein molecule to allow for the formation of intra- or intermolecular disulfide bonds, for example, by site-directed mutagenesis. In another embodiment, an intermolecular linkage is formed between apolipoprotein molecules using a chemical crosslinker to enhance the stability of the complex. Intermolecular crosslinking prevents or reduces the dissociation of apolipoprotein molecules from the complex and / or prevents replacement by endogenous apolipoprotein molecules in an individual to whom the complex is administered. In other embodiments, the apolipoprotein is modified by chemical derivatization of one or more amino acid residues or by site-directed mutagenesis to confer targeting ability or recognition ability for cell surface receptors.

[0147] By engineering receptor recognition properties into the apolipoprotein, the complex can be targeted to specific cell surface receptors. For example, the complex can be targeted to specific cell types known to contain a particular type of infectious pathogen, for example, by modifying the apolipoprotein to enable it to interact with receptors on the surface of the target cell type. For example, the complex can be targeted to macrophages by altering the apolipoprotein to confer recognition by macrophage class A scavenger receptor (SR-A). The apolipoprotein can be modified by site-directed mutagenesis to replace one or more positively charged amino acids with neutral or negatively charged amino acids to confer SR-A binding ability to the complex. SR-A recognition can also be conferred by preparing chimeric apolipoproteins that contain an N-terminal or C-terminal extension of a ligand with SR-A recognition or an amino acid sequence with a high concentration of negatively charged residues. Complexes containing apolipoproteins can also interact with apolipoprotein receptors, such as, but not limited to, the ABCA1 receptor, the ABCG1 receptor, Megalin, Cubulin, and HDL receptors (e.g., SR-B1).

[0148] 6.1.2.2. Apolipoprotein mimetics

[0149] Peptides, peptide analogs, and agonists that mimic the activity of apolipoproteins (collectively referred to herein as "apolipoprotein peptidomimetics") can also be used alone or in combination with one or more other lipid-binding proteins in the complexes described herein. Peptides and peptide analogs corresponding to apolipoproteins and mimetics of ApoA-I, ApoA-I M, non-limiting examples of agonists of ApoA-II, ApoA-IV, and ApoE activities are disclosed in U.S. Patent Nos. 6,004,925, 6,037,323, and 6,046,166 (issued to Dasseux et al.), U.S. Patent No. 5,840,688 (issued to Tso), U.S. Patent No. 6,743,778 (issued to Kohno), U.S. Publication Nos. 2004 / 0266671, 2004 / 0254120, 2003 / 0171277, and 2003 / 0045460 (Fogelman), U.S. Publication No. 2006 / 0069030 (Bachovchin), U.S. Publication No. 2003 / 0087819 (Bielicki), U.S. Publication No. 2009 / 0081293 (Murase et al.), and PCT Publication No. WO / 2010 / 093918 (Dasseux et al.), the disclosures of which are incorporated herein by reference in their entireties. These peptides and peptide analogs can consist of L-amino acids or D-amino acids or a mixture of L- and D-amino acids. They can also include one or more non-peptide or amide bonds, such as one or more well-known peptide / amide bioisosteres. Such apolipoprotein peptide mimetics can be synthesized or manufactured using any peptide synthesis techniques known in the art, including, for example, the techniques described in U.S. Patent Nos. 6,004,925, 6,037,323, and 6,046,166.

[0150] In some embodiments, the lipocalin molecule comprises an apolipoprotein peptide mimetic molecule and optionally one or more apolipoprotein molecules, such as those described above.

[0151] In some embodiments, the apolipoprotein peptide mimetic molecule comprises an ApoA-I peptide mimetic, an ApoA-II peptide mimetic, an ApoA-IV peptide mimetic, or an ApoE peptide mimetic, or a combination thereof.

[0152] 6.1.3. Amphiphilic Molecules

[0153] Amphiphilic molecules are molecules that have both hydrophobic (non-polar) elements and hydrophilic (polar) elements. Amphiphilic molecules that can be used in the complexes described herein include lipids (e.g., as described in Section 6.1.3.1), detergents (e.g., as described in Section 6.1.3.2), fatty acids (e.g., as described in Section 6.1.3.3), and non-polar molecules covalently linked to polar molecules (such as, but not limited to, sugars or nucleic acids) and sterols (e.g., as described in Section 6.1.3.4).

[0154] The complex can comprise a single class of amphiphilic molecules (e.g., a single type of phospholipid or a mixture of phospholipids), or can comprise a combination of multiple classes of amphiphilic molecules (e.g., phospholipids and detergents). The complex can comprise one amphiphilic molecule or a combination of amphiphilic molecules configured to facilitate the solubilization of lipid-binding protein molecules.

[0155] In some embodiments, the amphiphilic molecules comprised include phospholipids, detergents, fatty acids, nonpolar moieties covalently linked to sugars or sterols, or combinations thereof (e.g., selected from the types of amphiphilic molecules discussed above).

[0156] In some embodiments, the amphiphilic molecules include or consist of phospholipid molecules. In some embodiments, the phospholipid molecules include negatively charged phospholipids, neutral phospholipids, positively charged phospholipids, or combinations thereof. In some embodiments, the phospholipid molecules contribute 1 - 3 net charges per apolipoprotein molecule in the complex. In some embodiments, the net charge is a negative net charge. In some embodiments, the net charge is a positive net charge. In some embodiments, the phospholipid molecules consist of a combination of negatively charged phospholipids and neutral phospholipids. In some embodiments, the molar ratio of negatively charged phospholipids to neutral phospholipids ranges from 1:1 to 1:3. In some embodiments, the molar ratio of negatively charged phospholipids to neutral phospholipids is about 1:1 or about 1:2.

[0157] In some embodiments, the amphiphilic molecules comprise neutral phospholipids and negatively charged phospholipids in a weight ratio of 95:5 to 99:1.

[0158] 6.1.3.1. Lipids

[0159] Complexes based on lipid-binding proteins can include one or more lipids. In various embodiments, the one or more lipids can be saturated and / or unsaturated, natural and / or synthetic, charged or uncharged, zwitterionic or non-zwitterionic. In some embodiments, the lipid molecules (e.g., phospholipid molecules) can together contribute 1 - 3 (e.g., 1 - 3, 1 - 2, 2 - 3, 1, 2, or 3) net charges per lipid-binding protein molecule in the complex. In some embodiments, the net charge is negative. In other embodiments, the net charge is positive.

[0160] In some embodiments, the lipids include phospholipids. The phospholipids can have two identical or different acyl chains (e.g., chains with different numbers of carbon atoms, different degrees of saturation between the acyl chains, different branching of the acyl chains, or combinations thereof). The lipids can also be modified to include a fluorescent probe (e.g., as described at avantilipids.com / product-category / products / fluorescent-lipids / ). Preferably, the lipids include at least one phospholipid.

[0161] The phospholipid may have an unsaturated or saturated acyl chain having from about 6 to about 24 carbon atoms (e.g., 6-20, 6-16, 6-12, 12-24, 12-20, 12-16, 16-24, 16-20, or 20-24). In some embodiments, the phospholipid used in the complexes of the present disclosure has one or two acyl chains of 12, 14, 16, 18, 20, 22, or 24 carbons (e.g., two acyl chains of the same length or two acyl chains of different lengths).

[0162] Table 1 below provides non-limiting examples of acyl chains present in common fatty acids that may be included in phospholipids:

[0163]

[0164] Lipids that may be present in the complexes of the present disclosure include, but are not limited to, small alkyl chain phospholipids, lecithin phosphatidylcholine, soy phosphatidylcholine, dipalmitoyl phosphatidylcholine, dimyristoyl phosphatidylcholine, distearoyl phosphatidylcholine 1-myristoyl-2-palmitoyl phosphatidylcholine, 1-palmitoyl-2-myristoyl phosphatidylcholine, 1-palmitoyl-2-stearoyl phosphatidylcholine, 1-stearoyl-2-palmitoyl phosphatidylcholine, dioleoyl phosphatidylcholine, dioleoyl phosphatidylethanolamine, dilauroyl phosphatidylglycerol phosphatidylcholine, phosphatidylserine, phosphatidylethanolamine, phosphatidylinositol, phosphatidylglycerol, diphosphatidylglycerol, such as dimyristoyl phosphatidylglycerol, dipalmitoyl phosphatidylglycerol, distearoyl phosphatidylglycerol, dioleoyl phosphatidylglycerol, dimyristoyl phosphatidic acid, dipalmitoyl phosphatidic acid, dimyristoyl phosphatidylethanolamine, dipalmitoyl phosphatidylethanolamine, dimyristoyl phosphatidylserine, dipalmitoyl phosphatidylserine, cephalosphingomyelin, sphingomyelin, palmitoyl sphingomyelin, dipalmitoyl sphingomyelin, egg sphingomyelin, milk sphingomyelin, phytosphingomyelin, distearoyl sphingomyelin, dipalmitoyl phosphatidylglycerol salt, phosphatidic acid, galactocerebroside, ganglioside, cerebroside, dilauroyl phosphatidylcholine, (1,3)-D-mannosyl-(1,3)glycerol diester, aminophenyl glycoside, 3-cholesteryl-6'-(glycosylthio)hexyl ether glycolipid, cholesterol and its derivatives. Synthetic lipids, such as synthetic palmitoyl sphingomyelin or N-palmitoyl-4-hydroxysphinganine-1-phosphocholine (a phytosphingomyelin) can be used to minimize lipid oxidation.

[0165] In some embodiments, the lipocalin-based complex comprises two types of phospholipids: neutral lipids such as phosphatidylcholine and / or sphingomyelin (abbreviated as SM), and charged phospholipids (such as negatively charged phospholipids). "Neutral" phospholipids have a net charge of approximately zero at physiological pH. In many embodiments, the neutral phospholipids are zwitterionic, although other types of net-neutral phospholipids are known and can be used. In some embodiments, the molar ratio of charged phospholipids (such as negatively charged phospholipids) to neutral phospholipids ranges from 1:1 to 1:3, such as approximately 1:1, approximately 1:2, or approximately 1:3.

[0166] The neutral phospholipids can comprise, for example, one or both of phosphatidylcholine and / or SM, and can optionally comprise other neutral phospholipids. In some embodiments, the neutral phospholipids comprise phosphatidylcholine but not SM. In other embodiments, the neutral phospholipids comprise SM but not phosphatidylcholine. In still other embodiments, the neutral phospholipids comprise phosphatidylcholine and SM. All of these specific exemplary embodiments can comprise neutral phospholipids in addition to phosphatidylcholine and / or SM, but in many embodiments do not comprise such additional neutral phospholipids.

[0167] As used herein, the term "SM" includes sphingomyelin derived or obtained from natural sources, as well as analogs and derivatives of naturally occurring SM that are not affected by LCAT hydrolysis, just like naturally occurring SM. SM is a phospholipid with a structure very similar to phosphatidylcholine, but unlike phosphatidylcholine, it does not have a glycerol backbone and thus no ester bonds connecting acyl chains. Instead, SM has a ceramide backbone with amide bonds connecting acyl chains. SM can be obtained from, for example, milk, eggs, or the brain. SM analogs or derivatives can also be used. Non-limiting examples of useful SM analogs and derivatives include, but are not limited to, palmitoyl sphingomyelin, N-palmitoyl-4-hydroxysphinganine-1-phosphocholine (a phytosphingomyelin), palmitoyl sphingomyelin, stearoyl sphingomyelin, D-erythro-N-16:0-sphingomyelin and its dihydro isomer, D-erythro-N-16:0-dihydrosphingomyelin. Synthetic SM, such as synthetic palmitoyl sphingomyelin or N-palmitoyl-4-hydroxysphinganine-1-phosphocholine (phytosphingomyelin), can be used to produce a more homogeneous complex than sphingolipids from animal sources and with fewer contaminants and / or oxidation products. U.S. Publication No. 2016 / 0075634 describes methods for synthesizing SM.

[0168] Sphingomyelin isolated from natural sources can be artificially enriched with a specific saturated or unsaturated acyl chain. For example, milk sphingomyelin (Avanti Phospholipid, Alabaster, Ala.) is characterized by long saturated acyl chains (i.e., the acyl chain has 20 or more carbon atoms). In contrast, egg sphingomyelin is characterized by short saturated acyl chains (i.e., the acyl chain has fewer than 20 carbon atoms). For example, although only about 20% of milk sphingomyelin contains C16:0 (16 carbons, saturated) acyl chains, about 80% of egg sphingomyelin contains C16:0 acyl chains. Using solvent extraction, the composition of milk sphingomyelin can be enriched to have an acyl chain composition comparable to that of egg sphingomyelin, and vice versa.

[0169] SM can be semi-synthetic, and thus it has a specific acyl chain. For example, milk sphingomyelin can first be purified from milk, and then a specific acyl chain, such as a C16:0 acyl chain, can be cleaved and replaced with another acyl chain. SM can also be entirely synthesized, for example, by large-scale synthesis. See, for example, Dong et al., U.S. Patent No. 5,220,043, entitled Synthesis of D-erythro-sphingomyelin, issued June 15, 1993; Weis, 1999, Chem. Phys. Lipids 102 (1-2):3-12. SM can be fully synthetic, for example, as described in U.S. Publication No. 2014 / 0275590.

[0170] Optionally, the length and saturation of the acyl chains constituting the semi-synthetic or synthetic SM can be varied. The acyl chains can be saturated or unsaturated and can contain from about 6 to about 24 carbon atoms. Each chain can contain the same number of carbon atoms, or alternatively, each chain can contain a different number of carbon atoms. In some embodiments, the semi-synthetic or synthetic SM contains mixed acyl chains such that one chain is saturated and one chain is unsaturated. In such mixed acyl chain SMs, the chain lengths can be the same or different. In other embodiments, the acyl chains of the semi-synthetic or synthetic SM are either all saturated or all unsaturated. Again, the chains can contain the same or different numbers of carbon atoms. In some embodiments, the two acyl chains constituting the semi-synthetic or synthetic SM are the same. In certain embodiments, the chains correspond to the acyl chains of natural fatty acids (e.g., oleic acid, palmitic acid, or stearic acid). In another embodiment, SMs with saturated or unsaturated functionalized chains are used. In another specific embodiment, both acyl chains are saturated and contain 6 to 24 carbon atoms. Table 1 above provides non-limiting examples of acyl chains present in common fatty acids that can be included in semi-synthetic and synthetic SMs.

[0171] In some embodiments, the SM is palmitoyl SM, such as synthetic palmitoyl SM having a C16:0 acyl chain, or egg SM, which includes palmitoyl SM as a major component.

[0172] In a specific embodiment, functionalized SMs are used, such as phytosphingomyelin.

[0173] Lecithin can be derived or isolated from natural sources or can be obtained synthetically. Examples of suitable lecithins isolated from natural sources include, but are not limited to, egg phosphatidylcholine and soy phosphatidylcholine. Additional non-limiting examples of suitable lecithins include dipalmitoyl phosphatidylcholine, dimyristoyl phosphatidylcholine, distearoyl phosphatidylcholine, 1-myristoyl-2-palmitoyl phosphatidylcholine, 1-palmitoyl-2-myristoyl phosphatidylcholine, 1-palmitoyl-2-stearoyl phosphatidylcholine, 1-stearoyl-2-palmitoyl phosphatidylcholine, 1-palmitoyl-2-oleoyl phosphatidylcholine, 1-oleoyl-2-palmitoyl phosphatidylcholine, dioleoyl phosphatidylcholine, and their ether derivatives or analogs.

[0174] It is possible to enrich lecithin derived or separated from natural sources to include specific acyl chains. In embodiments using semi-synthetic or synthetic lecithin, the identity of the acyl chains can optionally be altered, as discussed above with respect to SM. In some embodiments of the complexes described herein, the two acyl chains on the lecithin are the same. In some embodiments of the complexes comprising SM and lecithin, the acyl chains of both SM and lecithin are the same. In certain embodiments, the acyl chains correspond to the acyl chains of myristic acid, palmitic acid, oleic acid, or stearic acid.

[0175] The complexes of the present disclosure can include one or more negatively charged phospholipids (e.g., alone or in combination with one or more neutral phospholipids). As used herein, a "negatively charged phospholipid" is a phospholipid that has a net negative charge at physiological pH. The negatively charged phospholipid can include a single type of negatively charged phospholipid, or a mixture of two or more different negatively charged phospholipids. In some embodiments, the negatively charged phospholipid is a negatively charged glycerophospholipid. Specific examples of suitable negatively charged phospholipids include, but are not limited to, 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)], phosphatidylglycerol, phosphatidylinositol, phosphatidylserine, phosphatidic acid, and salts thereof (e.g., sodium salt or potassium salt). In some embodiments, the negatively charged phospholipid includes one or more of phosphatidylinositol, phosphatidylserine, phosphatidylglycerol, and / or phosphatidic acid. In certain embodiments, the negatively charged phospholipid comprises or consists of a salt of phosphatidylglycerol or a salt of phosphatidylinositol. In another specific embodiment, the negatively charged phospholipid includes 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] or DPPG or a salt thereof or consists of 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] or DPPG or a salt thereof.

[0176] The negatively charged phospholipid can be obtained from natural sources or prepared by chemical synthesis. In embodiments using synthetic negatively charged phospholipids, the identity of the acyl chains can optionally be altered, as discussed above with respect to SM. In some embodiments of the complexes of the present disclosure, the two acyl chains on the negatively charged phospholipid are the same. In some embodiments, the acyl chains of all types of phospholipids included in the complexes of the present disclosure are the same. In certain embodiments, the complex comprises a negatively charged phospholipid and / or SM, both of which have C16:0 or C16:1 acyl chains. In certain embodiments, the fatty acid moiety of the SM is predominantly C16:1 palmitoyl. In one specific embodiment, the acyl chains of the charged phospholipid, lecithin, and / or SM correspond to the acyl chains of palmitic acid. In yet another specific embodiment, the acyl chains of the charged phospholipid, lecithin, and / or SM correspond to the acyl chains of oleic acid.

[0177] Examples of positively charged phospholipids that may be included in the complexes of the present disclosure include N1-[2-((1S)-1-[(3-aminopropyl)amino]-4-[bis(3-aminopropyl)amino]butylformamido)ethyl]-3,4-di[oleyloxy]-benzamide, 1,2-di-O-octadecenyl-3-trimethylammonium propane, 1,2-dimyristoyl-sn-glycero-3-ethylphosphocholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-ethylphosphocholine, 1,2-dioleoyl-sn-glycero-3-ethylphosphocholine, 1,2-distearoyl-sn-glycero-3-ethylphosphocholine, 1,2-dipalmitoyl-sn-glycero-3-ethylphosphocholine, 1,2-dimyristoyl-sn-glycero-3-ethylphosphocholine, 1,2-dilauroyl-sn-glycero-3-ethylphosphocholine, 1,2-dilauroyl-sn-glycero-3-ethylphosphocholine, 1,2-dioleoyl-3-dimethylammonium-propane 1,2-dimyristoyl-3-dimethylammonium-propane, 1,2-dipalmitoyl-3-dimethylammonium-propane, N-(4-carboxybenzyl)-N,N-dimethyl-2,3-bis(oleyloxy)propane-1-ammonium, 1,2-dioleoyl-3-trimethylammonium-propane, 1,2-dioleoyl-3-trimethylammonium-propane, 1,2-stearoyl-3-trimethylammonium-propane, 1,2-dipalmitoyl-3-trimethylammonium-propane, 1,2-dimyristoyl-3-trimethylammonium propane, N-[1-(2,3-dimyristyloxy)propyl]-N,N-dimethyl-N-(2-hydroxyethyl)ammonium bromide, N,N,N-trimethyl-2-bis[(1-oxo-9-octadecenyl)oxy]-(Z,Z)-1-propenylammonium methyl sulfate and its salts (such as chloride or bromide salts).

[0178] The lipids used preferably have a purity of at least 95% and / or a low content of oxidizing agents (such as, but not limited to, peroxides). Lipids obtained from natural sources preferably have a lower polyunsaturated fatty acid moiety and / or a fatty acid moiety that is less prone to oxidation. The level of oxidation in the sample can be determined using the iodometric method, which provides a peroxide value expressed as milliequivalents of iodine isolated per kilogram of sample, abbreviated as meq O / kg. See, for example, Gray, 1978, Measurement of Lipid Oxidation: A Review, Journal of the American Oil Chemists Society 55:539-545; Heaton, F.W. and Ur, Improved Iodometric Methods for the Determination of Lipid Peroxides, 1958, Journal of the Science of Food and Agriculture 9:781-786. Preferably, the oxidation level or peroxide level is low, such as less than 5 meq O / kg, less than 4 meq O / kg, less than 3 meq O / kg or less than 2 meq O / kg.

[0179] In some embodiments, the complex can include a small amount of additional lipid. Virtually any type of lipid can be used, including but not limited to lysophospholipids, galactocerebrosides, gangliosides, cerebrosides, glycerides, triglycerides, and sterols and sterol derivatives (such as, phytosterols, animal sterols (such as cholesterol), or sterol derivatives (such as cholesterol derivatives)). For example, the complexes of the present disclosure can contain cholesterol or cholesterol derivatives, such as cholesterol esters. Cholesterol derivatives can also be substituted cholesterol or substituted cholesterol esters. The complexes of the present disclosure can also contain oxidized sterols (such as, but not limited to, oxidized cholesterol or oxidized sterol derivatives (such as, but not limited to, oxidized cholesterol esters)). In some embodiments, the complex does not include cholesterol and / or its derivatives (such as cholesterol esters or oxidized cholesterol esters).

[0180] 6.1.3.2. Detergent

[0181] The complex may contain one or more detergents. The detergents can be zwitterionic, nonionic, cationic, anionic, or a combination thereof. Exemplary zwitterionic detergents include 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate (CHAPS), 3-[(3-cholamidopropyl)dimethylammonio]-2-hydroxy-1-propanesulfonate (CHAPSO), and N,N-dimethyldodecylamine N-oxide (LDAO). Exemplary nonionic detergents include D-(+)-trehalose 6-monooleate, N-octanoyl-N-methylglucamine, N-nonanoyl-N-methylglucamine, N-decanoyl-N-methylglucamine, 1-(7Z-hexadecenoyl)-rac-glycerol, 1-(8Z-hexadecenoyl)-rac-glycerol, 1-(8Z-heptadecenoyl)-rac-glycerol, 1-(9Z-hexadecenoyl)-rac-glycerol, 1-decanoyl-rac-glycerol. Exemplary cationic detergents include (S)-O-methyl-serine dodecylamide hydrochloride, dodecylammonium chloride, decyltrimethylammonium bromide, and cetyltrimethylammonium sulfate. Exemplary anionic detergents include cholesterol hemisuccinate, cholate, alkyl sulfates, and alkyl sulfonates.

[0182] 6.1.3.3. Fatty Acids

[0183] The complex may contain one or more fatty acids. The one or more fatty acids may include short-chain fatty acids having an aliphatic tail with 5 or fewer carbon atoms (e.g., butyric acid, isobutyric acid, valeric acid, or isovaleric acid), medium-chain fatty acids having an aliphatic tail with 6 to 12 carbon atoms (e.g., caproic acid, caprylic acid, capric acid, or lauric acid), long-chain fatty acids having an aliphatic tail with 13 to 21 carbon atoms (e.g., myristic acid, palmitic acid, stearic acid, or arachidic acid), very-long-chain fatty acids having an aliphatic tail with 22 or more carbon atoms (e.g., behenic acid, lignoceric acid, or cerotic acid), or combinations thereof. The one or more fatty acids can be saturated (e.g., caprylic acid, capric acid, lauric acid, myristic acid, palmitic acid, stearic acid, arachidic acid, behenic acid, lignoceric acid, or cerotic acid), unsaturated (e.g., myristoleic acid, palmitoleic acid, petroselinic acid, oleic acid, elaidic acid, vaccenic acid, linoleic acid, eleostearic acid, α-linolenic acid, arachidonic acid, eicosapentaenoic acid, erucic acid, or docosahexaenoic acid), or combinations thereof. Unsaturated fatty acids can be cis or trans fatty acids. In some embodiments, the unsaturated fatty acids used in the complexes of the present disclosure are cis fatty acids.

[0184] 6.1.3.4. Nonpolar Molecules and Sterols Linked to Sugars

[0185] The complex may contain one or more amphiphilic molecules, which include nonpolar molecules or moieties (such as hydrocarbon chains, acyl or diacyl chains) or sterols (such as cholesterol) linked to a sugar (such as a monosaccharide like glucose or galactose, or a disaccharide like maltose or trehalose). The sugar may be a modified sugar or a substituted sugar. Exemplary amphiphilic molecules containing nonpolar molecules linked to a sugar include dodecan-2-yloxy-β-D-maltoside, tridecan-3-yloxy-β-D-maltoside, tridecan-2-yloxy-β-D-maltoside, n-dodecyl-β-D-maltoside (DDM), n-octyl-β-D-glucoside, n-nonyl-β-D-glucoside, n-decyl-β-D-maltoside, n-dodecyl-β-D-maltopyranoside, 4-n-dodecyl-α,α-trehalose, 6-n-dodecyl-α,α-trehalose, and 3-n-dodecyl-α,α-trehalose.

[0186] In some embodiments, the nonpolar moiety is an acyl or diacyl chain.

[0187] In some embodiments, the sugar is a modified sugar or a substituted sugar.

[0188] 6.1.4. Formulation

[0189] The lipocalin-based complex can be formulated for the intended route of administration, for example, according to techniques known in the art (such as those described in Allen et al., eds., 2012, Remington: The Science and Practice of Pharmacy, 22nd Edition, Pharmaceutical Press, London, UK).

[0190] CER-001 for infusion administration can be formulated with sucrose and mannitol excipients in phosphate buffer, as described in, for example, WO 2012 / 109162.

[0191] 6.2. Subject Population

[0192] Subjects that can be treated according to the methods described herein are preferably mammals, most preferably humans.

[0193] In some aspects, the subject has a condition that includes inflammation, such as acute and / or excessive inflammation.

[0194] In some aspects, the subject has HLH or is at risk of developing HLH. In some embodiments, the subject has HLH. In other embodiments, the subject is at risk of developing HLH. HLH can be, for example, familial HLH, or HLH secondary to a malignant disease (such as acute leukemia or lymphoma) or a non-malignant disease (such as an autoimmune disease, such as rheumatoid arthritis, or an infection, such as a viral infection or a bacterial infection). In some embodiments, HLH is virus-induced HLH, such as caused by dengue infection, herpes simplex infection, or Epstein-Barr virus infection.

[0195] In some aspects, the subject has dengue hemorrhagic fever or dengue shock syndrome or is at risk of developing dengue hemorrhagic fever or dengue shock syndrome, such as the subject is infected with dengue (e.g., the subject has dengue). Dengue, dengue hemorrhagic fever, and dengue shock syndrome are described in Dengue haemorrhagic fever: diagnosis, treatment, prevention and control, 2 nd Edition, World Health Organization (1997) (the content of which is incorporated herein by reference in its entirety). A subject with dengue is at risk of developing more severe dengue hemorrhagic fever and even more severe dengue shock syndrome.

[0196] In some aspects, the subject has a herpes simplex infection.

[0197] In some embodiments, the subject has a Sequential Organ Failure Assessment (SOFA) score of 1 to 4 before treatment with the lipocalin-based complex, such as a score of 1, 2, 3, or 4 (see Vincent et al. 1996, Intensive Care Med, 22:707–710).

[0198] In some embodiments, the subject has acute kidney injury (AKI) or is at risk of developing AKI, such as due to a viral infection (e.g., dengue infection).

[0199] In certain aspects, the subject may have cytokine release syndrome (CRS) or be at risk of developing CRS, and / or require a reduction in the serum level of one or more inflammatory markers (such as IL-6). In some embodiments, the subject has CRS. In some embodiments, the subject has CRS secondary to an infection, such as a viral infection (e.g., dengue infection). In still other embodiments, the subject is at risk of developing CRS, such as due to an infection (e.g., dengue).

[0200] In another aspect, the subject is a subject in need of reducing the serum levels of one or more inflammatory markers, such as a subject having elevated levels of one or more inflammatory markers compared to normal levels. Exemplary inflammatory cytokines include interleukin 6 (IL-6), C-reactive protein, D-dimer, ferritin, interleukin 8 (IL-8), granulocyte-macrophage colony-stimulating factor (GM-CSF), monocyte chemoattractant protein (MCP) 1, and tumor necrosis factor α (TNFα). In some embodiments, one or more cytokines include IL-6. In some embodiments, the one or more cytokines include a combination of the foregoing, such as 2, 3, 4, 5, 6, 7, or all 8 of interleukin 6 (IL-6), C-reactive protein, D-dimer, ferritin, interleukin 8 (IL-8), granulocyte-macrophage colony-stimulating factor (GM-CSF), monocyte chemoattractant protein (MCP) 1, and tumor necrosis factor α (TNFα).

[0201] 6.3. Administration Regimen

[0202] The methods of the present disclosure generally require multiple administrations of the lipocalin-based complex (e.g., CER-001), e.g., 2 to 10 individual doses, although in some embodiments, a single dose may be used. In some embodiments, the administration regimen may include two or more, three or more, or four or more doses of the lipocalin-based complex (e.g., CER-001), e.g., five, six, seven, eight, nine, ten, eleven, twelve, or more than twelve doses. In some embodiments, the administration regimen includes or consists of a single dose. In some embodiments, the administration regimen includes or consists of two individual doses. In some embodiments, the administration regimen includes or consists of three individual doses. In some embodiments, the administration regimen includes or consists of four individual doses.

[0203] In some embodiments, the lipocalin-based complex is administered according to an induction regimen and an optional consolidation regimen, as described respectively in Sections 6.3.1 and 6.3.2. In some embodiments, the lipocalin-based complex may be administered in a single stage, e.g., according to the administration regimens described in this section. In some embodiments, the subject is not treated with the lipocalin-based complex according to a maintenance regimen, e.g., a regimen that includes long-term (e.g., one month or longer) administration of the lipocalin-based complex.

[0204] The administration regimen of the lipocalin-based complex (e.g., CER-001) may last up to one week, one week, or more than one week (e.g., two weeks).

[0205] For example, a dosing regimen for a lipocalin-based complex (e.g., CER-001) can include administering:

[0206] - Five doses of CER-001 within one week;

[0207] - Six doses of CER-001 within one week;

[0208] - Seven doses of CER-001 within one week;

[0209] - Ten doses of CER-001 within two weeks;

[0210] - Twelve doses of CER-001 within two weeks;

[0211] - Fourteen doses of CER-001 within two weeks.

[0212] In one embodiment, the method of the present disclosure includes administering seven doses of CER-001 within one week, such as on days 1, 2, 3, 4, 5, 6, and 7.

[0213] In some embodiments of the method of the present disclosure, the dosing intervals for multiple doses of the lipocalin-based complex (e.g., CER-001) are no more than one day. For example, in some embodiments, the dosing intervals for two or more individual doses are about 12 hours. In some embodiments, the dosing interval for two individual doses is about 12 hours. In other embodiments, the dosing interval for three individual doses is about 12 hours. In other embodiments, the dosing interval for two individual doses is about 12 hours and the dosing interval for the third individual dose is about one day. In other embodiments, the dosing intervals for three individual doses are about 12 hours and the dosing interval for the fourth individual dose is about one day.

[0214] In some embodiments of the method of the present disclosure, a lipocalin-based complex (e.g., CER-001) is administered to a subject at 0 and 12 hours (e.g., within a time period of 0.5 to 1 hour), for example, at a dose of 10 mg / kg or 15 mg / kg. In some embodiments of the method of the present disclosure, a lipocalin-based complex (e.g., CER-001) is administered to a subject (e.g., within a time period of 0.5 to 1 hour) at 0 and 12, 24 and 48 hours, for example, at a dose of 10 mg / kg or 15 mg / kg.

[0215] In some embodiments of the disclosed methods, the lipocalin complex (e.g., CER-001) is administered daily, e.g., for at least 5 days, at least 6 days, at least 7 days, or more than 7 days (e.g., up to one week or up to two weeks daily). In other embodiments, the lipocalin complex (e.g., CER-001) is administered less frequently, e.g., every other day, twice a week, three times a week, or once a week.

[0216] In practice, a dosing window can be provided, e.g., to accommodate minor variations in a multiple-dosing per week schedule. For example, a window of ±2 days or ±1 day around the dosing date can be used.

[0217] Based on the lipocalin complex (e.g., CER-001) for a predetermined period, e.g., one week. Alternatively, the lipocalin complex (e.g., CER-001) can be administered continuously until one or more symptoms of the disease (e.g., HLH or dengue shock syndrome) are alleviated, or until the serum levels of one or more inflammatory markers are reduced, e.g., reduced to normal levels or reduced relative to the baseline value of the subject, e.g., the baseline value measured before starting treatment with the lipocalin complex (e.g., CER-001). The reference or "normal" levels of various inflammatory markers are known in the art. For example, the Mayo Clinic Laboratories test catalog (mayocliniclabs.com / test-catalog) provides the following reference values: IL-6: ≤1.8 pg / ml; C-reactive protein: ≤8.0 mg / ml; D-dimer: ≤500 ng / mL fibrinogen equivalent units (FEU); ferritin: 24 - 336 mcg / L (male), 11 - 307 mcg / L (female); IL-8 < 57.8 pg / mL; TNF-α < 5.6 pg / mL.

[0218] The methods of the present disclosure generally include administering a high dose of the lipocalin complex (e.g., CER-001). The high dose can be a collection of multiple individual doses (e.g., two, three, four, five, six, seven, eight, nine, or ten individual doses), e.g., administered over one or more days (e.g., one day, two days, three days, four days, five days, six days, seven days, eight days, nine days, ten days, eleven days, twelve days, thirteen days, fourteen days, or fifteen days). In some embodiments, the individual doses of the high dose are administered daily, twice a day, or every two to three days.

[0219] In some embodiments, a high dose is an amount effective to increase the HDL and / or ApoA-I blood levels of a subject and / or improve the vascular endothelial function of the subject, e.g., as measured by circulating vascular cell adhesion molecule 1 (VCAM-1) and / or intercellular adhesion molecule 1 (ICAM-1) levels. In some embodiments, a high dose or a single dose is an amount that increases the HDL and / or ApoA-I levels of the subject by at least 25%, at least 30%, or at least 35% within 2 to 4 hours after administration.

[0220] In some embodiments, a high dose is an amount effective to reduce the serum levels of one or more inflammatory markers, e.g., one or more of IL-6, C-reactive protein, D-dimer, ferritin, IL-8, GM-CSF, and MCP1 TNF-α. In some embodiments, the serum levels of one or more inflammatory markers are reduced from an elevated range to a normal range, and / or reduced by at least 20%, at least 40%, or at least 60%.

[0221] The dose of the lipocalin-based complex (e.g., CER-001) administered to a subject (e.g., an individual dose that forms a high dose when aggregated with one or more other individual doses) can be in the range of 4 to 40 mg / kg based on protein weight (e.g., 10 to 40 mg / kg) (e.g., 5, 10, 15, 20, 25, 30, 35, or 40 mg / kg, or any range defined by any two of the foregoing values, e.g., 10 to 20 mg / kg, 15 to 25 mg / kg, 20 to 40 mg / kg, 25 to 35 mg / kg, or 30 to 40 mg / kg). As used herein, the expression "based on protein weight" means that the dose of the lipocalin-based complex (e.g., CER-001) to be administered to a subject is calculated based on the amount of ApoA-I in the lipocalin-based complex (e.g., CER-001) to be administered and the body weight of the subject. For example, a subject weighing 70 kg and to receive a dose of 20 mg / kg of CER-001 will receive an amount of CER-001 that provides 1400 mg of ApoA-I (70 kg × 20 mg / kg).

[0222] In other aspects, the lipocalin-based complex (e.g., CER-001) can be administered on a unit dose basis. In some embodiments, the unit dose used in the methods of the present disclosure can be 300 mg to 4000 mg per administration (e.g., 600 mg to 4000 mg) based on protein weight.

[0223] The dosage of the lipocalin-based complex (e.g., CER-001) is 600 mg to 3000 mg, 800 mg to 3000 mg, 1000 mg to 2400 mg, or 1000 mg to 2000 mg per administration (based on protein weight).

[0224] The high dosage (e.g., the total amount of multiple individual dosages) of the lipocalin complex (e.g., CER-001) is 600 mg to 40 g (based on protein weight). In certain embodiments, the high dosage is 3 g to 35 g or 5 g to 30 g (based on protein weight).

[0225] The lipocalin-based complex (e.g., CER-001) is preferably administered by intravenous infusion. For example, the stock solution of CER-001 can be diluted in physiological saline (e.g., normal saline (0.9% NaCl)) to a total volume between 125 and 250 ml. In some embodiments, the total volume for subjects weighing less than 80 kg is 125 ml, while the total volume for subjects weighing at least 80 kg is 250 ml. In some embodiments, the dosage of CER-001 is administered in a total volume of 250 ml. The lipocalin-based complex (e.g., CER-001) can be administered over a period of 1 hour to 24 hours. Depending on the needs of the subject, administration can be by slow infusion over more than one hour (e.g., up to 2 hours or up to 24 hours), rapid infusion for one hour or less, or a single bolus injection. In one embodiment, the lipocalin-based complex (e.g., CER-001) is administered within one hour, e.g., at a fixed rate of 125 ml / hr or 250 ml / hr using an infusion pump. In one embodiment, the dosage of the lipocalin-based complex (e.g., CER-001) is administered by infusion over 24 hours.

[0226] 6.3.1. Induction Regimen

[0227] In one embodiment, the induction regimen applicable to the methods of the present disclosure requires administration of multiple doses of the lipocalin-based complex (e.g., CER-001) on multiple consecutive days (e.g., three consecutive days).

[0228] In some embodiments, the induction regimen applicable to the methods of the present disclosure requires administration of the lipocalin-based complex (e.g., CER-001) twice a day, e.g., twice a day for multiple consecutive days. Administration twice a day can include, for example, two doses spaced approximately 12 hours apart or a morning dose and an evening dose (the interval can be greater than or less than 12 hours).

[0229] In one embodiment, the induction regimen comprises two doses of the lipocalin-based complex (e.g., CER-001) per day for three consecutive days.

[0230] The therapeutic dose range of the lipocalin-based complex (e.g., CER-001) administered by infusion in the induction regimen can be 4 to 40 mg / kg (e.g., 4 to 30 mg / kg) based on the protein weight (e.g., 4, 5, 6, 7, 8, 9, 10, 12, 15, 20, 25, 30, or 40 mg / kg, or any range defined by any two of the above values, e.g., 5 to 15 mg / kg, 10 to 20 mg / kg, or 15 to 25 mg / kg). In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used in the induction regimen is 5 mg / kg. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used in the induction regimen is 10 mg / kg. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used in the induction regimen is 15 mg / kg. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used in the induction regimen is 20 mg / kg. In some embodiments, the induction regimen comprises administering six doses of the lipocalin-based complex (e.g., CER-001) at a dose of 5 mg / kg, 10 mg / kg, 15 mg / kg, or 20 mg / kg over three days.

[0231] In other aspects, the lipocalin-based complex (e.g., CER-001) can be administered on a unit dose basis. The unit dose used during the induction phase can vary from 300 mg to 4000 mg (e.g., 300 mg to 3000 mg) per infusion (based on the protein weight).

[0232] In certain embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used during the induction phase is 300 mg to 1500 mg, 400 mg to 1500 mg, 500 mg to 1200 mg, or 500 mg to 1000 mg per infusion (based on the protein weight).

[0233] 6.3.2. Consolidation Regimen

[0234] The consolidation regimen applicable to the methods of the present disclosure requires administering one or more doses of the lipocalin-based complex (e.g., CER-001) after the induction regimen.

[0235] In one embodiment, the consolidation regimen comprises administering two doses of the lipocalin-based complex (e.g., CER-001). For example, the two doses can be administered approximately 12 hours apart, or as a morning dose and an evening dose (the interval can be greater or less than 12 hours).

[0236] In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) in the consolidation regimen can be administered on day 6 of the dosing schedule starting from the induction regimen on day 1. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) in the consolidation regimen can be administered on day 4 of the dosing schedule starting from the induction regimen on day 1. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) in the consolidation regimen can be administered on day 5 of the dosing schedule starting from the induction regimen on day 1. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) in the consolidation regimen can be administered on day 7 of the dosing schedule starting from the induction regimen on day 1.

[0237] The therapeutic dose range of the lipocalin-based complex (e.g., CER-001) administered by infusion in the consolidation regimen can be from 4 mg / kg to 40 mg / kg based on protein weight (e.g., 4 to 30 mg / kg) (e.g., 4, 5, 6, 7, 8, 9, 10, 12, 15, 20, 25, 30, or 40 mg / kg, or any range defined by any two of the above values, e.g., 5 to 15 mg / kg, 10 to 20 mg / kg, or 15 to 25 mg / kg). In some embodiments, the dose of the lipocalin complex (e.g., CER-001) used in the consolidation regimen is 5 mg / kg. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used in the consolidation regimen is 10 mg / kg. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used in the consolidation regimen is 15 mg / kg. In some embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used in the consolidation regimen is 20 mg / kg. In some embodiments, the consolidation regimen comprises administering two doses of the lipocalin-based complex (e.g., CER-001) in one day, with doses of 5 mg / kg, 10 mg / kg, 15 mg / kg, or 20 mg / kg respectively.

[0238] In other aspects, the lipocalin-based complex (e.g., CER-001) can be administered on a unit dose basis. The unit dose used during the consolidation phase can vary from 300 mg to 4000 mg per infusion (e.g., 300 mg to 3000 mg) (based on protein weight).

[0239] In certain embodiments, the dose of the lipocalin-based complex (e.g., CER-001) used during the consolidation phase is 300 mg to 1500 mg, 400 mg to 1500 mg, 500 mg to 1200 mg, or 500 mg to 1000 mg per infusion (based on protein weight).

[0240] The lipocalin-based complex (e.g., CER-001) can be administered during the consolidation phase in the same manner as described in Section 6.3, e.g., as a one-hour IV infusion.

[0241] 6.4. Combination Therapy

[0242] The lipocalin-based complex (e.g., CER-001) can be administered to the subjects described herein as a single therapy or as part of a combination therapy regimen. For example, the combination therapy may include a lipocalin-based complex (e.g., CER-001) in combination with the standard of care treatment for sepsis and / or AKI. See, e.g., Rhodes et al., 2017, Intensive Care Med 43:304–377; Dugar et al., 2020, Cleveland Clinic Journal of Medicine 87(1):53-64.

[0243] In some embodiments, the subject is treated with a lipocalin-based complex (e.g., CER-001) in combination with a fluid replacement therapy. In some embodiments, the subject is treated with a lipocalin-based complex (e.g., CER-001) in combination with an antibacterial agent. In some embodiments, the subject is treated with a lipocalin-based complex (e.g., CER-001) in combination with an antibiotic (e.g., ceftriaxone, meropenem, ceftazidime, cefotaxime, cefepime, piperacillin and tazobactam, ampicillin and sulbactam, imipenem and cilastatin, levofloxacin, or clindamycin). In some embodiments, the subject is treated with a lipocalin-based complex (e.g., CER-001) in combination with an antiviral agent. In some embodiments, the subject is treated with a lipocalin-based complex (e.g., CER-001) in combination with a blood pressure-elevating drug (e.g., norepinephrine or epinephrine). In some embodiments, the subject is treated with a lipocalin-based complex (e.g., CER-001) in combination with an immunosuppressant (e.g., tacrolimus or sirolimus).

[0244] In some embodiments, the combination therapy regimen can include one or more anti-IL-6 drugs and / or one or more other drugs for treating CRS, such as corticosteroids (e.g., methylprednisolone and / or dexamethasone). Exemplary anti-IL6 drugs include tocilizumab, cetuximab, olokizumab, elsilimomab, BMS-945429, sirukumab, rivacizumab, and CPSI-2364. In some embodiments, a lipocalin-based complex (e.g., CER-001) is co-administered with tocilizumab.

[0245] An antihistamine (e.g., diphenhydramine, cetirizine, fexofenadine, or loratadine) can be administered before administering the lipocalin-based complex (e.g., CER-001). The antihistamine can reduce the likelihood of an allergic reaction.

[0246] 7. Examples

[0247] 7.1. Example 1: Treatment of COVID-19 with CER-001

[0248] The SARS-CoV-2 virus can promote a life-threatening hyperinflammatory state in high-risk patients. Lipid profile remodelling, including a sharp decline in serum levels of apolipoprotein-AI (ApoA-I), is a hallmark of severe COVID-19. ApoA-I reduces pulmonary inflammation, modulates innate and adaptive immunity and prevents endothelial dysfunction and blood coagulation. This example describes a compassionate use trial in four subjects with COVID-19 cytokine storm who had progressive disease despite standard treatment. To raise ApoA-I to normal levels, the subjects received 2–4 infusions of CER-001 (10 mg / kg each). The injections were well tolerated and there were no serious adverse events. Three patients improved rapidly and were discharged 3–4 days after CER-001 infusion. The fourth patient received CER-001 during mechanical ventilation, had a transient improvement and then deteriorated with bacterial pneumonia. This study provides early safety and proof-of-concept data for the use of lipocalin-based complexes, such as CER-001, in the treatment of patients with virus-induced cytokine storm.

[0249] 7.1.1. Materials and methods

[0250] 7.1.1.1. Case history

[0251] Subject 1 is a 52-year-old male with the following medical history: IgA vasculitis, diabetes, and ischemic heart disease, and received a kidney transplant in 2018. He had received 3 doses of the mRNA COVID-19 vaccine but only developed weak anti-SARS-CoV-2 immunity (anti-spike antibody 15.5 BAU / mL). He developed COVID-19 symptoms (fever, diarrhea, and dyspnea) and was admitted to the transplant ward 8 days later. The oxygen saturation was 92%, and indoor air and oxygen supplementation (1 L / min) were initiated. Chest CT scan showed bilateral interstitial lung disease consistent with COVID-19 (parenchymal extension 25%), and SARS-CoV-2 (variant of concern (VOC) Delta) was identified by nasopharyngeal PCR. Blood tests showed hyperinflammation (ferritin 5,037 μg / L, C-reactive protein 34 mg / L), abnormal liver function tests (AST and ALT were 2.5 times and 3.5 times the upper limit of normal (ULN) values, respectively), and thrombocytopenia. Tacrolimus was administered, mycophenolate mofetil was discontinued, and dexamethasone (6 mg / day) and antibiotics were introduced. On day 2, blood tests showed pancytopenia and progression of hyperinflammation (ferritin 6,870 μg / L, C-reactive protein 55 mg / L). The subject received an infusion of the monoclonal anti-IL-6R antibody tocilizumab (8 mg / kg i.v.) and an infusion of a neutralizing monoclonal anti-SARS-Cov2 antibody (casirimivab / imdevimab). On day 4, the subject's ferritin increased to 19,219 μg / L, the subject's AST and ALT increased to 17 times and 14 times the ULN values, respectively, and the subject's arterial lactate was 2.7 mmol / L. Bone marrow aspirate showed hemophagocytosis. SARS-CoV-2 blood PCR was weakly positive. Worsening hypoxia required increased oxygenation (4 L / min; PaO2 62 mmHg), and CT scan showed progressive lung lesions typical of COVID-19 (50% parenchyma). Despite increasing dexamethasone to 10 mg / day, serum triglycerides and ferritin increased to 3.2 mmol / L and 27,394 μg / L, respectively, on day 6.

[0252] Subject 2 is a 38-year-old female with the following medical history: systemic lupus erythematosus, overweight, and kidney transplantation in 2011. She has received three doses of the mRNA COVID-19 vaccine but has not developed anti-SARS-CoV-2 immunity. She presented with COVID-19 symptoms (cough, chills, diarrhea, and fever) and was admitted to the transplant ward 10 days later. Nasopharyngeal PCR detected SARS-CoV2 VOC Omicron. At admission, SaO2 was 94%, and oxygen was administered at 9 L / min via a face mask. Chest CT scan showed typical COVID-19 lesions (50% extension). Blood tests revealed hepatitis with cytolysis and cholestasis (7 to 10 times the ULN values, respectively), acute kidney injury (KDIGO stage 1), and hyperinflammation (ferritin 2,000 μg / L, C-reactive protein 107 mg / L). High-flow oxygen therapy, awake prone position, dexamethasone (10 mg / day), tocilizumab (8 mg / kg once), and antibiotic treatment were initiated. Everolimus was discontinued, and tacrolimus was administered. On day 4, despite full treatment, high-flow oxygen therapy was still required, and the subject's hyperinflammatory state deteriorated (ferritin 2,800 μg / L).

[0253] Subject 3 is a 47-year-old female with the following medical history: diabetes, adrenal Cushing's syndrome, hypertension, and end-stage renal disease requiring chronic kidney replacement therapy since 2020. She has not been vaccinated against SARS-CoV-2 and had no anti-SARS-CoV-2 immunity at the time of admission. She presented with COVID-19 symptoms (cough, dyspnea, abdominal pain, and fever) and was admitted four days later. Nasopharyngeal PCR identified SARS-CoV2 VOC Omicron. Chest CT scan showed mild to moderate pulmonary lesions, which are typical of COVID-19 (10–25%). She did not require supplemental oxygen. Blood tests revealed a hyperinflammatory syndrome (ferritin 4,350 μg / L and C-reactive protein 55 mg / L), moderately elevated AST and ALT (2 and 1.5 times the ULN values, respectively), and mild thrombocytopenia and anemia. Dexamethasone (6 mg / day) was initiated. On day 3, the patient still had persistent hyperferritinemia (4,142 μg / L) and abnormal liver function tests and developed encephalopathy, so she was transferred to the intensive care unit.

[0254] Subject 4 is a 59-year-old male with the following medical history: hepatitis B, liver transplantation in 2006, HHV8-negative Kaposi sarcoma (complete remission), and end-stage renal disease requiring chronic renal replacement therapy since 2020. He had received 3 doses of the mRNA COVID-19 vaccine but did not develop anti-SARS-CoV-2 antibodies. Due to family exposure to SARS-CoV2, a nasopharyngeal PCR was performed, and the VOC Omicron was identified. He developed COVID-19 symptoms (fatigue), but initially had no respiratory symptoms. Chest CT scan showed mild to moderate pulmonary lesions typical of COVID-19 (10 - 25%). Dyspnea, cough, and fever developed two days later. At admission, PaO2 in room air was 54 mmHg, respiratory rate was 30 cycles / min, and body temperature was 38.5 °C. Blood tests showed hyperferritinemia (1,223 μg / L) and leukopenia (1,080 cells / mm 3 ). CT scan showed progression of the pulmonary lesions (25–50%). Oxygen inhalation, dexamethasone (10 mg / day), tocilizumab (8 mg / kg, once), antibiotics, and fresh frozen plasma from convalescent patients were administered. Mycophenolate mofetil was discontinued, and tacrolimus was administered. Acute respiratory failure developed on day 5, requiring orotracheal intubation and mechanical ventilation with neuromuscular blockade. Blood tests showed a state of hyperinflammation (ferritin 4,535 μg / L), and elevated AST and ALT (3 times the ULN value). At that time, bronchial alveolar fluid culture was negative, suggesting only critical COVID-19. The ratio of PaO2 to FiO2 was 150 to 180. Antibiotics were administered.

[0255] 7.1.1.2. Dosage

[0256] CER-001 was administered intravenously to Subject 1 at a dose of 10 mg / kg over 0.5 to 1 hour at doses of 0 and 12 hours. CER-001 was administered intravenously to Subjects 2 - 4 at a dose of 10 mg / kg over 0.5 to 1 hour at doses of 0, 12, 24, and 48 hours. Antihistamine prophylaxis (50 mg i.v.) with hydroxyzine was used before each dose of CER-001. All subjects also received dexamethasone treatment.

[0257] 7.1.2. Results and Discussion

[0258] 7.1.2.1. General Safety

[0259] Subjects 1, 2, and 3 did not experience any serious adverse events. Subject 4 developed two cases of ventilator-associated pneumonia (VAP; Klebsiella pneumoniae and Aspergillus fumigatus plus mucormycosis) and one case of bacteremia (Staphylococcus haemolyticus).

[0260] 7.1.2.2. Biological efficacy: lipid profile

[0261] At the first administration of CER-001, the serum ApoA-I levels of all four subjects (range 0.74 to 0.79 mg / L, normal value > 1.1 g / L)( Figures 2A - 2D ) and HDL levels (range 0.26 to 0.35, normal value > 0.45 g / L)( Figures 3A - 3D ) were very low, while the serum triglyceride levels were high (range 2.16 to 3.4 g / L, normal value < 1.5 g / L). After treatment with CER-001, the ApoA-I and HDL levels of all subjects returned to normal on day 2, but remained in the lower range of normal values in the subject with the most severe inflammation. In subject 4, who developed ventilator-associated pneumonia three days after starting CER-001, ApoA-I subsequently dropped below the normal value.

[0262] 7.1.2.3. Inflammatory kinetics

[0263] At baseline, IL-1β was normal in all individuals, IL-6 was increased in three subjects who had previously received tocilizumab, and IL-6 was normal in the fourth subject (3.3 to 1,295 pg / mL), while TNF-α was slightly increased (9.7 to 42.1 pg / mL). IL-8 was the only inflammatory cytokine that was generally increased (> 10 pg / mL; 14.8 to 64.5 pg / mL). After administration of CER-001, IL-8 returned to normal in subjects 1, 2, and 3. In subject 4, IL-8 decreased immediately after injection and increased again when ventilator-associated pneumonia developed. Six days after starting CER-001, the serum ferritin level decreased from 6,616 ± 8,696 to 1,712 ± 815 μg / L. Anti-IL6R antibody was administered to three of the four subjects before CER-001, precluding C-reactive protein analysis. The body temperature of all subjects remained below 37.5°.

[0264] 7.1.2.4. Clinical outcomes

[0265] After injection of CER-001, the clinical conditions of subjects 1, 2, and 3 improved rapidly and they were discharged 3 to 4 days after injection of CER-001( Figures 1A to 1D)。In Subjects 1 and 2, oxygen supply was discontinued 2 and 3 days after administration. In Subject 3, confusion disappeared within 2 days. In these 3 subjects, inflammatory parameters, liver function tests, and blood cell counts all improved until discharge. When CER-001 was introduced, Subject 4 had been on mechanical ventilation for 3 days. After the first-stage improvement over three days (discontinuation of neuromuscular blockers and reduction of sedation), he developed several episodes of ventilator-associated pneumonia secondarily and eventually died one month later.

[0266] 7.1.2.5. Discussion

[0267] In addition to having very good acute tolerance to CER-001, rapid improvement in respiratory status, a decrease in inflammatory parameters, and normalization of blood cell counts were observed in three out of four subjects after the use of CER-001, consistent with the normalization of ApoA-I levels. Despite the fact that they had been developing severe COVID-19-related cytokine storms, they were able to be discharged home without oxygen support 3 to 4 days after CER-001 infusion. After administration of CER-001, IL-8 in three subjects decreased rapidly with good results, consistent with clinical and biological improvement. In Subject 4, after the first-stage clinical improvement was accompanied by normalization of ApoA-I and a decrease in IL-8, ventilator-associated pneumonia and clinical deterioration were accompanied by an increase in C-reactive protein and IL-8, as well as a decrease in ApoA-I. In this study, the subjects received four injections of CER-001 (10 mg / kg), but the first injection at a higher dose (e.g., 15 mg / kg) might have helped to reach the optimal concentrations of ApoA-I and non-oxidized HDL more quickly to achieve the maximum therapeutic effect.

[0268] Without being bound by theory, it is believed that the results of this study can be extended to other hyperinflammatory states, such as virus-induced hyperinflammatory states, such as virus-induced hemophagocytic lymphohistiocytosis (HLH), dengue hemorrhagic fever, dengue shock syndrome, and herpes simplex infection, as well as other forms of HLH, such as familial HLH, and HLH secondary to other diseases (such as acute leukemia or lymphoma).

[0269] 8. Specific Embodiments

[0270] Aspects of the present disclosure are described in the embodiments set forth in the following numbered paragraphs.

[0271] 1. A method of treating a subject suffering from or at risk of suffering from a hyperinflammatory disorder, comprising administering to the subject a dose of a lipocalin-based complex, wherein the hyperinflammatory disorder is optionally hemophagocytic lymphohistiocytosis (HLH), dengue hemorrhagic fever, or dengue shock syndrome.

[0272] 2. A method of treating a subject having or at risk of having hemophagocytic lymphohistiocytosis (HLH), comprising administering to the subject a dose of a lipocalin-based complex.

[0273] 3. The method of embodiment 1 or embodiment 2, wherein the subject has HLH secondary to a non-malignant disorder or is at risk of having HLH secondary to a non-malignant disorder.

[0274] 4. The method of embodiment 3, wherein the non-malignant disorder is a viral infection.

[0275] 5. The method of embodiment 4, wherein the subject has HLH caused by dengue infection or is at risk of having HLH caused by dengue infection.

[0276] 6. The method of embodiment 5, wherein the individual has dengue.

[0277] 7. The method of embodiment 5, wherein the individual has dengue hemorrhagic fever.

[0278] 8. The method of embodiment 5, wherein the subject has dengue shock syndrome.

[0279] 9. The method of embodiment 4, wherein the subject has HLH caused by herpes simplex infection or is at risk of having HLH caused by herpes simplex infection.

[0280] 10. The method of embodiment 4, wherein the subject has HLH caused by Epstein-Barr virus infection or is at risk of having HLH caused by Epstein-Barr virus infection.

[0281] 11. The method of embodiment 3, wherein the non-malignant disorder is an autoimmune disease.

[0282] 12. The method of embodiment 1 or embodiment 2, wherein the subject has HLH secondary to a malignant disease or is at risk of having HLH secondary to a malignant disease.

[0283] 13. The method of embodiment 12, wherein the malignant disorder is leukemia or lymphoma.

[0284] 14. The method of embodiment 1 or embodiment 2, wherein the subject has familial HLH or is at risk of having familial HLH.

[0285] 15. The method of any one of embodiments 1 to 14, wherein the subject has HLH.

[0286] 16. The method according to any one of embodiments 1 to 14, wherein the subject is at risk of developing HLH.

[0287] 17. A method of treating a subject having a dengue infection, comprising administering to the subject a dose of a lipocalin-based complex.

[0288] 18. The method according to embodiment 1 or embodiment 17, wherein the subject has dengue.

[0289] 19. The method according to embodiment 1 or embodiment 17, wherein the subject has dengue hemorrhagic fever.

[0290] 20. The method according to embodiment 1 or embodiment 17, wherein the subject is at risk of developing dengue hemorrhagic fever.

[0291] 21. The method according to embodiment 1 or embodiment 17, wherein the subject has dengue shock syndrome.

[0292] 22. The method according to embodiment 1 or embodiment 17, wherein the subject is at risk of developing dengue shock syndrome.

[0293] 23. The method of treating a subject having a herpes simplex infection, comprising administering to the subject a dose of a lipocalin-based complex.

[0294] 24. The method of treating a subject having an Epstein-Barr infection, comprising administering to the subject a dose of a lipocalin-based complex.

[0295] 25. The method according to any one of embodiments 1 to 24, wherein the dose is a high dose.

[0296] 26. The method according to embodiment 25, wherein the high dose is administered over a period of one day to about two weeks, optionally wherein the high dose is administered over a period of one day, two days, three days, four days, five days, six days, seven days, eight days, nine days, ten days, eleven days, 12 days, 13 days, 14 days or 15 days.

[0297] 27. The method according to embodiment 25 or embodiment 26, wherein the high dose is the sum of two to ten individual doses, optionally wherein the high dose is the sum of three, four, five, six, seven, eight, nine or 10 individual doses.

[0298] 28. The method according to embodiment 27, wherein the multiple individual doses are administered daily or twice daily.

[0299] 29. The method according to embodiment 27 or embodiment 28, wherein a plurality of individual doses are administered at intervals of two to three days.

[0300] 30. The method according to embodiment 27, wherein the intervals between the plurality of individual doses are no more than one day.

[0301] 31. The method according to embodiment 30, which comprises administering two or more individual doses at intervals of about 12 hours.

[0302] 32. The method according to embodiment 31, which comprises administering two individual doses at intervals of about 12 hours.

[0303] 33. The method according to embodiment 31, which comprises administering three individual doses at intervals of about 12 hours.

[0304] 34. The method according to embodiment 32 or embodiment 33, which further comprises administering an individual dose about one day later.

[0305] 35. The method according to embodiment 27, which comprises administering three individual doses at intervals of about 12 hours and a fourth individual dose about one day later.

[0306] 36. The method according to embodiment 25, wherein the high dose is administered as a single individual dose.

[0307] 37. The method according to embodiment 25, wherein the high dose is the sum of two individual doses administered within one day.

[0308] 38. The method according to embodiment 37, wherein the intervals between the two individual doses are about 12 hours.

[0309] 39. The method according to any one of embodiments 27 to 38, wherein each individual dose is effective in increasing the HDL level of the subject.

[0310] 40. The method according to embodiment 39, wherein the high dose is effective in increasing the serum HDL level of the subject to a normal value (e.g., > 0.45 g / L).

[0311] 41. The method according to embodiment 39, wherein each individual dose effectively increases the HDL level of the subject by at least 25%, at least 30% or at least 35% within 2 - 4 hours after administration.

[0312] 42. The method according to embodiment 41, wherein each individual dose effectively increases the HDL level of the subject by at least 25%, at least 30% or at least 35% within 2 hours after administration.

[0313] 43. The method according to embodiment 41, wherein each individual dose effectively increases the HDL level of the subject by at least 25%, at least 30%, or at least 35% within 3 hours after administration.

[0314] 44. The method according to embodiment 41, wherein each individual dose effectively increases the HDL level of the subject by at least 25%, at least 30%, or at least 35% within 4 hours after administration.

[0315] 45. The method according to any one of embodiments 27 to 44, wherein each individual dose effectively increases the ApoA-I level of the subject.

[0316] 46. The method according to embodiment 45, wherein the high dose effectively increases the serum ApoA-I level of the subject to a normal value (e.g., > 1.1 g / L).

[0317] 47. The method according to embodiment 45, wherein each individual dose effectively increases the ApoA-I level of the subject by at least 25%, at least 30%, or at least 35% within 2 - 4 hours after administration.

[0318] 48. The method according to embodiment 46, wherein each individual dose effectively increases the ApoA-I level of the subject by at least 25%, at least 30%, or at least 35% within 2 hours after administration.

[0319] 49. The method according to embodiment 46, wherein each individual dose effectively increases the ApoA-I level of the subject by at least 25%, at least 30%, or at least 35% within 3 hours after administration.

[0320] 50. The method according to embodiment 46, wherein each individual dose effectively increases the ApoA-I level of the subject by at least 25%, at least 30%, or at least 35% within 4 hours after administration.

[0321] 51. The method according to any one of embodiments 25 to 50, wherein the high dose effectively improves the vascular endothelial function of the subject, optionally wherein the vascular endothelial function is measured by circulating VCAM-1 and / or ICAM-1.

[0322] 52. The method according to any one of embodiments 25 to 51, wherein the high dose effectively reduces the serum level of one or more inflammatory markers of the subject.

[0323] 53. The method according to embodiment 52, wherein the high dose effectively reduces the serum level of interleukin 6 (“IL-6”).

[0324] The method according to embodiment 52 or embodiment 53, wherein the high dose effectively reduces the serum level of C-reactive protein.

[0325] 55. The method according to any one of embodiments 52 to 54, wherein the high dose effectively reduces the serum level of D-dimer.

[0326] 56. The method according to any one of embodiments 52 to 55, wherein the high dose effectively reduces the serum level of ferritin.

[0327] 57. The method according to any one of embodiments 52 to 56, wherein the high dose effectively reduces the serum level of interleukin 8 (IL-8).

[0328] 58. The method according to any one of embodiments 52 to 56, wherein the high dose effectively normalizes the serum level of interleukin 8 (IL-8).

[0329] 59. The method according to any one of embodiments 52 to 58, wherein the high dose effectively reduces the serum level of granulocyte-macrophage colony-stimulating factor (GM-CSF).

[0330] 60. The method according to any one of embodiments 52 to 59, wherein the high dose effectively reduces the serum level of monocyte chemoattractant protein (MCP) 1.

[0331] 61. The method according to any one of embodiments 52 to 60, wherein the high dose effectively reduces the serum level of tumor necrosis factor α (TNF-α).

[0332] 62. The method according to any one of embodiments 52 to 61, wherein the high dose effectively reduces the serum level of one or more inflammatory markers from an elevated range to a normal range.

[0333] 63. The method according to any one of embodiments 52 to 62, wherein the high dose effectively reduces the serum level of one or more inflammatory markers by at least 20%, at least 40%, or at least 60%.

[0334] 64. The method according to any one of embodiments 1 to 63, wherein the subject has CRS or is at risk of developing CRS.

[0335] 65. The method according to embodiment 64, wherein the subject has CRS.

[0336] 66. The method according to embodiment 64, wherein the subject is at risk of developing CRS.

[0337] 67. The method according to any one of embodiments 25 to 66, wherein the high dose is effective in reducing the likelihood of a subject developing acute kidney injury (AKI).

[0338] 68. The method according to any one of embodiments 25 to 67, wherein the high dose is effective in delaying the onset of AKI.

[0339] 69. The method according to any one of embodiments 25 to 67, wherein the high dose is effective in preventing AKI.

[0340] 70. The method according to any one of embodiments 25 to 66, wherein the subject has acute kidney injury (AKI) or is at risk of developing acute kidney injury (AKI).

[0341] 71. The method according to embodiment 70, wherein the subject has AKI.

[0342] 72. The method according to embodiment 71, wherein the high dose is effective in reducing the severity of AKI.

[0343] 73. The method according to embodiment 70, wherein the subject is at risk of developing AKI.

[0344] 74. The method according to embodiment 73, wherein the high dose is effective in reducing the likelihood of the subject developing AKI.

[0345] 75. The method according to embodiment 73, wherein the high dose is effective in delaying the occurrence of AKI.

[0346] 76. The method according to embodiment 73, wherein the high dose is effective in preventing AKI.

[0347] 77. The method according to embodiment 73, wherein if the subject develops AKI, the high dose is effective in reducing the severity of AKI.

[0348] 78. The method according to any one of embodiments 1 to 77, wherein the subject has a SOFA score of 1 to 4 before administration of the lipocalin-based complex.

[0349] 79. The method according to embodiment 78, wherein the subject has a SOFA score of 2 to 4 before administration of the lipocalin-based complex.

[0350] 80. The method according to embodiment 78, wherein the subject has a SOFA score of 1 before administration of the lipocalin-based complex.

[0351] 81. The method according to embodiment 78, wherein the subject has a SOFA score of 2 before administration of the lipocalin-based complex.

[0352] 82. The method according to embodiment 78, wherein the subject has a SOFA score of 3 before administration of the lipocalin-based complex.

[0353] 83. The method according to embodiment 78, wherein the subject has a SOFA score of 4 before administration of the lipocalin-based complex.

[0354] 84. The method according to any one of embodiments 1 to 83, wherein the lipocalin-based complex is reconstituted HDL or an HDL mimetic.

[0355] 85. The method according to any one of embodiments 1 to 83, wherein the lipocalin-based complex is an Apomer or a Cargomer.

[0356] 86. The method according to any one of embodiments 1 to 85, wherein the lipocalin-based complex comprises sphingomyelin.

[0357] 87. The method according to any one of embodiments 1 to 86, wherein the lipocalin-based complex comprises a negatively charged lipid.

[0358] 88. The method according to embodiment 87, wherein the negatively charged lipid is 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG) or a salt thereof.

[0359] 89. The method according to embodiment 84, wherein the lipocalin-based complex is CER-001, CSL-111, CSL-112, CER-522 or ETC-216.

[0360] 90. The method according to embodiment 89, wherein the lipocalin-based complex is CER-001.

[0361] 91. The method according to any one of embodiments 1 to 90, wherein the lipocalin-based complex is administered systemically, optionally by infusion.

[0362] 92. The method according to any one of embodiments 1 to 91, wherein the lipocalin-based complex is administered until the serum level of one or more inflammatory markers is reduced.

[0363] 93. The method according to embodiment 92, wherein the lipocalin-based complex is administered until the serum level of one or more inflammatory markers is reduced to the normal range.

[0364] The method according to embodiment 92, wherein the lipocalin-based complex is administered until the serum level of one or more inflammatory markers is reduced to below the baseline level of one or more inflammatory markers measured before administration of the lipocalin-based complex.

[0365] The method according to any one of embodiments 1 to 94, wherein each individual dose of the lipocalin-based complex administered is 4 - 40 mg / kg (based on protein weight).

[0366] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 4 - 30 mg / kg (based on protein weight).

[0367] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 15 - 25 mg / kg (based on protein weight).

[0368] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 10 - 30 mg / kg (based on protein weight).

[0369] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 10 - 20 mg / kg (based on protein weight).

[0370] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 5 mg / kg (based on protein weight).

[0371] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 10 mg / kg (based on protein weight).

[0372] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 15 mg / kg (based on protein weight).

[0373] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 20 mg / kg (based on protein weight).

[0374] The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 5 to 15 mg / kg (based on protein weight).

[0375] 105. The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 10 to 20 mg / kg (based on protein weight).

[0376] 106. The method according to embodiment 95, wherein each individual dose of the lipocalin-based complex is 15 to 25 mg / kg (based on protein weight).

[0377] 107. The method according to any one of embodiments 25 to 106, wherein a high dose is administered according to an induction regimen, optionally followed by a consolidation regimen.

[0378] 108. The method according to embodiment 107, wherein the induction regimen comprises administering the lipocalin-based complex once or twice daily.

[0379] 109. The method according to embodiment 107 or embodiment 108, wherein the consolidation regimen comprises administering the lipocalin-based complex once daily or once every two days.

[0380] 110. The method according to any one of embodiments 1 to 109, wherein the subject is not treated with a maintenance regimen.

[0381] 111. The method according to any one of embodiments 107 to 110, wherein the consolidation regimen comprises administering to the subject one or more doses of the lipocalin-based complex one or more days after the final dose of the induction regimen.

[0382] 112. The method according to embodiment 111, wherein the first dose of the lipocalin-based complex administered during the consolidation regimen is administered two or more days after the final dose of the induction regimen.

[0383] 113. The method according to embodiment 111, wherein the first dose of the lipocalin-based complex administered during the consolidation regimen is administered three or more days after the final dose of the induction regimen.

[0384] 114. The method according to embodiment 113, wherein the first dose of the lipocalin-based complex administered during the consolidation regimen is administered three days after the final dose of the induction regimen.

[0385] 115. The method according to any one of embodiments 107 to 114, which comprises an induction regimen and a consolidation regimen, the induction regimen comprising administering the lipocalin-based complex twice daily on days 1, 2 and 3, and the consolidation regimen comprising administering two doses of the lipocalin-based complex on day 6.

[0386] The method according to any one of embodiments 107 to 115, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 4 - 40 mg / kg (based on protein weight).

[0387] 117. The method according to any one of embodiments 107 to 116, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 4 - 30 mg / kg (based on protein weight).

[0388] 118. The method according to any one of embodiments 107 to 116, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 15 - 25 mg / kg (based on protein weight).

[0389] 119. The method according to any one of embodiments 107 to 116, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 10 - 30 mg / kg (based on protein weight).

[0390] 120. The method according to any one of embodiments 107 to 116, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 10 - 20 mg / kg (based on protein weight).

[0391] 121. The method according to any one of embodiments 107 to 116, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 5 mg / kg (based on protein weight).

[0392] 122. The method according to any one of embodiments 107 to 116, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 10 mg / kg (based on protein weight).

[0393] 123. The method according to any one of embodiments 107 to 116, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 15 mg / kg (based on protein weight).

[0394] 124. The method according to any one of embodiments 107 to 116, wherein each individual dose of the lipocalin-based complex administered in the induction regimen is 20 mg / kg (based on protein weight).

[0395] 125. The method according to any one of embodiments 107 to 124, wherein the dose of the lipocalin-based complex administered in the consolidation regimen is 5 to 15 mg / kg (based on protein weight).

[0396] 126. The method according to any one of embodiments 107 to 124, wherein the dose of the lipocalin-based complex administered in the consolidation regimen is 10 to 20 mg / kg (based on protein weight).

[0397] 127. The method according to any one of embodiments 107 to 124, wherein the dose of the lipocalin-based complex administered in the consolidation regimen is 15 to 25 mg / kg (based on protein weight).

[0398] 128. The method according to any one of embodiments 107 to 124, wherein the dose of the lipocalin-based complex administered in the consolidation regimen is 5 mg / kg (based on protein weight).

[0399] 129. The method according to any one of embodiments 107 to 124, wherein the dose of the lipocalin-based complex administered in the consolidation regimen is 10 mg / kg (based on protein weight).

[0400] 130. The method according to any one of embodiments 107 to 124, wherein the dose of the lipocalin-based complex administered in the consolidation regimen is 15 mg / kg (based on protein weight).

[0401] 131. The method according to any one of embodiments 1 to 130, wherein each individual dose of the lipocalin-based complex administered is 300 mg to 4000 mg (based on protein weight).

[0402] 132. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is 300 mg to 3000 mg (based on protein weight).

[0403] 133. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is 300 mg to 1500 mg (based on protein weight).

[0404] 134. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is 400 mg to 4000 mg (based on protein weight).

[0405] 135. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is 400 mg to 1500 mg (based on protein weight).

[0406] 136. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is 500 mg to 1200 mg (based on protein weight).

[0407] 137. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is from 500 mg to 1000 mg (based on protein weight).

[0408] 138. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is from 600 mg to 3000 mg (based on protein weight).

[0409] 139. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is from 800 mg to 3000 mg (based on protein weight).

[0410] 140. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is from 1000 mg to 2400 mg (based on protein weight).

[0411] 141. The method according to embodiment 131, wherein each individual dose of the lipocalin-based complex administered is from 1000 mg to 2000 mg (based on protein weight).

[0412] 142. The method according to any one of embodiments 25 to 141, wherein the high dose of the lipocalin-based complex is from 600 mg to 40 g (based on protein weight).

[0413] 143. The method according to any one of embodiments 25 to 141, wherein the high dose of the lipocalin-based complex is from 3 g to 35 g (based on protein weight).

[0414] 144. The method according to any one of embodiments 25 to 141, wherein the high dose of the lipocalin-based complex is from 5 g to 30 g (based on protein weight).

[0415] 145. The method according to any one of embodiments 1 to 144, wherein the lipocalin-based complex is administered by infusion.

[0416] 146. The method according to embodiment 145, wherein each individual dose is administered within 1 to 24 hours.

[0417] 147. The method according to embodiment 146, wherein each individual dose is administered within 24 hours.

[0418] 148. The method according to embodiment 145, wherein each individual dose is administered within one hour or less.

[0419] The method according to embodiment 145, wherein each individual dose is administered over a period of half an hour to one hour.

[0420] 150. The method according to any one of embodiments 1 to 149, further comprising administering an antihistamine to the subject prior to each individual administration.

[0421] 151. The method according to embodiment 150, wherein the antihistamine comprises dextrochlorpheniramine or hydroxyzine.

[0422] 152. The method according to any one of embodiments 1 to 151, wherein the subject is receiving or has received one or more additional therapies and / or the method further comprises administering to the subject one or more additional therapies.

[0423] 153. The method according to embodiment 152, wherein the one or more additional therapies comprise one or more anti-IL-6 agents.

[0424] 154. The method according to embodiment 153, wherein the one or more anti-IL-6 agents comprise tocilizumab, cetuximab, olaratumab, asimadoline, BMS-945429, sirukumab, lebrikizumab, CPSI-2364, or a combination thereof.

[0425] 155. The method according to embodiment 154, wherein the one or more anti-IL-6 agents comprise tocilizumab.

[0426] 156. The method according to any one of embodiments 152 to 155, wherein the one or more additional therapies comprise one or more corticosteroids.

[0427] 157. The method according to embodiment 156, wherein the one or more corticosteroids comprise methylprednisolone, dexamethasone, or a combination thereof.

[0428] 158. The method according to any one of embodiments 1 to 157, wherein the lipocalin-based complex is CER-001.

[0429] 159. The method according to embodiment 158, wherein the CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids, wherein the weight ratio of ApoA-I: total phospholipid weight is 1:2.7 + / - 20%, and comprising the phospholipids sphingomyelin and DPPG, and the weight: weight ratio of sphingomyelin: DPPG is 97:3 + / - 20%.

[0430] The method according to embodiment 158, wherein the CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids, wherein the weight ratio of ApoA-I: total phospholipids is 1:2.7 + / - 10%, and comprising the phospholipids sphingomyelin and DPPG, wherein the weight ratio of sphingomyelin:DPPG is 97:3 + / - 10%.

[0431] 161. The method according to embodiment 158, wherein the CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids, wherein the weight ratio of ApoA-I: total phospholipids is 1:2.7, and the phospholipids sphingomyelin and DPPG, wherein the weight ratio of sphingomyelin:DPPG is 97:3.

[0432] 162. The method according to any one of embodiments 159 to 161, wherein the ApoA-I has the amino acid sequence of amino acids 25-267 of SEQ ID NO:2.

[0433] 163. The method according to any one of embodiments 159 to 162, wherein the ApoA-I is recombinantly expressed.

[0434] 164. The method according to any one of embodiments 159 to 163, wherein the CER-001 comprises natural sphingomyelin.

[0435] 165. The method according to embodiment 164, wherein the natural sphingomyelin is egg sphingomyelin.

[0436] 166. The method according to any one of embodiments 159 to 163, wherein the CER-001 comprises synthetic sphingomyelin.

[0437] 167. The method according to embodiment 166, wherein the synthetic sphingomyelin is palmitoyl sphingomyelin.

[0438] 168. The method according to any one of embodiments 158 to 167, wherein the CER-001 is administered in a formulation that is at least 95% homogeneous for CER-001.

[0439] 169. The method according to embodiment 168, wherein the CER-001 is administered in a formulation, wherein the CER-001 is at least 97% homogeneous.

[0440] 170. The method according to embodiment 168, wherein the CER-001 is administered in a formulation, wherein the CER-001 is at least 98% homogeneous.

[0441] The method according to embodiment 168, wherein the CER-001 is administered in the form of a formulation, and wherein the CER-001 is at least 99% homogeneous.

[0442] The method according to any one of embodiments 1 to 171, wherein the subject is human.

[0443] The method according to any one of embodiments 1 to 172, wherein when the CER-001 is administered for the first time, the subject is not mechanically ventilated.

[0444] The method according to embodiment 89, wherein the lipocalin-based complex is CSL-112.

[0445] The method according to any one of embodiments 163 to 173, wherein the ApoA-I is produced by a mammalian host cell.

[0446] The method according to embodiment 175, wherein the mammalian host cell is a Chinese hamster ovary (CHO) cell.

[0447] The method according to embodiment 176, wherein the CHO cell is a CHO-S cell.

[0448] The method according to any one of embodiments 175 to 177, wherein the ApoA-I has undergone post-translational processing (such as glycosylation) such that the ApoA-I has one or more structural features (such as glycosylation patterns) different from those of human ApoA-I purified from human plasma.

[0449] A method of treating a subject suffering from an over-inflammatory disorder or at risk of developing an over-inflammatory disorder, comprising administering to the subject a dose of an apolipoprotein AI ("ApoA-I") formulation, the apolipoprotein AI ("ApoA-I") formulation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex, optionally wherein the over-inflammatory disorder is hemophagocytic lymphohistiocytosis (HLH), dengue hemorrhagic fever or dengue shock syndrome.

[0450] A method of treating a subject suffering from hemophagocytic lymphohistiocytosis (HLH) or at risk of developing hemophagocytic lymphohistiocytosis, comprising administering to the subject a dose of an apolipoprotein AI ("ApoA-I") formulation, the apolipoprotein AI ("ApoA-I") formulation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

[0451] 181. The method according to embodiment 179 or embodiment 180, wherein the subject has HLH secondary to a non-malignant disorder or is at risk of developing HLH secondary to a non-malignant disorder.

[0452] 182. The method according to embodiment 181, wherein the non-malignant disorder is a viral infection.

[0453] 183. The method according to embodiment 182, wherein the subject has HLH caused by dengue infection or is at risk of developing HLH caused by dengue infection.

[0454] 184. The method according to embodiment 183, wherein the subject has dengue.

[0455] 185. The method according to embodiment 183, wherein the subject has dengue hemorrhagic fever.

[0456] 186. The method according to embodiment 183, wherein the subject has dengue shock syndrome.

[0457] 187. The method according to embodiment 182, wherein the subject has HLH caused by herpes simplex infection or is at risk of developing HLH caused by herpes simplex infection.

[0458] 188. The method according to embodiment 182, wherein the subject has HLH caused by Epstein-Barr virus infection or is at risk of developing HLH caused by Epstein-Barr virus infection.

[0459] 189. The method according to embodiment 181, wherein the non-malignant disorder is an autoimmune disease.

[0460] 190. The method according to embodiment 179 or embodiment 180, wherein the subject has HLH secondary to a malignant disorder or is at risk of developing HLH secondary to a malignant disorder.

[0461] 191. The method according to embodiment 190, wherein the malignant disorder is leukemia or lymphoma.

[0462] 192. The method according to embodiment 179 or embodiment 180, wherein the subject has familial HLH or is at risk of developing familial HLH.

[0463] 193. The method according to any one of embodiments 179 to 192, wherein the subject has HLH.

[0464] 194. The method according to any one of embodiments 179 to 192, wherein the subject is at risk of developing HLH.

[0465] A method of treating a subject having a dengue infection, comprising administering to the subject a dose of an apolipoprotein AI ("ApoA-I") preparation, the apolipoprotein AI ("ApoA-I") preparation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

[0466] The method according to embodiment 179 or embodiment 195, wherein the subject has dengue.

[0467] The method according to embodiment 179 or embodiment 195, wherein the subject has dengue hemorrhagic fever.

[0468] The method according to embodiment 179 or embodiment 195, wherein the subject is at risk of having dengue hemorrhagic fever.

[0469] The method according to embodiment 179 or embodiment 195, wherein the subject has dengue shock syndrome.

[0470] The method according to embodiment 179 or embodiment 195, wherein the subject is at risk of having dengue shock syndrome.

[0471] A method of treating a subject having a herpes simplex infection, comprising administering to the subject a dose of an apolipoprotein AI ("ApoA-I") preparation, the apolipoprotein AI ("ApoA-I") preparation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

[0472] A method of treating a subject having an Epstein-Barr infection, comprising administering to the subject a dose of an apolipoprotein AI ("ApoA-I") preparation, the apolipoprotein AI ("ApoA-I") preparation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

[0473] The method according to any one of embodiments 179 to 202, wherein the dose is a high dose.

[0474] The method according to embodiment 203, wherein the high dose is administered over a period of one day to about two weeks, optionally wherein the high dose is administered over a period of one day, two days, three days, four days, five days, six days, seven days, eight days, nine days, 10 days, eleven days, 12 days, 13 days, 14 days or 15 days.

[0475] The method according to embodiment 203 or embodiment 204, wherein the high dose is the sum of two to ten individual doses, optionally wherein the high dose is the sum of three, four, five, six, seven, eight, nine or 10 individual doses.

[0476] 206. The method according to embodiment 205, wherein multiple individual doses are administered daily or twice daily.

[0477] 207. The method according to embodiment 205 or embodiment 206, wherein multiple individual doses are administered at intervals of two to three days.

[0478] 208. The method according to embodiment 205, wherein the dosing interval of multiple individual doses does not exceed one day.

[0479] 209. The method according to embodiment 208, which comprises administering two or more individual doses at intervals of about 12 hours.

[0480] 210. The method according to embodiment 209, which comprises administering two individual doses at an interval of about 12 hours.

[0481] 211. The method according to embodiment 209, which comprises administering three individual doses at intervals of about 12 hours.

[0482] 212. The method according to embodiment 210 or embodiment 211, which further comprises administering an individual dose about one day later.

[0483] 213. The method according to embodiment 205, which comprises administering three individual doses at intervals of about 12 hours and a fourth individual dose about one day later.

[0484] 214. The method according to embodiment 203, wherein the high dose is administered as a single individual dose.

[0485] 215. The method according to embodiment 203, wherein the high dose is the sum of two individual doses administered within one day.

[0486] 216. The method according to embodiment 215, wherein the dosing interval of the two individual doses is about 12 hours.

[0487] 217. The method according to any one of embodiments 205 to 216, wherein each individual dose is effective in increasing the HDL level of the subject.

[0488] 218. The method according to embodiment 217, wherein the high dose is effective in increasing the individual serum HDL level to a normal value (for example, > 0.45 g / L).

[0489] The method of embodiment 217, wherein each individual dose effectively increases the HDL level of the subject by at least 25%, at least 30%, or at least 35% within 2 - 4 hours after administration.

[0490] The method of embodiment 219, wherein each individual dose effectively increases the HDL level of the subject by at least 25%, at least 30%, or at least 35% within 2 hours after administration.

[0491] The method of embodiment 219, wherein each individual dose effectively increases the HDL level of an individual by at least 25%, at least 30%, or at least 35% within 3 hours after administration.

[0492] The method of embodiment 219, wherein each individual dose effectively increases the HDL level of the subject by at least 25%, at least 30%, or at least 35% within 4 hours after administration.

[0493] The method of any one of embodiments 205 to 222, wherein each individual dose effectively increases the ApoA-I level of the subject.

[0494] The method of embodiment 223, wherein the high dose effectively increases the serum ApoA-I level of the subject to the normal value (e.g., > 1.1 g / L).

[0495] The method of embodiment 223, wherein each individual dose effectively increases the ApoA-I level of the subject by at least 25%, at least 30%, or at least 35% within 2 - 4 hours after administration.

[0496] The method of embodiment 224, wherein each individual dose effectively increases the ApoA-I level of the subject by at least 25%, at least 30%, or at least 35% within 2 hours after administration.

[0497] The method of embodiment 224, wherein each individual dose effectively increases the ApoA-I level of the subject by at least 25%, at least 30%, or at least 35% within 3 hours after administration.

[0498] The method of embodiment 224, wherein each individual dose effectively increases the ApoA-I level of the subject by at least 25%, at least 30%, or at least 35% within 4 hours after administration.

[0499] The method according to any one of embodiments 203 to 228, wherein the high dose effectively improves the vascular endothelial function of a subject, optionally wherein the vascular endothelial function is measured by circulating VCAM-1 and / or ICAM-1.

[0500] 230. The method according to any one of embodiments 203 to 229, wherein the high dose effectively reduces the serum level of one or more inflammatory markers in a subject.

[0501] 231. The method according to embodiment 230, wherein the high dose effectively reduces the serum level of interleukin-6 ("IL-6").

[0502] 232. The method according to embodiment 230 or embodiment 231, wherein the high dose effectively reduces the serum level of C-reactive protein.

[0503] 233. The method according to any one of embodiments 230 to 232, wherein the high dose effectively reduces the serum level of D-dimer.

[0504] 234. The method according to any one of embodiments 230 to 233, wherein the high dose effectively reduces the serum level of ferritin.

[0505] 235. The method according to any one of embodiments 230 to 234, wherein the high dose effectively reduces the serum level of interleukin 8 (IL-8).

[0506] 236. The method according to any one of embodiments 230 to 234, wherein the high dose effectively normalizes the serum level of interleukin 8 (IL-8).

[0507] 237. The method according to any one of embodiments 230 to 236, wherein the high dose effectively reduces the serum level of granulocyte-macrophage colony-stimulating factor (GM-CSF).

[0508] 238. The method according to any one of embodiments 230 to 237, wherein the high dose effectively reduces the serum level of monocyte chemoattractant protein (MCP) 1.

[0509] 239. The method according to any one of embodiments 230 to 238, wherein the high dose effectively reduces the serum level of tumor necrosis factor α (TNF-α).

[0510] 240. The method according to any one of embodiments 230 to 239, wherein the high dose effectively reduces the serum level of one or more inflammatory markers from an elevated range to a normal range.

[0511] The method according to any one of embodiments 230 to 240, wherein the high dose is effective in reducing the serum level of one or more inflammatory markers by at least 20%, at least 40%, or at least 60%.

[0512] 242. The method according to any one of embodiments 179 to 241, wherein the subject has CRS or is at risk of developing CRS.

[0513] 243. The method according to embodiment 242, wherein the subject has CRS.

[0514] 244. The method according to embodiment 242, wherein the subject is at risk of developing CRS.

[0515] 245. The method according to any one of embodiments 203 to 244, wherein the high dose is effective in reducing the likelihood of the subject developing acute kidney injury (AKI).

[0516] 246. The method according to any one of embodiments 203 to 245, wherein the high dose is effective in delaying the onset of AKI.

[0517] 247. The method according to any one of embodiments 203 to 245, wherein the high dose is effective in preventing AKI.

[0518] 248. The method according to any one of embodiments 203 to 244, wherein the subject has acute kidney injury (AKI) or is at risk of developing acute kidney injury (AKI).

[0519] 249. The method according to embodiment 248, wherein the subject has AKI.

[0520] 250. The method according to embodiment 249, wherein the high dose is effective in reducing the severity of AKI.

[0521] 251. The method according to embodiment 248, wherein the subject is at risk of developing AKI.

[0522] 252. The method according to embodiment 251, wherein the high dose is effective in reducing the likelihood of the subject developing AKI.

[0523] 253. The method according to embodiment 251, wherein the high dose is effective in delaying the occurrence of AKI.

[0524] 254. The method according to embodiment 251, wherein the high dose is effective in preventing AKI.

[0525] 255. The method according to embodiment 251, wherein if the subject develops AKI, the high dose is effective in reducing the severity of AKI.

[0526] 256. The method according to embodiments 179 to 255, wherein the subject has a SOFA score of 1 to 4 before administration of the formulation.

[0527] 257. The method according to embodiment 256, wherein the individual has a SOFA score of 2 to 4 before administration of the formulation.

[0528] 258. The method according to embodiment 256, wherein the individual has a SOFA score of 1 before administration of the formulation.

[0529] 259. The method according to embodiment 256, wherein the individual has a SOFA score of 2 before administration of the formulation.

[0530] 260. The method according to embodiment 256, wherein the individual has a SOFA score of 3 before administration of the formulation.

[0531] 261. The method according to embodiment 256, wherein the individual has a SOFA score of 4 before administration of the formulation.

[0532] 262. The method according to any one of embodiments 179 to 261, wherein the formulation is reconstituted HDL or an HDL mimetic.

[0533] 263. The method according to any one of embodiments 179 to 261, wherein the formulation is Apomer or Cargomer.

[0534] 264. The method according to any one of embodiments 179 to 263, wherein the formulation comprises sphingomyelin.

[0535] 265. The method according to any one of embodiments 179 to 264, wherein the formulation comprises a negatively charged lipid.

[0536] 266. The method according to embodiment 265, wherein the negatively charged lipid is 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG) or a salt thereof.

[0537] 267. The method according to any one of embodiments 179 to 266, wherein the formulation is optionally administered systemically by infusion.

[0538] 268. The method according to any one of embodiments 179 to 267, wherein the formulation is administered until the serum level of one or more inflammatory markers is reduced.

[0539] The method according to embodiment 268, wherein the formulation is administered until the serum level of one or more inflammatory markers is reduced to the normal range.

[0540] The method according to embodiment 268, wherein the formulation is administered until the serum level of one or more inflammatory markers is reduced to below the baseline level of one or more inflammatory markers measured before administration of the lipocalin-based complex.

[0541] The method according to any one of embodiments 179 to 270, wherein each individual dose of the formulation administered is 4 - 40 mg / kg (based on protein weight).

[0542] The method according to embodiment 271, wherein each individual dose of the formulation is 4 - 30 mg / kg (based on protein weight).

[0543] The method according to embodiment 271, wherein each individual dose of the formulation is 15 - 25 mg / kg (based on protein weight).

[0544] The method according to embodiment 271, wherein each individual dose of the formulation is 10 - 30 mg / kg (based on protein weight).

[0545] The method according to embodiment 271, wherein each individual dose of the formulation is 10 - 20 mg / kg (based on protein weight).

[0546] The method according to embodiment 271, wherein each individual dose of the formulation is 5 mg / kg (based on protein weight).

[0547] The method according to embodiment 271, wherein each individual dose of the formulation is 10 mg / kg (based on protein weight).

[0548] The method according to embodiment 271, wherein each individual dose of the formulation is 15 mg / kg (based on protein weight).

[0549] The method according to embodiment 271, wherein each individual dose of the formulation is 20 mg / kg (based on protein weight).

[0550] The method according to embodiment 271, wherein each individual dose of the formulation is 5 to 15 mg / kg (based on protein weight).

[0551] The method according to embodiment 271, wherein each individual dose of the formulation is 10 to 20 mg / kg (based on the weight of the protein).

[0552] 282. The method according to embodiment 271, wherein each individual dose of the formulation is 15 to 25 mg / kg (based on the weight of the protein).

[0553] 283. The method according to any one of embodiments 203 to 282, wherein the high dose is administered according to an induction regimen, optionally followed by a consolidation regimen.

[0554] 284. The method according to embodiment 283, wherein the induction regimen comprises administering the formulation once or twice daily.

[0555] 285. The method according to embodiment 283 or embodiment 284, wherein the consolidation regimen comprises administering the formulation once daily or once every two days.

[0556] 286. The method according to any one of embodiments 179 to 285, wherein the subject is not treated with a maintenance regimen.

[0557] 287. The method according to any one of embodiments 283 to 286, wherein the consolidation regimen comprises administering one or more doses of the formulation to the subject one or more days after the final dose of the induction regimen.

[0558] 288. The method according to embodiment 287, wherein the first dose of the formulation administered during the consolidation regimen is administered two or more days after the final dose of the induction regimen.

[0559] 289. The method according to embodiment 287, wherein the first dose of the formulation administered during the consolidation regimen is administered three or more days after the last dose of the induction regimen.

[0560] 290. The method according to embodiment 287, wherein the first dose of the formulation administered during the consolidation regimen is administered three days after the final dose of the induction regimen.

[0561] 291. The method according to any one of embodiments 283 to 290, which comprises an induction regimen and a consolidation regimen, the induction regimen comprising administering the formulation twice daily on days 1, 2 and 3, and the consolidation regimen comprising administering two doses of the formulation on day 6.

[0562] The method according to any one of embodiments 283 to 291, wherein each individual dose of the formulation administered in the induction regimen is 4 - 40 mg / kg (based on the protein weight).

[0563] 293. The method according to any one of embodiments 283 to 292, wherein each individual dose of the formulation administered in the induction regimen is 4 - 30 mg / kg (based on the protein weight).

[0564] 294. The method according to any one of embodiments 283 to 292, wherein each individual dose of the formulation administered in the induction regimen is 15 - 25 mg / kg (based on the protein weight).

[0565] 295. The method according to any one of embodiments 283 to 292, wherein each individual dose of the formulation administered in the induction regimen is 10 - 30 mg / kg (based on the protein weight).

[0566] 296. The method according to any one of embodiments 283 to 292, wherein each individual dose of the formulation administered in the induction regimen is 10 - 20 mg / kg (based on the protein weight).

[0567] 297. The method according to any one of embodiments 283 to 292, wherein each individual dose of the formulation administered in the induction regimen is 5 mg / kg (based on the protein weight).

[0568] 298. The method according to any one of embodiments 283 to 292, wherein each individual dose of the formulation administered in the induction regimen is 10 mg / kg (based on the protein weight).

[0569] 299. The method according to any one of embodiments 283 to 292, wherein each individual dose of the formulation administered in the induction regimen is 15 mg / kg (based on the protein weight).

[0570] 300. The method according to any one of embodiments 283 to 292, wherein each individual dose of the formulation administered in the induction regimen is 20 mg / kg (based on the protein weight).

[0571] 301. The method according to any one of embodiments 283 to 300, wherein the dose of the formulation administered in the consolidation regimen is 5 to 15 mg / kg (based on the protein weight).

[0572] The method according to any one of embodiments 283 to 300, wherein the dose of the formulation administered in the consolidation regimen is 10 to 20 mg / kg (based on the protein weight).

[0573] 303. The method according to any one of embodiments 283 to 300, wherein the dose of the formulation administered in the consolidation regimen is 15 to 25 mg / kg (based on the protein weight).

[0574] 304. The method according to any one of embodiments 283 to 300, wherein the dose of the formulation administered in the consolidation regimen is 5 mg / kg (based on the protein weight).

[0575] 305. The method according to any one of embodiments 283 to 300, wherein the dose of the formulation administered in the consolidation regimen is 10 mg / kg (based on the protein weight).

[0576] 306. The method according to any one of embodiments 283 to 300, wherein the dose of the formulation administered in the consolidation regimen is 15 mg / kg (based on the protein weight).

[0577] 307. The method according to any one of embodiments 179 to 306, wherein each individual dose of the formulation administered is 300 mg to 4000 mg (based on the protein weight).

[0578] 308. The method according to embodiment 307, wherein each individual dose of the formulation administered is 300 mg to 3000 mg (based on the protein weight).

[0579] 309. The method according to embodiment 307, wherein each individual dose of the formulation administered is 300 mg to 1500 mg (based on the protein weight).

[0580] 310. The method according to embodiment 307, wherein each individual dose of the formulation administered is 400 mg to 4000 mg (based on the protein weight).

[0581] 311. The method according to embodiment 307, wherein each individual dose of the formulation administered is 400 mg to 1500 mg (based on the protein weight).

[0582] 312. The method according to embodiment 307, wherein each individual dose of the formulation administered is 500 mg to 1200 mg (based on the protein weight).

[0583] The method according to embodiment 307, wherein each individual dose of the administered formulation is 500 mg to 1000 mg (based on protein weight).

[0584] 314. The method according to embodiment 307, wherein each individual dose of the administered formulation is 600 mg to 3000 mg (based on protein weight).

[0585] 315. The method according to embodiment 307, wherein each individual dose of the administered formulation is 800 mg to 3000 mg (based on protein weight).

[0586] 316. The method according to embodiment 307, wherein each individual dose of the administered formulation is 1000 mg to 2400 mg (based on protein weight).

[0587] 317. The method according to embodiment 307, wherein each individual dose of the administered formulation is 1000 mg to 2000 mg (based on protein weight).

[0588] 318. The method according to any one of embodiments 203 to 317, wherein the high dose of the formulation is 600 mg to 40 g (based on protein weight).

[0589] 319. The method according to any one of embodiments 203 to 317, wherein the high dose of the formulation is 3 g to 35 g (based on protein weight).

[0590] 320. The method according to any one of embodiments 203 to 317, wherein the high dose of the formulation is 5 g to 30 g (based on protein weight).

[0591] 321. The method according to any one of embodiments 179 to 320, wherein the formulation is administered by infusion.

[0592] 322. The method according to embodiment 321, wherein each individual dose is administered within 1 to 24 hours.

[0593] 323. The method according to embodiment 322, wherein each individual dose is administered within 24 hours.

[0594] 324. The method according to embodiment 321, wherein each individual dose is administered within one hour or less.

[0595] 325. The method according to embodiment 321, wherein each individual dose is administered within half an hour to one hour.

[0596] The method according to any one of embodiments 179 to 325, further comprising administering an antihistamine to the subject prior to each individual dose.

[0597] 327. The method according to embodiment 326, wherein the antihistamine comprises dextrochlorpheniramine or hydroxyzine.

[0598] 328. The method according to any one of embodiments 179 to 327, wherein the subject is receiving or has received one or more additional therapies and / or the method further comprises administering to the subject one or more additional therapies.

[0599] 329. The method according to embodiment 328, wherein the one or more additional therapies comprise one or more anti-IL-6 agents.

[0600] 330. The method according to embodiment 329, wherein the one or more anti-IL-6 agents comprise tocilizumab, cetuximab, olaratumab, elsilimomab, BMS-945429, sirukumab, rivocetumab, CPSI-2364, or a combination thereof.

[0601] 331. The method according to embodiment 330, wherein the one or more anti-IL-6 agents comprise tocilizumab.

[0602] 332. The method according to any one of embodiments 328 to 331, wherein the one or more additional therapies comprise one or more corticosteroids.

[0603] 333. The method according to embodiment 332, wherein the one or more corticosteroids comprise methylprednisolone, dexamethasone, or a combination thereof.

[0604] 334. The method according to any one of embodiments 179 to 333, wherein the formulation comprises ApoA-I and phospholipids in an ApoA-I weight:total phospholipid weight ratio of 1:2.7 + / - 20%, and sphingomyelin:DPPG in a weight:weight ratio of phospholipid sphingomyelin and DPPG of 97:3 + / - 20%.

[0605] 335. The method according to embodiment 334, wherein the formulation comprises ApoA-I and phospholipids, wherein the ApoA-I weight:total phospholipid weight ratio is 1:2.7 + / - 10%, and comprises phospholipid sphingomyelin and DPPG, wherein the sphingomyelin:DPPG weight:weight ratio is 97:3 + / - 10%.

[0606] The method according to embodiment 335, wherein the formulation comprises ApoA-I and phospholipids with a weight ratio of ApoA-I: total phospholipids of 1:2.7, and the phospholipid sphingomyelin and DPPG with a weight: weight ratio of sphingomyelin: DPPG of 97:3.

[0607] 337. The method according to any one of embodiments 334 to 336, wherein the ApoA-I has the amino acid sequence of amino acids 25-267 of SEQ ID NO:2.

[0608] 338. The method according to any one of embodiments 334 to 337, wherein the ApoA-I is recombinantly expressed.

[0609] 339. The method according to embodiment 338, wherein the ApoA-I is produced by mammalian host cells.

[0610] 340. The method according to embodiment 339, wherein the mammalian host cells are Chinese hamster ovary (CHO) cells.

[0611] 341. The method according to embodiment 340, wherein the CHO cells are CHO-S cells.

[0612] 342. The method according to any one of embodiments 339 to 341, wherein the ApoA-I has undergone post-translational processing (such as glycosylation) such that the ApoA-I has one or more structural features (such as glycosylation patterns) different from those of human ApoA-I purified from human plasma.

[0613] 343. The method according to any one of embodiments 179 to 342, wherein the formulation comprises natural sphingomyelin.

[0614] 344. The method according to embodiment 343, wherein the natural sphingomyelin is egg sphingomyelin.

[0615] 345. The method according to any one of embodiments 179 to 342, wherein the formulation comprises synthetic sphingomyelin.

[0616] 346. The method according to embodiment 345, wherein the synthetic sphingomyelin is palmitoyl sphingomyelin.

[0617] 347. The method according to any one of embodiments 179 to 346, wherein the formulation is at least 95% homogeneous.

[0618] 348. The method according to embodiment 347, wherein the formulation is at least 97% homogeneous.

[0619] 349. The method according to embodiment 347, wherein the formulation is at least 98% homogeneous.

[0620] 350. The method according to embodiment 347, wherein the formulation is at least 99% homogeneous.

[0621] 351. The method according to any one of embodiments 179 to 350, wherein the subject is a human.

[0622] 352. The method according to any one of embodiments 179 to 351, wherein the subject is not on mechanical ventilation at the time of the first administration of CER-001.

[0623] Although various specific embodiments have been illustrated and described, it should be understood that various changes can be made without departing from the spirit and scope of the present disclosure.

[0624] 9. Incorporation by reference

[0625] All publications, patents, patent applications, and other documents cited in this application are hereby incorporated by reference in their entirety for all purposes to the extent as if each individual publication, patent, patent application, or other document were specifically designated as being incorporated by reference for all purposes.

[0626] Any discussion in this specification regarding a document, act, material, device, article, etc. is provided only to give context to the present disclosure. It should not be considered that any or all of these constitute a part of the prior art base or an admission of common general knowledge in the relevant field of the present disclosure, as they existed prior to the priority date of this application.

Claims

1. A method of treating a subject suffering from or at risk of suffering from an excessive inflammatory disorder, the method comprising administering to the subject a dose of a lipocalin-based complex, optionally wherein the excessive inflammatory disorder is hemophagocytic lymphohistiocytosis (HLH), dengue hemorrhagic fever or dengue shock syndrome.

2. The method of claim 1, wherein the subject suffers from HLH secondary to a non-malignant disorder or is at risk of suffering from HLH secondary to a non-malignant disorder.

3. The method of claim 2, wherein the non-malignant disorder is a viral infection.

4. The method of claim 3, wherein the subject suffers from HLH caused by dengue infection or is at risk of suffering from HLH caused by dengue infection.

5. The method of any one of claims 1 to 4, wherein the subject suffers from dengue or dengue hemorrhagic fever.

6. The method of any one of claims 1 to 4, wherein the subject suffers from dengue shock syndrome.

7. The method of claim 3, wherein the subject suffers from HLH caused by herpes simplex infection or is at risk of suffering from HLH caused by herpes simplex infection.

8. The method of claim 3, wherein the subject suffers from HLH caused by Epstein-Barr virus infection or is at risk of suffering from HLH caused by Epstein-Barr virus infection.

9. The method of claim 2, wherein the non-malignant disorder is an autoimmune disease.

10. The method of claim 1, wherein the subject suffers from HLH secondary to a malignant disorder or is at risk of suffering from HLH secondary to a malignant disorder.

11. The method of claim 10, wherein the malignant disorder is leukemia or lymphoma.

12. The method of claim 1, wherein the subject suffers from familial HLH or is at risk of suffering from familial HLH.

13. The method of any one of claims 1 to 12, wherein the subject suffers from HLH.

14. The method of any one of claims 1 to 12, wherein the subject is at risk of suffering from HLH.

15. The method of any one of claims 1 to 14, wherein the dose is a high dose.

16. The method of claim 15, wherein the high dose is administered over a period of one day to about two weeks, optionally wherein the high dose is administered over a period of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14 or 15 days.

17. The method of claim 15 or claim 16, wherein the high dose is the sum of 2 to 10 individual doses, optionally wherein the high dose is the sum of 3, 4, 5, 6, 7, 8, 9 or 10 individual doses.

18. The method of claim 17, wherein multiple individual doses are administered daily or multiple individual doses are administered twice daily.

19. The method of claim 17 or claim 18, wherein multiple individual doses are administered at intervals of two to three days.

20. The method of claim 17, wherein multiple individual doses are administered at intervals of no more than one day.

21. The method according to claim 20, which comprises administering two or more separate doses at intervals of about 12 hours.

22. The method according to claim 21, which comprises administering two separate doses at intervals of about 12 hours.

23. The method according to claim 21, which comprises administering three separate doses at intervals of about 12 hours.

24. The method according to claim 22 or claim 23, which further comprises administering a separate dose after about one day.

25. The method according to claim 17, which comprises administering three separate doses at intervals of about 12 hours and administering a fourth separate dose after about one day.

26. The method according to claim 15, wherein the high dose is administered as a single separate dose.

27. The method according to claim 15, wherein the high dose is the sum of two separate doses administered within one day.

28. The method according to claim 27, wherein the two separate doses are administered at intervals of about 12 hours.

29. The method according to any one of claims 15 to 28, wherein each separate dose effectively increases the HDL level of the subject, optionally wherein the high dose effectively increases the serum HDL level of the subject to normal.

30. The method according to any one of claims 15 to 29, wherein each separate dose effectively increases the ApoA-I level of the subject, optionally wherein the high dose effectively increases the serum ApoA-I level of the subject to normal.

31. The method according to any one of claims 1 to 30, wherein the lipid-binding protein-based complex is reconstituted HDL or an HDL mimetic.

32. The method according to any one of claims 1 to 30, wherein the lipid-binding protein-based complex is an Apomer or a Cargomer.

33. The method according to any one of claims 1 to 33, wherein the lipid-binding protein-based complex comprises sphingomyelin.

34. The method according to any one of claims 1 to 33, wherein the lipid-binding protein-based complex comprises negatively charged lipids.

35. The method according to claim 34, wherein the negatively charged lipid is 1,2-dipalmitoyl-sn-glycero-3-[phospho-rac-(1-glycerol)] (DPPG) or a salt thereof.

36. The method according to any one of claims 1 to 30, wherein the lipid-binding protein-based complex is CER-001, CSL-111, CSL-112, CER-522 or ETC-216.

37. The method according to claim 36, wherein the lipid-binding protein-based complex is CER-001.

38. The method according to claim 37, wherein the CER-001 is a lipoprotein complex comprising ApoA-I and phospholipids with a weight ratio of ApoA-I:total phospholipids of 1:2.7, and a weight ratio of sphingomyelin:DPPG of 97:3 for the phospholipid sphingomyelin and DPPG.

39. The method according to claim 37 or claim 38, wherein the ApoA-I has the amino acid sequence of amino acids 25 - 267 of SEQ ID NO:

2.

40. The method according to any one of claims 37 to 39, wherein the ApoA-I is recombinantly expressed.

41. The method according to claim 40, wherein the ApoA-I is produced by a mammalian host cell.

42. The method according to claim 41, wherein the mammalian host cell is a Chinese hamster ovary (CHO) cell.

43. The method according to claim 42, wherein the CHO cell is a CHO-S cell.

44. The method according to any one of claims 41 to 43, wherein the ApoA-I has undergone post-translational processing (e.g., glycosylation) such that the ApoA-I has one or more structural features (e.g., glycosylation pattern) different from that of human ApoA-I purified from human plasma.

45. The method according to claim 36, wherein the lipocalin-based complex is CSL-112.

46. A method of treating a subject suffering from or at risk of developing an excessive inflammatory disorder, comprising administering to the subject a dose of an apolipoprotein AI ("ApoA-I") preparation, the apolipoprotein AI ("ApoA-I") preparation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex, optionally wherein the excessive inflammatory disorder is hemophagocytic lymphohistiocytosis (HLH), dengue hemorrhagic fever or dengue shock syndrome.

47. A method of treating a subject suffering from or at risk of developing hemophagocytic lymphohistiocytosis (HLH), comprising administering to the subject a dose of an apolipoprotein AI ("ApoA-I") preparation, the apolipoprotein AI ("ApoA-I") preparation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

48. A method of treating a subject suffering from a dengue infection, comprising administering to the subject a dose of an apolipoprotein A-I ("ApoA-I") preparation, the apolipoprotein A-I ("ApoA-I") preparation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex.

49. A method of treating a subject suffering from a herpes simplex infection, comprising administering to the subject a dose of an apolipoprotein A-I ("ApoA-I") preparation, the apolipoprotein A-I ("ApoA-I") preparation comprising ApoA-I and one or more lipids, wherein the ApoA-I and the lipids are in the form of a lipoprotein complex. Method for treating a subject having Epstein-Barr infection, comprising administering to the subject a dose of an apolipoprotein A-I ("ApoA-I") preparation, said apolipoprotein A-I ("ApoA-I") preparation comprising ApoA-I and one or more lipids, wherein said ApoA-I and said lipids are in the form of a lipoprotein complex.

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